Horticultural Plant Biotechnology

Uttam Kesri

HorticulturalPlant Biotechnology


Uttam Kesri

Published by Vidya Books,

305, Ajit Bhawan,

21 Ansari Road,

Daryaganj, Delhi 110002

Uttam Kesri

ISBN: 978-93-5429-538-6

© 2021 Vidya Books

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Contents

Chapter 1 Introduction ............................................................................ 1

Chapter 2 Trends in Horticulture Growth ........................................... 26

Chapter 3 Genetic Resources in Horticulture ...................................... 39

Chapter 4 Horticultural Biotechnology in Tissue Culture .................................................................................. 63

Chapter 5 Fruit Crop Cultivation ....................................................... 109

Chapter 6 Commercial Horticulture................................................... 133

Chapter 7 Greenhouse Crops System in Horticulture ...................... 141

Chapter 8 Biotechnology and Genetic Transformation .................... 191

Chapter 9 Plant Breeding .................................................................... 210

Chapter 10 Forestry Horticulture System ............................................ 234

1

Introduction

Horticulture is classically defined as the culture or growing of gardenplants. Horticulturists work in plant propagation, crop production, plantbreeding and genetic engineering, plant biochemistry, plant physiology, andthe storage, processing, and transportation of fruits, berries, nuts, vegetables,flowers, trees, shrubs, and turf. They improve crop yield, quality, nutritionalvalue, and resistance to insects, diseases, and environmental stresses. Geneticsis also used as a valuable tool in the development of plants that can synthesizechemicals for fighting disease (including cancers).

THE STUDY OF HORTICULTURE

Horticulture involves five areas of study. These areas are floriculture(includes production and marketing of floral crops), landscape horticulture(includes production, marketing and maintenance of landscape plants),olericulture (includes production and marketing of vegetables), pomology(includes production and marketing of fruits), and postharvest physiology(involves maintaining quality and preventing spoilage of horticultural crops).

Horticulturists can work in industry, government, or educationalinstitutions. They can be cropping systems engineers, wholesale or retailbusiness managers, propagators and tissue culture specialists (fruits,vegetables, ornamentals, and turf), crop inspectors, crop production advisors,extension specialists, plant breeders, research scientists, and of course,teachers.

College courses that complement Horticulture are biology, botany,entomology, chemistry, mathematics, genetics, physiology, statistics,computer science, and communications, garden design, planting design. Plantscience and horticulture courses include: plant materials, plant propagation,tissue culture, crop production, post-harvest handling, plant breeding,pollination management, crop nutrition, entomology, plant pathology,economics, and business. Some careers in horticultural science require amasters (MS) or doctoral (PhD) degree. Horticulture takes place in many

gardens and plant growth centres. Plants are often grown as seedlings withinplant nurseries. Activities in nurseries range from preparing seeds and cuttingsto growing fully mature plants. These are often sold or transferred toornamental gardens or market gardens.

INTRODUCTION TO HORTICULTURAL PLANTS

Horticulture is a broad field of study encompassing the production,maintenance, sales, and service of bedding, flowering, ornamental, andnursery plants and food crops. This major has four distinct concentrations.Floriculture Crop Production stresses greenhouse production. GeneralHorticulture allows students to design their own focus of study. Retail FloralDesign emphasizes the principles and practices which prepare students forthe successful operation of a retail floral business by concentrating on design,handling of plants, and business practices. Management of Woody Plantscovers plant production and management in a nursery and the operation of aretail garden centre.

Increasingly precise management of horticultural crops lends itself to theuse of mechanistic crop simulation models to predict growth and yield. In thischapter, we encourage systematic approaches for developing, parameterizing,and testing of dynamic crop growth models for horticultural crops. Using theCropgro model as an example, we describe crop development, photosynthesis,leaf area expansion, as well as the addition, growth, and maturation ofindividual fruits. An algorithm is described for predicting fruit fresh weight,size, and dry matter concentration from dry matter growth and physiologicalage of individual tomato (Lycopersicon esculentum Mill.) fruits. Parameterizingcrop models for response to climatic factors, especially temperature, requiresinterpretive use of literature information, along with testing against growth datafrom a range of growth conditions. A review of tomato literature for temperaturedependencies of processes is presented with a view to how this informationwould be used to parameterize the CROPGRO-tomato model.

In modern horticulture, growers are searching for reliable biosensors anddiagnostic tools to evaluate plant eco-physiological behaviour in order tooptimize plant growth and crop development. One of these biosensors is thelinear variable displacement transducer (LVDT) which accurately measuresstem diameter variations. The variations of stem diameter reflect thecombination of two main phenomena: irreversible radial stem growth anddaily shrinkage and swelling of the elastic stem cells due to changes ininternally stored water. A correct interpretation of the composite LVDT-signal,with respect to its practical application, requires an unambiguous distinctionbetween these two components of the signal. This chapter describes acomprehensive model for stem diameter variations which includes amechanistic description of radial stem growth and elastic stem diameter

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changes related to changes in internally stored water. Using the results of atwo-year-old beech tree (Fagus sylvatica), the practical application of themodel is illustrated with measured LVDT-signals where radial stem growthand elastic changes of the stem could be distinguished. As this approach isalso applicable to other ornamental or vegetable crops, the model can beconsidered as a very powerful tool for interpretation of the composite andcomplex origin of the variations in stem diameter as is observed with LVDTs.

APPLICATIONS OF BIOTECHNOLOGY IN HORTICULTURE

The requirement of fruits and vegetables is increasing proportionally withthe increasing population in the country. How do we keep horticulturalproduction on par with the burgeoning population? Although conventionalplant breeding techniques have made considerable progress in thedevelopment of improved varieties, they have not been able to keep pace withthe increasing demand for vegetables and fruits in the developing countries.Therefore an immediate need is felt to integrate biotechnology to speed upthe crop improvement programmes. Biotechnological tools haverevolutionized the entire crop improvement programmes by providing newstrains of plants, supply of planting material, more efficient and selectivepesticides and improved fertilizers. Many genetically modified fruits andvegetables are already in the market in developed countries. Modernbiotechnology encompasses broad areas of biology from utilization of livingorganisms or substances from those organisms to make or to modify a product,to improve plant or animal or to develop micro-organisms for specific use. Itis a new aspect of biological and agricultural science which provides new toolsand strategies in the struggle against world’s food production problem. Themajor areas of biotechnology which can be adopted for improvement ofhorticultural crops are.

T ISSUE CULT URE

One of the widest applications of biotechnology has been in the area oftissue culture and micro propagation in particular. It is one of the most widelyused techniques for rapid asexual in vitro propagation. This technique iseconomical in time and space affords greater output and provides diseasefree and elite propagules. It also facilitates safer and quarantined movementsof germplasm across nations. When the traditional methods are unable tomeet the demand for propagation material this technique can produce millionsof uniformly flowering and yielding plants. Micropropagation of almost allthe fruit crops and vegetables is possible now. Production of virus freeplanting material using meristem culture has been made possible in manyhorticultural crops. Embryo rescue is another area where plant breeders areable to rescue their crosses which would otherwise abort. Culture of excised

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embryos of suitable stages of development can circumvent problemsencountered in post zygotic incompatibility. This technique is highlysignificant in intractable and long duration horticultural species. Many ofthe dry land legume species have been successfully regenerated fromcotyledons, hypocotyls, leaf, ovary, protoplast, petiole root, anthers, etc.,Haploid generation through anther/pollen culture is recognized as anotherimportant area in crop improvement. It is useful in being rapid andeconomically feasible. Complete homozygosity of the offspring helps inphenotype selection for quantitative characters and particularly forqualitatively inherited characters making breeding much easier successfulisolation, culture and fusion of plant protoplasts has been very useful intransferring cytoplasmic male sterility for obtaining hybrid vigour throughmitochondrial recombination and for genetic transformation in plants.

In vitro germplasm conservation is of great significance in providingsolutions and alternative approaches to overcoming constrains in managementof genetic resources. In crops which are propagated vegetatively and whichproduce recalcitrant seeds and perennial crops which are highly heterozygousseed storage is not suitable. In such crops especially, in vitro storage is ofgreat practical importance. These techniques have successfully beendemonstrated in a number of horticultural crops and there are now variousgermplasm collection centers. In vitro germplasm also assures the exchangeof pest and disease free material and helps in better quarantine.

Plant breeders are continually searching for new genetic variability thatis potentially useful in cultivar improvement. A portion of plants regeneratedby tissue culture often exhibits phenotypic variation atypical of the originalphenotype. Such variation, termed somaclonal variation may be heritablei.e. genetically stable and passed on to the next generation. Alternatively, thevariation may be epigenetic and disappear following sexual reproduction.These heritable variation are potentially useful to plant breeders.

GENET IC ENGINEERING OF PLANT S

Using techniques of genetic engineering many useful genes have beenintroduced into plants and many transgenic plants have been developed inwhich the foreign DNA has been stably integrated and resulted in the synthesisof appropriate gene product. Transgenic plants have covered about 52.6 mhectares in the Industrial and developing countries upto 2001. Genes for thefollowing traits have been introduced to the crop plants.

Herbicide tolerance: Transgenic plants are developed that are resistantto herbicides allowing farmers to spray crops so as to kill only weeds but nottheir crops. Many herbicide tolerant plants have been developed in tomato,tobacco, potato, soybean, cotton, corn oilseed rape, petunia, etc. Glyphosateis one of the most potent broad spectrum environment friendly herbicideknown, it is marketed under the trade name Round up. Glyphosate kills plants

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by blocking the action of an enzyme (5-enolpyruvyl shikimate-3-phosphatesynthase) (EPSPS) an essential enzyme in the biosynthesis of aromatic aminoacids, tyrosine, phenylalanine and tryptophan. Amino acids are buildingblocks of protein.

Transgenic plants resistant to Glyphosate have been developed bytransferring gene of EPSPS that over prodoce this enzyme thus inhibiting theeffect of Glyphosate. A number of detoxifying enzymes have been identifiedin plants as well as in microbes. Some of these include glutahthione-s-transferase or GST in maize and other plants which detoxifies theherbicidebromoxynil and phosphinothricin acetyl transferase (PAT) whichdetoxifies the herbiside PPT (L-phosphinothricine). Transgenic plants usingbxn gene from Klebsiella and bar gene from Strepotomyces have been obtainedin potato, oilseed, sugarbeet, soybean, cotton and corn and are found to beherbicide resistance. These transgenic plants reduce the use of weeding labour,farmers cost and increase yield.

Engineering pathogen resistance: Viruses are the major pests of cropplants which cause considerable yield losses. Many strategies have beenapplied to control virus infection using coat protein and satellite RNA. Virusesare submicroscopic pockets of nucleic acid (DNA or RNA) enclosed in aprotein coat and can multiply within a host cell. Use of viral coat protein as atransgene for producing virus resistant plants is one of the most spectacularsuccesses achieved in plant biotechnology. Coat protein gene from tobaccomosaic virus (TMV) classified as a positive strand RNA virus has beentransferred to tobacco, making it nearly resistant against TMV. Using genefor nucelocapsid protein resistance has been introduced in crops like tomato,tobacco, lettuce, groundnut, pepper and in ornam ents like Impatiens, Ageratumand Crysnathemum against tomato spotted wilt virus. Use of satellite RNA(SATRNA) makes many transgenic plants resistant toCucumber Mosaic Virus(CMV). Transgenic resistant plants have also been developed against alfalfamosaic virus, potato virus X, Rice tungro virus, tobacco rattle virus and Papayaring spot virus.

During the last decade many resistance genes whose products areinvolved in recognizing the invading pathogens have been identified andcloned. A number of signaling pathways which follow the pathogen infectionhave been dissected. Many of the antifungal compounds synthesized by plantswhich combat fungal infections have been identified. The major strategiesfor developing fungal resistance have been production of transgenic plantswith antifungal molecules like proteins and toxins, and generation ofhypersensitive response through R genes or by manipulating genes of SARpathway. A ch itinase gene from bean plants in tobacco and Brassica napusshowed enhanced resistance to Rhizoctonia solani. In another case chitinasegene obtained from Serratia marcescens(soil bacterium) is introduced in tobaccomaking it resistant to Alternaria longipes which causes brown spot diseases.

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Acetyl transferase gene is introduced in tobacco making it resistanttoPseudomonas syringea, a causal agent of wild fire disease.

Stress resistance : A number of genes responsible for providing resistanceagainst stresses such as to water stress heat, cold, salt, heavy metals andphytohormones have been identified. Studies are also being conducted onmetabolites like proteins and betains that have been implicated in stresstolerance. Resistance against chilling was introduced into tobacco plants byintroducing gene for glycerol-1-phosphate acyl-transferase enzyme fromArabidopsis. Many plants respond to drought stress by synthesizing a groupof sugar derivatives called polyols (Mannitol, Sorbitol and Sion) . Plants thathave more polyols are more resistant to stress. Using a bacterial gene capableof synthesizing mannitols it is possible to raise the level of mannitol very highmaking plants resistant to drought.

Fruit Quality: Tomatoes which ripen slowly are helpful in transportationprocess. Transgenic tomato with reduced pectin methyl esterase activity andincreased level of soluble solids and higher pH increases processing quality.Tomatoes exhibiting delayed ripening have been produced either by usingantisense RNA against enzymes involved in ethylene production (Eg ACCsynthase) or by using gene for deaminase which degraded l-aminocyclopropane-l-carboxylic acid (ACC) an immediate precursor ofethylene. This increases the shelf life of tomatoes. These tomatoes can alsostay on the plant long giving more time for accumulation of sugars and acidsfor improving flavour. It is produced at commercial level in European andAmerican countries. Tomatoes with elevated sucrose and reduced starch couldalso be produced using sucrose phosphate synthase gene. Starch content inpotatoes has been increased by 20-40% by using a bacterial ADP glucosepyrophosphorylase gene.

Pest resistance : The insecticidal beta endotoxin gene (bt gene) has beenisolated from Bacillus thuringiensis the commonly occurring soil bacteria andtransferred to number of plants like cotton, tobacco, tomato, soybean, potato,etc. to make them resistant to attack by insects. These genes produceinsecticidal crystal proteins which affect a range of lepidopteran, coleopteran, dipteran insects. These crystals upon ingestion by the insect larva aresolubilised in the highly alkaline midgut into individual protoxins which varyfrom 133 to 136 kDa in molecular weight. Insecticidal crystal proteinproduced during vegetative growth of the cells (VIP)are also found to behighly effective against insect control. Bt resistant plants are already in themarket.

Male sterility and Fertility restoration: This is helpful in hybrid seedproduction. Transgenic plants with male sterility and fertility restorationgenes have become available in Brassica napus. It facilitates production ofhybrid seed without manual emasculation and controlled pollination as oftendone in maize. In 1990, Mariani and others from Belgium have successfully

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used a gene construct having another specific promoter from TA29 gene oftobacco and bacterial coding sequence for a ribonuclease gene from BacillusSp. (barnase gene) for production of transgenic plants in Brassica napus. Herethe translated gene prevented normal pollen development leading to malesterilily.

M OLECULAR DIAGNOST ICS

Nucleic acid probes:- It is now possible to detect the plant diseases evenbefore onset of symptoms by using cDNA probes. Probes are nucleic acidsequences of pathogen causing organisms labeled with certain markers. cDNAprobes corresponding to specific regions of the pathogens can be generatedusing standard recombinant DNA technique.

Monoclonal antibodies (McAb): Immunochemical techniques areextremely useful for the rapid and accurate routine detection of plantpathogens and ultimately the diagnosis of plant disease and their relatedness,The introduction of hybridoma technology has provided methods for theproduction of homologous and biochemically defined immunological reagentsof identical specificity which are produced by a single cell line and are directedagainst a unique epitope of the immunizing antigen. The great potential ofMcAbs in phytopathological diagnostics is essential because of homogeneousantibody preparations with defined activity and specificity can be producedin large quantities over long periods. Even though hybridoma technology isa laborious and expensive enterprise compared to standard immunizationprocedures it is going to be widely used for large scale diagnosis.

M OLECULAR M ARKERS

The possibilities of using gene tags of molecular makers for selectingagronomic traits has made the job of breeder easier. It has been possible toscore the plants for different traits or disease resistance at the seedling stageitself. The use of RFLP (Restriction Fragment Length polymorphism), RAPD(Random Amplified Polymorphic DNA) , AFLP (Amplified Fragment LengthPolymorphism) and isozyme markers in plant breeding are numerous. RFLPsare advantageous over morphological and isozyme markers primarily becausetheir number is limited only by genome size and they are not environmentallyor developmentally influenced. Molecular maps now exist for a number ofcrop plants including corn, tomato, potato, rice, lettuce, wheat, Brassica speciesand barley. RFLPs have wide ranging applications including cultivar fingerprinting, identification of quantitative trait loci, analysis of genomeorganization, germplasm introgression and map-based cloning. AFLP isbecoming the tool of choice for fingerprinting because of its reproducibilitycompared to RAPD. Microsatellile or simple sequence repeats (SSRS) markershave also become the choice for a wide range of applications in genotyping,genome mapping and genome analysis.

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DEVELOPM ENT OF M ICROBIAL INOCULANT S

Indiscriminate and injudicious use of chemical fertilizers and pesticidesfor the crop production and control of insect-pests has resulted in pollutionof the environment deterioration of soil health and development of resistanceby many insects and residue problems. Hence there is a great concern worldwide to use safer biofertilisers and biopesticdies in the integrated nutrientmanagement and pest management systems.

Biofertilizers are micro-organisms which fix atmospheric nitrogen orsolubilise fixed phosphorus in the soil and make more nutrients available tothe plant. Some of the organisms providing major inputs are the biologicalnitrogen fixing organisms like Rhizobium, Azotobacter, Azospirillum andph osphate so lubilising organism s lik e Bacillus polymyxa, B. magaterium,Pseudomonas striata and certain fungal species of Aspergillus and Penicillium.

The benefits of using micro-organisms as fertilizers are many fold. Theyare less expensive, nontoxic to plants, do not pollute the ground water norrender the soil acidic and unfit for growth of plants. Rhizobium forms noduleson the roots of leguminous plants and help in fixing nitrogen from theatmosphere to ammonium irons which get converted to amino acids in theplant system. Inoculation with this bacteria helps in reducing addition ofnitrogenous fertilizers to the soil. Azospirillum is also found colonizing intercellular spaces inside the root system. These bacteria also contributesubstantially to the nitrogen requirement of the plant.

Phosphate solubilising bacteria are another group of micro-organismswhich solubilise the insoluble phosphorus in the soil and make them readilyavailable to the crop.

Mycorrhiza is the symbiotic association of the roots of crop plants withnon-pathogenic fungus. They provide nutrients absorbed from deeper layersof soil to the plants. They help the plants in better plant establishment andgrowth when inoculated. Many fruit crops like papaya, mango, banana, citrus,pomegranate are found to be dependent on this association and are greatlybenefited by its inoculation in procuring higher phosphate and other nutrientfrom the soil. These mycorrhizal associations help the plants in overcomingpathogen attack also. They improve soil characters too.

Genetic modification of microbes: By using DNA recombinationtechnique it has been possible to genetically manipulate different strains ofthese bacteria suitable to different environmental conditions and to developstrains with traits with capacity for better competitiveness and nodulation.Biopesticides are biological organisms which can be formulated as that of thepesticides for the control of pests.

Biopesticides are gaining importance in agriculture, horticulture and inpublic heatlh programmes for the control of pests. The advantages of usingbiopesticides are many. They are specific to target pests and do not harm thenon target organisms such as bees, butterflies and are safe to humans and

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live stocks, they do not disturb the food-chain nor leave behind toxic residues.Som e of the m icrobial pesticid es used to control insect pests are Bacillusthuringiensis species to control various insect pests. Insecticidal property ofthese bacteria are due to crystals of insecticidal proteins produced duringsporulation. These proteins are stomach poisons and are highly insect specific.Bt toxins could kill plant parasitic nematode too. Number of baculoviruses(BV) nuclear polyhedrosis virus (NPV) is being developed as microbialpesticides both nationally and internationally, A few examples of these areHeliothis, Spodoptera, Plusia, Agrotis, Trichoplusia, etc.

Biocontrol agents : These are other microbes which are antagonistic toseveral pathogenic fungus and are good substitutes to fungicides or insecticide.These are Bacillus sps. Pseudomonas fluorescens, Trichoderma, Verticillium sp.,Streptromyces sps. etc. These organisms are commercially available.

TROPICAL AND SUB-TROPICAL FRUITS

BANANA

Banana (Musa paradisiaca L.) occupies over 1,64,000 hectares, mainly inTamil Nadu, West Bengal, Kerala, Maharashtra, Gujarat, Karnataka, Assam,Andhra Pradesh and Bihar. Though some inferior types of banana are foundgrowing as far north as the Himalayas, its commercial importance is mainlylimited to the more tropical conditions, such as those prevailing in central,southern and north-eastern India. It is a moisture- and heat-loving plant andcannot tolerate frost or arid conditions.

Varieties

Cultivated varieties are broadly divided into two groups: table andculinary. Among the former are ‘Poovan’ in Madras (also known as ‘KarpuraChakkarekeli’ in Andhra Pradesh); ‘Mortaman’, ‘Champa’ and ‘Amrit Sagar’in West Bengal; ‘Basrai’, Safed Velchi’, Lal Velchi’ and ‘Rajeli’ in Maharashtra;‘Champa’ and ‘Mortaman’ in Assam and Orissa; and ‘Rastali’, ‘Sirumalai’,‘Chakkarekeli’, ‘Ney Poovan’, ‘Kadali’ and ‘Pacha Nadan’ in southern India.‘Basrai’, which is known under different names, viz. ‘Mauritius’, ‘Vamankeli’,‘Cavendish’, ‘Governor’, ‘Harichal’, is also grown in central and southernIndia. Recently, the ‘Robusta’ variety is gaining popularity in Tamil Naduand Karnataka.

The ‘Virupakshi’ variety (Hill banana) is the most predominant varietyin the Palni Hills of Tamil Nadu. Among the culinary varieties, Nendranbananas, ‘Monthan’, ‘Myndoli’ and ‘Pacha Montha Bathis’ are the leadingcommercial varieties in southern India. ‘Gros Michel’ is a recent introductioninto southern India; it is suitable for cultivation only under garden-landconditions and is generally fastidious in its cultural requirements. It is not,therefore, in favour with the cultivation.

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Propagation and Planting

Propogation is by suckers or off-shoots which spring at the base of abanana-tree from underground rhizomes. Vigorous suckers, with stout base,tapering towards the top and possessing narrow leaves, are selected for plant.Each sucker should have a piece of underground stem with a few rootsattached to it.

Banana suckers can be planted throughout the year in southern India,except during summer, whereas in the rest of the country, the rainy season ispreferred. They are planted in small pits, each just enough to accommodatethe base of a sucker. The planting-distance varies from 2m×2m in the case ofdwarf varieties to 4m×4m in the case of very tall varieties.

Manuring

An application of 20 to 25 kg of farmyard manure, together with about 5kg of wood-ashes per plant is given at planting time. In southern India,ammonium sulphate is applied one month, five months and nine months afterplanting 20 kg per hectare each time.

In western India, a little over 2 kg of oilcake per stool is applied duringthe first three months after planting.

A complete fertilizer mixture may be applied to supply 100 to 200 kg ofN, 100 to 200 kg of P2O5 and 200 to 400 kg of K2O per hectare.

A green-manure crop is also considered beneficial. Trials at the IndianInstitute of Horticultural Research have shown that for the ‘Robusta’ variety,a fertilizer mixture comprising 180 g of N + 108 g of P2O5 + 225 g of K2O perplant is ideal.

After-Care

The removal of suckers, dry leaves and pseudostems, from which thefruits have been harvested, constitute the main after-care. Daughter-suckersshould be removed promptly until the mother-plant flowers, when onedaughter-sucker may be allowed to take its place. The removal of dry leavesand useless pseudostems requires to be done in time. After all the fruits areformed, the pendant portion of the remaining inflorescence along with theheart should be removed.

The propping of plants with bamboo poles, especially those which havethrown out bunches, is necessary wherever damage by wind is apprehended.Where the wind damage is recurring, dwarf varieties should be preferred.

Irrigation

The banana-plants require very heavy irrigation. Irrigation is given inmost places once in seven to ten days. Stagnation of water in the soils is notvery congenial to the proper growth of banana and, hence, the drainage ofsoil is also essential.

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Harvesting

Early varieties commence flowering in southern and western India aboutseven months after planting, and the fruits take about three months more toripen. In the Andhra Pradesh delta areas, the fruits are ready for harvestingabout seven to eight months after planting. The first crop of the ‘Poovan’variety matures in 12 to 14 months and the second in 21 to 24 months afterplanting. In other parts of India, the first crop is usually gathered a year afterplanting, whereas the succeeding crop may be ready in six to ten monthsthereafter.

The bunch is harvested just before it attains the ripening stage. When thefruits have reached the full size, they become plump, and mature with adistinct change in colour. For long transport, the bunch may be harvestedsomewhat earlier. The bunch is cut, retaining about 15 cm of the stem abovethe first hand. The yield varies considerably from 26,000 to 55,000 kg perhectare.

Curing and Marketing

The ripening of banana is done in several ways, e.g. exposing the bunchesto the sun, placing them over a hearth, wrapping them in closed godowns orsmoking them in various ways. One of the common ways is to heap the fruitsin a room and cover them with leaves, after which fire is lit in a corner and theroom is closed and made as air-tight as possible. Ripening takes place usuallyin 30 to 48 hours. In a cool store, the bunches ripen well at about 15o to 20oC.The application of vaseline, a layer of clay or coal-tar to the cut-ends of thestalks prevents rotting during ripening and storage.

Wrapping up the fruits and packing them in crates help to reduce thedamage during transport.

M ANGO

Mango (Mangifera indica L.) occupies nearly half of the total area underfruits in the country. It is adaptable to a wide range of soil and climaticconditions and grows well right from Assam to the southern-most limits ofthe country and from the sea-level up to about 1,500 metres.

Fig. Mango

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It withstands both fairly dry conditions and heavy rainfall, providedsevere and recurring frosts in winter do not endanger the young trees.

Varieties

The number of varieties is very large. Each variety has its own peculiartaste, flavour and consistently of pulp. Some of the important commercialvarieties grown in different regions are: ‘Bombay yellow’, ‘Alphonso’, ‘GopalBhog’, ‘Zafran’ (all early), ‘Langra’, ‘Desheri’, ‘Safeda Lucknow’, ‘SafedaMalihabad’, ‘Fajrizafrani’ (all mid-late). ‘Fajri’, ‘Same Bihisht’, ‘Chausa’,‘Taimura’ (all late). In Uttar Pradesh; ‘Bombai’, ‘Alphonso’, ‘Hemsagar’,‘Krishna Bhog’, ‘Aman Dasheri’, ‘Gulab Khas’ (all early), ‘Langra’, ‘AmanAbbasi’, ‘Khasul-Khas’ (all mid-late), ‘ Sinduri’, ‘Sukal’, ‘Taimuria’ (all late)in Bihar; ‘Bombai’ or ‘Maldah’, ‘Gopal Bhog’, ‘Hemsagar’ (all early), ‘KrishnaBhog’, ‘Zardalu’ (both mid-late), ‘Murshidabadi’, ‘Fazli Maldah’ (both late)in West Bengal; ‘Alphonso’, ‘Pairi’, ‘Cowsji Patel’, ‘Jamadar’ in Bombay;‘Swarnarekha’, ‘Benishan’, ‘Cherukurasan’, ‘Panchadarkalasa’,‘Desavathiyamamidi’, ‘Sannakulu’, ‘Nagulapalli’, ‘Irsala’ in Circars; ‘Rumani’,‘Neelum Benishan’, ‘Bangalore’, ‘Alampur Benishan’ in Rayalaseema;‘Murshidabadi’, ‘Mulgoa’, ‘Goabunder’, ‘Benishan’, ‘Neelam’, ‘Totapuri’ or‘Bangalora’ in Telengana; ‘Alphonso’, ‘Peter’, ‘Rumani’ in central districts;‘Mundappa’, ‘Neelam’, ‘Alphonso’, ‘Olour’, ‘Bennet Alphonso’, ‘Kalepad’,‘Peter’, ‘Fernandin’ in Coorg and Karnataka; and ‘Padiri’, ‘Alphonso’, ‘Peter’,‘Neelum’, ‘Bangalore’, ‘Rumani’ in Tamil Nadu. In Goa, some excellentvarieties like ‘Alphonso’, ‘Fernandin’, ‘Mankurad’ and ‘Moussorate’ are undercultivation. The new mango variety, ‘Mallika’ evolved at the IndianAgricultural Research Institute is now gaining popularity.

Other varieties, such as ‘Jehangir’ and ‘Himayuddin’, produce high-quality fruits, but are poor in yield and cropping tendencies. Attempts arebeing made to evolve hybrid progenies by crossing.

Propagation and Planting

Propogated vegetatively by inarching or budding in situ in the nursery,either by using Forkert or by using the T-method. The beginning of themonsoon in light-rainfall areas and the end of the monsoon in heavy-rainfallregions are the most suitable periods for inarching or budding. Recently,veneer-grafting has been found to be the best method of mango propagation.Grafted plant are ready for transplanting in the field after six to twelve months.Select straight-growing grafts and set them in pits filled with soil mixed withfarmyard manure (45 kg) and a fertilizer mixture containing 0.225 kg of N,0.45 kg of P and 0.225 kg of K per pit. The planting-distance is 7.5 to 9 metresin poor shallow soils and 15 to 17 metres in deep fertile soils. The beginningof the monsoon in low rainfall areas or the end of the monsoon in heavy rainfalltracts is the best time for planting. The graft-joint should be at least 15 cmabove the ground.

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Pruning

No systematic pruning is done. The removal of dead-wood and thethinning of over-crowded and mis-shapen branches after about four years isall that is necessary; flowers that appear during the first three or four yearsshould be removed.

Culture

Before planting, the field is ploughed, harrowed and levelled. Thereafter,it is ploughed and harrowed twice a year, once in the beginning of themonsoon and again at the close of the rainy season or in the cold-weather. Itis green-manured once every two or three years. Short-season intercrops, likevegetables, may be taken during the first four to five years. Young plantsrequire irrigation regularly. After five to six years, when they have establishedthemselves, the trees are able to grow and fruit satisfactorily without irrigationin most parts of Peninsular India. In northern India, they have to be irrigatedthroughout their life. Irrigation is usually withheld during the cold weatherbefore flowering, especially in deep retentive soils. Though the exact manurialrequirement is not known, regular manuring is beneficial. The doserecommended for the bearing trees is 45 to 70 kg of farmyard manure, 0.5 to0.7 kg of N, 0.7 kg to 1.0 kg of P and 1.2 to 1.5 kg of K per tree. Nitrogen andhalf of potash may be given before the monsoon, and farmyard manure,phosphate and half of potash in October or before flowering starts.

Crop Irregularity

Grafted mango-trees bear fruits from the fourth or fifth year onwardsand a full crop from the tenth or fifteenth year. The erratic bearing of mangois well known. It depends upon the variety, the weather and climaticconditions and cultural treatments. The selection of regular-bearing varieties,timely cultural practices and proper nutrition help to produce a regular crop.New growth in spring, on which flower-buds are produced during the nextwinter, can be encouraged by applying nitrogenous fertilizers (0.45 to 0.90 kgof N per tree). In the case of heavy late rains, an additional ploughing in winterhelps to produce flower-buds in January-February. In the case of individualtrees, ringing or girdling in August-September may also to help to force flower-buds the following winter. The application of Ethral (200 ppm) from Septemberonwards has been found to induce flowering in mango in Karnataka by theIndian Institute of Horticultural Research.

Improvement of Old and Seedling-Trees

Mango-trees of inferior varieties, so also those raised from seeddlings,can be converted into choice varieties by grafting them in situ either by crownor side-grafting. In crown-grafting, the trunk of the tree is cut down to abouthalf a metre from the ground and one or more scions of the selected variety

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are inserted into it between the bark and the wood by splitting open the bark.The scion should be a dormant, terminal shoot, about 12.5 mm in diameter,with a whorl of plump swollen buds at the top. In side-grafting, the procedureis the same as in crown-grafting, except that the trunk of the stock tree abovethe grafting joint is cut down after the scions have sprouted and haveestablished themselves properly. Old trees, having several branches, can besimilarly improved (top-worked) by crown-grafting on each branch at asuitable height. Sometimes, the grafting is done by inarching, but the processis cumbersome, expensive and not very satisfactory.

Harvesting and Marketing

The fruit takes five to six months to mature. Depending upon the onsetof flowering, the mature fruits are ready for harvesting from April to May inwestern India, from May to June in the Deccan, from February to March inMalabar, from April to July in the coastal Andhra Pradesh, from May toAugust in Mysore and Rayalaseema, and from June to August in northernIndia. The mature fruits are harvested by severing the stalks to which theyare attached, when they are still green and hard. The signs of maturity varywith different varieties. As a mango tree usually bears flowers in three orfour distinct flushes lasting over a month, it is preferable to harvest the fruitsas they mature. The fruits, so harvested, can be transported after packing themin baskets or wooden crates, properly padded with straw, wood-shavings orwool, to long distances. For overseas markets, they are packed in a single layerin specially designed wooden crates.

For ripening, the fruits are spread out on rice straw in a single layer. Twoor three such layers are built one above another in a well-ventilated room.The mangoes are ready for disposal after they change colour.

Yield varies considerably with the variety, vigour of growth, flowering,etc. A grafted tree yields about 300 to 500 fruits in the tenth year, about 1,000in the 15th year and 2,000 to 5,000 from the 20th year onwards.

HORTICULTURE AND ANTHROPOLOGY

The origins of horticulture lie in the transition of human communitiesfrom nomadic hunter gatherers to sedentary or semi-sedentary horticulturalcommunities, cultivating a variety of crops on a small scale around theirdwellings or in specialized plots at some remove (such as the "milpa" or maizefield of mesoamerican cultures). In forest areas such horticulture is oftencarried out in swiddens ("slash and burn" areas). A characteristic ofhorticultural communities is that useful trees are often to be found plantedaround communities or specially retained from the natural ecosytem.

Horticultural communities may be distinguished from agricultural onesby:

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(1) the small scale of the cultivation, using small plots of mixed cropsrather than large field of single crops.

(2) the use of a variety of crops, often including fruit trees.(3) the encouragement of useful native plants alongside direct

cultivation.(4) continued use of other forms of livelihood.In pre-contact North America the semi-sedentary horticultural

communities of the eastern woodlands (growing maize, squash and sunflower)contrasted markedly with the mobile hunter gatherer communities of thePlains people. In central America, Mayan horticulture involved augmentationof the forest with useful trees such as papaya, avocado, cacao, ceiba andsapodilla. In the cornfields, multiple crops were grown such as beans (usingcornstalks as supports), squash, pumpkins and chili peppers, in some culturestended mainly or exclusively by women.

PEST S AND DISEASES OF ROSES

Roses Rosa sp. are susceptible to a number of pests, diseases anddisorders. A large number of the problems affecting roses are seasonal andclimatic. Certain varieties of roses are naturally more resistant or immunethan others to certain pests and diseases.

Cultivation requirements of individual rose species and cultivars, whenobserved, often assist in the prevention of certain pests, diseases and disorders.

Pests

Pests are often considered to be the insects that affect roses:• Aphids (Greenfly) (Order Hemiptera: Family Aphididae) Macrosiphum

Rosae: Likely to be found on new shoots and buds, aphids are softbodied insects 1-2mm long. Often green but occasionally light-brown, and sometimes with wings, they may cover (in a colony)the complete growing tip of the plant. Aphids are most active inspring and summer and multiply at a prodigeous rate feeding onthe sap of the plant by piercing the plant cells via a proboscis. Inlarge quantities they may seriously retard the growth of the plantand ruin buds. They are particularly damaging to the new shootswith subsequent damage to the emerging leaves which becomemalformed with much the same appearance as leaf-curl in peaches.

• Two-spotted Mite (Spider-mites or Red spider mite) (Order Acari: FamilyTetranychidae) Tetranychus Urticae: Previously known as red-spidermite these arachnids prefer the underside of leaves and are difficultto see with an unaided eye. Evidence of their presence is silveringof leaves where the mites have destroyed individual leaf cells. Finewebbing and eggs on the undersides of leaves is further evidenceof the presence of Tetranychus urticae.

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• Thrips (Order Thysanoptera): Thrips are slim-winged insects 1mm inlength, resembling fine black slivers of wood. Preferring light-coloured blooms and often appearing in plague numbers flowersare often left looking bruised and lustreless.

• Caterpillars (Order Lepidoptera) See also List of Lepidoptera which Feedon Roses: The tortryx (tortrix) moth Lozotaenia forsterana is aprominent pest of roses, although not the sole pest. The caterpillarsare green, up to 15mm long, and can be found boring into buds orwithin curled leaves. When disturbed the caterpillars move swiftly,dropping to the ground on a fine thread. Damage is chewn leavesand flowers and buds with "shot holes".

Cottony Cushion Scale (Order Hemiptera : Family Coccoidea) Icerya

purchasi

This scale infests twigs and branches. The mature female is oval in shape,reddish-brown with black hairs, 5 mm long. When mature the insect remainsstationary and produces an egg sac in grooves, by extrusion, in the body whichencases hundreds of red eggs. The insect causes little damage but producescopious honeydew (frass) that can cause damaging sooty mould.

California Red Scale (Order Hemiptera : Family Coccoidea) Aonidiella

Aurantii

A hard scale, orange to orange-pink, the female covering being less than1.5mm across. Often in plague numbers this scale infests upper surfaces offoliage causing yellowing, leaf fall, and twig and branch dieback. Seriousinfestations can cause plant death.

Rose Scale (Order Hemiptera : Family Coccoidea) Aulacaspis rosae

Mainly found on the stems and branches of the plant, lack of control willallow the pest to spread to flower stalks and petioles. At this point the plantwould be stunted, spindly and with a white, flaky crust of scales on the bark.Female Aulacaspis rosaemay live for 1 year and may lay 80 eggs each withseveral overlapping generations living within milliimetres of the originalparent.

• Leaf cutting bee (Order Hymenoptera : Family Megachilidae) Megachilespp.

Leafcutter bees are 6-16mm long and mostly black with bands of light-coloured hair. They chew pieces from the edges of leaves. The pieces areregular in shape, circular or oval. Damage is not often significant.

• Nematodes (Eelworms)(Order Tylenchida: Family Heteroderidae).Root-knot nematode Meloidogyne spp.See - Root-knot nematode - symptoms of Meloidogyne infestation in roses

is stunting, slow-growth, pale green leaves and wilting in mild weather.• Metallic flea-beetles (Order Coleoptera: Family Chrysomelidae) Altica spp.

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The small, shiny and metallic Altica beetles have thickened hindlegsadapted to jumping, similar to fleas. The insects are 3mm long and chew holesof irregular shapes in young leaves and buds. As the leaves enlarge so do theholes.

Diseases

Fungal, bacterial and viral diseases affect roses:

Fungal Disease

• Black Spot (Class Leotiomycetes: Family Helotiales) Marssonina rosae syn.Diplocarpon rosae : Marssonina rosae causes black spots on leaves.The spots, which may be as much as 12mm across, are generallycircular and have an irregular edge often with a yellow halo. Leavesfrequently turn yellow and fall early. Sometimes new leaves areproduced, and these may also become affected.Continual defoliation will cause weakness, die-back or death of theplant. Some very susceptible species may have stems affected witha considerable reduction in plant vigour.

• Powdery Mildew Oidium sp. : Oidium produces a very fine, powderycoating on the surface of buds and leaves. Significant cases havestems and particularly thorns, infected. Attacks on young leavesand buds will cause deformity with retardation of growth. Infectedbuds will fail to open. The disease is likely in hot, humid weather,with fungal spores overwintering on the stems and fallen leaves.

• Downy mildew (Class Oomycetes : Family Peronosporaceae) Peronosporasparsa : Peronospora causes purple-red to dark-brown spots on theleaves with irregular margins, however, often angular. Stems,petioles and flower stalks can split and spotted with purple marks.Buds, sepals, petals and calyces can be affected and will presentpurple spots. New growth affected will be deformed. The diseaseis spread by wind.

• Rust Phragmidium mucronatum : Rose rust appears as yellow patcheson the surface of leaves, with orange pustules of spores underneaththe leaf. The fungus is spread by wind. Affected leaves fall prior tohealthy ones and plants may be defoliated in serious infections.

• Anthracnose Sphaceloma rosarum : Spots caused by this fungusoriginate from a point where leaves are watersoaked, ususallyunnoticeable at first, until they turn black with a very distinctdefined edge. As the spots enlarge the centre becomes gray andmay fall out resulting in a shot-hole appearance. Defoliation mayoccur but is often not serious.

• Grey mould (Class Leotiomycetes: Family Sclerotiniceae) Botrytis cineria:On roses grey mould is prmiarily a disease of the flowers and buds,

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leaves are infrequently attacked. Infected buds rot on the stem andinfection may progress down the stem. On petals botrytis cineriaproduces pink rings.

• Verticillium wilt (Class Incertae sedis: Family Verticillium) Verticilliumdahliae.

• Sooty moulds Alternaria spp. : Sooty mould appears as black, drypowder on leaves similar to chimney soot. Many sooty moulds growon the honeydew (frass) produced by sap-sucking insect such asaphids and soft scales. Alternaria does no direct damage to plantsbut surface cover of leaves will reduce the plants capacity tophotosynthesise and may create an unsatisfactory plant appearance.

• Canker Leptosphaeria coniothyrium and Cryptosporella umbrina : Cankerspresent as small yellowish or reddish spots on bark slowlyincreasing in size. Leptosphaeria coniothyrium turns brown,increases in size, and may eventually girdle the stem. The tissuewithin the infection begins to dry out and shrink, presenting ashrivelled appearance. If the disease infects only part of the stem,growth above the canker will continue. If it girdles the stem,however, growth will cease and the stem will die.

Viral Disease

• Rose Mosaic : This diesase is caused by a complex or viruses and ischaraterised by yello patterns on the leaves. The patterns varyconsiderably, ranging between all-over fine blotches to patterns oflines in waves. The patterns may appear on a few or many leaves.Plants are infected by this virus at propagation using infected plantmaterial.

• Rose Wilt : Rose wilt is a complex of viruses and is referred to asdieback in some areas. The disease can be spread by vectors suchas aphids. Symptoms are variable and range from stunted growthto curled young leaves. The soft tissue symptoms are more evidentin spring and new leaves will reflex towards their own petioles.The affected leaves are brittle and easily fall from the plant. Fullyformed leaves will 'wilt' as if the plant were water stressed.

Bacterial Disease

• Crown gall rot (Class Alpha Proteobacteria: Family Rhizobiaceae)Agrobacterium rhizogenes : This disease is characterised by largelumps at the base of the plant stem or on roots. Galls may appearhigher on stems as the disease progresses. Galls are soft comparedto surrounding plant tissues. The pathogenic bacteria enter the plantvia a wound. If the disease affects the plant whilst it is young theplant may be affected to the degree where it will not produceblooms. All affected plants wilt readily and grow poorly.

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Environmental Disorders

• Frost : Frost will destroy fresh growth causing stems and leaves towilt, turn black and fall away from the plant. Timing pruning topromote growth after the threat of frost is a means to avoid frostdamage.

• Salinity : Salinity will present in roses as limp and light brown leaveswith dry leaf margins. Soil may require testing to determine salinitylevels. Symptoms will present if salinity is greater than 1200 partsper million.

• Herbicide Damage : Overspray or soil leaching of herbicidal sprayscan present with several symptoms: Prolonged exposure tooverspray of glyphosatewill cause yellow leaves and new leaveswill be small and elongated.

Hormone weedsprays (e.g. 24-D & 245-T) may cause grotesque newgrowth with thin twisted leaves and distorted buds. Plants may die in severecases.

Pre-emergent herbicides contacting the plants' rootsystem via the soil willcause yellowing foliage. Effects of soil borne herbicide may take several yearsto clear.

NURSERY (HORT ICULT URE)

A nursery is a place where plants are propagated and grown to usablesize. There are retail nurseries which sell to the general public, wholesalenurseries which sell only to other nurseries and to commercial landscapegardeners, and private nurseries which supply the needs of institutions orprivate estates. Some retail and wholesale nurseries sell by mail.

Nurseries grow annuals, perennials, and woody plants (trees and shrubs).These have a variety of uses: decorative plants for flower gardening andlandscaping, garden vegetable plants, and agricultural plants.

Nurseries often grow plants in a greenhouse, a building of glass or inplastic tunnels, designed to protect young plants from harsh weather(especially frost), while allowing access to light and ventilation. Moderngreenhouses allow automated control of temperature, ventilation and lightand semi-automated watering and feeding. Some also have fold-back roofsto allow "hardening-off" of plants without the need for manual transfer tooutdoor beds.

Some nurseries specialize in one phase of the process: propagation,growing out, or retail sale; or in one type of plant: groundcovers, shade plants,fruit trees, or rock garden plants.

Nurseries remain highly labour-intensive. Although some processes havebeen mechanised and automated, others have not. It remains highly unlikelythat all plants treated in the same way at the same time will arrive at thesame condition together, so plant care requires observation, judgement and

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manual dexterity; selection for sale requires comparison and judgement. AUK nurseryman has estimated that manpower accounts for 70% of hisproduction costs.

Business is highly seasonal, concentrated in spring and autumn. There isno guarantee that there will be demand for the product - this will be affectedby temperature, drought, cheaper foreign competition, fashion, etc. A nurserycarries these risks and fluctuations.

Annuals are sold in trays (undivided containers with multiple plants),flats (trays with built-in cells), peat pots, or plastic pots. Perennials and woodyplants are sold either in pots, bare-root or balled and burlaped and in a varietyof sizes, from liners to mature trees.

Plants may be propagated by seeds, but often desirable cultivars arepropagated asexually by budding, grafting, layering, or other nurserytechniques.

CLASSIFICAT ION OF PLANT S

Plants are classified in several different ways, and the further away fromthe garden we get, the more the name indicates a plant's relationship to otherplants, and tells us about its place in the plant world rather than in the garden.Usually, only the Family, Genus and species are of concern to the gardener,but we sometimes include subspecies, variety or cultivar to identify aparticular plant.

Starting from the top, the highest category, plants are classified as follows.Each group has the characteristics of the level above it, but has somedistinguishing features. The further down the scale you go, the more minorthe differences become, until you end up with a classification which appliesto only one plant.

IM PORTANT FEAT URES OF FLOWERING PLANT S

Flowering plants (Angiospermae) represent one of the largest groups ofprimary producers. Their contribution to the production of oxygen as well asthat to the nutriment of animals and man is consequently very large. Allfeatures reviewed in this chapter refer to seed-producing plants, also calledspermatophytes.

Typically, flowering plants are organized into an underground root anda shoot above ground that consists itself of a stem and leaves. The organs ofa plant that serve sexual reproduction are the flowers. Part of the pollinatedflower ripens and becomes the fruit.

In contrast to many other plant groups, flowering plants are striking,numerous and common. They are the most important group of the so-calledprimary producers that generate the prerequisite for life on earth: oxygen.Green plants have the ability to convert solar energy into chemical energy(photosynthesis) producing the oxygen necessary for all other organisms as a

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by-product. The useable plants among the flowering plants are - directly orindirectly - the basis of human existence; they are, too, an importanteconomical factor. A basic knowledge of flowering plants should thereforebe among everybody's general knowledge.

Much has been written about flowering plants and every reader of thischapter will miss something that he regards worth knowing, while he mightfind other information trivial. But everybody will understand that it isimpossible to review in a few lines a theme about which an extensive, partlypopular scientific literature exists. And although this term may sometimes beused in a disparaging way, most of the popular scientific literature isscientifically correct, lucid and, above all, very well illustrated.

To get an idea of the variety of existing plants and to get to know specialspecies, it is necessary to identify them. Many books on classification withdifferent approaches exist. In many popular books, colour photos or drawingsare used and often is the colour of the flower a primary feature of recognition.Most of the so-called scientific books on classification work on dichotomickeys, i.e. the user is asked a lot of questions in succession and has at each oneto decide between two answers. This procedure is continued until the planthas been identified.

The "scientific nature" is mostly based on the completeness of the varietiespresent in a book, since almost all of the illustrated books contain only themost common or most striking plants. Recently, electronic tools have beenused to figure out identification keys for plants.

The research into the flora of Central Europe has a tradition that goesback for centuries. It is mirrored in the nearly complete modern floras andbooks on classification. Incomplete, if existing at all, are books on theclassification of less well discovered regions, like tropics, subtropics and manymountain areas.

The question of the origin of the wealth of forms (evolution) is discussedelsewhere where it is also shown that mountains with their rather small andisolated areas provide ideal conditions for the coming into being of newspecies. This is the reason, why even very experienced botanists equippedwith renowned books on classification of the Central European flora willsometimes and in some places fail (Alps).

This chapter will deal with the characteristics of flowering plants thatproduce seed (phanerogames or spermatophytes) only. Many of the structurespresent in this plant group can be found with other non-flowering plants,too. But mosses, ferns and algae miss some features, like flowers or seeds whileothers, like roots or leaves exist in an incomplete way or are replaced by otherorgans.

The body of vegetation of many-celled algae (and mosses) is called thallus,that of flowering plants, ferns and fern-like plants (pteridophytes) is calledcormus. The latter are therefore summed up as cormophytes. The special

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features of the different plant groups. The body of vegetation of a "typical"flowering plant consists of an underground root and a shoot above ground.The shoot is organized into stem and leaves. Each of these basic organs canexist in many variations and these again can be combined in many differentways. The almost unlimited ability of combination is one of the main reasonsof the existence of such a high number of species while at the same time, theidentification of the relations of the species is aggravated.

If seemingly different organs with different functions can be traced backto the same basic organ, they are called homologous. It is also spoken ofhomology. Contrasting is analogy where organs with a similar look andfunction have descended from different organs.

T HE ST EM

Typically, flowering plants are organized into an underground root anda shoot above ground that itself consists of a stem and leaves. The stem is themain supporting axis of a flowering plant. It is made up of nodes, the partwhere one or more leaves are borne and of the usually longer, in-betweenlying internodes. Stems are characterized by their type of branching, by theirsurface and their symmetry. Special cases of stems are those of climbing plants,succulence, runners and rhizomes.

Shoot and root can be clearly distinguished within a seed. Aftergermination, they do develop into the shoot that grows towards the sun andinto the root which grows towards the earth's score. The shoot that developsduring the first stage of development is called the main- or the primary shootand depending on the plant, it does bear one, two, or more cotyledons.Growing and sprouted shoots are organized into nodes and the in-betweenlying internodes.

The nodes are the parts where the leaves are inserted. At the tip of agrowing shoot sits the terminal bud of the shoot apex that is surrounded bythe leaf (or flower) primordias. In the area of the nodes are axillary buds wherethe axillary shoots insert.

Vegetable shoots are either annual or perennial. The realised alternativedepends usually on the species and in some species on the quality of theenvironment.

The shoot of annual species is mostly fleshy and is usually called a stalkwhile perennial plants lignify and thus form trunks and branches. Somespecies, like Graminaceae or Poaceae have a hollow stalk that is called a bladeof grass.

To characterize a shoot, its type of branching and its surface (smooth,hairy, with or without grooves, round or squared) is used, as well as the ratioof heights to diameter.

It can generally be distinguished between monopodial and sympodialbranching. Monopodial branching exists, if a continuous main shoot has feeble

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apical shoots. Sympodial branching is given, if apical shoots of differentgenerations exist at the same time since the apical meristem of each sympodialbranch has either died or is differentiated into a flower, a thorn or a tendriland has thus lost its ability to grow. The respective shoot tip is nearer to theground than that of the next apical shoot. If only one of the apical shoots hasthe same direction as the main shoot would have had, then a monochasiumresults which may sometimes be difficult to distinguish from monopodialbranching.

But if two buds of a sympodial branch sprout at the same time, a more orless fork-shaped pattern, a dichasium, is formed. Hardly ever do several budsput forth at the same time (pleiochasium).

An extreme ratio of heights to diameter is found in climbing plants wherethe diameter (and the stability) is very small compared to the length so thatupright growing is only possible with the support of other plants or growingaids. Succulence is another extreme, that is typical for plants living in arid orsaline habitats.

Most shoots are of a radial symmetry but with a few species, the apicalshoots may be changed into flat, leaf-like phylloclades. Often, especially withwooden plants, the axes are differentiated into long and short shoots. Theinternodes of the long shoots that cause elongation with trees and shrubs arelonger than the short shoots. These again are often specialized apical shootswith shortened internodes. The thorns of white-horn (Crataegus), for example,or those of sloe (Prunus spinosa) correspond to such short shoots.

Apical shoots may, too, be changed into runners or rhizomes. Runnersare horizontally growing shoots with greatly prolonged internodes that areeither underground or above ground and are sometimes bearing leaves. Theinternodes produce sometimes roots at the part that is nearest to the mainshoot (shoot-borne roots). Some parts of these runners can switch to an uprightgrowth and thus behave like a normal shoot. They serve to propagatevegetatively (strawberries are a typical example). Rhizomes are mostlyunderground and grow in a horizontal (plagiotrop) manner. They are ratherenduring shoots whose internodes usually stay short. They serve as survivalorgans and are often at the same time used for storage; sometimes, they endin specially developed storage tubers like in potatoes. Except for small, scale-like leaves, rhizomes do usually have no leaves. Shoots which happen to comeinto touch with a moist soil surface may start to develop leaves and flowers.

ROOT S

Roots, the lower, underground part of a plant are characterized by theirlack of leaves. They are the structures that function to take up water,nutriments and minerals from the soil. The primary root is the main axis of aroot system. Mostly, it bears thinner side-roots. Special types of primary rootsare tap roots or turnips. Adventitious roots are roots originating directly from

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the stem. The most striking morphological feature of roots is their lack ofleaves. Roots are organized into primary roots and side-roots. The latter maydevelop a frequently branched and extensive root system. Among theprominent types of roots are the tap roots where the primary root is dominantwhile the side-roots are hardly developed at all. The primary root grows deepinto the soil. Swollen roots that have developed into storage organs are calledturnips. Their formation may involve the hypocotyl. Swollen side-roots canalso become storage organs (tubers).

The root-system of many monocots (like, that of the poaceae,for example)is completely formed from adventitious roots. Adventitious roots are rootsthat originate directly at the shoot. Rhizomes and shoots that touch the soilare able to form adventive roots. This fact is used to propagate plantsvegetatively through cuttings (parts with nodes in them).

Epiphytes do often develop air roots. They are, just like usual roots ableto take up minerals and rain water and sometimes they can even performphotosynthesis.

The picture shows a two-dimensional model of a root seeking water inthe soil during its development. The initial water distribution has beenpredetermined, forming an S-shaped zone of high concentration indicated bythe light colour. The growing tips of the main root and rootlets absorb waterthat diffuses in the soil. The decreased water concentration is indicated bydark areas that emerge around the root system. In areas with insufficient waterconcentration the rootlets cease to grow before they have reached theirpotential full length.

A leaf is an aerial and lateral outgrowth of the stem of a usually flat anddorsiventral anatomy. It functions mainly to manufacture food byphotosynthesis and consists typically of a stalk also called petiole, a flattenedblade, the lamina, and the leaf base. Strands of conducting and strengtheningtissues, the veins, run through it. Their pattern, also called leaf venation, is afeature of characterization. Leaves may be simple, i.e. undivided or compound(composed of several parts called leaflets). The blade margin and the leafarrangement at the stem are further features of characterization.

While potentially unlimited growth is a feature of the shoot, that of leavesis usually limited. They are almost always flat and have a dorsiventralanatomy, i.e. their upper and their undersurface are constructed differentlyand have different functions. This morphological and anatomical peculiarityis strongly connected to the main function of leaves, photosynthesis. Withmost conifers, the leaves are very thin and often tough (needles).

A typical leaf is organized into blade (lamina), stalk or petiole and leafbase. In some plant species, the leaf base is plainly set off against the petiolewhile the transition is invisible in others. Sometimes, it is merely indicatedby a faint widening. Occasionally, lateral excrescences develop. Most specieshave clearly recognizable petioles. If they are missing, the leaves are said to

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be sessile in which case the leaf base may sometimes surround the shoot. Theextremes are laminas with fused basal rims evoking the impression of a leavethat the shoot grows through (Claytonia perfoliata). The basis of the leaf stalkmay even grow down on the shoot a little bit further. With many monocots,like Poaceae or Graminaceae, the shoot is completely surrounded by a sheath-forming petiole.

The blade is radiated by leaf veins that contain the fibres. It isdistinguished between parallel, arched, pinnated or netlike vein structures.

Just as variable as the vein structure is the shape of the leaves. Thefundamental difference is that between simple and compound leaves. Thecompound leaves are built from several small leaves or from pinnations thatsit in a regular organisation at the undivided or branched rhachis.

Cataphylls are simply organized, scale-like leaves. They can be found atthe sprout, at the basis of sprouting shoots or underneath the normal leavesand in wooden plants also as a protection for the buds.

Finally, bracts have to be mentioned that live above the region of the usualleaves, for example at the inflorescence as subtending bracts for the flowers.But they can also have very different functions. Remarkable are the strikinglycolored bracts of Bougainvillea spectabilis and Poinsettia pulcherrima(poinsettia) that add strongly to the attractivity of the rather inconspicuousflowers. Another example are the Cuckoo-pint and its relatives where the bractencloses the inflorescence and is at the same time a refined trap for pollinatinginsects (Arum maculatum).

Some species, especially those that live in water have differently shapedleaves (heterophylly). An example is Ranunculus aquatilis (water crowfoot)which has pinnation-shaped, submerse living leaves and flat swimming leaves.

Another feature of recognition is the blade margin that can be shapedvery differently. It can be plain, serrated, double serrated, crenated or sinuated.Leaves with deep clefts are called pinnatified, pectinated, palmated or lobed.

Quite a few species have structures called leaf metamorphosis (variations),because they are homologous to leaves. Among these are thorns where themoulding of the blade does not occur and the leaf veins have beenstrengthened by the deposit of a though material, as well as leaf tendrils thatalso miss the lamina but have an extremely pliable and tension-proof petiole.There may, too, be succulence of leaves.

Leaves are always organized around the shoot in a regular pattern thatcan be recognized best when looked at from above. It is obvious in most casesthat the leaves are arranged in the way of a screw and that there is always thesame angle measurable between subsequent leaves (angle of divergence).

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2

Trends in Horticulture Growth

INTRODUCTION

Tamil Nadu is one of the leading horticulture States in India contributing7.7 per cent to the National Horticultural production with 5.7 per cent of thenational level area. Tamil Nadu has been blessed with diversified agro-climaticconditions, suitable for a wide range of horticulture crops like fruits,vegetables, spices, plantation crops, flowers and medicinal plants. A largeextent of wastelands and under-utilized lands are available in the State forhorticulture development. Tamil Nadu has a long coastal belt of 1000 km.suitable for crops like cashew, coconut, tropical orchids etc. The southern partof Tamil Nadu has the potential for growing off-season mangoes and grapes.A lot of awareness has been created among the farmers of Tamil Nadu aboutcultivation of high value horticulture crops. It is aimed to achieve 8 per centannual growth rate during X Five-Year Plan in the horticultural sector.

The area coverage in 2003-04 was 8.24 lakh hectares, a little less than thenormal area of 8.52 lakh hectares. During 2004-05, despite the failure of themonsoon, the area coverage would still be around 8.91lakh hectares. In theyear 2005-06, an area of 9.73 lakh hectares is expected to be covered.

TAMIL NADU HORTICULTURE DEVELOPMENT MISSION

The Government have setup a Mission for Horticulture development inTamil Nadu vide G.O. Ms. No.155, Agriculture (H1-2) Department, dated28.4.2003 with a mandate to give impetus to production, processing for valueaddition and marketing of Vegetables, Fruits, Flowers and Medicinal Plants.Tamil Nadu is the first state in India to setup a separate Mission fordevelopment of Horticulture in the year 2003. It is aimed to double theproduction of horticulture crops by the year 2011-12 in Tamil Nadu. Thepresent level of productin of 99.47 L.MT will be doubled to 187.80 L.MTsduring the year 2011-12.

OBJECT IVES OF T HE M ISSION

1. Improving production through balanced nutrition management.2. Evolving suitable mechanism for regulating the production of

quality planting materials and giving impetus to need basedresearch.

3. Establishing adequate infrastructure for post harvest managementespecially preservation and marketing.

4. Encouraging active involvement of farmers associations in adoptionof modern technological practices.

A Governing Body has been constituted to monitor the effectiveimplementation of the mission under the Chairmanship of the HonourableMinister for Agriculture with the Vice- Chancellor of Tamil Nadu AgriculturalUniversity, Secretaries to Government and Senior officers of variousdepartments and Nominees of Central Government agencies like NationalHorticulture Board, APEDA (Agricultural and Processed Food ProductsExport Development Authority) as members. The Chief Secretary toGovernment is the Vice Chairperson of the Governing body.

Mango Development Mission

Under Mango Development Mission it has been programmed to layoutdemonstration plots in the farmer’s field to enable them to have a visual impactof various new technologies involved in the cultivation of Mango crop. A sumof Rs.22.55 lakhs has been proposed for this purpose. National HorticultureBoard has agreed to sanction Rs. 10.00 lakhs out of which released a sum ofRs.5.00 lakh for implementation of this scheme. On receipt of GovernmentOrder, this scheme will be implemented in the selected Mango growingdistricts.

Cashew Development Mission

Under Cashew Development Mission it has been programmed to layoutdemonstration plots in the farmer field to increase the productivity of cashewwith high yielding varieties and hi technologies. A sum of Rs.23.00 lakhs hasbeen proposed for this scheme. National Horticulture Board has agreed tosanction Rs.10.00 lakhs out of which released a sum of Rs. 5.00 lakh forimplementation of this scheme. On receipt of Government Order this schemewill be implemented in the selected cashew growing districts.

Training to Farmers

With a view to expose the farmers to various hitech aspects ofHorticulture, farmers are taken to various research stations in out of state ontour-cum-training basis. About 10,000 Nos. of farmers are given trainingduring the year 2004-2005. Apart from this farmers were given awareness invarious horticulture technologies of Medicinal Plants, Flower cultivation and

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Vegetable Cultivation through the conduct of District level seminars andExhibition.

The extension officers and field level workers of this department are alsogiven training in the latest technologies of horticulture from the ResearchStations situated inside and outside the State.

STAT E PLAN SCHEM ES

The following State Plan Schemes have been implemented during2004-05.

Integrated Horticulture Development Scheme (IHDS)

In G.O. Ms. No.180, Agriculture (H2) Department, dated 28.5.2004,Integrated Horticulture Development Scheme was sanctioned with an outlayof Rs.766.26 lakhs. Under this scheme, planting materials, high yielding/hybridvegetable seeds are distributed to horticultural farmers at 50% subsidised cost.

Integrated Tribal Development Programme (ITDP)

In G.O. Ms. No.189, Agriculture (H2) Department, dated 28.7.2004,Integrated Tribal Development Programme was sanctioned with an outlay ofRs.39.325 lakhs. Under this scheme, horticulture inputs are distributed toTribal farmers in Salem, Namakkal, Dharmapuri, Tiruvannamalai, Vellore,Trichy and Villupuram districts at subsidised cost. Tour-cum-Training is alsogiven to the Tribal farmers under this scheme.

Western Ghats Development Programme (WGDP)

In G.O. Ms. No.52, Plannting, Development and Special Initiatives (TC-1) Department, dated 22.7.2004, Western Ghat Development Programme wassanctioned. Under this scheme, horticulture inputs are distributed to thehorticultural farmers in Theni, Madurai, Dindigul, Virudhunagar, Coimbatore,Erode, Tirunelveli and Kanyakumari districts at 25% subsidised cost.

Hill Area Development Programme (HADP)

In G.O. Ms. No.60, Planning, Development and Special Initiatives (TC 1& 2) Department, dated 5.8.2004, Hill Area Development Programme wassanctioned with an outlay of Rs.315.00 lakhs. Under this scheme, horticulturalinputs are distributed to the farmers of Nilgiris district at 25% subsidised cost.

The above Schemes will be continued during the year 2005-06 also. Inthe above schemes:-

• Focus is given for cultivation of high yielding / hybrids andvarieties, adoption of efficient water and nutrient management andIntegrated Pest Management.

• Adoption of location specific and crop specific technologies isencouraged.

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• Mango, Amla, Guava, Sapota and Vanilla are given importanceunder fruits and spices categories. High yielding / export potentialvarieties are given priority.

• Under vegetables, focus is given on hybrids / high yielding varietieswhich fetch good return to the farming community.

• Under Integrated Horticulture Development Scheme, horticultureplants are distributed at 50% subsidy for a maximum of 1 hectarefor fruits and 0.5 hectare for seeds of vegetables and flowers toeach beneficiary.

• In the Tribal areas of Tamil Nadu, viz., Shevroy hills,Arunoothumalai, Pachamalai, Kolli hills, Sitheri hills, Javvadu hills,Kalrayan hills, and the tribal farmers are encouraged to developindividual orchards by optimally utilizing the available waterresources.

• Considering the economic status of the tribal farmers, thehorticulture plants are distributed at 75% subsidy under theIntegrated Tribal Development Programme (ITDP).

• Watershed approach is followed in the Western Ghat DevelopmentProgramme and the Hill Area Development Programme.

• Focus is given on prevention of soil erosion and preserving the Eco-System through appropriate cropping pattern.

• Planting materials and seeds are distributed at 50% cost underWestern Ghat Development Programme and Hill Area DevelopmentProgramme.

• The State is maintaining 55 State Horticulture Farms in whichPedigree and Hybrid planting materials are produced and suppliedto the farmers under various schemes.

• To update the technical knowledge of the extension officers of theHorticulture Department, in-service training is given at CentralHorticulture Training Centre, Kudumianmalai in Pudukkottaidistrict.

• A two-year diploma course is being conducted at the HorticultureTraining Centre, Madhavaram (Chennai) with an intake of 40students every year.

Promotion of Alternate Cropping in the Nilgiris

Promoting alternate crops is considered as an important measure toovercome the crisis facing the tea growers. Steps are being taken to popularizemulti-tier cropping, viz. cultivation of silver oak, pepper, cardamom, mandarinorange, etc., along with tea plantation. Considering the prevailing agro-climatic condition in The Nilgiris, new kinds of crops like Macadamia andPeacan nuts have been identified for introduction. It is proposed to raise thesecrops in the State Horticultural Farms at Nanjanad and Colegrain in Nilgiris

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district on trial basis. Subsequently, efforts would be made to introduce thesecrops to the farmers in select pockets. A sum of Rs.50.00 lakhs has beensanctioned vide G.O. Ms. No.244, Agriculture (H2) Department, dated17.6.2004 by the Government for promoting alternate cropping in Nilgirisdistrict and the scheme will be continued during 2005-06 also.

Tamil Nadu Precision Farming Project

As announced during last budget session, Government have sanctionedan Innovative Scheme by name “Tamil Nadu Precision Farming Project” tobe implemented in Dharmapuri District and newly formed Krishnagiri districtcovering an area of 400 Ha. of farmers’ land with Micro Irrigation andFertigation vide G.O. Ms. No.24, Agriculture (H1) Department, dated23.1.2004. This scheme will be implemented with a total cost of Rs.720.60 lakhsover a period of 3 years.

It is programmed to cover an area of 100 Ha. in the first year, 200 Ha. inthe second year and 100 Ha. in the third year at financial allocation of Rs.208.59lakhs, 316.16 lakhs and Rs.195.85 lakhs respectively.

The Tamil Nadu Agricultural University will undertake this project as aTurnkey project and implement it with the co-operation of the Departmentsof Horticulture, Agricultural Engineering, Agriculture, Agricultural Marketingand Agri-Business and the District Administration. High value like gherkins,hybrid tomatoes, capsicum, paprika, babycorn, white onion, bhendi, cabbageand cauliflower are proposed to be cultivated under the scheme. Under thisproject, 100% subsidy will be given on the cost of cultivation of the first cropto the farmers selected during first year. 10% of the cost of cultivation will becollected from the farmers selected during second year. 20% of the cost ofcultivation will be collected from the farmers selected during third year.

During the year 2005-06, the above scheme will be implemented in sixmore districts viz., Theni, Vellore, Cuddalore, Erode, Tirunelveli andThanjavur with an financial outlay of Rs.10.00 crores. An area of 100 Ha. willbe covered in each district.

SCHEM ES

Under Part II Scheme a sum of Rs.120.00 lakh was sanctioned vide G.O.Ms. No.398, Agriculture (H1) Department, dated 6.8.2004 to improve theirrigation facilities in 28 State Horticulture Farms and the work is entrustedwith Public Work Department and Agriculture Engineering Department andwill be completed before 31.3.2005.

Tamil Nadu Horticulture Development Agency (TANHODA)

An independent nodal agency viz., The Tamil Nadu HorticultureDevelopment Agency (TANHODA) has been formed vide G.O. Ms. No.250,Agricultue (H1) Department, dated 18.6.2004 and Rs.20.00 lakhs wassanctioned under Part II 2004-05 with the following objectives.

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1. To promote Hi-Tech Horticulture and Precision Farming.2. To promote Public Private Partnership (PPP) for horticulture

development.3. To disseminate knowledge on modern technologies in horticulture

to the farming community as well as extension personnel.4. To manage production and distribution of pedigree planting

materials through State Horticulture Farms.5. To implement the horticulture development schemes including the

schemes under the Tamil Nadu Horticulture Development Missionand the National Horticulture Mission by accessing funds from theagencies concerned.

This agency will serve as a special purpose vehicle (SPV) for theimplementation of various horticultural development programmes.

Part II Schemes (2005-06)

The following new schemes will be implemented during the year 2005-06.

1. Scheme for provision of infrastructure facilities in 25 StateHorticulture Farms (Farm equipments like Power Tillers withAccessories and Multipurpose Tanker) - Rs.50.00 lakhs.

2. Organic farming vegetable cultivation with market linkage in 15State Horticulture Farms - Rs.10.00 lakhs.

6. Water Management - Command Area Development Programmes(Centrally sponsored scheme - shared between State and Centre).

Command Area Development Programme

The Command Area Development Programme is implemented by theAgricultural Engineering Department as a centrally sponsored and equallyshared programme on 50: 50 basis between the State Government andGovernment of India.

At present, the programme is implemented in Cauvery Command,Parambikulam Aliyar Project, Tambirabarani River Basin Project, Palar-Poranthalar Project, Krishnagiri Reservoir Project, Gadana RamanadhiIrrigation System, Nambiyar River Basin System, Patchaiyar River BasinSystem, Manimuthar Irrigation System, Aanaimaduvu System, ChinnarResevoir system and Maruthanadi System.

The programme has been completed in Lower Bhavani Project, CumbumValley Project, Periyar - Vaigai Project, Sathanur Reservoir Project,Amaravathy Reservoir Project, Kothayar-Chittar project and ThoppaiyarProject. The remaining projects will be covered as per the schedule approvedby the Government of India.

The six major components of Command Area Development Programmeare as follows:

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a. Construction of Field Channel.b. construction of Field drain.c. Reclamation of Water logged Saline and alkaline soil areas.d. Rotational Water Supply.e. Participatory Irrigation Management.f. Management Subsidy.The Agricultural Engineering Department has also developed a water

management strategy for the area below the sluice, which is crucial foragricultural production. Water management holds the key for optimisationof water use and maximisation of crop production.

Construction of Field Channel

Construction of Field channel is done by considering the needs of thehead reach farmers so that their interference is eliminated and the tail endfarmers also get adequate irrigation water. This brings in equity in distributionof water to all the farmers in the entire sluice command. This has been achievedby using optimum flow concept.

The unit cost is Rs.10000/- per ha. in all New Project areas includingCauvery Command. For old ayacut of Cauvery Command the unit cost ofconstruction of field channel works is Rs.4400/- per ha. An area of 9.87 lakhha. has been covered till 31.3.2004 out of total cultivable command area of10.97 lakh ha. with Construction of field channel component in 12 commandareas.

During 2004-2005, this work is being carried out in an area of 25695 ha. Itis programmed to continue the scheme during 2005-2006 also.

Construction of Field Drains

Under the construction of field drains component, drains from individualfields to Government drains (or) natural drains outside the outlet are executed.The expenditure on field drains is inclusive of costs of earthwork, road cuttingsand drop structures, if needed to prevent erosion. The unit cost per ha. forthis component is Rs.4000/-.

Reclamation of Water Logged, Saline & Alkaline Soil Areas

In this component, the area affected by Waterlogging, saline and alkalineproblems within the command area are identified and measures to reclaimthose patches are takenup. At present this component is taken up inTamirabarani River Basin Project and Krishnagiri Reservoir Project at an unitcost of Rs.15000/- per ha.

Rotational Water Supply Works

Rotational Water Supply or Warabandi is a system of equitable waterdistribution, by turns according to a predetermined schedule specifying the

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day, time and duration of supply to each farmer in proportion to his holdingsize in an outlet command.

The rotational water supply schedule is prepared by the staff ofAgricultural Engineering Department after executing the on farmdevelopment works and handed over to farmers for implementation.

Rotational water supply was first introduced in Lower Bhavani Project,Periyar - Vaigai Project and Cauvery Command in the year 1984-’85.

An area of 8.92 lakh ha. has been covered under rotational water supplyin Tamil Nadu in 12command areas till March 2004. During 2004-2005, thiswork is being taken up in an area of 52750 ha. It is programmed to continuethe scheme during 2005-2006 also.

Participatory Irrigation Management

To implement On Farm Development Programmes and for ensuing equityin distribution of water, farmers participation is important.

Agricultural Engineering Department is intensively encouraging itself inthis work for the last 20 years.

The task of handing over the responsibilities to the farmers is beingimplemented. In orders to ensure farmers participation, farmers associationsare formed on a three tier basis as follows.

Farmers’ Association - at sluice command level.Farmers’ Council - at distributory level.Farmers’ Federation - at project level.

Management Subsidy

In order to sustain the farmers associations/ councils already formed andfor the maintenance of the on-farm infrastructures created and to continueRWS (Warabandhi) for equitable water distribution, a financial assistance isprovided in the form of one time functional grant to registered Farmers’Councils at Rs.540.00 per ha. shared equally by the Central and the StateGovernments. To avail this, farmers have to contribute Rs.60.00 per ha.

Out of the 632 Farmer’s Councils registered, an amount of Rs.1478.95 lakhshas been released as management subsidy till March 2004 to 597 registeredFarmers’ Councils to maintain the infrastructure created under the CommandArea Development Programme.

World Bank Aided Model Rehabilitation Project in Hanumanadhi Sub

Basin

The World Bank aided IAIP pilot project was implemented inHanumanadhi sub basin covering 135 ha. in the command area, which is fedby Adavinainarkoil system tanks, at a cost of Rs.81.00 lakhs. This project isconstrued as a threshold for a broader Model Rehabilitation Project inHanumanadhi sub basin. The World Bank aided model Rehabilitation Project

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in Hanumanadhi sub basin of Tamirabarani river basin in Tirunelveli districthas been successfully completed by September 30th 2004. The project has beenexecuted and implemented in a multi disciplinary project approach involvingPWD-WRO, Departments of Agricultural Engineering, Agriculture,Horticulture, Fisheries, Agricultural Marketing and TNAU. The funds ofRs.62.5 Crores was provided vide GO MS No. 33 PW (WR 1), Dated 30-1-2004, out of which Rs. 22.55 Crores was allotted to Agricultural EngineeringDepartment. The project area encompasses Sengottai, Tenkasi and V.K. Pudurtaluks.

The ayacut irrigated by 50 tanks and 14 anicuts is spread over an area of3000 ha. in 15 villages namely Ayikudi, Elathur, Kilankadu, Kodikurichi,Kulayaneri, Kutthukalvalasai, Nainaragaram, Neduvayal, Panpoli,Rajagopalaperi, Sambavar Vadagarai, Surandai, Sundarapandiyapuram,Verakeralampudur and Vadagaraikilpidagai. An area of 2194 Ha. has beencovered with an expenditure of Rs. 16.35 crores by the AgriculturalEngineering Department.

Objectives

• Improving productivity of water and crop production by sharingof water and Mechanized farming.

• Uniform and optimum application of water and fertilizer by hightech micro irrigation systems. Rain water harvesting and rechargeof ground water through rejuvenation of wells and farm ponds.

• Training, effecting attitudinal changes among the stakeholders andimproving their socio - economic status with measures likeAquaculture in farm ponds.

High Tech Drip and Sprinkler facility

In the pre project period only two crops were cultivated in a year andmost of lands were kept fallow for want of water in the third season Afterimplementation of the project it is made possible to cultivate an additionalthird crop using the water conserved through micro irrigation techniques.After the project implementation an intensive cropping scheme one like belowis followed.

I. Crop : 28 Ac-inch water required crops like groundnut, vegetables.II. Crop : Paddy.III. Crop: 18 Ac-inch water requirement crops like Pulses, fodder

cholam etc.The introduction of drip and sprinkler irrigation systems in the project

area on a 10 Ha block basis is estimated to save atleast 30 - 40 % total irrigationwater. Sprinkler irrigation system are provided to the individual farmers whodo not have wells and new borewells are sunk for these farmers. Drip irrigationsystems are provided to the farmers who have atleast one well.

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Under ground PVC pipes are laid to convey water to the micro irrigationsystems from the sources of water. However open field channels shall carrywater during the paddy season. An area of 2194 Ha., is covered with anexpenditure of Rs.1065.72 crores under high tech micro irrigation systemincluding pipe laying.

GIS and Survey

The geographical information survey is used to get precise informationon land holding, water bodies, vegetation etc., The GIS format creation in1:1000 scale, high resolution satellite data like IRS PAN merged 1:10,000 scale,GIS in 3-D, aerial photographs are some of the advanced survey and mappreparation techniques.

ISDN / Lease line connections are installed at offices of water users groups,AED office in Tenkasi, Tirunelveli and MDPPP office AED, Commissioner ofAgricultural and Commissioner of Agricultural marketing, secretariat inChennai to provide latest information on farm practices and market trends tothe farmers. These facilities have been created with Rs. 2.00 crores.

Water Harvesting Structures

Farm ponds are executed for collection of rainwater. 52 ponds have beenconstructed.

The pond water is used for domestic, irrigation and aquaculture purposes.The defunct and low yielding wells are rejuvenated with vertical andhorizontal in - well bores to improve their yield. Shallow and medium borewells are sunk and the new bore wells are provided with energized motors.An amount of Rs. 2.84 crores has been incurred on these water-harvestingmeasures.

Farm Mechanization

Modern agricultural machineries like power tillers, paddy transplantersetc. have been procured at the cost of Rs.0.53 crores and are hired to thefarmers.

Training to Farmers and Officers

Trainings on high tech micro irrigation systems, water-harvestingmethods and farm mechanization are imparted to the water user groups anddepartmental officers with Rs. 0.34 crores. Documentation, video graphy andevaluation studies of the project are also included in the programme.

Farmers’ Participation

The participation of the stakeholders - the farmers, is emphasized rightfrom the decision-making and implementation stages. The farmers contributedin cash, kind and labour at various stages of the project. The progressive

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farmers, the women farmers and rural youth participated in every sphere ofactivity. The farmers will be sharing a portion of the project cost in the formof betterment levy.

Follow up on Project

With the funds from the ongoing schemes, state and other organizationslike NABARD etc., the uncovered areas in this project are proposed to becovered.

JAT ROPHA

It has been proposed to taken up Jatropha Cultivation in 200 Ha. fundedby NOVOD Board during the year 2004-05 in the following 11 StateHorticulture Farms.

1 G.O. Vallathirakottai, Pudukottai District 45 Ha.2 G.O. Karumanthurai, Salem District 50 Ha.3 SHF, Jeenur, Krishnagiri District 20 Ha.4 SHF, Kudumianmalai, Pudukottai District 10 Ha.5 SHF, Navlock, Vellore District 20 Ha.6 SHF, Mulluvadi, Salem District 10 Ha.7 G.O. Srivilliputhur, Virudhunagar District 12 Ha.8 SHF, Melkadirpur, Kancheepuram District 10 Ha.9 SHF, Vichanthangal, Kancheepuram District 10 Ha.10 SHF, Thagarakuppam, Vellore District 10 Ha.11 SHF, Kannampalayam, Coimbatore District 5 Ha.Total 200 ha.NOVOD has sanctioned a sum of Rs.75.00 lakhs to the Managing Director,

Tamil Nadu Watershed Development Agency, Chennai 32 towardsimplementation of this programme.

T ARGET S AND ACHIEVEM ENT S

The financial allocation for the year 2004-05 and the expenditure incurredupto February 2005 and the proposed outlay for the year 2005-2006 for theState Plan Schemes and Schemes shared between State and Centre arefurnished below.

Allocation for the year 2004-2005 and Proposed outlay for 2005-2006.

Unit: Rs. in lakhs

Sl. Scheme 2004 – 2005 2005-2006

No Amount Acht. upto Proposed

Sanctioned Feb. 05 Outlay

A. State Plan Schemes

1 Integrated HorticultureDev. Scheme 766.260 615.080 769.400

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2 Integrated Tribal Dev. Programme 39.330 35.135 41.7903 Western Ghat Dev. Programme 135.480 105.158 132.0804 Hill Area Development Programme 315.000 267.750 314.9405 Precision Farming inDharmapuri Dt. 232.950 203.000 1120.850

6 Promotion of alternatecropping(Nilgiris Dt) 50.000 48.090 50.000

7 Part-II Schemes 120.000 105.000 60.0108 TANHODA 20.000 0 20.000

Total 1679.020 1379.213 2509.0 0B. Schemes shared between State and Centre

1 Integrated Prog. for Dev.of Cashew 155.200 141.847 273.320

2 Integrated Prog. for Dev. of Fruits 218.000 186.389 318.3503 Integrated Prog. for Dev. of Spices 60.000 47.946 180.1304 Integrated Prog. for Dev.

of Vegetables 219.000 191.644 310.0005 Integrated Prog. for

Development of Medicinaland Aromatic Plants 52.500 48.350 52.700

6 Integrated Prog. for Dev. Cocoa 12.000 11.190 12.0007 Dev. of Commercial Floriculture 145.820 111.200 165.0008 Development of Mushroom cultivation 10.000 9.500 10.0009 Development of Horticulture

through plasticulture interventions 14.500 13.030 14.50010 Innovative Programmes for

Development of Horticulture 113.900 43.455 90.000

Total 1000.920 804.471 1426.000

Grand Total (A+B) 2679.940 2183.684 3935.070

PROGRAMME IMPLEMENTATION FOR 2005-06

• Thrust on Hi-Tech Horticulture and precision farming.• Expansion of area under micro-irrigation and fertigation.• Stabilizing the area of water loving crops and expanding the area

under dry land crops with a focus on effective water management.• Area expansion of horticulture crops through various Waste Land

Development programmes.• Strengthening the system of production of pedigree and hybrid

planting materials in both public and private sectors.• Promotion of Organic farming with focus on export market.• Promotion of Agri Export Zones (AEZ) for specified crops.

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• Building up of Public and Private Partnership.• Promotion of Contract / Corporate farming.• Empowerment of farmers with special focus on farm women.• Effective transfer of technologies by tour-cum-training to farmers.• E-Governance and Human Resources Development through

effective training for extension officers.• Linkage with Agro Processing Industries with New Anna

Marumalarchi Thittam (NAMT).• Post harvest management and reduction of post harvest losses.

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3

Genetic Resources in Horticulture

The rural poor depend upon biological resources for an estimated 90%of their needs. In the industrialised world, access to diverse biologicalresources is necessary to support a vast array of industrial products. In thecontinuing drive to develop efficient and sustainable agriculture for manydifferent conditions, these resources provide raw material for plant and animalbreeding, as well as for the new biotechnologies. Genetic diversity inagriculture enables crops and animals to adapt to different environments andgrowing conditions. The ability of a particular variety to withstand droughtor inundation, grow in poor or rich soil, resist one of the many insect pests ordiseases, give higher protein yields or produce a better-tasting food are traitspassed on naturally by its genes. This genetic material constitutes the rawmaterial that plant and animal breeders and bio-technologists use to producenew varieties and breeds. Without this diversity we would lose the ability toadapt to ever-changing needs and conditions. Sustainable agriculture couldnot then be achieved in many of the world’s different food productionenvironments.

GENETIC DIVERSITY

Diversity among individual plants and animals, species and ecosystemsprovides the raw material that enables human communities to adapt to change- now and in the future. Deprived of bio-diversity, the ability of humankindto meet the challenges resulting, for example, from climate change would beseverely limited. The diversity found within the small number of plant andanimal species which form the basis of world agriculture and food productionremains a small but vital part of the earth’s bio-diversity.

Genetic diversity is not evenly distributed throughout the world; it is infact concentrated in tropical and sub-tropical areas, where the majority ofdeveloping countries are located. Plant genetic resources are of an inestimablevalue, and will continue to be so in the future, independently of whetherscientists use them by means of conventional plant breeding or modern genetic

engineering. The importance of plant genetic resources as the ultimate sourceof food is enormous. Their loss constitutes a serious threat to world foodsecurity. The conservation of genetic resources goes far beyond the salvationof species. The objective must be to conserve sufficient diversity within eachspecies to ensure that its genetic potential will be fully available in the future.

Within the Plant Kingdom, around 350,000 species have been classified,out of which about 80,000 have been classified, out of which about 80,000 havebeen found to be edible. In the course of history, mankind has utilised about7,000 of these plant species for food. Today, only 150 plant species arecultivated and of these, the so-called major crops can be contemplated in about30 plant species which are producing about 95% of the world’s calories andproteins. About 75% of food consumption comes from only 12 plant speciesand 5 animal species. Half of this food comes from only 4 plant species (rice,maize, wheat and potato) and 3 major animal species (cattle, swine andpoultry).

Over millennia, men and women farmers have developed within eachdomesticated food species thousands of landraces (farmers varieties) andbreeds adapted to local conditions and needs. This “humanmade”agrobiodiversity is now seriously endangered. Today the world populationamounts to about 5 billion people. An increase of about 60% is expected inthe next thirty years, which will bring the world population to some 8 billionpeople. Today, 800 million people are chronically malnourished. On the basisof recent estimates, in the developing countries alone, food production willneed to increase by about 60% in the next 25-30 years in order to keep pacewith the expected demographic growth.

PLANT GENETIC RESOURCES

Despite the useful progress made so far, some major questions are stillcurrently being debated in the FAO fora that deal with plant genetic resourcesfor food and agriculture, i.e.:

• Who owns plant genetic resources?• Whose responsibility is it to conserve them?• Who benefits from their use?• What form the benefits deriving from their use should take, and

how should they be shared?• How can the benefits of biotechnology be directed to the largest

number of people and countries?• How can measures be taken to prevent certain applications of

biotechnology endangering the livelihoods of large numbers ofpeople in developing countries? (for example by new productsreplacing the agricultural commodities that provide theirlivelihood)?

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• How can biotechnological research be directed towards themaximum benefit to world society?

• What measures can be taken to prevent modern, intensiveagriculture, with homogeneous varieties, stamping out the world’srich inheritance of plant germplasm conserved in the heterogeneouslocal varieties in farmers’ fields?

In June 1996, the Fourth International Technical Conference on PlantGenetic Resources, which was attended by 150 countries and 54intergovernmental and nongovernmental organisations, was held in Leipzig,Germany. The Conference was convened at the request of the FAOCommission on Plant Genetic Rthe request of the FAO Commission on PlantGenetic Resources, and was endorsed in Agenda 21 of the United NationsConference on Environment and Development, and at the Nairobi Conferencefor the Adoption of the Agreed Text of the Convention on Biological Diversity.

The Conference adopted a Global Plan of Action for the Conservationand Sustainable Utilisation of Plant Genetic Resources, together with theLeipzig Declaration, and it also considered the first Report on the State of theWorld’s Plant Genetic Resources.

The Global Plan of Action was prepared through a participatory,countrydriven process, involving a wide variety of stakeholders:governments, nongovernmental and industry organisations, and individualscientists. A total of 158 governments prepared Country Reports, assessingthe status of their plant genetic resources, as well as their capacity to care forand utilise these resources. Twelve regional and subregional meetings wereheld, where governments considered regional problems and opportunities,and made recommendations for the Plan. The Report on the State of theWorld’s Plant Genetic Resources is the first comprehensive worldwideassessment of the state of plant genetic resource conservation and use,identifying the urgent priorities for action which are, in turn, addressed inthe Global Plan of Action. The Report and the Plan are now two strategicelements of the Plan are now two strategic elements of the FAO Global Systemfor the Conservation and Utilisation of Plant Genetic Resources.

The Plan is intended to provide a coherent strategy for action in severalfields: in-situ and ex-situ conservation, sustainable utilisation of plant geneticresources, and institutional and capacity building. Therefore, the GPA needsto be taken as a whole and considered in a comprehensive manner embracingall operational, programmatic, policy and political aspects.

The Leipzig Declaration asserts that “our primary objective must be toenhance world food security through conserving and sustainably using plantgenetic resources”. This Declaration commits the Governments to takingnecessary steps to implement the Global Plan of Action.

On a different, albeit related, plane, the world’s political leaders, meetingin Rome at the 1996 World Food Summit, have made a public commitment to

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end hunger. Through the Plan of Action adopted by the Summit, governments,international organisations and all sectors of civil society are encouraged tojoin forces in a concerted effort to ensure access at all time to the food requiredfor a healthy active life for all the world’s people. Meeting the same month,in November 1996, the Third Conference of the Parties of the Convention onBiological Diversity held in Buenos Aires focused on agricultural bio-diversity,and its res focused on agricultural bio-diversity, and its role in ensuring atransition towards sustainable agricultural practices.

All of these inter-governmental meetings recognised the importance ofplant genetic resources for food security and called on countries to implementthe Global Plan of Action for the Conservation and Sustainable Use of PlantGenetic Resources for Food and Agriculture.

The needs highlighted at Leipzig, Rome and Buenos Aires, and therecommendations agreed upon by governments in the Global Plan of Actionfor Plant Genetic Resources present many challenges to all crop scientists, interms of developing or adapting suitable technologies and approaches for thesustainable utilisation of plant genetic resources for food and agriculture(PGRFA). The concept of sustainable utilisation is defined in the Conventionon Biological Diversity as the use of genetic resources in a productive mannerwhich does not lead to the long term loss of such resources. This provides foran approach which integrates both conservation and utilisation and whichspans all aspects of the use of PGRFA, whether by scientists or farmers.

Plant genetic resources can be described as the part of bio-diversity thatnurtures people and is nurtured by people. Horticultural crops, althoughconstituting only one part of plant genetic resources for food and agriculture,comprise a very wide range of species of vital importance to ensuring age ofspecies of vital importance to ensuring food security.

All governments have now recognised that the utilisation of plant geneticresources is the key to improving agricultural productivity and sustainabilityand can contribute to socio-economic development, food security and thealleviation of poverty. To achieve these desirable goals, the Global Plan ofAction promotes an integrated strategy for the conservation and sustainableutilisation of PGRFA with the following features: productivity - greater use ofplant genetic resources will be required to contribute to the productivityincreases needed to meet growing populations. This will require continuedaccess to, and exchange of, the world’s plant genetic resources; sustainability -there is a need to ensure that such use of plant genetic resources is coupledwith the conservation of plant genetic resources, both in-situ and ex-situ.

This will require, inter alia, approaches to crop improvement which allowthe maintenance of higher levels of genetic diversity and resources inproduction systems, thereby contributing to reduced genetic vulnerability andless genetic erosion. Additionally, conservation programmes should be clearlylinked with utilisation efforts and the sharing of benefits, in order to reinforce

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the sustainability of such programmes; equity - those responsible for conservingand developing plan those responsible for conserving and developing plantgenetic resources should be able to participate fully in the benefits derivedfrom their use. There is thus a need to develop crop improvement approachesto enable farmers in marginal areas, as well as those in high-productivity areasto benefit fully from the utilisation of plant genetic resources.

In meeting these challenges of harnessing plant genetic resources forsustainable agriculture and future food security, the role of crop scientistswill be crucial.

GLOBAL NETWORKS

The importance of conserving plant genetic resources has been stressedin several international agreements, such as the International Undertakingon Plant Genetic Resources and the Convention on Biological Diversity andits Agenda 21. The successful conservation and sustainable utilisation of plantgenetic resources for food and agriculture involves action by a wide range ofpeople in every country: policy makers, planners, scientists, genebank curators,breeders, rural communities and farmers. Strong coordination mechanismsare required at the national level to enable all these players to participateconstructively.

In response to th is need , through its international netw orks on ex-situand in-situ conservation, FAO is pursuing efforts in order to coordinate, underthis umbrella, the conservation and utilisation of er this umbrella, theconservation and utilisation of plant genetic resources of many agriculturalcrops and to facilitate the exchange of genetic material, information andtechnology.

The need for germplasm conservation is recognised for all crops andplants of agricultural importance. This, in turn, dictates the need forprogrammes in evaluation and documentation, and these activities must becarried out in cooperation with institutions, scientists and technicians withcrop-specific competence. In this context, we consider it important to fosterthe establishment of crop-related networks with special competence andresponsibilities related to genetic resources of specific crops.

The development of clearer policy lines related to the conservation ofgenetic variability of crop species, through regional and global inter-countrycooperative programmes, was one of the principal aims of the LeipzigTechnical Conference on Plant Genetic Resources. The Global Plan of Actionon PGR is now under implementation through a partnership mechanisminvolving inter-country crop-related networks, as well as several national andinternational institutions and organisations, including non-governmentalorganisations. The Global Networks which have been recently launched bythe FAO are thus elements of the Global Plan of Action.

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It is being demonstrated that inter-country networking constitutes a usefulinstrument to facilitate policy itutes a useful instrument to facilitate policydevelopment and to provide guidance and global coordination for inter-institutional research work concerning crop genetic resources. It should beunderlined that the crop-related networks that operate under the aegis of theFAO have been constituted on the basis of voluntary and self-fundingparticipation of interested countries, institutions and scientists.

In facilitating the implementation of the Global Plan of Action on PGR,and recognising the important role of horticultural crops for agriculture andfor food security, FAO has concentrated efforts on promoting regional, inter-regional and global crop-related networks, particularly oriented towards thehorticultural sector, as instruments to facilitate scientific exchange, informationsharing, technology transfer, and research collaboration. The networks alsoprovide a forum for determining ways of sharing responsibility for thecollection, conservation, evaluation, characterisation and utilisation ofhorticultural genetic resources.

These crop-related networks bring together different types of specialiststo set collaborative research objectives, define policy priorities and strengthenactivities on the conservation and utilisation of genetic resources in relationto specific groups of crops. Most of the partners of these inter-countrycooperative programmes are internationally recognised scientists andmembers of the Internatignised scientists and members of the InternationalSociety for Horticultural Science which further strengthens collaborationbetween the ISHS and FAO.

The innovative aspect of these networks is that they promote a co-ordinated approach to identifying, evaluating and conserving the geneticvariability of selected crop species, with the aim of improving cultivars andadapting them to farmers’ needs. The networks combine a thoroughknowledge of the agricultural conditions of farmers in network membercountries, with an understanding of the genetic potential of the crop speciesin question, and use a “farmers-to-farmers” basis for the development ofconservation and utilisation activities. That involves the collection fromfarmers of locally adapted germplasm, its improvement and its return to thesame farmers, or farmers in similar biotopes. The work of several of thesenetworks is also guided by an economic intelligence function (analysis ofmarket factors), which helps farmers to improve quality and suit theirproduction to market requirements.

The distinguishing characteristic of the networks’ components on plantgenetic resources consists in identifying the genetic variability within eachparticular crop species, and the development of methodologies forcharacterisation, evaluation and conservation of different genotypes in viewof their utilisation by present and future generw of their utilisation by presentand future generations of farmers. As such, the networks are regarded as very

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useful instruments for facilitating the process of implementing the Global Planof Action, and their expansion, in terms of both crops and regions covered, isbeing pursued.

The crop-related networks also provide a useful forum for updating theReport on the State of the World’s Plant Genetic Resources. Computeriseddata bases are being developed for several of the networks and the informationbeing gathered is available to be systematically included in the WorldInformation and Early Warning System.

The many crop-specific networks and working groups may operate atsub-regional, regional, inter-regional or global level. The networks that FAOhas developed over the last seven years, with the aim of promoting acoordinated approach to identifying, evaluating and conserving the geneticvariability of selected crop species, include the Global Mushroom GermplasmConservation Network; the Olive Genetic Variability Conservation Network;the International Network on Cactus Pear; the Mediterranean and Inter-American Citrus Networks and the Global Citrus Germplasm Network; theInter-regional Cooperative Network on Nuts; the Global Network on Tropicaland Subtropical Fruit Genetic Resources, and the Network on TraditionalCrops of Southern Africa.

THE INTERNATIONAL MUSHROOMGERMPLASM CONSERVATION

Fungi are regarded as being the second largest group of organisms inthe biosphere after the arthropods. The total number of fungal species isevaluated as 1,500,000 in the world. Only 5% of theses species are describedand catalogued. Therefore our knowledge on the worldwide fungal speciesis poor. Out of the 70% described species of fungi, there are about 10,000species of mushrooms in which about 2,000 species from more than 30 generaare regarded as prime edible mushrooms, but only a few dozen of them arecultivated. In the particular of tropical and sub-tropical areas, the fungal andmushroom genetic resources are certainly the most abundant of the worldand the least known.

This network was established to strengthen international collaborationamong specialised institutions, with a view to constituting a coordinatedglobal system of mushroom germplasm collections under the aegis of FAO,and to facilitate technical communication and the exchange of strains ofcultivated mushroom, and of germplasm of other mushroom species of interestfor food and agriculture. The network also has the aim of establishing a morecomprehensive and coordinated information mechanism, which would helpto make mushroom strains, together with appropriate productiontechnologies, available to as many interested countries and growers aspossible.

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T HE USE OF BIOT ECHNOLOGY

Through modern biotechnologies that are helping to overcome some ofthe limitations of conventional breeding, wild diversity can be more readilyincorporated into crops and thus contribute to world agriculturaldevelopment.

It is recognised that production must be intensified, productivityincreased and productive natural systems must be suitably managed all ina sustainable manner. This requires the combined application of new and oldtechnologies, including innovative approaches to plant breeding and tofarming practices. The success of such endeavours will depend on thesustainable utilisation of a broader range of species and genetic material withineach species, including the wild relatives of domestic species.

Many of the recent advances in agricultural technology, includingbiotechnologies, have been made in industrialised countries where they haveoften been developed on a commercial basis. In this sense, the potential forthe new biotechnologies to improve the livelihood and food security situationof the world’s poor is largely under-realised.

The new biotechnologies do provide excellent tools for manipulating thegenetic diversity of crop varieties and animal breeds among the alreadydomesticated species and further increasing productivity and stabilspeciesand further increasing productivity and stability. Through the developmentof plants and animals that are more adapted to ecological conditions, thestability of production can be increased. The genetic manipulation of otherbiological elements in crop production systems, such as soil bacteria associatedwith nitrogen fixation, or natural predators of pests (especially insects and tosome degree pathogens and weeds) is also a tool that can be used for enhancingsustainability in agriculture.

Ours is the first generation with the immense biotechnological power toradically change the evolutionary processes of natural and artificial selection.Ours, too, is the responsibility that goes with this power.

However it is recognised that there are also risks involved in theapplication of biotechnologies; these risks include misuse by humans, andenvironmental accidents. To address these and other issues, a draft Protocolon Bio-safety is now being negotiated by those countries which are signatoriesto the Convention on Biological Diversity. With the advent of geneticengineering, the private sector biotechnology industry has promoted theextension of industrial patenting regimes to living organisms an approachpopularly known as life patenting.As a result, genes, microorganisms and varieties of higher species (plants

and animals), either discovered in nature or genetically manipulated by erdiscovered in nature or genetically manipulated by breeders or geneticengineers, can and have become the intellectual property of individuals orindustry in countries in which “life patenting” has been legalised. Between

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1981 and 1985 at least 1175 patents for human genes were granted worldwide;and of these patent applications, some 75% were from the private sector.Perhaps for reasons guided by the market, the socioeconomic and food

security needs of the poorer sections of society, especially in developingcountries, have not yet been sufficiently targeted by biotechnologyprogrammes, or have not been adequately considered in shaping researchpolicy.

This is an important point for decision-making in relation to the allocationof public sector funding for agricultural research, since it is estimated thatthere are now at least 1,400 million people in developing countries who aredependent upon resource-poor farming systems which have not benefitedfrom technological advances in plant or animal breeding or biotechnology.

It can also be debated whether current intellectual property right systemsallow appropriate incentives for directing public sector agricultural researchso that the social and economic needs of these poor countries can be adequatelymet. This issue becomes particularly worrying when it is considered that thelimitation of access for the world’s poor farmers to improved genetics for theworld’s poor farmers to improved genetic materials could become a constraintto increased food production.

T HE FAO GLOBAL SYST EM

Within the programmes of the Food and Agriculture Organisation of theUnited Nations (FAO), the inter-related questions of who benefits from theuse of genetic resources and who owns genetic resources stored ininternational seedbanks, in farmers’ fields, or in nature, became the focal pointof international debates during the 1980s.

To confront and address these problems, the member countries of FAOagreed on the need to develop a global system which might ensure theconservation and sustainable use of plant genetic resources for food andagriculture, as well as the just and equitable sharing of the benefits andresponsibilities derived from them.

The development of the Global System on Plant Genetic Resources forFood and Agriculture began in 1983 with the establishment of its governingbody, the Commission on Plant Genetic Resources, now the Commission onGenetic Resources for Food and Agriculture, a permanent intergovernmentalforum for discussion and consensus building between countries. Governmentsalso adopted, that same year, an international nonbinding agreement, thefirst of its kind, called the International Undertaking on Plant GeneticResources, which provides the formal frameant Genetic Resources, whichprovides the formal framework for the Global System. The Commission hassince coordinated, overseen and monitored the development of a GlobalSystem for the Conservation and Utilisation of Plant Genetic Resources forFood and Agriculture.

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Countries continue to negotiate and develop, through the Commission,other important elements of the System which include various internationalagreements, scientific regulations, technical cooperation mechanisms andglobal instruments which are in different stages of development.

The availability of information about plant genetic resources of cropspecies is fundamental to being able to harness the genetic variabilityconserved in collections and critical for evaluating the usefulness o f in-situconservation initiatives to counter the risk of genetic erosion. In response tothis need, the FAO World Information and Early Warning System on PlantGenetic Resources (WIEWS) was created five years ago, within the overallframework of the FAO Global System on Plant Genetic Resources. Its primarypurpose is to facilitate access to information so as to allow a more coherentutilisation of available genetic resources for plant improvement work.

GENETIC DIVERSITY, BREEDING AND UTILIZATION OFCITRUS FRUITS

Citrus fruits are the third largest of the fruits grown in India with anestimated production of 2979.11 thousand m.t. from an area of 369.56 thousandha. The country is the home of many citrus species, namely, Citrusinchangensis, C. latipes, C. macroptera, C. assamensis, C. aurantium, C.jambhiri, Citrus limonia, C. karna, C. pennivesiculata and C. maderaspatana.The diverse ecogeographical distribution and the occurrence of spontaneousmutations and natural hybridization have given rise to a wide range ofvariability in citrus and related genera. Genetic resources of citrus in Indiahave been well reviewed.

In India, more than 500 citrus varieties are available of which, mandarin,sweet orange and acid lime are commercially grown with regional preferenceconsidering the agroecological conditions, resulting in their increased areaand production. Increase in production over the base year of 1961 is over242.59% with an annual growth rate of 7.82%. Citrus fruits thus occupy 11.53%of the total area under fruits in India. However, genetic base of cultivars isvery narrow and enrichment of germplasm needs greater attention.

DIST RIBUT ION OF DIVERSIT Y

There are three major centres of diversity. The types found include thepapeda, pummelo and their hybrids, Citrus indica and many types of citron,lemons and mandarins. These include interesting types like Nemerutenga,Soh-Systeng, a sour fruit similar to the sweet lime and Soh-Siem, a sourmandarin. Diversity of this region is well studied and described byBhattacharya and Dutta (1956).

In South India, the indigenous types include Gajanimma or Baduvapuli,Kichli and some wild mandarin types, viz., Kodakithuli, Billikichili, Nakoor

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lemon, Mole puli (sour orange type), etc. In western India, at the foot of theHimalaya, the hill lemon (galgal) and Attani are common. Citron, Citronlemon, Karna Khatta, Rough lemon, Rangpur lime, acid lime, hybrid pummeloand various types of mandarins are found all over the country.

Wild types or plants growing without any attention are largely found inthe foot hill regions of northwestern India, in the northeast, in south in Malabarhills and in the Western Ghats under varying ecological conditions.

Commercial Cultivars

Although citrus fruits have been grown in India since long, theircommercial production is only a few decades old. These grow in area whereannual rainfall ranges from 2000-3000mm as well in the regions withprecipitation of 500 to 800 mm. They are commercially grown in AndhraPradesh, Arunachal Pradesh, Assam, Bihar, Gujarat, Karnataka, MadhyaPradesh, Maharashtra, Meghalaya, Orissa, Punjab, Rajasthan, Sikkim, Tripuraand West Bengal. Among the commercially known citrus fruits, mandarintops with respect to area and production followed by sweet orange, acid lime,lemons, grapefruit, pummelo and others. The citrus species and varieties triedas rootstock.

Germplasm Collection

Exotic collection of citrus germplasm was started in 1940. One of thecollections, Kinnow mandarin is now a leading cultivar in northwest Indiawhich has replaced local cultivars. Besides Kinnow, the other exotic collectionswere Valencia Late, Washington Navel, Jaffa, Malta Blood Red, Pineapple,Shamouti, Ruby Orange, Satsuma, Dancy tangerine, Clementine, Cleopatra,Wilking, Temple, Duncan, Marsh Seedless, Lisbon lemon, Sporalime, Trifoliateorange, Inchang lemon, C. hystrix and Microcitrus australis. Large numberof exotic collections were established at IARI, Pusa; Fruit Research Station,Abohar; Citrus Experiment Station, Gonikoppal/Chethalli; and CitrusExperiment Stations at Srirampur and Tirupati during the fifties and sixties.Recently, large number of exotic materials have been introduced from USA,Italy, Spain and Japan at NRCC, Nagpur and HRS, Yercaud, especially ofmandarins, sweet orange and rootstocks. Many exotic cultivars, viz., Valencia,Jaffa and Hamlin which proved commercially successful have beenrecommended in specific regions. Cleopatra mandarin and Troyer citrangehave been found to be promising as rootstocks for mandarin and sweet orangerespectively. Tahiti lime has became commercially important in Tamil Naduand many parts of the country.

Explorations, besides saving the endangered species, help in collectingof material including wild species and the locally developed primitive andimproved cultivars for specific research needs. Northeastern region is a majorregion of diversity of citrus where collections were attempted during the later

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part of 19th century and with the turn of 20th century, surveys were carriedout for collecting of herbarium specimens and identification. First systematicattempt to explore and collect the samples from the region and conserve them,and to characterize them were made by Bhattacharya and Dutta (1956). Allthe areas of Assam province were explored in about ten years, valuable specieswere identified, specimens of both mature fruits and inflorescences with leavesand, wherever available, both seedling and clonal progenies of each specimenwere collected and maintained. Surveys were later undertaken by Singh andSingh (1967) in different parts of the country and some unknown/new specieswere identified (Singh 1967, 1977). Tanaka also explored and described Citrusspecies of India. Singh (1981) explored the northeastern region to locate sitefor in situ gene sanctuary of citrus especially for the endangered species.

Chakrawar et al. (1988) identified two promising clones of acid lime,Vikram and Pramalini, in Maharashtra. Seedless form of Santra (MudkhedSeedless) was selected from the variability of the local cultivar. Several formsof Musambi have been identified from Central Maharashtra. Singh et al. (1994)collected a thornless and seedless acid lime from Central Maharashtra. AtNagpur, seedless santra has been selected which has commercial potentiality.At Periyakulam, PKM-1 acid lime was selected which gives high yield ofquality fruits. Short juvenile Poncirus was also spotted during a survey. AtRahuri, 150 accessions of acid lime were collected with desirable traits of yield,resistance to canker and leaf miner and higher summer yield. Five of theseaccessions have potential for commercial exploitation. One of the accessionshas been selected and released as Sai Sarbati for its cultivation in Maharashtraregion. Similarly, exploration of citrus orchards have also resulted in selectionof superior clones of acid lime and sweet oranges at Tirupati and Rahuri.

During 1988, as a result of systematic exploration by NBPGR, in thenortheastern region, C. indica and many endangered species were collectedfor conservation. From another mission, 94 accessions were collected from allover Orissa and these were characterized and grouped into 12 known Citrusspecies. However, it is apparent that efforts were concentrated on selectionof commercial types and many of the species, varieties which may prove tobe the donor source of important gene and not a cultivar of commercialsignificance, are not getting due attention. Resultantly, there is a likely chancefor the loss of valuable germplasm. It is of concern that many of the valuableaccessions collected in the past have been lost due to heavy disease pressure.Therefore, it is essential to give attention for the collection of diversity whichis available in northeastern region, central India, in the Western Ghats andthe western regions.

Characterization

Characterization of citrus genetic resources was first attempted byBonavia (1890) who described the citrus germplasm available in India and

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Lushington (1910) described the Citrus species of northeastern region. Otherworkers have also described the Citrus germplasm. These studies were basedon taxonomic observation of herbarium specimen. Systematic characterizationand documentation of citrus in northeastern India was done by Bhattacharyaand Dutta (1956) following the descriptors of Swingle and studying minutestdetails of the samples. Hodgson et al. (1963a,b,c), Singh (1967, 1977), Singhand Singh (1967, 1968), and Singh and Nath (1969) also described Citrusspecies. However, these morphological descriptions could not remove theconfusion in classification but on the contrary, the divergent views complicatedthe Citrus taxonomy.

Further, the chemotaxonomic approaches have been used to confirm thecharacterization. With use of pictorial scatter diagram for characterization,Singh and Singh (1983) used numerical classification using 145 characters. Forassessing diversity, electrophoresis of proteins and enzymes has proved tobe an authentic tool in characterization of citrus germplasm at gene level.Biochemical markers may help in eliminating duplicates in genepool resultingfrom variations in phenotypic expression due to environmental conditions.

Evaluation

Citrus genepool has been evaluated for agri-horticultural characteristics.The evaluation at different locations has helped to identify cultivars/rootstocksfor different agroclimatic situations as well as identification of suitablerootstocks. Variability in mandarin and acid lime was studied to understandthe genotype × environment interaction.

Several citrus species types have been evaluated for their suitabilty asrootstocks, cultivars resistant to insect pest and disease and for their processingqualities. So, by effective evaluation, it has been possible to identify anddomesticate the exotic and indigenous collection for commercial adoption.Kinnow mandarin growing commercially in Punjab is one of the best exampleof successful introduction.

Documentation

Although documentation of genetic resources of Citrus has been donefor better accessibility of information, database with central monitoring systemis still lacking.

Utilization

There are ample evidences to show that genetic resources of citrus havebeen well utilized. In fact, the germplasm collected from exotic or indigenoussources have been the basis of commercial cultivation in different citrusgrowing regions in the country. By and large, the commercial cultivars arethe result of selection from indigenous diversity. Owing to inherent problemsof breeding, only limited attempts have been made for the transfer of desired

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genes through hybridization. However, natural variability existing incommercial cultivars or related species have been utilized. Extensive surveyof acid lime orchards in Maharashtra state resulted in selection of twocommercial cultivars viz. Pramalini and Vikram which are superior to existingacid lime cultivars. Ten clones of acid lime were assembled and evaluated atPeriyakulam for 15 years, which resulted in selection of superior types andrelease for commercial cultivation. At Rahuri, 150 accessions collected fromdifferent lime growing regions were evaluated which has enabled to identify5 superior types with respect to yield, fruit quality, bearing period and diseaseresistance. One of the accessions, Selection-49 has been released for growingin Maharashtra as Sai Sarbati.

Exotic germplasm viz. Kinnow, Seedless lime, Jaffa, Valencia Late, Marshand Grape fruit have become important commercial cultivars in different citrusgrowing regions. Unshiu introduced from Japan has also promise with its shortduration and dwarf canopy. Indigenous collection, Nepali Oblong lemon hasproved to be a best donor for imparting resistance to canker in hybridizationprogramme. The efforts have resulted in development of acid lime havingresistance to citrus canker, a serious problem of acid lime.

The reviews of rootstock situation in India suggest that Rangpur lime isthe most promising for mandarin and sweet orange in central and South India.Performance of Cleopatra mandarin has also been highly satisfactory.However in Punjab, Jatti Khatti (C. jambhiri) and Rangpur lime for Kinnow;Rangpur lime and Cleopatra mandarin for Blood Red, Jatti Khatti andCleopatra mandarin for Jaffa have shown promise. A 25-year rootstock trialsuggested Belladakithuli (C. maderaspatna) as the best for Coorg mandarinwith respect to fruit yield but was not recommended due to bottleneck growthat graft union. Thus, this rootstock needs further testing. Feronia limoniaproved to be highly dwarfing and precocious at Shrirampur and Tirupati,and is thus of interest for high density planting. Since, rootstocks currentlyemployed have one or other defect, work to develop ideal rootstock havingresistance to root rot, nematode, salinity with good agronomic characters, bycitrus breeding was started at Chethalli wherein trifoliate orange was usedfor imparting resistance.

Trifoliate orange has been found to be a donor for the resistance to rootrot and nematode. Several hybrids having tolerance to diseases and nematodealso have promising growth characters. These hybrids are being tested fortheir suitability as rootstock for mandarin and sweet orange.

Taxonomic Status

The commonly grown citrus species belong to three genera, Citrus,Fortunella and Poncirus. All these genera are closely related and have inter-generic fertility. These genera are grouped under the subtribe Citrinae, tribeCitreae, sub-family Aurantioideae and family Rutaceae.

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Divergent views on classification of Citrus have been expressed. Swingleand Reece (1967) recognised 16 species, while Tanaka (1954) proposed as manyas 145 and later 159. Tanaka (1961) classified the forms of Citrus into twosub-genera, eight sections, fifteen sub-sections, nine groups, two sub-groups,two micro-groups and 159 species. Swingle's classification had recognized thesub-genera Papeda (containing six species) and Citrus (formerly Eucitrus) withten species. Bhattacharya and Dutta (1956) published a monograph onclassification based on the study of a large number of specimens of citrus fruitscollected during 1938-48 from wide range of soil and climatic conditions upto 2000 m altitude in Assam. They followed the classification of Swingle in ageneral way, but accepted C. nobilis Lour., C. karna Raf., C. limetta (Risso)Lush., C. jambhiri Lush., C. megaloxycarpa Lush, and species of Tanaka thatwere not accepted by Swingle (1948). Their classification found C.pennivesiculata Tanaka, and C. limonia Osbeck to be invalid. They named anew species, C. assamensis Dutta and Bhattacharya, which is known asAdajamir in Assam. Validity of this species was, however, questioned by Singhand Nath (1969) as it differs from the C. pennivesiculata Tanaka essentiallyin the nature of the essential oil of peel. This was identified by Tanaka as C.medica var. alata.

Subsequently, large exotic introd uctions were stud ied by Hodgson et al.who recognized twenty three species of Tanaka. Singh (1967) comparedcharacters used by Swingle and Tanaka, and proposed a key to theidentification of Citrus compromising the two systems and recognized C.maderaspatana Tan. (kichli), C. unshiu Tan., C. deliciosa, C. pennivesiculataHort. ex Tan., C. semiperflorens and C. limonia Osbeck as valid species besidesa new species, believed to be a complex natural hybrid of acid lime (C.aurantifolia Swingle) and papeda group parentage. Being distinct with leastresemblance to any other form of citrus(it was designated as Citrus nakoorD. Singh sp. novo).

Considering the complexity of classification based on morphological datawhich is difficult to comprehend in terms of pattern of variation arising dueto natural hybridization and mutation, numerical taxonomy has beenattempted. It has been found eminently suitable in assessing the similaritiesbetween different species for their classification into groups and theirphylogenetic relationship. Use of pictorial scatter diagram in clarifying theposition of hybrids, probably of bispecific origin, is also advocated. Polygonalmethod of presentation (after Hutchinson) of morphological data was founduseful in characterising different citrus types. To confirm this, chemicalconstituents have been proposed as additional tools. By combination ofmodified almen test, ammonium molybdate test and ferric chloride test, itwas possible to distinguish between Citrus limon and C. jambhiri, C.limettioides and C. aurantifolia; C. grandis and C. paradisi. Usefulness offlavonoid composition of leaves, bark and ripe fruit peel in identifying the

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taxa of doubtful origin was also emphasised. Chemotaxonomic studies wereemployed to study the variability in lemon, mandarin and related citrus types.Study of 145 morphological characters, affinity relationships, chemical dataand breeding behaviour favoured the contention that only three species, C.medica, C. grandis and C. reticulata with a sexual reproduction system are inEucitrus group.

Prasad (1995) studied 71 cultivars belonging to 31 species for geneticdiversity and genotype affinity using D2 analysis and leaf peroxidase isozyme.The genotypes exhibited wide variability for morphological characters.Numerical analysis and polygraph suggested some affinity at intra-specificlevel. Very narrow relationship between Rangpur lime, to very widerelationship between pummelo and sour orange revealed the broad nature ofgenetic divergence in citrus. Based on minimum generalized distances, the71 genotypes fell into four distinct clusters. Leaf peroxidase isozyme bandingpattern proved the genetical differences and distinct genotypes. The isozymesresolved the differences between seemingly similar types of citrus, particularlyat intra-specific level. He also noted the close affinity among mandarin orange,sweet orange, acid lime and Rangpur lime inspite of enormous diversity.

Conservation

Germplasm is maintained under different agroclimatic conditions atCentral Horticultural Experiment Station, Chethalli, Karnataka; IndianInstitute of Horticultural Research, Bangalore, Karnataka; Regional FruitResearch Station, Abohar, Punjab; Horticultural Experiment Station, Bhatinda,Punjab; Division of Fruits and Horticultural Technology, IARI, New Delhi;Department of Horticulture, Rahuri, Maharashtra; Citrus ImprovementProject, Tirupati, Andhra Pradesh; Citrus Experiment Station, Katol,Maharashtra; Horticultural Experiment Station, Periyakulam, Tamil Nadu;Citrus Experiment Station, Tinsukia, Assam and others. At these centres,germplasm is maintained as working collections.

Citrus germplasm is conserved in field genebanks at different locations.Though substantial diversity is conserved in field genebanks, it does notcontain total representation of the variability available in citrus. Many of thegermplasm collected by Bhattacharya and Dutta (1956) from northeasternIndia are currently not available in any of the collections. At the same time,many of the germplasm are safely duplicated at few centres owing to the factthat germplasm of one region behaves differently at other region.

In Vitro Conservation

Maintenance of citrus germplasm in field genebank requires large areaand funds, and plants are also affected by insect pests and diseases. Therefore,in vitro conservation of the citrus germplasm may require attention. In vitroconservation technique is also used for revitalization of citrus germplasm

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affected by virus and virus-like diseases as meristem culture technique isfound to eliminate many of the viruses. However, to conserve the germplasmin vitro, there is need for the research with respect to genetic stability andduration of storage, etc. Proliferation in callus was observed even after severalyears by sub-culturing in medium supplemented with 5 per cent sucrosealongwith 2,4- D, NAA and kinetin each of 0.25 mg l-1. In vitro root tip culturescontinued to retain the ability to form shoot buds which were maintained at25°C for 8-10 months and regenerated to plants.

BIOT ECHNOLOGY, GENET IC ENGINEERING AND T HE SEED

“Biotechnology,” deals with live organisms. These may be plants oranimals, visible to the naked eye, or microorganisms seen only through amicroscope. Agricultural biotechnology—our focus here, encompasses variousmethods used to modify (or create new or modified) plant life. Such life formsmay range from simple one-celled algae or complex multi-celled trees. Asearlier noted, research scientists in this field use biological processes involvingorganisms or natural substances to develop or modify products. Their aim isto change the genetic code of living cells or seeds which is carried in themolecule known as DNA (deoxyribonucleic acid). This is the process we call“genetic engineering.”

DNA is composed of two alternating chemicals joined together by twoother base chemicals. The sequence of the latter determines the instructionsthat direct cells to make the various proteins that support life. By manipulatingthis sequence, corporate scientists can enable these cells or seeds to performnew functions or to produce new substances. This is done by decoding the orderor “sequencing” of a gene’s building blocks (i.e., its chemical units), and thenchanging that order. The latter process is known as “splicing”. Through thisseries of steps, scientists can add or remove traits from a plant that aredesirable or undesirable. They can make crops resistant to drought orpoisonous to pests and weeds. The process of adding or removing traits to orfrom plants becomes most controversial when it is done across species. Thatis, foreign genetic material is inserted into the cells of organisms of a differentspecies. This is how “transgenic organisms” are formed. The product may besomething as simple as a carrot containing milk proteins or as complex as agenetically manipulated animal and even human forms.

No one doubts that biotechnology’s potential for good is immense — inboth medicine and agriculture. Progress has been made in finding medicinesthat can cure heretofore fatal diseases. Methods of “gene therapy” are alreadyin use whereby a “sick” gene is replaced by a healthy one. But in addition tocurative ends, experiments may be undertaken for eugenic purposes, e.g., todetermine the height, weight, hair colour and other traits of a fetus. Indeed, apopular test in India and elsewhere enables parents to discard unwantedfemale fetuses until the prized male fetus materializes. A patent pending in

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the U.S. patent office proposes to genetically change sperm cells so that theresulting altered animals can pass down the new traits to subsequentgenerations. Such experiments in animal eugenics will undoubtedly pave theway to human eugenics. Hitler’s dream of a “superior” race may be realizedwithin a century of his death

The industries most active in these kind of genetic engineering methodsare in the biotech, pharmaceutical and chemical fields. The top companies inthese industries include, in addition to Monsanto, DuPont, Bayer, Upjohn andothers. Once they have identified and patented plant genes (as well as animal),they will be able to control agriculture and the increasingly powerful foodprocessing industry which manufactures much of the food products that weput on our tables, especially in the Western world. But as these giant TNCsexpand their operations and with that their influence, in the economies ofThird World countries, their agricultural production and food consumptionpatterns will undoubtedly reflect that influence and power. For they will haveacquired patent monopolies of not only the “discovered” genes but also ofthe genetic engineering methods used to manipulate these genes. Inherent inthis kind of experimentation is the familiar dilemma of humankind of “lettingthe genie out of the bottle.” It’s difficult to put it back in. This dangerousscientific game with genes will create transgenic forms that will not onlyreproduce, but in the process of reproducing could be transformed, and havingbeen transformed, could literally move on — by wind, rain, pollen and othernatural ways, to other crops or to wild plants. Eventually, these novelorganisms could displace the existing forms, homogenizing agricultural life.

It is safe to say that it is in the agricultural field that genetic engineeringis most popular. The easily anticipated reason is the profits it promises.Corporate scientists in agribusiness stress the remarkable increase in yieldthat they claim will end starvation and malnutrition. But as with any newtechnology—from the wheel to nuclear power—there is a fine line betweengood and evil. It is that fine line that is stressed by critics who fear that theevil consequences may overwhelm the good. Whether they are pessimists, assome say, or realists, as viewed by others, the end results they foresee do notbode well for humanity and especially for the “wretched of the earth.”

Since the bulk of research money comes from agribusiness, the researchwill focus on foods that sell well in the DC markets. As has already happenedin many countries in the South, arable land is increasingly allocated to cashcrops that can be exported and earn foreign exchange. Moreover, farmers whoused to grow several products are now inclined to grow one or two crops onthe same large landholdings. The promise of huge yields from geneticallyengineered seeds lures even the small-scale farmers who make up the bulk ofthe farming community in the Third World. But once hooked on such seeds,they are locked into the contract they were required to sign when they boughtthese seeds. Cargill “generously” gave free hybrid seeds to farmers in the

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Philippines. Instead of the increased yields and lower use of chemicalspromised by Cargill, quite the opposite happened. The seeds were not suitedto the moist climate of the Philippine Islands, and more pesticides, weed killersand fertilizers had to be applied. Worse still, owing to the restrictive terms ofthe contract signed, they were prevented from using their neighbours’ seeds,as traditionally done when a crop failed. Even worse was the contract sold toIndian farmers by Monsanto which demanded a hefty ‘technical fee’ per acrein addition to the amount paid for the seed. But they were also locked intoMonsanto’s chemicals as well, and were subject to a fine if they used someoneelse’s chemicals. Activist Vandana Shiva’s description of Monsanto’s methodsas totalitarian is clinched by the provision that allows Monsanto inspectorsto visit these farmers (even in the absence of the land owner) in order todetermine whether they have violated any of the terms of the contract.

Beneath a façade of official concern about the effect of invading “seedgiants” on Indian agriculture, there is a lot of support on the part of Indianagribusiness and allied interests with ties to outside agricultural ventures foropening up India to foreign agribusiness. It is argued that such ties will bringthe technology and capital needed to make Indian agriculture more productivethan it is now. Mixed signals have come from Seattle regarding the IndianDelegation’s policy stance during the doomed Conference. What bearswatching is the evolution of the BJP Government’s agriculturalpronouncements in the aftermath of the Seattle debacle. Thus far, these havenot been reassuring. The status of a proposed law, the Protection of PlantVarieties and Farmers’ Rights Bill, remains unclear. It has been reported bythe Press Information Bureau of India that this Bill was introduced in the LokSabha on December 3rd. But the same agency announced on December 15,1999 that this Bill was “likely to be introduced shortly in Parliament. “Nevertheless, the draft that has been circulated has raised more questions thanit has answered. One critic states that this bill “will end up benefiting thelarge seed corporations, some very large farmers, and corporate farmingagencies.” Such a Bill, if enacted, may well signal the nature of relations wecan anticipate between India and powerful industrial nations like the UnitedStates — the principal architect of the new corporate order. Will India submitto “the inevitable” or will it stand and fight?

KEEP OUT PAT ENT S ON CONVENT IONAL SEEDS AND ANIM ALS

For several years, patents on genetically modified seeds and animals havebeen granted worldwide. The damaging impacts on farmers, who are deprivedof their rights to save their seeds, and on breeders who can no longer use thepatented seeds freely for further breeding, are well known.

In Canada and the U.S., for example, the multinational seed companyMonsanto has sued many farmers for alleged patent infringements. The samecompany has also filed court cases against importers of Argentinean soy to

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Europe. Furthermore, the possibility of patenting seeds has fostered a highlyconcentrated market structure with only 10 multinational companiescontrolling about half of the international seed market. Many farmersorganisations and NGOs around the world are fighting against these patents.Because genetically modified organisms (GMOs) are still not grown in mostcountries, or only used in a small number of crops, the negative impacts ofthese patents are not being felt everywhere.

However, there is an alarming new trend for patents not only to beclaimed on GMOs (such as Round-up ready soybeans), but also onconventional plants. For example, patent claims have been made for soy beanswith a better oil quality covering parts of the plant genome when used inconventional breeding and technologies to improve conventional breeding(such as marker assisted breeding).

Some of the most threatening examples in this context are patentapplications from Syngenta which claim huge parts of the rice genome andits use in breeding of any food crops that have similar genomic informationto rice (such as maize and wheat).

The European Patent Office has also granted a patent on aphid resistantcomposite plants which are based on marker assisted breeding. Other recentpatent applications by Monsanto on pigs are also related to normal breedingmethods, indicating the increasing danger of agricultural genetic resourcesbecoming monopolised by a few multinationals on a global scale.

Soon the Enlarged Board of Appeal of the European Patent Office willdecide on another patent of this kind - for a method of increasing a specificcompound in Brassica species.

This decision will determine the patentability of conventional seeds inEurope.

Whereas patents on conventional plant varieties are normal practice inthe U.S., many other countries, especially developing countries, do not grantpatents on plants or animals. But as the recent history shows, the standardsdefined and used at the European, Japanese and U.S. patent offices influenceinternational regulations (the WTO agreement on trade related aspects ofintellectual property rights, TRIPS, and the World Intellectual PropertyOrganisation, WIPO). Patent offices all over the world are pushed to adapttheir regulations and practices either through the international regulationsor by bilateral agreements. India, for example, has just passed a third patentamendment in order to adapt its law to the TRIPS regulations.

This frightening new trend in patent policy will affect many more farmersand breeders, than has been the case with GMO patents. Any remainingfarmers rights and breeders’ access to plant varieties and animal breeds forbreeding purposes, will disappear everywhere. These patents will destroy asystem of farmers’ rights and breeders’ privileges that has been shown to becrucial for the survival of farmers and breeders, for food sovereignty, and for

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the preservation of bio-diversity in agriculture. The vast majority of farmersin developing countries are small-scale farmers, completely reliant on savingand exchanging their seeds.

In order to secure the continued existence of independent farming,breeding and livestock keeping and hence the food security of futuregenerations, we, the undersigned farmers, researchers, breeders and civilsociety organisations from all over the world, restate our rejection of anypatents on life, and urge policy makers and patent offices to act swiftly tostop any patents being granted on conventionally bred plants and animalsand on gene sequences for use with conventional breeding technique, as wellas on methods for the conventional breeding of plants and animals. We alsourge companies not to apply for any patents of this kind.

Savage Garden: Slack-Potting the Dewy Pine

Over twenty years ago, British nurseryman Adrian Slack originated arather clever way to successfully grow the often temperamental dewy pine(Drosophyllum lusitanicum). Glaringly omitted from my own book, The SavageGarden, I wish to correct that oversight here, for this method, called “Slack-potting”—a term coined by Barry Rice—is an ingenious method of cultivatingDrosophyllum with long-term ease.

We must remember that dewy pines are not wetland plants like mostother carnivores. Instead, they are typically found on sandy, gravel slopesthat are dry most of the year, and usually alkaline in pH. The plant is foundin widely scattered sites in Portugal, Spain, and Morocco. The climate is warm-temperate and Mediterranean, which means that most of the rainfall occursin the cool winter months, with very rare frosts. Summers are warm and dry,but plants near the coast can experience early morning fog drip, which maybe absorbed by their leaves.

Seed is usually the only way to propagate Drosophyllum, and since plantsdespise root and stem disturbance they are never sold through the mails. Seedis usually produced in late spring or early summer and can germinate readilyif very fresh. Older seed benefits from several methods of pretreatment. Youcan scarify the hard seed coating by rubbing the black, egg-shaped seeds withsandpaper until the whitish interior becomes slightly visible. You can alsosoak the seed for a day or two in a cup of water in which you have added atiny bit of powdered gibberellic acid—about 0.5 cm (1/8 inch) collected onthe tip of a toothpick works well. I have also scarified the seed and then soakedthem in a solution of one-drop Superthrive™ per cup of water.

Slack warns against moving freshly germinated seedlings about, andprefers to sow a few seed directly in their permanent pots, allowing only oneplant to grow by removing any others that germinate. This can be a hassleand waste of seed if you have more than just a few to sow, but Slack’s warningis legitimate since the first tiny root to appear from a seed will rapidly plunge

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into the soil medium and can be killed if disturbed. I prefer to sow numerousseed on damp vermiculite (they will germinate on any damp substance, evena sponge) and I check them daily after a week or two.

Now this is where the Slack-potting begins. Take a 10-15 cm (4-6 inch)pot made of unglazed terra-cotta clay. Place a wad of damp, long-fiberedsphagnum moss through its drainage hole, totally blocking it. As a soilmedium, I use one part each perlite, vermiculite, and horticultural sand to beexcellent. Slack includes a portion of John Innes Compost #2 (a humus-basedhouse plant soil), but I find this component to be unnecessary. (It is onlyavailable in Britain.) Dampen the mix with purified water and fill the claypot firmly with it to the brim. Gently place your germinated seed on the centresurface of this, one seedling per pot. Place the pot in a shallow water tray ina sunny place with good air circulation, keeping the soil damp to wet forseveral weeks as the plant establishes itself.

Let the baby plant grow for several weeks, as Slack recommends, beforeproceeding with phase two. The plant will grow rapidly and when it hasseveral leaves you are ready to Slack-pot it.

You will need another pot with drainage holes at least two inches largerthan the pot containing the plant. Slack recommends another clay pot, but inhotter, drier climates like California, I use plastic. Block the drainage holeswith long-fibered sphagnum. Fill the pot with the perlite-vermiculite-sandmedium only as deep as to allow the smaller potted plant to sit comfortablyon the medium and have its rim at the same level (or just slightly above) asthe larger pot. In the gap between the two pots, tightly pack long-fiberedsphagnum.

The whole reason for this elaborate double-potting is to keep the soilaround the stem and upper roots on the dry side, or just barely damp, sincethe prostrate stems tend to rot if left on permanently wet soils. Initially youcan keep the whole double pot in a shallow tray of water as well as gentlywatering the plant itself, keeping the soil damp. But in a very few months theroots of your dewy pine will enter the soil of the larger pot through thesphagnum-crocked hole of the smaller. You can then water the plant via thesphagnum-moat that separates the two pots. Alternatively, you can set theSlack-potted plant in a shallow water tray, and water by the tray system onlyenough to keep the sphagnum-moat damp. Just enough moisture willpermeate the inner clay pot to keep your dewy pine happy, and the deepestroots will find plenty of moisture in the medium of the larger container. Thisprobably recreates the natural situation well, for in the wild dewy pines haveroots that are extensive, obtaining moisture from deep underground. Somealso believe the plants absorb moisture in summer from condensing earlymorning fog, but misting the plant is not required. Slack-potting a dewy pineworks best for greenhouse cultivation, particularly in humid climates withlong, cool winters, such as Slack’s own England. It is not quite as necessary if

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you live in a Mediterranean climate similar to the dewy pine’s. I have seenphotographs of huge Drosophyllum plants grown in giant clay pots outdoorsin Australia. Geoff Wong’s prize winning plant photographed in The SavageGarden was a judiciously watered specimen grown in a plastic pot of primarilyperlite and a tiny bit of peat, then placed in that lovely glazed ceramicornamental. I myself grew one outdoors on my deck in northern California,using a plain clay pot. The plant survived months of winter rain, a hard freeze,and baking summer heat. So while not for everyone, for many growers Slack-potting is ideal. At my nursery just this past week we Slack-potted severaldewy pines. In my imagination, I visualized Adrian visiting us in his whitesuit and Panama hat, nodding his approval as we shared a glass of wine.

Experiments with Highland Nepenthes Seedlings

One of the most difficult things to describe to someone is how to grow aplant that is restricted in its tolerances. Partly to blame is our human perceptionof these conditions. What exactly is “bright light”? What is “clean water” andhow acidic is “acid”? Highland Nepenthes are often one of the more perplexinggroups to work with. What seems to work for one person fails when someoneelse tries to reproduce the cultural environment. Nepenthes villosa, for example,is not an easy plant to grow from seed; it has a narrow range of conditionsnecessary to grow well. Beans are easy to grow, highland Nepenthes are not.

In order to get better insight into the needs of several species, I conductedfive years worth of experiments aimed at quantifying their needs. Instead ofsubjective terms, I used tools to measure environmental conditions in all testgroups. A light meter, conductivity meter, pH meter, recording thermometersand a high quality humidity meter were employed to put some numbersbehind the findings. (Cheap meters are dangerous to rely on, so only metersthat could be calibrated and tested were used.) This way, anyone could nearlyduplicate the conditions that were successful, and avoid potential disaster bymeasuring variables before plants get put into a bad environment.

Th e fo llow ing Nepenthes were used in this study: N. burbidgeae, N.edwardsiana (Tambuyukon type), N. fusca, N. stenophylla, N. tentaculata (Mt.Tambuyukon type and Mt. Kinabalu type), and N. villosa. All were raised fromdonated seed or very tiny seedlings (cotyledon spread 0.63 cm., 0.25 inch).Dead or struggling plants were considered indicators of exceeding the speciestolerance to one or more variables within their environment. The bane andbeauty of using seedlings is that they do not take long to indicate improperenvironments; they die quickly.

Several points deserve mention, before getting into the “meat” of thischapter. First, plants grow in a system, where the elements of culture play offeach other. For example, higher light levels require greater nutrient levels, asplants need more nitrogen to eliminate photosynthetic (waste) byproducts.Stresses from lower relative humidity might be offset by lower light or more

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available moisture at the root zone. Thus, I am not suggesting one rigidmethod, when explaining a successful technique. My intention is to givebenchmarks and warning signs.

Second, this chapter was written as a distillation from five years (1996-2001) worth of records. A full description of these experiments would fillseveral volumes of this newsletter with tedious detail. My intention is topresent a simplified summary of what happened during these experiments.One example of this practical focus was to simplify the complex science oflight. The usual grower purchases a readily available light source or usessunlight. The intensity of the illumination striking the plants is determinedby the light source, the distance from the source to the plants, and/or theamount of shading. The simplest way to get a reasonably accuratemeasurement of intensity is to use a light meter, specifying the unit of measureand the source (very important). For spectral analysis of artificial lights usedherein, contact the manufacturer.

Finally, it is important to note that, unlike formal experimental design,there were no control groups, per se. In many of these species, seedling deathis the norm. Other growers, who worked with many of the same seed lots,reported complete losses in difficult species like N.edwardsiana, and N. villosa.Thus, it was nearly impossible to assign a control group without first figuringout how to keep it alive! The main support for these findings is thatsignificantly large numbers of plants were used. The results herein arepreliminary and not conclusive; other factors (such as bacterial infection, seedviability, genetics of the donated seed, etc.) could have had an effect on thesefindings. Readers should consider this chapter a detailed summary by aprofessional horticulturist, rather than a classic, formal experiment. As far asI am aware, this is the first time that meters have been used to quantify themajor (growing) environmental parameters, and that these measurementshave been united in print.

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4

Horticultural Biotechnology inTissue Culture

DEVELOPMENT OF PLANT BIOTECHNOLOGY

The development of plant biotechnology has been related to the discoveryof the growth regulators and their progressive introduction in tissue culture.In 1934, in the States, white was producing continuous growth of in vitrotomato roots. After that, went and Thiman’s work on auxin, Gautheret inFrance (1935) and white in US (1938) were auxin, Gautheret in France (1935)and white in US (1938) were able to obtain division of isolated cells from Salixand tobacco, and finally to maintain undefined cell lines.

In 1957, Skoog and Miller demonstrated that 6-furfurylaminopurine, acytokinin isolated from a DNA hydrolysate, could induce some caulogenesis,but also that the cytokinin/auxin balance could start different morphologicalprogrammes. A ratio higher than one unit induced caulogenesis, a ratioinferior at this unit induced roots. On this basis, a multibillion dollar industrywas created. Hundreds of small and large nurseries and biotech labsthroughout the world are propagating in vitro more than 1000 different plantspecies. This micropropagation technique offers not only means for masspropagation, but also plays an important role to conserve elite or rare plantsthat are threatened with extinction. Moreover, meristem tip culture offers thepossibility of virus eradication. These technologies started about 20 to 25 yearsago, especially for ornamental plants, and for a few other crops: potato,strawberry, bananas. Propagated clonally, it was very important to have atrue to type propagation technique.

On the other hand, it’s also possible to use tissue or cell culture to increasegenetic variability. Undifferentiated cells obtained from callus, cells orprotoplasts culture are produced and submitted to an selective pressure toimprove a and submitted to an selective pressure to improve and fix thesomaclonal variants.

HORTICULTURE RESEARCH

India has a wide variety of climate and soil on which a large range ofhorticultural crops such as, fruits; vegetables, potato and other tropical tubercrops; ornamental, medicinal and aromatic plants; plantation crops; spices,cashew and cocoa are grown. After attaining independence in 1947, majoremphasis was laid on achieving self sufficiency in food production.Development of high yielding wheat varieties and high productiontechnologies and their adoption in areas of assured irrigation paved the waytowards food security ushering in green revolution in the sixties.

It, however, gradually became clear that horticultural crops for whichthe Indian topography and agro climates are well suited is an ideal methodof achieving sustainability of small holdings, increasing employment,improving environment, providing an enormous export potential and aboveall achieving nutritional security. As a result, due emphasis on diversificationto horticultural crops was given only during the last one decade.

RESEARCH INFRAST RUCT URE

The Indian Council of Agricultural Research is the premier agency whichpioneered systematic research on agricultural crops in the country.Horticulture research in India received very little attention till the 3rd FiveYear Plan.

The establishment of the Indian Institute of Horticultural Research atBangalore and starting of eight All India Coordinated Crop ImprovementProjects to cover different horticultural crops was a landmark in the historyof horticulture in 4th Five Year Plan (1969-74). Rapid expansion ofinfrastructure took place in 7th and 8th Plans. Today, the horticultural researchin the country is being carried out at eight ICAR institutes (with 26 regionalstations), 10 National Research Centres (on major crops) and a ProjectDirectorate on Vegetable crops.

Area specific, multi-disciplinary research is also being conducted under14 All India Coordinated Research Projects each on Tropical, Subtropical andArid Fruits; Vegetables, Potato, Tuber Crops and Mushrooms; OrnamentalCrops, Medicinal and Aromatic crops; Palms, Cashew, Spices and Betel vine;and Post Harvest Technology at 215 centres located at various researchInstitutes, and State Agricultural Universities.

In addition, four net work projects each on hybrid research in vegetablecrops, drip, irrigation in perennial horticultural crops, protected cultivationof ornamental crops and Phytophthora diseases of horticulture crops are nowin operation. Research on horticulture is also being undertaken at severalmulti-crop, multi-disciplinary Institutes. Departments of Horticulture in 24Agricultural Universities, one deemed to be University and one full fledgedUniversity of Horticulture and Forestry are also engaged in horticultural

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research. Besides 280 adhoc schemes supported from Agriculture Produce CessFund and a number of foreign-aided projects have also been in operation onspecific problems of different horticulture crops. As a result, the country nowhas a sound research infrastructure in horticulture to meet the growing needsand expectations of the fast developing horticulture industry.

Budgetary Support

The investment in horticulture research by the ICAR in the Central sectorhas increased significantly in the last two Plans. The Plan allocation forhorticultural crops started in 4th Plan (1969-74) with a modest allocation ofRs. 34.78 million and was enhanced to Rs. 319.56 million in the 7th Plan (1985-90) and to Rs. 1047 million in the 8th Plan (1992-97). Non-Plan expenditurealso increased from Rs. 73.55 million in the 5th Plan to Rs. 768 million in 8thPlan.

Overall increase in Plan investment in 25 years has been of the order of2775.21 per cent. The per cent budget allocation for horticulture research outof the total budget for agriculture research rose from 6.1 in 5th to 6.5, 6.67and 7.7 in 6th, 7th and 8th five year plans, respectively. Similarly, expenditurefor Central Sector Schemes of the Department of Agriculture & Cooperationfor horticulture crop development also rose tremendously from Rs. 20.5million (4th Plan) to Rs. 76.18 million (5th Plan), Rs. 146.37 million (6th Plan),Rs. 250 million (7th Plan) and Rs 10,000 million (8th Five Years Plan).

Table : Budgetary Support (Million Rupees)

Five year

plans

Total for Agriculture

Research

Share of Horticulture

(%)

Total Development

Support

IV 1969-74 6105 N.A 20.50

V 1974-78 7292 6.10 76.18

VI 1980-85 10684 6.50 146.37

VII 1985-90 8445 6.67 250.00

VIII 1992-97 15165 7.70 10,000.00

Manpower

Nearly one sixth of the total strength of 5906 scientists working in ICARis allocated for horticulture research in ICAR Institutes. Besides,approximately, 560 scientists are working in State Agricultural Universitiesin ICAR funded All India Coordinated Projects. In addition, a large numberof scientists are working on horticultural crops in State AgriculturalUniversities.Total Scientists in ICAR 5906

Scientists working in the Division of Horticulture 832

Scientists working in other ICAR Institutes 150

% of Total 15

Scientists working in SAUs in ICAR funded projects 560

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Research Achievements

Several new crops have been introduced for commercial cultivation, e.g.:• Kiwi fruit in sub-mountain areas of North India.• Olive in mid hills of North Western Himalayas.• Low chilling stone fruits in the North Western plains.• Oilpalm in coastal states of Karnataka, Andhra Pradesh, etc.• Gherkin in south and west India.• Baby corn and sweet corn in certain specific pockets.• Broccoli, Brussels’ sprouts, asparagus, celery, parsley near the cities.

Crop Improvement

• A large number of high yielding varieties developed in severalhorticultural crops e.g. fruits (76), vegetables (160), potato (29), othertuber crops (24), ornamental crops (300), palms (20), spices (51),cashew (33) and betel vine (1).

• First seedless variety of mango developed.• 40 F1 hybrids developed in brinjal, tomato, chillies, cauliflower,

carrot, capsicum and muskmelon.• Self incompatible lines in cauliflower, gynodioecious lines in

cucumber and muskmelon, genetic male sterile lines in tomato andtemperature tolerant strains of button mushroom developed.

Propagation of Quality Planting Material

• Standardized propagation technique for many fruits hithertopropagated by seed. e.g., aonla, bael, ber, black pepper, cardamom,cashew, cassia, cinnam on, clove , custard apple, jack fruit, jamun,nutmeg, sapota and walnut.

• Standardized Seed Plot Technique resulting in successful diseasefree potato seed production in the tropics and sub tropics of thecountry. Standard ized m ethod of m icro-propagation and in vitromicro-tuber production in potato.

• Identification of suitable parental lines for production of True PotatoSeed and Standardized technology for raising commercial crops.

• Micropropagation protocols developed in banana, black pepper,betel vine, cardamom, ginger and turmeric.

• Production of coconut hybrids through establishment of Seedgardens of Tall (T) x Dwarf (D) and D x T hybrids.

• Standardized rootstocks in citrus, grape and apple.

Agrotechniques

• Standardized high density plantations in banana, citrus, mango andpineapple and high production technology in several crops e.g.,pineapple, black pepper and cardamom.

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• Year round production technology in tomato and “off season”cultivation of onion and cauliflower developed.

• Standardized use of several plant growth regulators and chemicalsnow commercially employed in production and qualityimprovement of horticultural crops e.g., paclobutrazol for inductionof flowering in mango; gibberellic acid for improving berry sizeand quality in grape; Maleic hydrazide for preventing sprouting inonion and potato; Dormex for hastening bud burst in grapes; andBoron and Calcium for changing flower cycle in some cucurbits.

Crop Protection

• Developed improved disease detection techniques such as ELISAand ISEM for improving seed quality and tissue culture techniquefor rapid multiplication of potato.

• Developed IPM for fruit borer in brinjal, diamond back moth incabbage, thrips in chillies, phytophthora foot rot in black pepper,“Katte”, rhizome rot in cardamom, rhizome rot in ginger, late blightand bacterial wilt in potato.

• Developed biological control measures for mealy bug in grape, fruitborer in tomato and okra.

Post Harvest Management

• Preharvest treatments to control post harvest losses in citrus, mangoand grape standardized.

• Chemical treatment for regulation of ripening in mango, sapota andbanana standardized.

• Optimum storage temperatures worked out for several fruits,vegetables and tuber crops.

• A mango harvester, fruit peeler, hand and pedal operator cassavachipping machine, harvesting tools (5-14 times efficient); Implementsfor mechanization of potato cultivation e.g., oscillating tray typepotato grader, fertilizer application cum line marker, potato culti-ridger, soil crust breakers, potato digger and automatic potatoplanter/diggers developed.

• Low cost environment friendly storage system for fruits, vegetables,potato and onion developed.

TISSUE CULTURE TECHNIQUES

SOM ACLONAL VARIAT IONS

The production of plantlets by callus regeneration, cell suspensions,protoplast cultures, could present some deviations with regard to the mother

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plant. This is a way to increase the genetic variability. Associated with aselective pressure (stress to toxins, pH, salinity, cold) it’s used to obtainresistant lines. Indeed, after regeneration, plants can express new potentialitiesrarely obtained another way. Stable and profitable variants are sale anotherway. Stable and profitable variants are selected and introduced in breedingprogrammes. In 1976, a Pelargonium cv Velvet Rose was created by thistechnique. Later, other applications were described, as in sugarcane, tomato,red pepper. Following Sibi (1994), extrachromosomic systems must beinvolved to explain the genetic behaviour of some new expressed tissue culturecharacteristics. She uses the term “epigenic” to evoke a whole of hereditaryelements which belongs to the cytoplasmic or nuclear compartments, but arenot transmitted by mendelian rules.

Haplodiploidy Station

Many species are able to produce haploids through different in vitrotechniques. The oldest was anther culture of androgenesis. To date, the resultsvary considerably from one species to another. Solanae (datura, tobacco, redpepper, eggplant, petunia), cereals (wheat, barley, rice, triticale, maize) orcrucifers (soybean, cabbage) are species easy to regenerate by anther culture.To the contrary, tomatoes, leguminous, or Compositae are recalcitrant.

Ovule culture, or gynogenesis, was a successful technique for Gerbera,beet, courgette. However, the most common technique is pollination withirradiated pollen, in order to induce in vivo parthenogenesis. The oospheresdeveloped in embryos without fertilisation are saved by embryo rescue.

The haploid plantlets (n chromosomes) are tree. The haploid plantlets (nchromosomes) are treated with colchicine to produce fertile homozygous lines,called “double haploid lines”. This haplodiploidisation technique could giveimmediately new elite genotypes, hybrid parents after in vitro propagation,as asparagus supermales (MM), or useful genetic material to establish genemapping. A good synthesis book concerning all these possibilities waspublished by Bajaj (1990).

PROT OPLAST CULT URE

Protoplasts are the smallest units able to regenerate a whole plant.Therefore protoplasts cultures can serve to enlarge genetic variability byintroducing somaclonal variation. However, the main interest of protoplasts,is their capacity to fuse and to produce hybrids or cybrids, new organismsmost often unknown in nature. Indeed, protoplasts allow to transgress thebarrier of the botanical species or genus. Naked protoplasts can accept withoutrejection external elements: nuclei, cytoplasmic corganelles, liposomes,containing genetic information. The first protoplast fusion application was acytoplasmic transfer from one genotype to another to induce male sterility(CMS) from mitochondrial origin. These male sterile hybrids are interesting

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to produce F1 hybrids. Another cytoplasmic transfer concerns the introductionof chloroplastic DNA to induce herbicide (atrazine) resistance, from Solanumnigrun duce herbicide (atrazine) resistance, from Solanum nigrun to tomatofor instance.

Many intergeneric and interspecific hybridations are possible but notalways profitable because these hybrids are too often infertile. This problemdoes not exist if we can vegetatively propagate the hybrids, as in the case ofCitrus rootstocks. In Florida, Grosser (1990) was able to analyse severalthousand Citrus hybrids.

CONTRIBUTIONS OF MOLECULAR BIOLOGY INHORTICULTURE

More and more pathologists are using molecular probes for earlydetection of diseases. However, the main contributions of modernbiotechnology remain probably in the hands of the plant breeders. Indeed,molecular markers are of great use in detecting desirable new genes, or toidentify important QTL. This marker-assisted selection is very useful for manyvegetable plants. Other applications are DNA - fingerprinting and geneticengineering. This recent technology aims to insert and to express new specificgenes into a selected plants.

M ARKER-ASSIST ED SELECT ION

Molecular markers and genome mapping will reach a large extension inthe next breeding programmes of tomatoes, peas, cabbages, melons, potatoes,

It will be possible to improve the speed and the incorporation efficiencyof desirable new genes by utilizing of closely linked selectable molecularmarkers genes by utilizing of closely linked selectable molecular markers. Thenumber of backcross will be reduced. Tanksley et al. (1981) show that onlywith 12 markers, one on each tomato chromosome, the composition of therecurrent parent genome is similar to that observed in the absence of selectionafter the third backcross two years later. Therefore, it’s easy to understandthe high improvement that we can get with the 700 tomato markers alreadyknown in 1990 for this crop.

The utilization of molecular markers could also identify importantquantitative trait loci (QTL). Recently Foolad and Chen (1998) have identified13 RAPD markers at eight genomic regions, that were associated with QTLsaffecting salt tolerance during germination in tomato.

DNA Fingerprinting

The DNA polymorphism observed by RFLP, AFLP, or RAPD, allowscultivar identification of fruit trees, apple, citrus, or other vegetativelypropagated plants. Although some mutants, as coloured fruits, escape these

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identification techniques, some people would evaluate the genetic uniformityof regenerated plants in this way.

Genetic Engineering

Genetic engineering consists of introducing a foreign gene into a plantgenome to create a new function. Or at the contrary, it could be to reduce orsuppress an existing gene function, as in the antisense strategy.

1. Historic: In this field, progress were very rapid. In 1974 thepathogenicity of Agrobacterium tumefaciens, due to a large plasmidcalled Ti, was demonstrated. In 1977, Chilton et al. demonstrate astable incorporation of plasmid Ti-DNA into higher plant cells. In1983, the team of Van Montagu and Schell in Gent succeeded inproducing a transgenic plant, and eleven years later the firsttransgenic cultivars were available in the market.

2. Transgenic Plants with New Agronomic Traits: Two differenttechniques are commonly used to introduce genetic information(DNA) inside cells, protected by pectocellulosic walls. A particlebombardment process, called biolistic, is used with success formonocots gynmosperm, squash, peas, Today, this technique allowsthe bombardment of meristems and avoids the difficulties ofregeneration. But the mediation of Agrobacterium remains till nowthe most routinely system to transform dicotyledonous plants.The main agronomic traits introduced in horticultural plants andalready commercialised are Bt toxin and herbicide resistance. Otherstudies concern virus resistance, male sterility.

3. Antisense Strategy: Calgene created in 1994 the first commercialtransgenic plant, a long shelf life tomato, by the suppression ofpolygalacturonase activity due to an antisense gene. However, tactivity due to an antisense gene. However, this Flavr Savr tomatovariety was removed from the trade 3 years later, because of itsdisease susceptibility and its lack of productivity.Later, other tomato varieties with long storage qualities wereobtained by the utilization of an antisense RNA inhibition of ACCsynthase or ACC oxydase, two ethylene precursors.The antisense technique was also used to reduce the lignificationof woody plants, by blocking the enzymes involved in the precursorof lignin biosynthesis. Another interesting application was theinduction of white flowers in petunia and in different otherornamental plants by the suppression of chalcone synthase activity.

4. Transmission of the New Traits: These traits induced by the transferredDNA are transmitted to the progeny as a dominant Mendeliancharacter. Nevertheless in some cases, transgenic plants with astrong gene expression can generate progenies with only a faint nogene expression. This problem is not clearly elucidate.

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5. Marker, Reporter, Promotor and Expression Genes: To select thetransformed cells, some marker genes, very often resistant toantibiotics or to herbicides, are attached to the coding sequence, asa promoter or an expression gene. Those allow the new gene toexpress in the whole plant or in a specific plant tissue.Some reporter genes are used to follow the evolution one.Some reporter genes are used to follow the evolution of thetransformed cells. The most frequent reporter genes are the gene“gus” or the gene lux (luciferase).

6. Horticultural Transgenic Crops already released in USA: The Flavr Savrtomato was the first genetically engineered whole food approvedfor commercial sale in 1994. Four other transgenic tomatoes with adelayed ripening were approved later (1995 and 1996).

In 1995, potatoes with Bt genes and a squash cultivar resistant to twoviruses were released. Two years later, another squash cultivar resistant to 3viruses and a papaya line also resistant to viruses were approved. Two otherhorticultural crops are waiting the authorities approval: red hearted cichory(Radicchio) with male sterility and resistance to herbicide, and a tomato withBt toxin.

It there are so few commercial transgenic plants in horticulture, it’sprobably due to the relatively low acreage of these horticultural crops incomparison with other agricultural crops. In 1997, in the world 5 million hawere planted with glyphosate resistant soybeans released by Monsanto alone.

Nowadays, it’s true also that many transgenic plants exist in researchlaboratories, awaiting authorization for field testing, as they are veryinteresting model plants for learning physiology: lettuce with less nitrate byincreasing nitrate reductase gene active with less nitrate by increasing nitratereductase gene activity, a yeast ribonuclease gene to reduce viroid infectionon potato, modification of lignin synthesis in plant to produce trees adaptedto paper industry, or timber, or biomases production, regulation genes tocontrol tree architecture.

Situation of Biotechnologies in the World: Development and Risks

Situation in the Young Countries or in Developing Countries: A generaluse of in vitro micropropagation in the developing countries is now a reality,as it was predicted by Albert Sasson (1993). African countries for instance areproducing potatoe, banana, cassava.

Even the production of transgenic plants is no longer a prerogative ofthe Northern countries. Different experiments are now starting in the South,specially in the international laboratories from CGIAR. Although CGIAR’sexpenditures on biotechnology for 1997 raise only 24.2 million US$ on anannual budget of 345 million US$ this Consultative Group on InternationalAgricultural Research plays a very important role as contributor to agricultural

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research for developing countries. The large international research centres IITA(Ibadan, Nigeria), CIP (Lima, Peru), CIAT (Cali, Columbia), belong to theCGIAR. Their roles and NGO’s are primordial. The genetic transformationtheir roles and NGO’s are primordial. The genetic transformation of cassavais an excellent example. Its tuberous roots provide food for over 500 millionpeople, mostly small-scale farmers. Unfortunately, till recently this integralplant for food security in developing countries has been recalcitrant totransformation approaches. However, thanks to the participation ofinternational institutes located in cassava growing countries, CIAT and IITAwith the help of four development associations from Swiss, UK, TheNetherlands and USA, were able to initiate procotols for cassavatransformation and regeneration.

Other networks, like the Cassava Biotechnology Network (CBN)previously described, exist. REDBIO, a technical cooperation network on plantbiotechnology, is supported by FAO to promote a best use of scarce manpower,equipment and other resources in Latin America.

On the other hand, more and more private laboratories exist in thedeveloping countries which have a high scientific and technical level (China,Singapore, Taiwan, India, Brasil, Mexico, Chile). In these countries,European, American or Japanese agrochemical companies are investing. Alsothese countries will have their chance to develop their own technologies, andto export to the Northern markets.

Meanwhile, genetic engineering techniques are being applied mainly tocrops which are important for g applied mainly to crops which are importantfor the industrialized world, not crops on which the world’s hungry depend.Therefore, it is unrealistic when Monsanto writes “the experts said thatbiotechnological innovation will increase the crop productivity withoutoccupation of new lands, saving tropical forest of best quality and animalhabitat”. However, the World Bank was promising in a 1997 publicationBioengineering of crops that” transgenic crops could improve food yield byup to 25 per cent in the developing countries and could help to feed anestimated additional three billion people over the next 30 years”.

But, not all people are confident in this prospective. To the contrary, someare afraid that the poorest countries, where more than 700 million people arechronically undernourished will miss the biotechnological revolution.

Nevertheless, there is some hope to transfer to the developing countrieshigh technology, which don’t undermine the environmental and socialnetwork. So, the Agricultural Biotechnology for Sustainable ProductivityProject (ABSP project) is managed by Michigan University. Four Americanuniversities, two research centres from France and USA, and two Americanprivate companies are also involved in this project. The ABSP’s objectives arethe reduction of losses due to pathogens and the reduction of losses due topathogens and pests by using transgenic plants (sweet potatoes, potatoes,

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cucurbits, tomatoes) and by cloning commercial value-plants (bananas,pineapple, coffee).

Situation in the Northern Countries

1. Mass Propagation: In 1988, Pierik (1991) predicted a European invitro propagation of 212 million plantlets/year. He was alsopredicting an important development of this activity for the future.Following Pierik this quantity accounted for only 5 % of the plantsable to be propagated by vegetative means. However a few yearslater Pierik’s prediction was not confirmed. The world productionwas estimated at 600 million. In USA, Zimmerman (1997) wasrecording a production of 121 million plantlets for a total of 110laboratories, half produced in 11 laboratories, and 6 labs werepropagating each more than 6 million plants per year.In Europe, only the Dutch production of micropropagated plants isprecisely known. In 1990 the total production was 95.5 million. In1995, the Dutch in country production reached only 53,7 million,whereas importations increased 77,3 million. (37 million fromPoland and 17,1 million from India) (Pierik, data unpublished).These laboratory relocations to low-cost manpower countries arecommonly observed, and put the question of the qualitymanagement.

2. Transgenics: The GMOs invade progressively all the US lands: 1.6% in 1996, 15 % in 1997, 50 % in 2000. It will represent anestimated market of 3,9 million US$ in 2003 ! The largesttagrochemical companies are merging to become giants ofagrobusiness such as Novartis, Monsanto, Zeneca.Following Monsanto in 1996, about 950 T insecticides were savedby introduction of resistant cotton, id est a net savings of 81 US$/ha for the grower. Glyphosate-resistant soybeans (4 million ha in1997) allow a savomgs of 44 to 49 US$/ha. (Information fromcultivar, suppl. n 436, Feb. 16, pp. 24-37, 1998).

3. Perspectives, Limitations and Environmental Risks: Ecological impactrelated with the introduction of GMOs is always an open debate.The application to agriculture of these new technologies certainlyopens interesting perspectives, but also raises potential problems.The risk of crop transgene spreading has been demonstrated. Aresearcher of Clemson University in South Carolina reported “thatin a population of wild strawberries growing within 50 meters of astrawberry field, more than 50 % of the wild plants containedmarker genes from the cultivated strawberries”.

A Danish team have shown a possible rapid spread of genes from oilseedrape to the weedy relative Brassica campestris.

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There are other risks. Brassica campestris.There are other risks. The introduction of Bt gene allows a drastic

reduction in the use of toxic chemicals for crop protection. But a poorlycontrolled use of Bt-technology can destroy more effectively the predatorsthan the pests.

Or when” many crop plants are transformed with similar effective traits,in such situation, many polyphagous pest species, which by nature are moreflexible evolutionarily than those that have a narrower diet, are likely toovercome Bt resistance very quickly”.

Therefore, before releasing a transgenic plant, any risk has to be weighedagainst the benefit of the transgenic crops. We must not forget that annuallyin the world 500.000 acute pesticide poisonings, with 5000 deaths, areobserved.

DURING THE MICROPROPAGATION PROCESS: THE GENETIC STABILITY

During the micropropagation process, the genetic stability of the newshoots is dependent upon their origin. Axillary shoots issue from pre-existingbuds and are normally true to type, indeed the meristematic cells aregenetically very stable.

Adventitious shoots, such as somatic embryos, are neo-formed budsdeveloped directly on some organs, or indirectly through a callus phaseformed on this organ.

So, if the mother plant presents a cell mosaic or chimaeric tissues, risksof genetic variation exists. It is similar in the case of an indirect regeneration,when the callus phase is too long.

PROPAGAT ION BY AXILLARY SHOOT ING

This technique has proved to be the most applicable and reliable methodof in vitro propagation. Axillary shoot growth is stimulated by overcomingapical meristem dominance.

Commercial tissue culture laboratories are now able to propagate a largenumber of herbaceous ornamental species and several woody plants in thisway. However, the propagation of Pelargonium, Howea, and a few otherhorticultural plants are always difficult to propagate by axillary branching.

Propagation by Direct or Indirect Organogenesis

Adventitious shoots could arise directly from the tissue of explantswithout callus formation. Several plants of the family gesneriaceae(Saintpaulia, Streptocarpus) regenerate directly buds on leaf explants,likewise Lilium rnerate directly buds on leaf explants, likewise Liliumregenerates on scales.

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However more often, like for Ficus lyrata, adventitious buds appear oncallus. While coffee, cocoa trees, and many conifers are produced by somaticembryogenesis developed on callus or cell suspensions.

Constraints of in Vitro Micropropagation

The establishment of axenic culture could be difficult when the explantsare coming from hot and humid countries.

Moreover, in the course of micropropagation several authors observed asudden appearance of endogeneous bacteria. Nevertheless only Holland andPollaco (1994) have demonstrated the presence of this kind of bacteria in morethan 70 different species. For Leifert and Woodward (1997), the major sourceof contamination is the initial explant. They add that microbial contaminationin commercial plant tissue culture laboratories is the most important cause oflosses. For this reason, they support the introduction of a microbiologicalproduction control strategy.

A second important constraint during the initial phase mainly concernswoody plants. A mature tree must be rejuvenated to recover its morphogeneticcompetence. Franclet (1981) described various treatments, micrografts, cascadegrafts, cytokinin treatments to reverse the adult phase to juvenile. Themicropropagation of Acacia Senegal was only possible after rejuvenation bymicrografting (Palmnly possible after rejuvenation by micrografting.

A distinction between chronological, ontogenic and physiological age isneeded to understand the rejuvenation concept. This was definitivelydemonstrated after a complete reversion of Sequoia sempervivens.

Improvement of Axillary Branching

The cost of micropropagated plantlets is also an important limitation ofthe techniques. In New Zealand, where they produce 2-3 millionmicropropagated radiata pine per annum, the relative cost of micropropagatedplanting stock had dropped from 13,8 times the cost of seedlings in 1988 to6,9 by 1993.

To reduce manpower costs, several improvements have been proposed.The more simple method was in vitro layering developed by Wang (1977) toclone PVX-free potato plants. The first plantlets placed on the medium in ahorizontal position developed axillary shoots. They are harvested by cuttingone centimetre above the medium surface, at 3 weeks intervals. A similartechnique called ‘hedging system ‘ by Aitken Christie and Jones (1987) waslater used to produce Pinus radiata.

Ziv (1990) proposed for corm plants, Gladiolus and Nerine, a very rapidpropagation system. She reduces the internodes and leaves by introductionof an anti-gibberellin agent in the medium. Finally, only aggregates of budsare formed, then they finally, only aggregates of buds are formed, then theyare divided and introduced in bioreactors for mass production. Similar

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systems were developed in Gembloux, to propagate carub trees and asparagus.Since 1988, Duhem was producing very large quantities of Eucalyptus plantletsin Petri dishes without anti-gibberellin but in complete darkness. Transfersfrom one Petri to another is made by a simple squashing.

Somatic Embryogenesis Propagation

For genetically stable species, somatic embryogenesis offers a very fastscaling-up system, especially when it’s possible to produce embryos inbioreactors. Unfortunately this production via fermentors was not as simpleas first envisaged. And today, only a few model plants are successfullyproduced by such technology: carrot, celery. Other applications remain at theexperimental stage: coffee, oil, palms, conifers, Euphorbia pulcherrima, andseveral other horticultural species.

Several bottlenecks limit the use of this interesting technology. One ofthe main problems is genetic stability. So, despite the clonal nature of nucellarembryos, different morphological anomalies can occur among mango somaticembryos, as it was also observed in Citrus plants derived from nucellarcultures.

Another difficulty is the loss of embryogenic capacity over time, aphenomenon observed with different species. It is also a phenomenonobserved with different species. It is also important to note that somaticembryogenic lines of conifers are always originated from immature embryos.

Artificial Seeds

Another very interesting possibility of the somatic embryogenesistechnology has been developed during the past fifteen years by Redenbaughand his team (1991, 1993). They were able to encapsulate somatic embryos byhydrogel coatings (sodium alginate), producing single embryo artificial seeds.To date, some improvements offer the possibility to directly plant the artificialseeds in the greenhouse on special substrates (vermiculite, sans). Thismethodology will provide in the future a good technique to reduce the costof transplants.

Gene Bank

Tissue culture methods offer the opportunity for in vitro collecting, rapidmultiplication and distribution of important, elite, or rare plants that arethreatened with extinction. The two major in vitro storage strategies are slowgrowth and cryopreservation. Since the first results of Seibert (1976), who wasable to initiate shoots from carnation shoot apices frozen to -196C, thistechnique is now successful for m any of horticultural species. Dereuddre etal. (1991) have provided a very simple technology to freeze encapsulatedmeristems in dried alginate beads. It works for pear, strawberry, eucalyptus,potato.

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Some Ior Pear, Strawberry, Eucalyptus, Potato

Some international institutions have very large collections of old andcurrent varieties, available for exchange and introduction in crop improvementprogrammes. The International Potato Centre (CIP) in Lima, Peru, has a largeworld potato collection. Germplasm of sweet potato and cassava is at theInternational Institute of Tropical Agriculture (IITA), Ibadan, Nigeria.

The movement of germplasm involves the risks of accidentallyintroducing plant quarantine pests along with the host plant material. To limitthese risks, the plant material should be transferred from one country toanother as in vitro cultures through a transit Centre, where it should beindexed. For bananas, in the framework of INIBAP, the transit Centre is theCatholic University of Leuven in Belgium where a very large in vitrogermplasm exists.

RESEARCH PREPAREDNESS FORACCELERATED GROWTH OF HORTICULTURE

The horticulture scenario of the country is rapidly changing. Theproduction and productivity of horticultural crops have increased manifold.Production of fruits and vegetables has tripled in the last 50 years. Theproductivity has gone up by three times in banana and by 2.5 times in potato.Today horticultural crops cover about 25 per cent of total agricultural exportsof the country. The corporate sector is also showing greater interest inhorticulture. A major shift in consumption pattern of fresh and processed fruitsand vegetables is expected in the coming century. There will be greatertechnology adoption both in traditional horticultural enterprise as well as incommercial horticulture sectors. Diversification and value addition will bethe key words in the Indian horticulture in the 21st Century.

Horticulture research in India is about four decades old. Systematicresearch on fruit, vegetable and ornamental crops began in 1954 with theinitiation of independent institutions and programmes. The research agendais designed relevant to national plans and priorities for the horticulturedevelopment. Today, eight ICAR institutes with 27 regional stations, 1 projectdirectorate, 10 national research centres, 16 all India coordinated researchprojects (AICRPS) with 223 research stations, 1 full-fledged university ofhorticulture, 25 state agricultural universities and 7 multi-disciplinaryinstitutes of the ICAR are engaged in horticulture research. In addition, a fewR&D establishments of crop/commodity boards and private sectors areproviding research support to Indian horticulture. Research system inhorticulture is now geared to provide necessary technological support to theexpanding horticultural industry. The research efforts in the past were mainlyconcentrated on crop improvement, propagation of seed/planting material,agrotechniques, crop protection and post harvest management.

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VARIETAL DEVELOPM ENT

Among the fruit crops, improved high yielding mango varieties, Mallika,Amrapali, Ratna, Sindhu, Arka Aruna, Arka Puneet, Dashehari-51 and hybridsnamely CISH-M-1 and CISH-M-2 have been developed. Mallika is comingup in southern states like Karnataka and Amrapali is performing well ineastern India. Dashehari-51, a regular bearing cultivar with about 38 % higherproductivity than the normal Dashehari, has been identified after 14 years ofrigorous selection. In guava, three selections, namely Lalit, CISH-G-1 andCISH-G-2 have been developed for domestic and export markets.

The fruits of Lalit are of medium size weighing about 150 g each andsuitable for both table and processing purposes. In banana, high yieldinghybrids like FHIA-01 and FHIA-03 are promising for replacing varieties,Panchananda and Bluggoe, respectively. Cultivar, Saba is found promisingunder sodic soils. In addition, high yielding varieties like Col, Hl and H2 havebeen developed. In grape, superior and high yielding varieties have beendeveloped e.g. Beauty Seedless and Pusa Seedless and early ripening variety,Perlette for cultivation under North Indian conditions and Anab-e-Shahi,Dilkhush, Thompson Seedless, Tas-A-Ganesh, Sonaka, Bangalore Blue andPachadraksha for south Indian conditions.

In citrus, high yielding and cluster bearing varieties of acid lime havebeen developed. e.g., Rough Lemon, Rangpur Lime, Pramaini, Vikram PKM-1. Trifoliate oranges, namely Flying Dragon and Rich 16-6 are dwarfing types.In papaya, high yielding superior varieties both for table purpose and papainproduction have been developed. e.g., Co-1 to Co-7, Coorg Honey Dew, PusaDelicious, Pusa Majesty, Pusa Giant and Pusa Nanha. In apple, superiorhybrids have been developed. e.g., Lal Ambari, Sunehari. Red Spur, StarCrimson, Golden Spur, Red Chief, Oregon Spur, Skyline Supreme and VanceDelicious have been identified. Tissue culture protocols for micro-propagationof two commercial varieties have been developed.

Among the vegetable crops, more than 130 open pollinated varieties, 36hybrids, 3 synthetics and 29 resistant varieties of 20 vegetable crops have beendeveloped and released for cultivation in different agro-climatic regions. Theseinclude 40 in tomato, 45 in brinjal, 13 in cauliflower, 12 in chillies, 20 in pea, 9in musk melon, 16 in onion and 44 in other crops.

In potato, 33 high yielding varieties have been developed indigenouslyfor large scale cultivation in different regions. Kufri Ashoka and Kufri Pukhrajmature in 75 days. Kufri Jawahar and Kufri Satluj are field resistant to lateblight. Kufri Jawahar has most ideal plant type for inter-cropping. KufriChipsona-1 and Kufri Chipsona-2 have been developed with excellentprocessing attributes, comparable to exotic varieties. Kufri Swarna resistantto golden nematode is ideal for Nilgiri Hills.

In tuber crops, improved varieties of different tuber crops have beenrecommended/released for commercial cultivation. These includes 9 varieties

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of cassava, 15 varieties of sweet potato, 6 varieties of colocasia, 3 varietieseach of greater yam and lesser yam, 1 each of Amorphophallus, taro andyambean. Cassava varieties, Sree Visakham and Sree Prakash are popular inKerala. Triploid clone, Sree Harsha with high dry matter and starch contentis suitable for industrial belt of Tamil Nadu. Two early maturing varietiesSree Jaya and Sree Vijya have been released for culinary purposes. Elephantfoot yam variety, Am-15 has been released with high yield potential of 41 t/ha.

Among the plantation and spice crops, India is the first country to exploithybrid vigour in coconut. Twelve hybrids involving tall and dwarf parentsand 4 varieties have been released for commercial cultivation. These varietiesyield 21 to 89 % more than the local cultivars. Some of the released varietieslike Chandra Kalpa and Pratap (Banawali Green Round) are receiving wideacceptance of farmers. Chowghat Green Dwarf variety is good for tendercoconut purpose. In arecanut, 4 high yielding varieties, namely, Mangala,Sumangala, Sreemangala and Mohitnagar have been developed, giving about30 % higher yield than the local cultivars.

In oil palm, first efforts for improvement were made by producing Tenerahybrids using Pisifera pollen imported from Nigeria. Dura x Pisifera hybridsare field tested in East and West Godavari districts, Khammam and Krishnadistricts of Andhra Pradesh, with yield potential of 20-25 tonnes/ha of FFBfrom the fifth year. In cashew, 22 region specific selections and 12 hybridswith yield potential of 1.5-2 tonnes of raw nuts/ha have been produced andreleased for commercial cultivation.

The present standards fixed for cashew varieties include export gradekernels of W-210 to W-240 and at least one tonne per ha yield with 30 percent shelling. In black pepper, 6 varieties, namely, Sreekara, Subhakara,Palode-2, Panniyur-2, Panniyur-4, Panchami and Pournami, and 2 hybridsviz. Panniyur-1 and Panniyur-4 have been developed. In cardamom, a numberof improved varieties have been developed and released for commercialcultivation e.g., CCS-1, Mudigere-1, PV-1, ICRI-1 and ICRI-2. In ginger,varieties like Suprabha, Suruchi, Suravi and Varada have been developed. Inturmeric, several varieties viz., Co-1, Krishna, Sugandham, BSR-1, Suvarna,Roma, Suroma, Rajendra Sonia, Sugana, Sudarshana, Ranga, Rasmi, Prabha,Prathiba, Mega Turmeric and RCT-1 have been developed with yield potentialof up to 44 tonnes of fresh rhizomes per ha. Three high yielding cinnamonlines, namely, Navashree, Nithyashree and Konkan Tej have been releasedfor cultivation.

Agrotechniques

In fruit crops, improved agrotechniques developed have helped thefarmers in improving the productivity and quality of produce. Soil applicationof paclobutrazol (4 g/tree) increase flowering and fruiting in mango on

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commercial scale in coastal Maharashtra. It also controls irregular bearing incultivar Dashehari. Spray of NAA @ 200 ppm in October is recommended forcontrol of malformation. Heavy fruit drop at maturity in cultivar Langra canbe controlled by spraying NAA (20 ppm). In guava, double spray of 10 and20 % urea on cultivars, Allahabad Safeda and Sardar twice at bloom eliminatepoor quality rainy season crop and increases winter season yield by 3 and 4times, respectively. Application of neem coated urea (800 g/plants) yields 98kg fruits/plant in variety, Sardar compared to 37 kg from untreated plants. Inbanana, high density planting (4550 plants/ha) yield up to 174 t/ha.

Adoption of improved technology in Maharashtra has resulted in fruityield increase up to 52 t/ha. In citrus, two grafting methods using inverted‘T’ cut and apical triangle cut have been developed with overall success ofaround 36% using either method. For accelerating the survival of growth, shoottip grafts, the successful grafts are double grafted (side grafted) on vigorousgreenhouse grown Rough Lemon and Rangpur Lime seedlings. Rangpur Limerootstock is found superior for sweet oranges and mandarins. In papaya, closerspacing of 1.4 x 1.4 m is recommended for high yield. Drip irrigationtechniques have been standardized. In banana, it has resulted in productiongain (60-70%) and early harvesting (40-50 days), besides improved waterefficiency. Likewise, in grapes higher yields have been obtained with betterwater use efficiency (11 %).

In vegetable crops, improved production technology has been developedfor major crops. Drip irrigation is economical in tomato and brinjal. Incucumber, replenishment of evaporation loss through irrigation resulted inmaximization of yield of quality fruits. Drip irrigation in watermelon provided33% higher yield with a water saving of 40%. Nutrient requirements andfertilizer schedules have been worked out crop-wise and recommended fordifferent agro-climatic regions. In leguminous vegetables, high N depressesnodulation. The VAM fungi increases P availability to plants.

In all leguminous vegetables, inoculation of the VAM fungi along withRhizobium culture is beneficial. Production technologies for kharif onion innorthern India and long day type onions for high altitudes have beenstandardized. Pendimethalin (Stomp) has been found effective in controllingweeds in tomato, brinjal, chilli, bell pepper and okra. In potato, a number ofpotato-based multiple and inter cropping systems have been developed fordifferent potato growing regions. Intercrop combinations with sugarcane inMaharashtra, wheat in Chhota Nagpur area and linseed in central UttarPradesh are found remunerative. A suitable method of urea application hasalso been worked out.

In tuber crops, short duration legumes viz., groundnut and French beanand cowpea can be successfully inter-cropped with cassava. Short-durationcassava, Sree Prakash is ideal in double cropped rice fields. Studies on cassava-based multiple cropping systems involving banana, coconut, Leucaena and

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Eucalyptus, have shown banana-cassava combination to give maximum rootyield. Banana and coconut combination reduces soil loss and surface run-offconsiderably. Dioscorea and elephant foot yam with banana, Nendran cangenerate an additional income of Rs 20,000/- over the sole crop of banana. ForInter-cropping Dioscorea in coconut garden, the ideal planting density is 9000plants/ha. When Amorphophallus is raised as an inter-crop in coconut garden,one third dose of recommended fertilizers is sufficient. Inoculation with VAMfungi in cassava give about 15-20% increase in yield.

Among the plantation and spice crops, density of 175 coconut palms/ha(7.5 m x 7.5 m spacing) is found ideal. In general, NPK application of500:320:1200 g/palm/year is found optimum. A multi-storied cropping systeminvolving black pepper trained on coconut trees, and cocoa in between therows of coconut and pineapple in the ground floor has been found ideal forexploiting light, soil and air spaces. In arecanut, application of NPK (100:40:140g) and green leaf (14 kg) per palm per year is recommended for coastal regionsof Kerala and Karnataka and for plains of West Bengal, Karnataka and Assam.

In oil palm, application of NPK (1200:600:1200 g/palm) is found to give17.1 tonne of FFB per ha. In black pepper, rapid methods for production ofrooted cuttings of pepper have been developed and a commercial protocolhas been standardised for micropropagation of black pepper. Ginger yieldcould be increased up to 33 percent by application of neem cake at the rate oftwo tonnes per ha and the fertilizer schedule of 75 kg each of N, P205 and K20.Technology for storage of ginger seed rhizome is standardised andrecommended.

Protection Technologies

For major fruit crops, plant protection schedules have been developedfor the control of significant insect-pests for wider adoption. In the recent years,research efforts are directed to devise eco-friendly, economical and long lastingcontrol measures. Success has been achieved in biological control of mealybugs in mango and guava. The Beauveria bassiana has been found killing mangom ealy bug and hopper. In grapes, integrated m anagem ent o f Spodopteracaterpillar involving light and pheromone traps, NPV and neem basedinsecticides and biological control o f m ealy bug by the beetle Cryptolaemusmontrouzieri have been standardized. Studies on pesticide residues haveresulted in working out of safe-waiting periods for harvesting andconsumption of fruits.

In vegetable crops, about 50 improved measures for efficient managementof diseases and 23 for insect-pests have been worked out and popularized indifferent agro-climatic regions in the country. Integrated pest management(IPM) for controlling diamond back moth on cabbage through a trap croplike mustard has been demonstrated. Fruit borer (H. armigera) on tomato canbe controlled by the release of Trichogramma pretiosum alone and in

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combination with HaNPV. In potato, integrated management schedules forcontrol of bacterial wilt and tuber moth have been developed. A late blightforecasting system has been developed for the hills.

Among the tuber crops, the major diseases affecting tuber crops arecassava mosaic and brown leaf spot in cassava, Phytophthora leaf blight incolocasia, Fusarium wilt in elephant yam and virus diseases of sweet potato.

Foliar sprays of Bavistin (0.1%) combined with disodium and dipotassiumphosphates (100 ppm) and calcium sulphate at 15-day interval was found tocheck foliar diseases in sweet potato. The major pests include spider mites,scale insects and white fly on cassava, weevil on sweet potato, defoliators,aphids and mites on colocasia, and scales and mealy bugs on yams andelephant yam. Cultural methods for weevil control include clean cultivation,destruction of alternate hosts and timely harvest. An effective IPM packageusing synthetic sex pheromone has been developed. Control measuresinvolving insecticides have been evolved for the control of pests of other tubercrops.

Among the plantation and spice crops, bud rot of coconut caused byPhytophthora palmivora can be effectively controlled by spraying Bordeauxmixture. Calyxin root feeding and drenching of soil with 1% Bordeaux mixturealong with neem cake application @ 5 kg per palm per year is recommendedfor controlling Thanjavur wilt disease reported in Tamil Nadu, AndhraPradesh and Karnataka.

A package of practices has been developed for managing mycoplasmalike organisms (MLOs) in root wilt affected coconut palms in Kerala andThatipaka disease affected palms in Andhra Pradesh.

Eradication of all root wilt affected palms is recommended. In cashew,tea mosquito bug (TMB) can be effectively controlled through a schedule ofspray coinciding with flushing, flowering and fruiting. For effective controlof stem and root borer infestation, constant monitoring and adoption of strictsanitation in the plantations coupled with prophylactic application of coaltar and kerosene in the ratio of 1:2 on trunks are recommended. In blackpepper, spraying Bordeaux mixture (1 %) and drenching the soil with copperoxychloride (0.2 %) is found effective in managing Phytophthora foot rot.

Seed/planting Material Propagation

In most of the fruit crops, vegetative propagation techniques have beenstandardized. Soft wood grafting has been standardized for mango, sapota,custard-apple and jackfruit. Other vegetative propagation techniques havebeen developed for ber, aonla, jackfruit, custard-apple and bael. In mango,veneer grafting and stone-grafting is practised commercially. Mango variety,Vellaikolumban is suitable semi-dwarfing rootstock for Alphonso. Oldunproductive mango trees can be rejuvenated successfully by pruning the4th order branches during December-January. Flowering and fruiting are

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regular in pruned trees. For mandarin orange, Rangpur lime is a droughthardy rootstock. In grapes, Dogridge and Salt Creek (Ramsey) are suitablefor minimizing adverse effects of soil salinity on Thompson Seedless.

A tissue culture technique for mass multiplication of Dogridge has beenstandardised. In banana, sword suckers of 700-1000 g are optimum. Rhizomeswith active lateral buds and dead central buds are preferred for distanttransportation in western India. Double paring and shade drying followedby dipping in Monocrotophos (0.5 %) and Bavistin (0.2 %) is recommendedto disinfect nematodes and soil borne fungi. Tissue cultured banana plantsare now commercially adopted for their uniformity in flowering and produce.Shoot-tip grafting technique in citrus has been considerably advanced.

In vegetable crops, seed production of over 120 open pollinated highyielding varieties of different vegetables has been well established in thecountry. Hybrid seed production has become easier with the development ofmale sterile lines in tomato, self incompatible lines in cauliflower andgynoecious lines in cucumber and muskmelon. In brinjal, functional malesterility controlled by a single recessive gene has been identified.

Temperature barrier in cole crops (cabbage and cauliflower) has beenovercome by developing heat tolerant hybrids. It is now possible to cultivatecabbage and cauliflower in southern India. Development of tomato varietiesresistant to bacterial wilt has made their cultivation successful in non-traditional areas. Onion seed production technology for cultivation in kharifseason has been developed for north Indian states especially, Haryana, Punjaband western Uttar Pradesh. Seed Plot Technique has been developed forproduction of disease-free potato seed in plains. It is widely adopted byfarmers. A new technology for raising commercial crop of potato using ‘TruePotato Seed’ (TPS) has been developed and standardized as supplementarytechnology to the traditional tuber grown crop. Two TPS populations, TPS-C-3 and HPS-113 are recommended for commercial production in Bihar,Gujarat, Tripura and West Bengal. Micropropagation protocols have beendeveloped in banana, oil palm, cashew, black pepper, ginger, etc. Seed gardensof Tall (T) x Dwarf (D) and D x T hybrids have been established for productionof coconut hybrids.

Post Harvest Management Technologies

The post harvest handling of fruits and vegetables accounts for 20-30%of losses at different stages of storage, grading, packing, transport and finallyat marketing as a fresh produce or in processed form. A number of improvedtechnologies have been developed for commercial exploitation. An on-farm,low cost, environment friendly cool chamber, Zero Energy Cool Chamber hasbeen developed using locally available material.

The principle of evaporative cooling reduces the inside temperature byas much as 17-18 oC and keeps the relative humidity above 90% during peak

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summer. The chamber increases the shelf life and reduces PLW of banana,mango, orange lime, grape fruit, tomato and potato in different situations inIndia. Maturity standards for mango, guava, grape, litchi and ber and chemicaltreatments for regulation of ripening in mango, sapota and banana have beenstandardised.

Optimum storage temperatures worked out for several fruits, vegetablesand tuber crops. A mango harvester, fruit peeler, hand and pedal operatorcassava chipping machines, harvesting tools (5-14 times efficient), coconutdehusking machine, implements for mechanization of potato cultivation andother crops have been developed. A number of improved technologies havebeen developed for commercial exploitation viz., tent type foldable solar dryer,packaging boxes for distant transportation of apple, mango, citrus and plum,production of value added products-pectin from peel and flour from mangofruit kernel, production of fruit post carbonated beverages etc.

Production Constraints

In spite of great strides made, the productivity of horticultural crops, ingeneral, is still quite low and the post harvest losses particularly of perishablecommodities, are considerable. Improvement in quality standards of theproduce and their marketing are essential to increase our share in the globalmarket. The research agenda in horticulture is by design relevant to nationalplans and priorities and research programmes are normally formulatedkeeping in view the thrust areas in development. The major technology relatedconstraints contributing to low productivity of horticultural crops and inferiorquality of produce are:

• Vast majority of holdings are small and un-irrigated.• Large tracts of low and unproductive plantations needing

replacement/rejuvenation.• Low productivity of crops due to inferior genetic stocks and poor

management.• Inadequate supply of quality planting materials of improved

varieties.• High incidence of pests and diseases.• Heavy post harvest losses and low utilization in processing sector.For addressing the above constraints, research institutions are engaged

in both basic and applied research. While formulating research strategies someof the inherent weaknesses associated with perennial tree crops and certainperpetual problems in Indian horticulture must be kept in mind. They are:

• Long period required for development of improved genotypes.Application of biotechnological tools/methods in horticultural cropsis still in its early stage of development in the country.

• Chronic production problems due to major disorders like alternatebearing, malformation and spongy tissue in mango, guava wilt,

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citrus decline, root wilt in coconut, viral disease in vegetables,Phytophthora diseases in large number of crops etc. remained largelyunresolved.

• Lack of advanced technologies for post harvest handling, processingand marketing of produce.

Losses caused by biotic stresses are very high and due to pesticide residueproblems development of eco-friendly IPM strategy is more relevant inhorticulture. There is a threat for loss of valuable genetic resources, if measuresare not taken for their conservation.

Wastelands and hilly terrains being the potential future expansion areas,matching technologies for dry land and hill horticulture need to be developed.Counter seasonal advantages from diverse agro-climatic situations providestrength for extended availability of horticultural crops round the year andsuch potentials can be harnessed only with relevant research support.

PLANT TISSUE SYSTEMS

ST RUCT URE AND FUNCT ION

Although the architecture varies considerably among roots, stems, andleaves, all plant organs are comprised of 3 basic tissue types: dermal tissue,vascular tissue, and ground tissue. A tissue is simply an assemblage of cellsthat work together as a structural or functional unit. In this lab, we willexamine the general arrangements of those tissues among different plantorgans. You will find that the distribution of tissues varies somewhat amongdifferent plants as well as among different organ systems. For example, oneof the key differences between monocots and dicots has to do with thearrangement of vascular tissues in leaves and stems. In next week's lab, wewill continue our examination of plant tissues by exploring some of thedifferent cell types that form these three plant tissue sytems.

DERM AL T ISSUE

The dermal tissue system is derived from the protoderm* and consists of 2 tissuetypes:

• Periderm: Periderm includes cork and cork-producing tissues andis only found in plants with secondary growth.

• Epidermis: The epidermis constitutes the outmost layer of cells ofroots, stems and leaves. Epidermal cells are usually covered by awaxy layer, called a cuticle that serves for waterproofing the cells.

VASCULAR T ISSUE

Vascular tissues, derived from the procambium*, are the tissuesresponsible for transporting substances from one point in the plant body to

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another. As is the case with animals, the vascular tissues are responsible forthe movement of dissolved gases, sugars, and minerals throughout theorganism.

In plants, there are 2 types of conducting tissues:• Xylem: Xylem is the tissue specialized for the conduction water and

dissolved minerals. Cells of the xylem are dead at maturity and actas the structural medium for the movement of water. Individualxylem cells have multiple holes in the walls to allow for the passageof water between adjacent cells. The bulk of wood consists of xylemtissue.

• Phloem: Phloem conducts dissolved sugars throughout the plant.The main conducting cells of phloem, called"sieve elements", arealive at maturity, but are highly modified for carrying out thefunction of transport. For example, many of the organelles youmight expect to find in a living plant cell, such as nuclei, ribosomes,Golgi apparati, cytoskeleton, and vacuoles are absent from the sieveelements in order to make more space for fluid transport. The smallamount of protoplast in sieve elements is distributed in a thin layeralong the periphery of the cell wall. In some cases, you may beable to observe smaller cells called"companion cells" distributedamong the sieve elements. Companion cells contain all theorganelles you would expect to find in a typical plant cell and carryout many of the necessary life functions for the sieve elements whichlack organelles.

GROUND T ISSUE

Derived from the ground meristem*, ground tissues are distributedbetween and among the dermal and vascular tissues of roots, stems, andleaves.

Ground tissues include a wide variety of tissue types that perform amultitude of functions within the plant.

Some of the more familiar ground tissues include the following:• Cortex: the ground tissue distributed between the epidermis and

vascular tissues in roots and stems. Cortex is usually modified forstorage, especially of starches. In herbaceous green stems, cortexoften contains chloroplasts and thus is further modified for carryingout photosynthesis.

• Pith: like cortex, pith is generally modified for starch storage, butis only found to the interior of the vascular tissues in stems androots.

• Mesophyll: the ground tissue of leaves found between the two layersof epidermis. Leaf mesophyll is the tissue most adapted forphotosynthesis and thus is generally rich with chloroplasts.

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In addition to the functions indicated above, ground tissues also have animportant role in wound healing and the regeneration of damaged or deadcells throughout the plant.

PLANT T ISSUES ARE DERIVED FROM M ERIST EM S

Unlike animals, most plants grow (increase the number and size of theircells) throughout their lifetime. This ability to grow indeterminately is due toa special form of tissue known as a"meristem".

A meristem is a region of tissue that is perpetually embryonic - that is, itis relatively undifferentiated and divides continuously, giving rise to all newcells and tissues that make up the plant body. Can you think of an analogy tothe plant's meristem in animals/humans?

There are several meristems distributed throughout the plant body whichare named for their location and/or for the tissue systems that are derivedfrom them.

APICAL (PRIM ARY) M ERIST EM S

Are located in the tips (or, apex) of the growing roots and shoots of allplants. Primary meristems produce primary tissues and account for all theprimary growth of plants. Primary growth is growth that contributes toelongation of the plant (height and root lengthening) and for the establishmentof its basic organ structure (roots, stems, leaves).

LAT ERAL (SECONDARY) M ERIST EM S

Are organized as concentric cylinders (think of a can inside a can inside acan) within the plant body and are responsible for secondary growth.Secondary growth contributes to the thickening of a plant structure, orincreases in girth. With few exceptions, lateral meristems tend to be presentonly in woody plants, such as trees, shrubs, and some vines.

As cells in the apical meristem divide and give rise to new cells, the tipsof roots and shoots extend outward, away from the central plant body. As aconsequence of this elongation due to the addition of new cells, the slightlyolder meristematic cells become distributed into different regions within theroot and shoot. These slightly more mature cells still retain their meristematicproperties, but are somewhat differentiated from the true apical meristem.These regions of meristematic tissue derived from the apical meristem arereferred to as"primary meristems" and are named for the tissues that theywill give rise to:

• Protoderm: Gives rise to dermal tissues,• Ground Meristem: Gives rise to ground tissues, and• Procambium: Gives rise to vascular tissue (aka,"cambial tissue").The organization of primary meristems also varies among plant types

and organs.

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Like other organisms, the cells in a plant are grouped together into varioustissues. These tissues can be simple, consisting of a single cell type, or complex,consisting of more than one cell type.

DERM AL T ISSUE SYST EM

The dermal tissue system consists of the epidermis and the periderm.The epidermis is a single layer of closely packed cells. It both covers andprotects the plant. It can be thought of as the plant's"skin." Depending on thepart of the plant that it covers, the dermal tissue system can be specialized toa certain extent. For instance, the epidermis of a plant's leaves secretes a coatingcalled the cuticle that helps the plant retain water.

EPIDERM IS CELLS

The periderm, also called bark, replaces the epidermis in plants thatundergo secondary growth. The periderm consists of cork cells and protectsthe plant from pathogens, prevents excessive water loss and providesinsulation for the plant. Xylem and phloem throughout the plant make upthe vascular tissue system. It allows water and other nutrients to betransported throughout the plant.

GROUND T ISSUE SYST EM

The ground tissue system synthesizes organic compounds, supports theplant and provides storage for the plant. It is mostly made up of parenchymacells but can also include some collenchyma and sclerenchyma cells as well.

PLANT TISSUE CULTURE

Practically any plant transformation experiment relies at some point ontissue culture. There are some exceptions to this generalization, but the abilityto regenerate plants from isolated cells or tissues in vitro underpins most planttransformation systems.

PLAST ICIT Y AND T OT IPOT ENCY

Two concepts, plasticity and totipotency, are central to understandingplant cell culture and regeneration. Plants, due to their long life span, have

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developed a greater ability to endure extreme conditions and predation thanhave animals. Many of the processes involved in plant growth anddevelopment adapt to environmental conditions. This plasticity allows plantsto alter their metabolism, growth and development to best suit theirenvironment.

Particularly important aspects of this adaptation, as far as plant tissueculture and regeneration are concerned, are the abilities to initiate cell divisionfrom almost any tissue of the plant and to regenerate lost organs or undergodifferent developmental pathways in response to particular stimuli. Whenplant cells and tissues are cultured in vitro they generally exhibit a very highdegree of plasticity, which allows one type of tissue or organ to be initiatedfrom another type. In this way, whole plants can be subsequently regenerated.This regeneration of whole organisms depends upon the concept that all plantcells can, given the correct stimuli, express the total genetic potential of theparent plant. This maintenance of genetic potential is called ‘totipotency’. Plantcell culture and regeneration do, in fact, provide the most compelling evidencefor totipotency.

The Culture Environment

When cultured in vitro, all the needs, both chemical and physical, of theplant cells have to met by the culture vessel, the growth medium and theexternal environment (light, temperature, etc.). The growth medium has tosupply all the essential mineral ions required for growth and development.In many cases (as the biosynthetic capability of cells cultured in vitro may notreplicate that of the parent plant), it must also supply additional organicsupplements such as amino acids and vitamins.

Many plant cell cultures, as they are not photosynthetic, also require theaddition of a fixed carbon source in the form of a sugar (most often sucrose).One other vital component that must also be supplied is water, the principalbiological solvent. Physical factors, such as temperature, pH, the gaseousenvironment, light (quality and duration) and osmotic pressure, also have tobe maintained within acceptable limits.

Plant Cell Culture Media

Culture media used for the in vitro cultivation of plant cells are composedof three basic components:

(1) essential elements, or mineral ions, supplied as a complex mixtureof salts;

(2) an organic supplement supplying vitamins and/or amino acids; and(3) a source of fixed carbon; usually supplied as the sugar sucrose.For practical purposes, the essential elements are further divided into

the following categories:(1) macroelements (or macronutrients);

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(2) microelements (or micronutrients); and(3) an iron source.

Media Components

It is useful to briefly consider some of the individual components of thestock solutions.

Macroelements

As is implied by the name, the stock solution supplies those elementsrequired in large amounts for plant growth and development. Nitrogen,phosphorus, potassium, magnesium, calcium and sulphur (and carbon, whichis added separately) are usually regarded as macroelements. These elementsusually comprise at least 0.1% of the dry weight of plants.

Nitrogen is most commonly supplied as a mixture of nitrate ions (fromthe KNO3) and ammonium ions (from the NH4NO3). Theoretically, there isan advantage in supplying nitrogen in the form of ammonium ions, as nitrogenmust be in the reduced form to be incorporated into macromolecules. Nitrateions therefore need to be reduced before incorporation.

However, at high concentrations, ammonium ions can be toxic to plantcell cultures and uptake of ammonium ions from the medium causesacidification of the medium.

In order to use ammonium ions as the sole nitrogen source, the mediumneeds to be buffered. High concentrations of ammonium ions can also causeculture problems by increasing the frequency of vitrification (the cultureappears pale and ‘glassy’ and is usually unsuitable for further culture). Usinga mixture of nitrate and ammonium ions has the advantage of weaklybuffering the medium as the uptake of nitrate ions causes OH¯ ions to beexcreted.

Phosphorus is usually supplied as the phosphate ion of ammonium,sodium or potassium salts. High concentrations of phosphate can lead to theprecipitation of medium elements as insoluble phosphates.

Microelements

These elements are required in trace amounts for plant growth anddevelopment, and have many and diverse roles. Manganese, iodine, copper,cobalt, boron, molybdenum, iron and zinc usually comprise themicroelements, although other elements such as nickel and aluminium arefrequently found in some formulations.

Iron is usually added as iron sulphate, although iron citrate can also beused. Ethylene Diamine Tetra acetic Acid (EDTA) is usually used inconjunction with the iron sulphate. The EDTA complexes with the iron so asto allow the slow and continuous release of iron into the medium.Uncomplexed iron can precipitate out of the medium as ferric oxide.

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Organic Supplements

Only two vitamins, thiamine (vitamin B1) and myo-inositol (considereda B vitamin) are considered essential for the culture of plant cells in vitro.However, other vitamins are also added to plant cell culture media.

Amino acids are also commonly included in the organic supplement. Themost frequently used is glycine (arginine, asparagine, aspartic acid, alanine,glutamic acid, glutamine and proline are also used), but in many cases itsinclusion is not essential.

Amino acids provide a source of reduced nitrogen and, like ammoniumions, uptake causes acidification of the medium. Casein hydrolysate can beused as a relatively cheap source of a mix of amino acids.

Carbon Source

Sucrose is cheap, easily available, readily assimilated and relatively stableand is therefore the most commonly used carbon source. Other carbohydrates(such as glucose, maltose, galactose and sorbitol) can also be used, and inspecialized circumstances may prove superior to sucrose.

Gelling Agents

Media for plant cell culture in vitro can be used in either liquid or ‘solid’forms, depending on the type of culture being grown. For any culture typesthat require the plant cells or tissues to be grown on the surface of the medium,it must be solidified (more correctly termed ‘gelled’).

Agar, produced from seaweed, is the most common type of gelling agent,and is ideal for routine applications.

However, because it is a natural product, the agar quality can vary fromsupplier to supplier and from batch to batch. For more demandingapplications, a range of purer (and in some cases, considerably moreexpensive) gelling agents are available.

These components, then, are the basic ‘chemical’ necessities for plant cellculture media. However, other additions are made in order to manipulatethe pattern of growth and development of the plant cell culture.

PLANT GROWTH REGULATORS

We have already briefly considered the concepts of plasticity andtotipotency. The essential point as far as plant cell culture is concerned is that,due to this plasticity and totipotency, specific media manipulations can beused to direct the development of plant cells in culture.

Plant growth regulators are the critical media components in determiningthe developmental pathway of the plant cells. The plant growth regulatorsused most commonly are plant hormones or their synthetic analogues.

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Classes of Plant Growth Regulators

There are five main classes of plant growth regulator used in plant cellculture, namely:

(1) auxins;(2) cytokinins;(3) gibberellins;(4) abscisic acid;(5) ethylene.Each class of plant growth regulator will be briefly looked at.

Auxins

Auxins promote both cell division and cell growth The most importantnaturally occurring auxin is IAA (indole-3-acetic acid), but its use in plantcell culture media is limited because it is unstable to both heat and light.Occasionally, amino acid conjugates of IAA (such as indole-acetyl-L-alanineand indole-acetyl-L-glycine), which are more stable, are used to partiallyalleviate the problems associated with the use of IAA. It is more common,though, to use stable chemical analogues of IAA as a source of auxin in plantcell culture media. 2,4-Dichlorophenoxyacetic acid (2,4-D) is the mostcommonly used auxin and is extremely effective in most circumstances. Otherauxins are available, and some may be more effective or ‘potent’ than 2,4-Din some instances.

CYTOKININS

Cytokinins promote cell division. Naturally occurring cytokinins are alarge group of structurally related (they are purine derivatives) compounds.Of the naturally occurring cytokinins, two have some use in plant tissueculture media. These are zeatin and 2iP (2-isopentyl adenine). Their use isnot widespread as they are expensive (particularly zeatin) and relativelyunstable. The synthetic analogues, kinetin and BAP (benzylaminopurine), aretherefore used more frequently. Non-purine-based chemicals, such assubstituted phenylureas, are also used as cytokinins in plant cell culture media.These substituted phenylureas can also substitute for auxin in some culturesystems.

Gibberellins

There are numerous, naturally occurring, structurally related compoundstermed ‘gibberellins’. They are involved in regulating cell elongation, and areagronomically important in determining plant height and fruit-set. Only afew of the gibberellins are used in plant tissue culture media, Gibberelic Acid3 (GA3) being the most common.

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Abscisic Acid

Abscisic acid (ABA) inhibits cell division. It is most commonly used inplant tissue culture to promote distinct developmental pathways such assomatic embryogenesis.

Ethylene

Ethylene is a gaseous, naturally occurring, plant growth regulator mostcommonly associated with controlling fruit ripening, and its use in plant tissueculture is not widespread. It does, though, present a particular problem forplant tissue culture.

Some plant cell cultures produce ethylene, which, if it builds upsufficiently, can inhibit the growth and development of the culture. The typeof culture vessel used and its means of closure affect the gaseous exchangebetween the culture vessel and the outside atmosphere and thus the levels ofethylene present in the culture.

Plant Growth Regulators and Tissue Culture

Generalisations about plant growth regulators and their use in plant cellculture media have been developed from initial observations made in the1950s. There is, however, some considerable difficulty in predicting the effectsof plant growth regulators: this is because of the great differences in cultureresponse between species, cultivars and even plants of the same cultivar grownunder different conditions.

However, some principles do hold true and have become the paradigmon which most plant tissue culture regimes are based. Auxins and cytokininsare the most widely used plant growth regulators in plant tissue culture andare usually used together, the ratio of the auxin to the cytokinin determiningthe type of culture established or regenerated.

A high auxin to cytokinin ratio generally favours root formation, whereasa high cytokinin to auxin ratio favours shoot formation. An intermediate ratiofavours callus production.

CULTURE TYPES

Cultures are generally initiated from sterile pieces of a whole plant. Thesepieces are termed ‘explants’, and may consist of pieces of organs, such as leavesor roots, or may be specific cell types, such as pollen or endosperm. Manyfeatures of the explant are known to affect the efficiency of culture initiation.Generally, younger, more rapidly growing tissue (or tissue at an early stageof development) is most effective. Several different culture types mostcommonly used in plant transformation studies will now be examined in bitdetail.

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CALLUS

Explants, when cultured on the appropriate medium, usually with bothan auxin and a cytokinin, can give rise to an unorganised, growing anddividing mass of cells. It is thought that any plant tissue can be used as anexplant, if the correct conditions are found. In culture, this proliferation canbe maintained more or less indefinitely, provided that the callus is subculturedon to fresh medium periodically.

During callus formation there is some degree of dedifferentiation (i.e. thechanges that occur during development and specialization are, to some extent,reversed), both in morphology (callus is usually composed of unspecialisedparenchyma cells) and metabolism. One major consequence of thisdedifferentiation is that most plant cultures lose the ability to photosynthesise.This has important consequences for the culture of callus tissue, as themetabolic profile will probably not match that of the donor plant. Thisnecessitates the addition of other components—such as vitamins and, mostimportantly, a carbon source—to the culture medium, in addition to the usualmineral nutrients.

Callus culture is often performed in the dark (the lack of photosyntheticcapability being no drawback) as light can encourage differentiation of thecallus. During long-term culture, the culture may lose the requirement forauxin and/or cytokinin. This process, known as ‘habituation’, is common incallus cultures from some plant species (such as sugar beet). Callus culturesare extremely important in plant biotechnology. Manipulation of the auxinto cytokinin ratio in the medium can lead to the development of shoots, rootsor somatic embryos from which whole plants can subsequently be produced.Callus cultures can also be used to initiate cell suspensions, which are usedin a variety of ways in plant transformation studies.

CELL-SUSPENSION CULT URES

Callus cultures, broadly speaking, fall into one of two categories: compactor friable. In compact callus the cells are densely aggregated, whereas in friablecallus the cells are only loosely associated with each other and the callusbecomes soft and breaks apart easily. Friable callus provides the inoculum toform cell-suspension cultures. Explants from some plant species or particularcell types tend not to form friable callus, making cell-suspension initiation adifficult task. The friability of callus can sometimes be improved bymanipulating the medium components or by repeated subculturing. Thefriability of the callus can also sometimes be improved by culturing it on ‘semi-solid’ medium (medium with a low concentration of gelling agent). Whenfriable callus is placed into a liquid medium (usually the same compositionas the solid medium used for the callus culture) and then agitated, single cellsand/or small clumps of cells are released into the medium. Under the correctconditions, these released cells continue to grow and divide, eventually

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producing a cell-suspension culture. A relatively large inoculum should beused when initiating cell suspensions so that the released cell numbers buildup quickly. The inoculum should not be too large though, as toxic productsreleased from damaged or stressed cells can build up to lethal levels. Largecell clumps can be removed during subculture of the cell suspension. Cellsuspensions can be maintained relatively simply as batch cultures in conicalflasks. They are continually cultured by repeated subculturing into freshmedium. This results in dilution of the suspension and the initiation of anotherbatch growth cycle. The degree of dilution during subculture should bedetermined empirically for each culture. Too great a degree of dilution willresult in a greatly extended lag period or, in extreme cases, death of thetransferred cells.

After subculture, the cells divide and the biomass of the culture increasesin a characteristic fashion, until nutrients in the medium are exhausted and/or toxic by-products build up to inhibitory levels—this is called the ‘stationaryphase’. If cells are left in the stationary phase for too long, they will die andthe culture will be lost. Therefore, cells should be transferred as they enterthe stationary phase. It is therefore important that the batch growth-cycleparameters are determined for each cell-suspension culture.

Protoplasts

Protoplasts are plant cells with the cell wall removed. Protoplasts are mostcommonly isolated from either leaf mesophyll cells or cell suspensions,although other sources can be used to advantage. Two general approaches toremoving the cell wall (a difficult task without damaging the protoplast) canbe taken—mechanical or enzymatic isolation. Mechanical isolation, althoughpossible, often results in low yields, poor quality and poor performance inculture due to substances released from damaged cells. Enzymatic isolationis usually carried out in a simple salt solution with a high osmoticum, plusthe cell wall degrading enzymes. It is usual to use a mix of both cellulase andpectinase enzymes, which must be of high quality and purity. Protoplasts arefragile and easily damaged, and therefore must be cultured carefully. Liquidmedium is not agitated and a high osmotic potential is maintained, at least inthe initial stages. The liquid medium must be shallow enough to allow aerationin the absence of agitation. Protoplasts can be plated out on to solid mediumand callus produced. Whole plants can be regenerated by organogenesis orsomatic embryogenesis from this callus. Protoplasts are ideal targets fortransformation by a variety of means.

ROOT CULTURES

Root cultures can be established in vitro from explants of the root tip ofeither primary or lateral roots and can be cultured on fairly simple media.

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The growth of roots in vitro is potentially unlimited, as roots are indeterminateorgans. Although the establishment of root cultures was one of the firstachievements of modern plant tissue culture, they are not widely used in planttransformation studies.

SHOOT T IP AND M ERIST EM CULT URE

The tips of shoots (which contain the shoot apical meristem) can becultured in vitro, producing clumps of shoots from either axillary oradventitious buds. This method can be used for clonal propagation. Shootmeristem cultures are potential alternatives to the more commonly usedmethods for cereal regeneration as they are less genotype-dependent and moreefficient (seedlings can be used as donor material).

Embryo Culture

Embryos can be used as explants to generate callus cultures or somaticembryos. Both immature and mature embryos can be used as explants.Immature, embryo-derived embryogenic callus is the most popular methodof monocot plant regeneration.

Microspore Culture

Haploid tissue can be cultured in vitro by using pollen or anthers as anexplant. Pollen contains the male gametophyte, which is termed the‘microspore’. Both callus and embryos can be produced from pollen. Two mainapproaches can be taken to produce in vitro cultures from haploid tissue.

The first method depends on using the anther as the explant. Anthers(somatic tissue that surrounds and contains the pollen) can be cultured onsolid medium (agar should not be used to solidify the medium as itcontainsinhibitory substances). Pollen-derived embryos are subsequentlyproduced via dehiscence of the mature anthers. The dehiscence of the antherdepends both on its isolation at the correct stage and on the correct cultureconditions. In some species, the reliance on natural dehiscence can becircumvented by cutting the wall of the anther, although this does, of course,take a considerable amount of time. Anthers can also be cultured in liquidmedium, and pollen released from the anthers can be induced to formembryos, although the effi-ciency of plant regeneration is often very low.Immature pollen can also be extracted from developing anthers and cultureddirectly, although this is a very time-consuming process.

Both methods have advantages and disadvantages. Some beneficial effectsto the culture are observed when anthers are used as the explant material.There is, however, the danger that some of the embryos produced from antherculture will originate from the somatic anther tissue rather than the haploidmicrospore cells. If isolated pollen is used there is no danger of mixed embryoformation, but the efficiency is low and the process is time-consuming. In

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microspore culture, the condition of the donor plant is of critical importance,as is the timing of isolation. Pretreatments, such as a cold treatment, are oftenfound to increase the efficiency. These pretreatments can be applied beforeculture, or, in some species, after placing the anthers in culture.

Plant species can be divided into two groups, depending on whether theyrequire the addition of plant growth regulators to the medium for pollen/anther culture; those that do also often require organic supplements, e.g. aminoacids. Many of the cereals (rice, wheat, barley and maize) require mediumsupplemented with plant growth regulators for pollen/anther culture.

Regeneration from microspore explants can be obtained by directembryogenesis, or via a callus stage and subsequent embryogenesis.

Haploid tissue cultures can also be initiated from the female gametophyte(the ovule). In some cases, this is a more efficient method than using pollenor anthers.

The ploidy of the plants obtained from haploid cultures may not behaploid. This can be a consequence of chromosome doubling during theculture period. Chromosome doubling (which often has to be induced bytreatment with chemicals such as colchicine) may be an advantage, as in manycases haploid plants are not the desired outcome of regeneration from haploidtissues. Such plants are often referred to as ‘di-haploids’, because they containtwo copies of the same haploid genome.

PLANT REGENERATION

Having looked at the main types of plant culture that can be establishedin vitro, we can now look at how whole plants can be regenerated from thesecultures.

In broad terms, two methods of plant regeneration are widely used inplant transformation studies, i.e. somatic embryogenesis and organogenesis.

SOM AT IC EM BRYOGENESIS

In somatic (asexual) embryogenesis, embryo-like structures, which candevelop into whole plants in a way analogous to zygotic embryos, are formedfrom somatic tissues. These somatic embryos can be produced either directlyor indirectly. In direct somatic embryogenesis, the embryo is formed directlyfrom a cell or small group of cells without the production of an interveningcallus. Though common from some tissues (usually reproductive tissues suchas the nucellus, styles or pollen), direct somatic embryogenesis is generallyrare in comparison with indirect somatic embryogenesis.

In indirect somatic embryogenesis, callus is first produced from theexplant. Embryos can then be produced from the callus tissue or from a cellsuspension produced from that callus. Somatic embryogenesis usuallyproceeds in two distinct stages. In the initial stage (embryo initiation), a high

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concentration of 2,4-D is used. In the second stage (embryo production)embryos are produced in a medium with no or very low levels of 2,4-D.

In many systems it has been found that somatic embryogenesis isimproved by supplying a source of reduced nitrogen, such as specific aminoacids or casein hydrolysate.

Organogenesis

Somatic embryogenesis relies on plant regeneration through a processanalogous to zygotic embryo germination.

Organogenesis relies on the production of organs, either directly froman explant or from a callus culture. There are three methods of plantregeneration via organogenesis. The first two methods depend on adventitiousorgans arising either from a callus culture or directly from an explant.Alternatively, axillary bud formation and growth can also be used toregenerate whole plants from some types of tissue culture. Organogenesisrelies on the inherent plasticity of plant tissues, and is regulated by alteringthe components of the medium. In particular, it is the auxin to cytokinin ratioof the medium that determines which developmental pathway theregenerating tissue will take. It is usual to induce shoot formation by increasingthe cytokinin to auxin ratio of the culture medium. These shoots can then berooted relatively simply.

PLANT T ISSUE ANALYSIS

Crop nutrient uptake is influenced by many factors other than the soiltest. Soil testing and plant analysis are designed to work together. Soil testingidentifies the soils nutrient reserves and predicts the nutrient needs, whileplant analysis identifies the actual nutrient uptake.

Plant tissue analysis is a useful diagnostic tool and can greatly help withfertility management. From emergence through the first six weeks of growth,plant analysis is helpful in identifying nutrient uptake. Testing corn leafsamples at this early stage may indicate where additional nitrogen should beapplied. This is a way to determine the cost-effectiveness of the additionalapplication of fertilizer.

Plant analysis may be utilized at the reproductive stage (full grown) todetermine nutrient uptake for that season. This allows for determination offertility requirements for the upcoming growing season. Tissue sampling andanalysis is a key tool in determining the soil’s ability to meet the sufficientnutrient requirements of the crop, thereby reducing any possible nutrientstress that may result in a potential yield drag or loss. A variety of tests areavailable, including phosphorus, potassium, calcium, magnesium, sulfur,sodium, zinc, iron, manganese, copper, nitrogen, and boron.

Remember: test results are only as good as the sample. The most commonmistake in plant sampling is not obtaining enough tissue for an accurate

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analysis. Several leaves from several plants will ensure a more representativesampling and ensure enough plant material. Fertility management samplesshould be collected from representative areas of the field. Always takecomposite samples. One plant is not a sample. Generally, select only uppermature leaves for your tissue sample. When samples are being collected fornutrient review, specific leaves are normally selected depending upon theplant. Here are a few examples:

• Corn-at tasseling, collect the leaf at the ear node,• Soybeans-at flowering collect the most mature trifoliant from the

top of the plant,• Wheat and small grain-at beginning of head set collect the flag leaf,• Alfalfa-at flowering collect the top 6 inches of the plant.When troubleshooting is the issue, it is extremely important to take leaves

from several plants experiencing the same problem and use this compositefor one sample. Compare this analysis with a composite sample taken fromthe non-affected area of the field. Sample #2 is the control sample. This givesthe grower an idea of the sufficient nutrient level for that crop in that field atthat sampling period.

For example: a portion of a corn field is exhibiting chlorosis (yellowing)and stunted growth. Plant tissue analysis is the best way to determine thereason for the chlorosis and abnormal growth. A composite sample of uppermature leaves from 6 to 10 chlorotic plants and a composite sample takenfrom the upper mature leaves from 6 to 10 plants located in the non-affectedarea of the field are submitted for analysis.

Where to Sample

Avoid leaves that were heavily preyed upon by insects or mechanicallydamaged. Avoid taking samples from areas located in border rows or heavyshade. Do not collect tissue that is already dead. Do not take samples fromcontaminated or chemically altered plants. Take diagnostic samples from theaffected site for one composite sample and one from the non-affected area fora second composite sample. If the entire field is distressed, collect a compositesample over the entire area and note that the entire field is affected.

When to Sample

Timing of diagnostic sampling depends on the first sign of trouble. Theearlier the problem is detected, the easier it is to treat with fertilizer or limeapplication. If nutrient application is not feasible due to plant growth, theseresults, along with soil analysis, can be used in fertilizer recommendationsfor the following growing season. Fertility management sampling is done atthe time of flowering or pollination (reproductive stage). Avoid taking samplestoo late in the growing season. Take a single composite sample at the onset ofthe reproductive stage to measure nutrient uptake throughout the growing

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season. Take samples when the plant is still viable and growing. It is too lateonce it has started to produce seed. These values determine the followinggrowing season’s fertility needs.

SUBMITTING THE SAMPLE

Place each dried composite sample in a paper bag. Do not use plasticbags for plant samples because the plants mold. Label each bag with the fieldname and sample number. Please label them good, bad, affected or non-affected. Please include information that contains: name, address, telephonenumber, field names, number of samples per field and the type of test.

A plant’s ability to take up adequate nutrients is influenced by manyfactors such as soil fertility level, soil temperature, soil compaction, soilmoisture, nutrient balance in the soil, fertilizer applications, plant genetics,and many other factors. A chemical soil test cannot accurately account for allof these factors. A producer must use plant analysis if he expects to identifyand understand the nutrient efficiency and needs of his crops and soils. Onlyplant analysis can tell us the actual nutrient status of the crop. Plant analysiscan be used to detect low nutrient levels in plants before they are seen by thehuman eye. Once visual symptoms are visible, top yields and quality havealready been compromised. Plant analysis is an important part of planningfuture fertility programmes.

Sampling Guide

Our plant sampling guide contains information on most plant species. Ifyou need furthur assistance, contact the lab for specific information on yourcrop.

USES OF PLANT ANALYSIS

• Plant analysis can be used by turf managers to;• Confirm suspected nutrient deficiency symptoms;• Verify toxicities;• Reveal hidden hunger (i.e. plants show no visible symptoms, but

the nutrient content is low enough to reduce growth or affect qualitycharacteristics);

• Aid in evaluating the efficiency of applied fertilisers;• Assist in formulating fertilisation practices; • Monitor the nutrient status of plants throughout the growing season

to determine whether each nutrient is present in sufficientconcentration for optimum growth characteristics.

Plant analysis is a proven and effective means of predicting fertilizer needsfor many plant species. However, it does not completely replace a soil test.Soil and plant analysis serve different purposes and when properly used they

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compliment each other in providing detailed information for maximizing theefficiency of fertility programmes. Soil testing is based on the concept thatthe concentration of a particular nutrient in a given volume of soil reflectswhether or not the nutritional level of that soil is adequate for optimum plantgrowth or production. Plant analysis is based in part on the concept that theamount of a specific nutrient in the plant tissue is related to the plantavailability of that element in the soil. Plant analysis also reflects nutrientuptake conditions in the soil. Soil properties such as compaction, root disease,impervious layers and/or poor drainage may inhibit the uptake of nutrientsby plants. Or a low concentration of one nutrient in the plant may result fromthe excessive application of another nutrient. Conversely, favourable soilphysical properties and optimum soil moisture may accentuate nutrientuptake even though the soil may not have an abundant supply of nutrients.

As a result of these soil-plant interactions, there are certain instances whencontradictions occur between soil and plant analysis results. For example,assume turf is growing on a soil in which the soil tests revealed a mediumlevel of extractable Magnesium. A plant analysis from the area a few weekslater indicates that Magnesium is low. Immediately, the validity of the testresults are questioned, which is an absolutely normal response. However, acloser examination of the plant analysis results revealed that the Calcium andPotassium concentrations of the turf were high.

Upon checking the information accompanying the plant analysis results,it was noted that Lime (CaCO3) had been applied at 500g/m2 and a high rateof Potassium was applied in the fertilizer programme. As a result of thesetwo management practices, the level of Calcium and Potassium in the soilwas sufficient to reduce the uptake of Magnesium. This is one example ofhow soil testing and plant analysis can be used together for making betternutrient management decisions. Plant analysis can also be used to supplementa soil testing programme. It is particularly useful in distinguishing betweenNitrogen and Sulphur deficiencies in turf, as deficiency symptoms of the twoelements are similar. Plant analysis offers an excellent means of delineatingwhich element is deficient (which cannot be ascertained through soil testing).If this distinction is not made properly and the wrong corrective treatment isapplied, plant growth can be affected appreciably.

In the case of most turf grass species, a soil analysis prior to active growthin spring makes it possible to determine whether lime, dolomite, gypsum and/or specific fertilizer applications will be needed. Plant analysis of the turfduring the growing season will indicate if the applied materials were effectiveand whether the predictions based on the soil analysis were correct.

A& L EAST ERN LABS-SAM PLING FOR PLANT ANALYSIS

Plant analysis assesses nutrient uptake while soil testing predicts nutrientavailability. The two tests are complementary as crop management tools. Plant

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analysis will detect unseen hidden hunger and confirm visual deficiencysymptoms. Toxic levels may also be detected. If it is done early, plant analysiswill allow a corrective fertilizer application in the same season. A basicknowledge of plant structure is necessary before collecting samples. A leaf ismade up of a leaf “blade” and a “petiole”. The petiole is the stalk attached tothe blade. A compound leaf may have several “leaflets” attached to it. In somecases, only terminal “leaflets” may be sampled, as in the case of walnuts andpistachios. A common error in tomatoes is when only leaflets are sampledinstead of the whole compound leaf. This shows the importance ofunderstanding proper sampling technique.

The most recent mature leaf (MRML) is the first fully expanded leaf belowthe growing point. It is neither dull from age nor shiny green from immaturity.For some crops, the most recent mature leaf is a compound leaf. The mostrecent mature leaf on soybean and strawberry, for example, is a trifoliatecompound leaf: three leaflets comprising one leaf.

For cotton, grape, potato and strawberry, petioles provide an additionalindication of nitrogen status. When sampling these crops, collect most recentmature leaves and their petioles. Detach leaves from petioles in the field tostop the translocation of nutrients. Put petioles in a separate bag. “Midribs”are the middle ribs to large leaves such as corn, lettuce, and cabbage, andwould equate to a petiole sampling.

Deciding When to Sample

To monitor plant nutrient status most effectively, sample during therecommended growth stages for your specific crop Take samples weekly orbiweekly during critical periods, depending on management intensity andcrop value. However, to identify a specific plant growth problem, take sampleswhenever you suspect the problem.

The best time to collect samples is between mid-morning and mid-afternoon. Nitrate nitrogen varies with time of day and prevailing conditionsbut generally not enough to alter interpretation. Sampling during dampconditions is okay but requires extra care to prevent tissue from decomposingduring shipping. Keep samples free of soil and other contaminants that canalter results.

The appropriate part of the plant to sample varies with crop, stage ofgrowth, and purpose of sampling. When sampling seedlings less than 4 inchestall, take whole plants from 1 inch above the soil line. For larger plants, themost recent mature leaf is the best indicator sample.

Taking A Representative Sample

Proper sampling is the key to reliable plant analysis results. A samplecan represent the status of one plant or 20 acres of plants. In general, acommon-sense approach works well. When problem solving, take samples

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from both “good” and “bad” areas. Comparison between the two groups ofsamples helps pinpoint the limiting element. Comparative sampling also helpsfactor out the influence of drought stress, disease, or injury. Take matchingsoil samples from the root zones of both “good” and “bad” plants for themost complete evaluation.

When monitoring the status of healthy plants, take samples from auniform area. If the entire field is uniform, one sample can represent a numberof acres. If there are variations in soil type, topography, or crop history, takemultiple samples so that each unique area is represented by its own sample.

Choosing Sample Size

The actual laboratory analysis requires less than one gram of tissue.However, a good sample contains enough leaves to represent the areasampled. Therefore, the larger the area is, the larger the sample size needs tobe.

Sample size also varies with crop. For crops with large leaves, like tobacco,a sample of three or four leaves is adequate. For crops with small leaves, likeazalea, a sample of 25 to 30 leaves is more appropriate. For most crops, 8 to15 leaves is adequate. For crops requiring petiole analysis, collect at least 15to 20 leaves.

Submitting the Sample

Send the completed information sheet and proper fee with each sample.Use permanent ink or pencil on sample forms and bags Avoid numberingsamples simply as 1,2,3 as it may lead to confusion later. Give each samplea unique identifier that will help you remember the plants or area itcorresponds to-such as HOUSE1, 15B, GOOD, or BAD. You can use up to sixletters and/or numbers. Put the identifier on both the information sheet andthe sample envelope. Pay attention to detail when filling out the informationsheet. Note any conditions-drought, disease, injury, pesticide or foliar nutrientapplications-that might be relevant. Indicate the analysis desired and providevery specific information on stage of growth and plant part if an interpretationis required. The laboratory does not automatically provide an interpretation,as some clients prefer to make their own.

Diagnostic interpretations require more details than predictive. Whensending matching soil, solution, or waste samples, indicate the matchingsample ID in the designated areas on the information sheet. Be sure the growername and address are exactly the same on all matching information sheets.Ship all matching samples in the same container. Ship the tissue sample in apaper envelope or cardboard box so it can begin drying during transport.Samples put in plastic bags will rot, and decomposition may alter test results.

If samples are very wet, air-dry to a workable condition before packaging.Otherwise, decomposition or molding will occur. Include a completed plant

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analysis information sheet or cover letter with instructions within the samepackage. Processing will be delayed if sent separately. Also, include paymentif you do not have an established account. Samples should be shipped by acarrier such as UPS or FEDEX, or by first class mail.

Interpreting the Report

Samples are analyzed the next day of their arrival. The promptturnaround makes it possible for growers to take any corrective action neededto salvage the current crop. The report can be emailed or post mailed to thegrower.

PLANT ANALYSIS AS A GUIDE FORMINERAL NUTRITION OF SUGAR BEETS

Nitrogen is the nutrient most likely to limit growth and sucroseproduction in sugar beets. Profitable responses to P fertilization are becomingcommon. Potassium fertilization is necessary in certain soils of the Sacramento-San Joaquin Delta. Sulfur is deficient in some soils of the upper SacramentoValley. Micronutrient deficiencies of sugar beets are uncommon in California.

Research and practical experience have shown that plant analysis canserve as an effective guide in the fertilization of sugar beets. A plant analysisprogramme, properly carried out, will help optimize income for the producer,provide quality beets for processing, conserve fertilizer, and minimize theleaching of nutrients to underground water supplies.

CRIT ICAL CONCENT RAT IONS

Critical concentrations of most of the essential nutrients have beendetermined for various parts of the sugar beet plant. These values along withconcentration ranges found in healthy plants and levels in plants withdeficiency symptoms.

The critical concentration for any nutrient is a relatively constant valuewhich changds little with sugar beet variety, climate, or soil conditions.Nutrient concentrations, on the other hand, are changed by conditions thataffect plant growth. For example, petioles of mature leaves of plants infectedwith the beet yellows virus may be higher in N03-N concentration than arehealthy plants. This is because diseased plants grow more slowly, and nitrateaccumulates in their petioles.

Potential users of sugar beet leaf analysis sometimes express concern thatdifferences in plant moisture content at different times of the day maysignificantly alter the concentrations of nutrients in plant tissues and thus affectthe interpretation of analytical results. In field experiments we observed littleor no change in dry-weight N03-N concentration in petioles samples hourlyfrom sunrise to sunset. Nor were there important differences with moderate

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(slight wilting) compared to no moisture stress. Only when plants wereseverely stressed was there a significant decrease in petiole nitrate. Plantsmoderately stressed compared to those severely stressed (loss of many leaves)had N03-N concentrations of 5, 1 00 and 3,600 ppm, respectively.

Differences in the nutrient concentration due to variety, weather, moisturestress, disease and the like, are largely related to the kind and extent of rootdevelopment or to other growth factors, and not to differences in internalrequirements.

SAM PLING AND EVALUAT ION

For the sugar beet, recently matured leaves (the youngest leaves that havejust attained maximum size) are usually the most appropriate part to sample.Petioles are most satisfactory for evaluating the plant’s nutritional status forN, P, and Cl, while the blade is most satisfactory for the other essentialelements.

A series of four samplings should be made throughout the growingseason: one at thinning time, the second at early mid-season, the third at laternid-season, and the fourth just before harvest. By comparing the analysis ofthese samples to critical nutrient concentrations, it is possible to detect andcorrect impending nutrient deficiencies before they occur, and to change thetiming or method of fertilization to assure that fertilizer needs of the crop arefully met.

Large amounts of N are required to insure adequate top and root growth,but if storage roots are to be high in sucrose concentration the plants must beN deficient for 4 to 8 weeks before harvest. An estimate of the amount offertilizer N needed for a specific crop should be made according to past historyof the field or by soil tests.

This amount should be applied to the crop early in the growing season,not later than thinning time. If an error is made, it is best to make it on theside of under-fertilization, because such a mistake can be corrected. If,however, the initital fertilizer application is more than the crop can use,fertilizer will be wasted, sucrose concentration (and, consequently, crop value)will be depressed, and excess nitrate will possibly be leached to the groundwater.

Adequacy of the initial fertilizer application can be ascertained byanalyzing petiole samples taken at 4 and 2 weeks before mid-season-the datemidway between the dates of emergence and harvest. (To determinemidseason for a crop to be overwintered, determine the date midway betweenemergence and November 1.) Indication of an impending deficiency of morethan 10 weeks before harvest can be corrected by an addition of from 40 to 80pounds of N per acre. These two early-season petiole samples can alsocontribute to the overall evaluation of the fertilizer programme as outlinedabove.

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Abnormal Plant Growth

Frequently, plants in a field or part of a field show abnormal growth. Aknowledge of nutrient deficiency symptoms gained from previous experienceor a printed visual guide may lead to a tentative diagnosis which can beconfirmed or discarded by analyses of comparable petioles and blades ofabnormal and normal appearing plants. A useful visual guide is: Sugar BeetNutrient Deficiency Symptoms, A Colour Atlas and Chemical Guide (Ulrichand Hills, 1969). To confirm a diagnosis of deficiency symptoms by chemicalanalysis, plants should be sampled soon after the symptoms appear. After aprolonged period of stress, the concentration of certain nutrients increases inphysiologically mature tissue, especially if new supplies of the nutrient becomeavailable to the plant.

Guide to Harvesting

When there is a choice of fields to be harvested, the results of a systematicplant analysis programme can serve as a guide to scheduling of beet harvest.Those fields depleted of N first would be scheduled for an early harvest; thosehigh in N would not be harvested until the N is depleted or would be held aslong as possible before harvesting. Root pulp as well as petioles may beanalyzed to determine depletion of N before harvest. Pulp from preharvestroot samples may be analyzed for nitrate by a semi-quantitative spot test usingthe diphenylamine reagent or, more accurately, by other quantitative methods.To maximize the sucrose concentration of beet roots, petioles and/or rootsshould be below critical N03-N levels about 4 to 8 weeks before harvest.

Nutrient Survey

A nutrient survey of sugar beet fields in an area can result in detection ofdeficiencies and identification of fields on which future fertilizer trials shouldbe located. Surveys conducted by Farm Advisors or other interestedagricultural groups can do much to improve fertilizer practices in an area.

Sampling Procedure

To serve as a measure of variability within a field, a minimum of two,but preferably four, samples should be taken from each field. From 25 to 50leaves are usually sufficient to reasonably estimate the true mean of thenutrient concentration in the plants represented by the sample. A convenientsampling system is to divide a field into imaginary quarters. The samplerwalks across the centre of each quarter at right angles to the plant rows andcollects from 25 to 50 leaves per quarter of the field. Samples are placed in apaper bag of convenient size and taken to the laboratory for processing assoon as possible. Delays of more than 48 hours at ordinary temperaturesshould be avoided, and storage in closed compartments of automobiles duringwarm weather should not be allowed. If samples must be stored before drying

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they should be kept at 5’C; at this temperature, sugar beet petioles changevery little in nitrate, phosphate, and K concentration.

For microelement analyses, blades must be free of dust and should bewashed for 30 seconds in a 0. I N HC1 bath followed by two successive rinsesin distilled water.

A sample is reduced in size by cutting it into small pieces (approximately1/8 inch), thoroughly mixing the pieces and then placing a sub-sample of about100 grams (a large handful) of fresh material in a cut-down paper bag or othersuitable container and drying it overnight in an oven, preferably with a forceddraft, at 70’C to 80’C (158'- 1 76’F). After drying, samples are ground to passa 40-mesh screen and stored. Samples treated in this way have been kept formany years without significant change in their mineral composition.

INTERPRETATION AND LIMITATIONS

With plants in the field, the likelihood of a growth response from theaddition of a nutrient to the soil will depend upon whether the nutrientconcentration of the plants is above or below the critical level. When thenutrient concentration is above the critical level and remains there throughoutthe growing season, there is little chance of a response in growth from theaddition of more of the nutrient. When the nutrient concentration of the plantsfalls below the critical level, the chance of a growth response becomes greaterthe longer the plants remain deficient, provided other growth factors arefavourable (e.g., climate is favourable for rapid growth, and growth is notseriously limited by disease, water stress, and the like). If the deficiencyappears late in the growing season, there is little chance for a significant yieldincrease with the addition of more of the nutrient as time is required for anydeficiency to cause a measurable decline in crop yield.

When plant material for a sample is taken from a large area, the samplemay differ considerably in nutrient concentration within itself. When theaverage nutrient concentration of the sample is at or near the critical level,the material collected will represent both deficient and nondeficient plants. Ifthere are areas in a field differing in plant-growth characteristics, these areasshould be sampled separately.

Analysis of plant tissue usually reveals only one deficiency at a time. Asecond nutrient, or even a third, may be in short supply, but due to reducedgrowth caused by the primary nutrient deficiency, other nutrients willaccumulate in the tissue. However, when the primary deficiency is corrected,the increased growth will decrease the concentration of the second nutrient,and most likely the second nutrient will become deficient rather soon. Anexception to this general rule is with regard to P, deficient surgar beets which,particularly as seedlings, are also occasionally deficient in N, because Pdeficiency retards nitrate uptake.

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In the interpretation of the results for multiple sampling dates, the sharperthe drop in nutrient concentrations and the earlier in the season it occurs, thegreater the degree of deficiency. Even when the results of several samplingdates indicate the plants have been deficient for a considerable period, theexact degree of deficiency is not known. At harvest, however, the degree ofdeficiency for a given field can be estimated in terms of the maximum yieldfor the area, and fertilization for the next crop is then adjusted to this finding.For example, a field that produced only 25 percent of the potential yield ismuch more deficient and will require more fertilizer than one that produced90 percent of its potential.

One of the greatest values of plant analysis is the prevention of deficienciesrather than their correction. Trends in nutrient content of leaves followed overa period of years can be a valuable guide in delineating the kinds, amounts,frequency, and methods of fertilization for more efficient crop production ona given field. Nutrient deficiencies can be predicted or at least detected assoon as they occur, and corrective measures can be taken before serious croploss takes place.

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5

Fruit Crop Cultivation

Fig. Crop Cultivation.

CROPS GROWN

India grows crops like apple (Malus pumila Mill.), pear (Pyrus communisBerm.f.), peach/nectarine (Prunus persica (L). Bats. Ch.), plum (Prunus domesticaL.), apricot (Prunus armeniaca L.), sweet cherry (Prunus avium L.) and sourcherry (Prunus cerasus L.) on a commercial scale.

CULT IVARS

The promising cultivars of different temperate fruits in 3 major deciduousfruit growing States of India. Some relevant information on cultivars andcultivar selection is indicated below:

Apple: Over 700 accessions of apple, introduced from USA, Russia, U.K.,Canada, Germany, Israel, Netherlands, Australia, Switzerland, Italy andDenmark have been tried and tested during the last 50 years. The deliciousgroup of cultivars predominates the apple market.

The areas covered under Delicious cultivars are: 83% of the area underapple in H.P., 45% in J&K and 30% in U.P. hills.

In more recent times improved spur types and standard colour mutantswith 20-50% higher yield potential are favoured.

The important selections are:• Spur types-Red spur, Starkrimson, Golden spur, Red Chief and

Oregon spur.• Colour mutants-Vance Delicious, Top Red, Skyline Supreme.• Low chilling cultivars-Michal, Schlomit.• Early cultivars-Benoni, Irish Peach, Early Shanburry, Fanny.• Juice making cultivars-Lord Lambourne, Granny Smith, Allington

Pippin.• Scab resistant cultivars-Co-Op-12, Florina, Firdous, Shirean.• New Hybrids-Lal Ambri (Red Delicious X Ambri), Sunehari (Ambri

X Golden Delicious), Amred (Red Delicious X Ambri), ChaubatiaAnupam & Chaubatia Princess (Early Shanberry X Red Delicious)developed in India.

In H.P. monoculture of a few cultivars such as Royal Delicious, RedDelicious and Richared have started showing negative impact on the appleindustry. Serious problems of apple scab disease and outbreak of prematureleaf fall and infestation of red spider mite are causing great concern. U.P. Hills,particularly the Kumaon hills division, have the unique advantage of earlyharvest of apple, mainly due to cultivation of early maturing varieties likeEarly Shanburry, Fanny and Benoni. The early maturing varieties areharvested 2-3 weeks before the arrival of fresh apple from H.P. and J&K, andhence fetch very remunerative prices.

Pear: In pear for higher altitude conditions high chilling requirementvarieties (like Bartlett) are mainly grown. In more recent years, red-colourstrains of pear like Max Red Bartlett, Red Bartlett and Starking are replacingyellow coloured cultivars. In warmer sub-mountainous areas of H.P. and sub-tropical Punjab oriental pear cultivars like Baghugosha, Kieffer, China andsand pear Patharnakh are cultivated commercially both for table andprocessing purpose.

Apricot: Generally there are two types of apricot, namely, sweet kerneltype and bitter kernel type. About 81 exotic accessions and 19 indigenouscultivars were collected and evaluated. The local types Halman andRakhaikarpo have been popular, whereas exotic introductions namely, Nari,Kaisha, Shakarpa and New Castle are promising. These cultivars arerecommended for dry cold areas. The USA variety Nugget is self-fruitful andbears sweet and attractive coloured fruits.

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Peach: For colder conditions the peach cultivars July Elberta, Elberta,Peshwari, Quetta, Burbank and Stark Earliglo are well adopted. Low-chillingcultivars viz. Flordasum, Flordared, Shan-e-Punjab, Sharbati and Sunred(nectarine) have become popular in subtropical belts of U.P. and Punjab States.

Plum: A large number of cultivars (283) have been introduced into thecountry. European plums performed better in the hills, while Japanese plumsare more adopted in sub-mountainous lower elevations. Leading cultivar inthe hills is Santa Rosa. In the north-Indian plains small fruited cultivars likeTitron, Kala Amritsari, Kelsey, and Alubukhara showed better performance.A good number of low-chilling Florida hybrids (Fla-1-2, Fla 73-4, Fla 85-2, 85-3, Fla 86-4) Sungold, Redgold etc., are under evaluation.

Cherry: Many cultivars of sweet cherry have been introduced fromEurope, USSR and British Columbia. Promising exotic cultivars like BigarreanNapoleon, Black Heart, Guigne Noir for J&K and cultivars like Black TartarianBing, Napoleon (white) Sam, Sue (White), Shella for H.P. have been identified.For warmer climate, cultivars like Summit, Sunburst, Lapins, Compacat andStella have been found to be promising.

PROCESSING IN FRUIT CROPS

Seed Extraction

In general, fruits or seeds for storage should be harvested at fully maturedstage, otherwise it affects the seed longevity. Tree seeds should be collectedfrom a single plant if not sufficient, then seeds from neighbouring plants withsame age be collected. It is safer to carry the fruits than seeds. Moist extractedseeds may germinate or begin to germinate during shipment.

Also extracted seeds may lose moisture rapidly and damage the seedquality especially in recalcitrant types and may rot. It is desirable to store thefruit temporarily when delay is expected for seed storage. A relatively smallquantity of seeds are required for long term storage. It is desirable to removeseeds manually, followed by soaking in water and fermentation and chemicaltreatments.

Seed Drying

Moisture content of seed is most important factor in determining the seedlongevity. After removal of seeds, simple washing is sufficient to remove anysubstance adhering to the seeds. Further, seeds are to be dried based on theirstorage behaviour.

It is desirable to dry seeds to 5-7% moisture in orthodox group. Seeds oftropical species should be dried at cool temperature of about 15°C. Delay indrying or drying at higher temperature considerably affect the viability inorthodox seeds especially those rich in oils. Silica gel is an effective desiccantcommonly used for seed drying.

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Seed moisture is determined as per the guidelines prescribed byInternational Seed Testing Association (ISTA). For large seeds, moisturecontent determination of about 20-25 individuals seeds is recommended.Separate determination of the moisture content of embryo or embryonic axes,cotyledons or endosperm and the seed covering structure may also be helpfulin interpreting the results.

Seed Packing

Seed storage in suitable containers will prevent the direct contact of seedswith storage environment and also protect from pests and diseases. Containerslike paper, cloth, polyethylene bags, glass bottle, cans and aluminium foillaminated pouches are being used as containers. The choice of containersdepends on kind and quantity of seeds, duration and condition of storage.Paper and cloth bags are cheap and can be used for short term storage.Aluminium cans and foil laminated pouches are effective for retaining theviability for longer period.

Storage Environments

Storage temperatures, humidity and oxygen are important factorsassociated with storage of seeds. The metabolic activity and incidence of pestsand diseases increase at high seed moisture and high storage temperaturesresulting in loss of viability. Thus, low moisture at low temperature is effectivein maintaining seed viability especially for orthodox type of seeds. Such seedsare being stored in moisture proof containers at -20°C for very long period.However, seeds can be stored at 5 to 10°C for medium term storage. Whileundried seeds of recalcitrant species are to be packed in semi-moistureresistant containers along with measures to prevent attack of saprophytic fungiand stored at medium temperature (10-15°C), to keep seed alive for shorterperiod.

SEED ST ORAGE IN CERTAIN FRUIT CROPS

Mango

Mango seeds are protected by hard stony endocarp. Seed is ex-albuminous having two fleshy cotyledons with testa and tegmen representedby two papery layers. Polyembryony is observed in certain cultivars. Seedexhibits recalcitrant type of storage behaviour wherein seed loses the viabilityon drying after extraction and undried seeds cannot be preserved long at lowtemperatures. Reduction of seed moisture to below 25% in Alphonso and 32%in Totapuri low ered the germ ination percentage and seed ling vigour. Patil etal. (1986) obtained 40% of germination in Alphonso stones after 90 days ofstorage at 25°C in polyethylene bags with charcoal powder. It declined from75 to 12% after 60 days when stored in open while those stored in polyethylene

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bags at 8°C did not germinate at all. The moist storage appears to be successfulfor preservation of recalcitrant seeds. Here seeds are imbibed with eithergermination inhibitor or fungicides for preventing germination and pathogensduring moist storage. Seed treatment with 1% 8-hydroxy quinoline sulphateand storage at 15°C could preserve the seed viability for one year.

Citrus

Citrus seeds are either mono- or polyembryonic in nature. They are largelyUtilized for raising rootstocks as well as in breeding programme. Lime (Citrusaurantifolia) seeds are small, oval and polyembryonic with white cotyledons.Lemon (Citrus limon) seeds are ovoid and polyembryonic with 10-15%nucellar embryos and white cotyledons. Sweet orange (Citrus sinensis) seedsare obovoid, polyembryonic with white embryos. Mandarin (Citrus reticulata)seeds are small polyembryonic with green embryos. Pummelo (Citrus grandis)seeds are large ridged yellowish and monoembryonic. Grape fruit (Citrusparodisi) seeds are white, polyembryonic with white cotyledons.

Seeds of many Citrus species are viable for a short period under ambientconditions. The seeds belong to orthodox group and/or show intermediatestorage behaviour and require special procedure for preservation. Seed storagein moisture proof containers at low temperature checks the deterioration byreducing metabolism. Seed storage with calcium chloride in polyethylene bagsat 8°C resulted in 100% viability in Citrus karna and rusk citrange and 84% inCitrus jambhiri and Citrus limonia after 150 days (Krishna and Shankar 1974).

Acid lime seeds were successfully stored for 10 years at 5 and -20°C. Grapefruit seeds retained viability for 3 months when stored with charcoal at roomtemperature and for 4 months at 5-8°C. In Poncirus trifoliata and Citrusgrandis, germination was the highest after 80 days in seeds stored inpolyethylene bags at 4.4°C and 56-58% RH. Seeds could be stored in fruit itselffor short term preservation. Fruits of some lemon cultivars were successfullypreserved for 40 days at ambient temperature and for 60 days at 5°C. Bajpaiet al. (1963) also reported that seed storage in whole fruit of kagzi lime andsweet orange is better than in polyethylene bags.

Rambutan

Seeds are difficult to store under normal conditions and at roomtemperature they lose their viability within a matter of days. They can bestored for a few weeks in moist saw dust or charcoal to which some aril juicehas been added at 21-28°C.

Jackfruit

Seeds are large oblong about 3 x 2 cm in size having thick gelatinousyellow covering and belong to recalcitrant group. Seeds remained viable for15 days and gave 40% germination after 30 days of storage.

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Litchi

Litchi seeds are dark brown, covered with white fleshy, juicy andtranslucent aril. Seeds are recalcitrant and should not be allowed to dry. Whensown immediately after extraction, their germination was 100% which reducedjust after one day storage in water and it was 75% on the seventh day and30% on 15th day after storage. Seeds stored in moisture proof containerssprouted within 15 days and those stored for 10 days in polyethylene bagsshowed 50.7% germination. However, seeds can be preserved in fruit for shortperiods. Seed viability was preserved for 24 days by storing fruits in sealedpolyethylene bags after treatment with benomyl (0.05%) and 6% wax emulsion.

Mangosteen

Seeds are recalcitrant and lose their viability rather quickly, especiallywhen the thin membrane around the seed is damaged or when seed is placedin ambient or cool atmosphere. Storage of seeds in charcoal or moss at roomtemperature retains the viability for a few months.

Conclusions

Seeds of several tropical fruit crops show recalcitrant storage behaviourand they do not withstand drying or are able to keep alive at low temperatures.Thus, they are difficult to store for longer period. In this context, more researchis needed for extending storage life. However, it is desirable to preserve thegermplasm in field as fruit repository. To supplement the conservationprocess, wherever, as far as possible seeds could be conserved by use ofavailable techniques like moist storage, etc., for certain period.

CARE AND MANAGEMENT OF ORCHARDS

T RAINING AND PRUNING

Modified central leader system of training has been recommended forboth apple and pear on seedling rootstocks. The proportional heading backand thinning out system of pruning is adopted after the juvenile phase ofplant growth. Spur pruning encourages vegetative growth and helps in newspur development in old plantations.

For high density planting on semi-dwarfing and dwarfing rootstocksspindle bush, dwarf pyramids and cordon system of training are suggested.Spindle bush on M7 and modified central leader on MM106 rootstocks aresuccessful.

Peach is generally trained to open centre system, while in sweet cherrymodified central leader system is followed. Pruning is done in such a way sothat 20-50 cm new growth in young trees and 25-30 cm in older trees aresecured every year. In sweet cherry about 10% of fruit bearing area should be

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removed annually, whereas in apricot slightly heavier pruning should bepractised to keep the spur system renewed.

Manures and Fertilizers

Nutrient requirement varies from place to place. In Himachal Pradeshwidespread deficiencies of N, P, K, Ca, Mn, Zn and B have been recorded. Afertilizer dose of 700:350:700 g of N, P and K for full grown bearing apple treehas been recommended.

It has been observed that VAM (Vesicular arbuscular mycorrhizea) fungiincreases P uptake by apple roots. The following corrective measures fornutrient deficiencies have been suggested for apple.

In peach cv. Sharbati, 17.5 g N/tree/year and in apricot 70-100 g N/tree/year have been found to be optimum. For peach, N has been the main limitingelement in H.P, while for plum, maximum fruit yield was recorded when leafN, P and K contents were 2.89, 0.28 and 0.89 respectively.

Weeding and Mulching

In apple orchards, grass mulching (10 cm thick) with one application ofpost-emergence herbicide Glyphosate (0.8 kg/ha) was found to be effective.The shrubby weeds were, however, best controlled by 500 ppm Gramaxone +100 ppm 2,4,5 -T application.

Moisture conservation was maximum under grass mulching and,therefore, recommended for apple in H.P. Mulching with oak leaf was effectivein U.P.

Supplementary Irrigation

Trickle irrigation at 75 per cent of field capacity results in better treegrowth and higher fruit yield in apple when raised on semi-dwarfingrootstocks. For apple, most critical period of water requirement is April toAugust and peak water requirement is just after fruit set. 114 cm of waterduring whole year through 19 irrigations have been recommended. Dripirrigation saves water considerably. Field trials indicated a total irrigationrequirement of 3840 liters water per tree under conventional system ofirrigation; under drip system 1695 liters of water was enough. The applicationefficiency under drip system is about 2.27.

In Santa Rosa plum, irrigation at 50% field capacity gave better growthand economic returns.

Pest and Disease Control

Pests: In apple and pear about a dozen pests are causing serious damageto the crops. Most important ones are: San Jose Scale, Woolly apple aphid,Root borer, Blossom thrips, Coddling moth and European red mite. San JoseScale can be effectively controlled with eco-friendly miscible spray oils at 2%

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concentration when applied during February-March. Woolly aphids can becontrolled through soil application of Phorate or Carbofuran granules duringMay and October/November. For coddling moth, pheromone trapping wasfound to be effective, and certain bio-control agents have also been identifiedin apple orchards for controlling certain insect pests.

European red mite is becoming a very serious pest for apple in H.P. andJ&K. The pest attack causes premature leaf fall. Late dormant sprays ofmiscible oil provide effective control of eggs. The mite can also be controlledby sprays of Dicotol (0.05%) followed by Malathion (0.05%).

In peach, leaf curl aphid can be controlled by pre-bloom sprays (at pinkbud stage) of Dimethoate (0.03%) or Monocrotophos (0.04%).

Fruit fly in peach, apricot and plum can be controlled through foliar sprayof baits consisting of Malathion (0.1%) + 1% sugar.Stem and shoot borerscausing damage to peach, plum, apricot, almond and cherry can be controlledby inserting 0.5g of PDCB (paradichlorobenzene) into the holes and pluggingthem with mud.

Diseases: Apple scab caused by Venturia inaequalis is a serious diseasecausing maximum economic loss. A sound forecasting and early warningsystem has been developed for prediction of scab attack. Also, a judiciousfungicide spray schedule has been devised. Under high disease pressure,systematic fungicides performed better, while under low disease pressureErgosterol biosynthesis inhibiting (EBI) fungicides were as good as protectants.Ascosporic inoculum produced by over-wintered apple leaves could besubstantially reduced by giving post-harvest applications of Bavistin (0.1%)and EBI chemicals Penconazole (0.5%) and Flusilazole (0.01%) as pre-harvestfungicidal sprays control scab during storage. Powdery mildew disease canbe kept under check by pruning and spraying wettable sulphur (0.2-0.3%) orKarathane (0.05%) during dormancy, bud swell and petal fall stages.

Collar rot and white root rot diseases in apple occur mainly in poorlydrained soils. Proper drainage of orchards and soil drenching with 0.1%Carbendazim for white root rot and with 0.3% Mancozeb or 0.3% RidomilMZ for collar rot are effective.Virus and virus-like diseases such as mosaic,chlorotic leaf spot, rubbery wood and others have been reported. Viruscleaning through tissue culture and supply of virus free bud wood materialare being pursued to contain further spread of the viral diseases in apple.

ESTABLISHMENT OF ORCHARDS

LAND PREPARAT ION AND PLANT ING

Fertile sandy-loam to clay loam soils with pH range from 5.5 to 7.5 andfree from water logging conditions are suitable for establishing deciduousfruit orchards. In flat lands/valleys, square, rectangular or triangular planting

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may be adopted. In hill slopes, planting on contours or terraces isrecommended. In shallow slopy lands, small terraces (half-moon terrace), maybe made to establish the plants and large scale disturbance of surface soil needto be avoided.

Planting of deciduous fruit plants is done during winter months fromthe end of December to mid March. Planting is done in the centre of pits(1x1x1m cube or circle), prepared a month before planting. While refillingthe pit, 50g of aldrin powder (15%) is mixed with the soil.

After planting, the young plants are supported with stakes and basinsare kept free of weeds. Mulching with dry grass or polyethylene is advisable.Irrigation is provided after planting.

Spacing

Spacing varies from species to species and depends also on the type ofrootstocks used. For apple raised on seedlings of crab or other commercialvarieties of apple, a planting distance of 6x6m or 7x7m accommodating 277-205 plants per ha is recommended for J&K hills. On clonal rootstocks likeM4, M7, M26, MM106 a spacing of 4.5m x 4.5m (555 plants/ha) is suggested.

For spur type varieties and standard coloured mutants of apple, highdensity planting on dwarfing rootstocks like M9, M4, M7 and MM107 hasbeen found to be feasible. Fruit yield of 30-35 tons/ha has been achieved in 12year old orchards of colour mutants of apple on MM106 under a plantingdensity of 2222 plants/ha (3m x 1.5m) in cooler hills of H.P.Optimum Plant Spacing for Spur Types and Standard Colour Mutants of Apple.

Type Rootstock Spacing (m×m) No. of trees/ha

Spur Type Seedling (Crab) 5×5 400

Spur Type MM111, MM109 4×4 625

Standard Type MM106, MM109 5×5 400

Spur Type MM106, M7 3×3 1111

Standard Type M9 2×2 2500

Source: Awasthi, R.P. and Chauhan, P.S., 1997. Apple and Pear. In: 50 years of CropScience Research, ICAR Publication, New Delhi.

In pear, for trees on seedling rootstock under normal conditions, spacingof 5x5m is recommended, but on clonal rootstocks (Quince A) spacing can bereduced to 3x3m. In peach, a spacing of 4.5 to 5m and for plum, apricot andcherry a spacing of 6m are followed.

PRODUCTION OF PLANTING MATERIAL

There is a large number of government and private nurseries engaged inmultiplication of planting material of deciduous fruit crops. In addition, the

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State Agricultural Universities and the Research Institutions multiply plantingmaterial of improved cultivars for sale and distribution to the farmingcommunities. The Government of India had supported establishment of a largenumber of fruit nurseries, both in public and private sectors as plannedactivities. During the 8th Five Year Plan (1992-97) it is estimated that over 55million nursery plants of different perennial crops, including temperate fruits,have been produced and distributed under the scheme. There was a targetfor the establishment of 85 big nurseries, 587 small nurseries, and 37 tissueculture units (20 by the private sector and 17 by the Government) of differentfruit crops. Exact number of these nurseries engaged in production of pomeand stone fruits is not known. However, the existing nurseries (more than600), covering both public and private Institutions, are sufficient to meet therequirement of planting material of deciduous fruit crops.

PROPAGAT ION AND ROOT ST OCK INFORM AT ION

In all the pome and stone fruits vegetative propagation techniques ofbudding or grafting are followed for multiplication of planting material onstandard rootstocks, raised both from seeds or through clonal methods.Cropwise details are as follows:

Apple: For raising rootstock seedlings, seeds of crab apple or commercialcultivars are stratified during December for 2-3 months at 2-5°C. One yearold seedlings are used for budding/grafting.

Clonal rootstocks are raised through mound or stool layering. The motherplants are allowed to grow for one season and cut back to 3 cm from theground level just before the growth begins. When new growth is about 10cm, the shoots are covered with soil leaving the growing parts exposed. Rootedlayers are cut off close to the ground level and planted in nursery beds forgrafting/budding.

In H.P. hills, different types of budding/grafting are recommended:• Chip budding in mid-June and mid-September.• Whip and Tongue and cleft grafting in February-March.• T-budding in May-June.Pear: Seedlings of Kainth (Pyrus pashia Linn.) or Shiara (P. serotiana Rehd)

are stratified in moist sand for 35-45 days at 2-5°C. Tongue grafting inFebruary-March or T-budding during June-July is recommended.

Others: Plum and apricot are generally grafted on wild apricot seedlingrootstocks. Peach is also used as a rootstock.

Peach is budded/grafted on wild peach and peach-almond hybridrootstocks. Hard wood cuttings (treated with 500 ppm IBA) can also beemployed for raising rootstocks.

For apricot, peach, plum and cherry shield budding during July-Augusthas been recommended for the hills of J&K. Micro-propagation through tissueculture technique has not yet been commercialized for temperate fruits,

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although tissue culture protocols have been developed at research Institutes/Universities. Colt, a rootstock for cherry can be multiplied in large numbersby following the tissue culture technique. Shoot buds are cultured in MSmedium containing 1-2 mg/litre benylaminopurine. Individual shoots arerooted on MS medium containing 1 mg/litre Indole Butyric Acid (IBA). Therooted plants are transferred to sterilized sand + soil (1:1) mixture and coveredfor 7-10 days for hardening.

CONSTRAINTS IN DECIDUOUS FRUIT PRODUCTION

Large number of old orchards (more than 30 years old) are showingdecline in terms of growth and fruit yield. Such old trees do not produceadequate extension growth. Large scale replanting is therefore needed.Delicious group of cultivars constitute the major share (about 83% in H.P.) ofapple production in the country. These cultivars are self unfruitful and needcross pollination to ensure good fruit set. Interplanting pollinizer cultivars(Golden Delicious, Jonathan, Red Gold, Lord Lambourne etc.) in theproportion of 25 to 33 percent is necessary for good fruit set, and choice ofwrong pollinizers and their inadequacy in number often result to lowproductivity.

In many countries, Delicious group has been replaced or is in the processof replacement with more promising cultivars. The need for injecting newblood into the apple industry through spread of new cultivars (spur types,colour mutants, strains of Gala, Red Fuji; scab resistant cultivars, bud sportselections of Royal Delicious, and some of the promising hybrids) is urgentlyfelt. Some of the spur type and coloured mutants are already popular withfarmers and high density planting has also caught the imagination ofdevelopmental departments and agencies both in H.P. and J&K. The researchsystem has already identified Early, Mid and Late cultivars for different agro-climatic regions.

The low chilling cultivars of stone fruits have also covered large tracts ofthe sub-tropical plains of Punjab, U.P and H.P. For the hills, promisingcultivars identified need further spread.

Generally, apple is grown in marginal land and fertilizers are not appliedaccording to the requirements of the trees. The water and fertilizer useefficiency is generally poor. Also, spring frost and hailstorms are adverseweather parameters leading to low productivity. Research results have shownthat through proper orchard management practices (soil and waterconservation and fertilizer application) the fruit yield can be doubled in theexisting orchards. The adoption of improved production technologydeveloped by the research system can bring visible and perceptible changesin the temperate fruit industry in India. Technologies like use of clonalrootstocks, introduction of renewal pruning techniques and micro nutrient

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applications have not been transferred and adopted at a satisfactory level.Apple scab disease has been the major plant protection problem in apple inJ&K and H.P, whilst U.P hills are comparatively free from the disease. Applescab forecasting system developed and the chemical control scheduleprescribed have helped in reducing loss due to apple scab to a considerableextent.

Apple growers are adopting the prescribed schedule of chemical spraysto control the disease. For checking entries of diseased material in the freeareas of U.P. and North-Eastern Hills, strict quarantine and selection of elitedisease-free mother plants are very essential.

Often it is not followed strictly. Some of the virus diseases have also beenreported in apple for which biological and serological indexing/detectionprocedures have been developed. Limited quantity of virus-free budwood isalso being supplied. Extreme care is now required to be taken to multiplyquality planting material (in apple alone approximately 2 million plants/year)for establishing new plantations.

Most of the orchardists still sell their crop at flowering to contractors asthere is no well organized marketing system. Transportation in the hills itselfis problematic. Post-harvest management problems originating from poorharvesting (strip picking) and improper packing system (non CFB boxes) andlack of proper pre-cooling and cold storage facilities result in huge (25-30%)loss of fruits. Capacity of the processing sector is also inadequate. Productdiversification, value addition and market infrastructure development wouldrequire very substantial investment. The existing processing units are quiteold and they require modernization for which substantial investment isrequired. CA storage trials have shown good promise. Its extension in largergrowing areas is needed. Technology for storage of apple is now known, as aresult of which apple is now available throughout the year.

GOVERNMENT POLICIES AND PLANS

Research on pome and stone fruits is conducted mainly by three StateAgricultural Universities, namely: (a) Sher-e-Kashmir University forAgriculture Science & Technology, J&K; (b) Y.S. Parmar University forHorticulture and Forestry, H.P.; and (c) G.B. Pant University for Agriculture& Technology, U.P. A good number of research stations of these Universitieslocated in major pome and stone fruit growing belts are engaged in temperatefruit research. The Indian Council of Agricultural Research (ICAR) throughAll India Coordinated Research Projects on Fruits and Post-harvestTechnology and another Network project on Apple Scab disease has furtherstrengthened the research activities in deciduous fruits. A few long establishedtemperate fruit research stations namely at Shalimar Bagh in J&K, atMashoobra in H.P. and at Chaubatia in U.P. hills have made commendable

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progress in temperate fruits. Both State Governments and the ICAR providefinancial support to strengthen fruit research in these Universities. A largenumber of ad-hoc research projects on pome and stone fruits funded by ICARare also generating good information on these crops.

During the 8th Government Developmental Plan (1992-1997) the ICARestablished a Central Institute for Temperate Horticulture (CITH) with itsheadquarters at Srinagar in the J&K State, with a regional station atMukteshwar in the U.P. Hills. Both these research stations will workexclusively on temperate fruit crops. This new Institute will receive majorsupport during the 9th Plan period (1997-2001). In the North-Eastern Hillsregion, the ICAR Research Complex for NEH Region with its headquarters inMeghalaya State and regional units in each of the 5 other States are engagedin fruit research.

On the developmental side the State governments are engaged in nurseryproduction of quality planting material. For example, in H.P. alone currentlythere are 600 nurseries in private and public sector producing and distributingmore than 0.8 million plants of apple alone, every year. There are ambitiousprogrammes in all the States to further expand/replant with new improvedcultivars. Apple scab disease control and post-harvest processing sectors aregetting focused attention in Government developmental plans. The Directorateof Marketing and Inspection of the Government of India has framed gradestandards for apple, plum and William pear. The organizations like NationalHorticultural Board (NHB), National Cooperative Development Corporation(NCDC), Agricultural and Processed Food Products Export DevelopmentAuthority (APEDA) etc., are providing incentives to traders and exporters toimprove their infrastructural facilities like grading and packaging centres,refrigerated transport, setting up of pre-cooling, cold storage, auctionplatforms etc. The NCDC is undertaking procurement and marketing of appleon a limited scale. The NHB has set up a market information service for thebenefit of growers.

Deciduous fruits, covering pome and stone fruits contribute significantlyto the horticulture economy of India. Apple production dominates the sceneand systematic cultivation and marketing of apple can change the ruraleconomy in the hills of North-Western India. New vision and concerted effortsare required for change in variety mix, supply of quality planting materialfrom elite clones on indexed clonal rootstocks. High density planting, watermanagement including micro-irrigation, integrated plant nutrientmanagement and IPM strategy for plant protection are some of the areas whichneed greater R&D focus. Adoption of post-harvest management practices andinfrastructure development for grading, packaging, pre-cooling and storageof the produce needs focused developmental attention. Value addition andexport promotion, particularly of apple are drawing due attention of thedevelopmental agencies in India.

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Ginger, Fat and Fibrinolysis

Ginger (Zingiber officinale) is a popular food spice, and is used as amedicine to treat various ailments from time immemorial in different partsof the world. It occupies an important place in Ayurvedic and Graeco-Arabicsystems of medicine where it is commonly used as carminative, digestive andto treat chronic rheumatism and gout. It is customary in India to add gingerin the diet rich in fat. It is reported to contain anti-histaminic and antioxidantfactors.

Earlier, it has been reported that dietary condiments and spices such asgarlic, onion and asafoetida neutralize fat induced alteration in blood lipids,platelet functions and fibrinolysis. Ginger has also been evaluated regardingits effect on blood lipids and platelet aggregation both in healthy individualsand patients of coronary artery disease, However, its effect on fibrinolyticactivity has not been studied. The present study therefore, was envisaged toobserve the effect of ginger on fibrinolytic activity (FA) modified by high fatdiet in healthy individuals.

Material and Methods

The study was conducted on 30 healthy adult males, between the age of30 to 50 years who were not consuming tobacco in any form. After a writtenconsert, fasting blood samples were collect and they were immediatelyadministered 50 gm of butter with four slices of slices of bread. Blood sampleswere again collected after 4 hours. During the subsequent week, a similarprocedure was adopted on the same subjects on 2 days, but one of thefollowing was administered randomly in addition to the butter: (a) Ginger 5gm in gelatin capsules or (b) placebo in matched gelatin capsules. Bloodsamples were again collected. Thus all 30 patients received 50 gm of butter t5 gm of ginger on one occasion and 50 gm butter + placebo on other occasion.Therefore, all served their own control.

All the blood sample were analysed for fibrinolytic activity which wasdetermined by employing a method described by Buckell & Elliot. The methodis based on the following principle: the euglobin fraction of plasma containsplasminogen activator, plasminogen and fibrinogen. Normally occurringinhibitors of conversion of plasminogen to plasmin are not present in thisplasma fraction. The euglobulib fraction is clotted with thrombin and the timetaken for clot lysis is estimated and expressed in units by multiplying thereciprocal of the lysis time in minutes by 10,000. The results were statisticallyanalysed with student’s ‘t’ test for paired data.

Preparation of Ginger Capsules: Dry ginger rhizomes were powdered inthe mortar and pastle. The dry powder was filled in gelatin capsules. Eachcapsule contained 625 mg of dry ginger powder. Eight capsules wereadministered with butter. Placebo capsules contained same amount of lactosepowder.

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Results

Fibrinolytic activity decreased by 18.8 per cent after administration offatty meal. Addition of ginger along with the fat not only prevented the fallin fibrinolytic activity but actually increased it by 6.7 per cent. As comparedwith fatty meal, ginger has actually increased increased fibrinolysis by 31.5per cent. The placebo has not prevented the fat induced decrease in fibrinolyticactivity.

Discussion

Administration of 50 gm of butter has lead to a significant fall in FA (P<0.001). Addition of ginger to a fatty meal in the dose of 5 gm not onlyprevented the fat induced decrease in FA. But kept the fibrinolysis above thenormal level. The change is significant (P< 0.001) as compared to controlexperiment in which fatty meal was administered alone to the same subjects.Preliminary trials have lesser amount of ginger would give inconsistant resultstherefore 5 gm dose has been used throughout this study. No side effects wereobserved even in this dose.

Dietary fat administration leading to significant fall in fibrinolytic hasbeen observed by many workers. A rich fatty meal if consumed frequentlyfor long time even in apparently healthy individuals may be harmful becauseit may lead to poor tendency to clot lysis. Moreover, this defective fibrinolysismay be an important factor in the genesis of coronary artery disease as hasbeen stressed by several workers. Therefore, addition of ginger may prove tobe a convenient and safe dietary measure for every day use in personspredisposed to such situations.

Chemically ginger contains several classes of compounds. The chemicalcomposition of dried ginger is as follows: starch 40-60%, proteins 10%, fats10%, fibres 5%, inorganic material 6%, residual moisture 10% and essentialoil (oleorasin) 1-4 per cent. The essential oil of ginger contains various terpinsand sesquiterpenes. The predominant sesquiterpene hydrocarbon iszingiberene. The characteristic pungent odour is due to its oleoresin contentwhich is an oily liquid containing oxymethyl phenols like shogaol, zingeroneand gingerol etc. In all more than 200 different volatile substances have beencharacterised in the essential oil fraction; out of all which compound(s) isexactly responsible for frbrinolysis is not known.

Ginger has been a popular food spice possessing many medicinalproperties. Recently it has drawn a lot of scientific attention. It has beenreported to have components (gingerols, shogaols) which are potent inhibitorof platelet aggregation and inhibit cyclooxygenase and 5lypoxygenase invitro. It has also shown amelioration in symptoms (pain and inflammation)in patients with musculokeskeletal disorders and migraine However, it doesnot affect blood lipids, blood sugar and fibrinogen in patients with coronaryartry disease. The present study, therefore, brings out a new property of ginger,

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not reported earlier, whereby it neutralizes the altered fibrinolytic stateinduced by fatty meal in healthy individuals.

We have earlier reported that ginger inhibits platelet aggregation inhealthy individuals and patients with coronary artery disease. The effect onfibrinolytic activity further adds to its therapeutic value. Moreover it alsoconfirms the rationale of the age old Indian tradition of adding variouscondiments to a diet rich in fat in order to neutralize the deleterious effects ofa fatty meal. Ginger may therefore prove useful in situations which areassociated with altered state of fibrinolysis.

SOIL STRUCTURE AND CROP GROWTH

Soil physical properties affect root and shoot growth directly andindirectly, the latter for example through poor drainage causing pores to fillwith water and plants to suffer from anaerobiosis. Root growth has beendescribed under various soil physical conditions, but relationships have onlyrarely been established between features such as crop yield, root growth andsoil pore size distribution or conductivity, a more aggregate measure.

The difficulty in establishing simple relationships does not mean that soilhas little influence upon root growth; rather it points to the complexity of theinteractions and the internal homeostasis which plants maintain. Roots bothelongate and proliferate and spread laterally as they grow and age.Concomitantly some roots die and others become suberised and function asconduits, but not as absorbers, of water and nutrients. Roots can elongatedownwards as fast as 8 cm/d, as for example, soybean growing in a silt loamin a rhizotron. Deep-rootedness and maximum rooting depth reflect soilproperties (for example, roots will not grow through pores that they cannotdeform to a larger diameter than the root).

However, relationships are not often reported. Maximum rooting depthvaries with species and soil type. For example, wheat roots penetrated to 0.8m in heavy-textured soils and to 1.2 m in a loamy sand but it is often foundthat a variety will have a consistent rooting depth across similar soil types ina particular year or in one soil across several years. Angus et al. found thatrice and six dryland crops (mung bean, cowpea, soybean, groundnut, Maizeand sorghum) extracted different amounts of stored soil water (ranging from100 mm for rice to 250 mm for groundnut) and that extraction was, in part,related to rooting depth.

The spread of roots with age can be related to the growth (increase inweight) of the whole plant, and to accumulated temperature or growing day-degrees (GDD); indeed, there is some evidence that temperature influencesthe direction of newly-appeared roots as well as the rate of appearance andextent of growth. Clearly, however, there are factors other than plant size,temperature and soil which influence root proliferation. Otherwise the plants

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sown at three different times of year in the same soil in Figure would aligntheir root growth along a single growth-GDD relationship. These other factors,of which day length is probably particularly important, tend to maskunderlying relationships between growth and soil structure. That the tillagecan affect root length, though in this case its effects took three years to develop.Measurable differences in soil porosity developed under two tillagetreatments: in the first year both root growth and water infiltration (Kapproximately 5 mm/h) were the same under minimum tillage andconventional tillage. By the third year, when differences were measuredbetween roots, infiltration rates were 84 mm/h in minimum tillage and 0.2mm/h under conventional tillage. Despite the differences in root growth therewere no substantial differences in grain yield, reflecting the overall constraintof climate in the semi-arid environment.

Increases in soil density or strength retard root penetration and thus limitthe volume of soil exploited by the crop and the water available. It is difficultto quantify the relationships between these soil parameters and plant growth.In the cases of bulk density and strength, particularly, a gross measure of eitherfor an undisturbed mass of soil can give only a remote indication of what aroot encounters. A determination of gross bulk density does not assess whethera root is growing within a pore (in which case it may deform surrounding soilbefore its radial environment reaches the density or strength of the gross soil)or if it is growing within the soil material, in which case it has already exerteda radial force equivalent to that measured for the gross soil.

This problem of scale-what is measured in a gross estimate cannot assessthe micro-environment of the root, to which it (and through hormonal signals,the plant top) responds—does not invalidate some general hypotheses aboutplant behaviour in response to soil compaction and structural arrangement.Roots stop growing when they are unable to deform their micro-environment.This probably occurs at suctions about 60 kPa when they cannot generateadequate internal turgor. It is thought that root elongation declines curvilinearlywith increasing either bulk density or shear strength. In the case of bulk density,little effect is noted until a ‘critical value’ and root elongation ceases within afurther 10 per cent increase in bulk density. As explained earlier, these valuesvary with soil type. Root elongation declines asymptotically with increasingsoil strength though the actual critical values would be expected to vary withsoil type, water content and the method of measurement.

Studies of the effect of bulk density or strength on other plant processes,particularly germination and shoot elongation, also suffer from the sametechnical problem of scale. Shoots are able to explore macropores withoutbeing subject to the gross values of the soil they are in. The actual local valueswhich inhibit shoot elongation appear to be quite small, for example, 0.76kPa. These values from controlled situations contrast sharply with gross fieldvalues and with the large number of inconsistent correlations which arise from

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attempting to correlate crop performance with grossly-measured soil structure.Perhaps, most encouragingly, studies of genetic variation in the sensitivity of cropsto soil strength suggest that there is appreciable range in plant sensitivity. Therelative ranking of genotypes is, however, the same when under near-critical stressas when growing with virtually no mechanical stress. Genotypes suited to stressfulsituations may be selected, therefore, by screening at a single soil strength.

CROP-LIVESTOCK INTEGRATION

This issue is central to improving land productivity in the SAZ. It is integralto labour intensification, for which the necessary condition is population growth.A large literature supports the thesis that rural population density explains ahigh proportion of the observed variation in smallholder farming intensity(defined in terms of frequency of cultivation cycles and labour inputs per ha)in tropical Africa. In the SAZ, livestock are usually a central component in suchintensification under smallholder conditions.

McIntire et al argue strongly that ‘farming intensity and crop-livestockinteractions increase with population growth and with market infrastructure. Theintensification of animal production allows more interactions: farmers invest incattle, herders manage them, stock eat more crop residues and byproducts, andproduce more manure’. Crop-livestock interaction follows an inverted U-patternthrough time. ‘First, specialised farming and herding societies that trade productsgive way to mixed farming societies, in which cropping and animal activities arein the same management unit. This movement to mixed farming, which we callthe first transition, occurs when opportunities for using less labour intensivetechniques of soil fertility maintenance are exhausted as population densitiesincrease, and as the opportunity cost of labour rises. The latter encourages farmmechanisation, usually via animal traction; as draft power becomes more valuable,crop farmers start to manage livestock and herders begin to cultivate. As exogenousmarkets and technologies develop further, there is a reverse movement away fromintegration and towards specialisation, which we call the second transition. Thesetechnical changes-fertilizers replacing manure, tractors replacing animals, andsupplements replacing fodder crops and pastures-eliminate the cost advantagesfor a mixed enterprise to provide some of its own inputs. As population densityrises, causing land pressure, resource competition occurs within the farm whichinduces further specialisation’.

CROP COEFFICIENT CURVE

After the selection of the calculation approach, the determination of thelengths for the crop growth stages and the corresponding crop coefficients, acrop coefficient curve can be constructed. The curve represents the changesin the crop coefficient over the length of the growing season. The shape of thecurve represents the changes in the vegetation and ground cover during plant

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development and maturation that affect the ratio of ETc to ETo. From the curve,the Kc factor and hence ETc can be derived for any period within the growingseason.

Single Crop Coefficient

Shortly after the planting of annuals or shortly after the initiation of newleaves for perennials, the value for Kc is small, often less than 0.4. The Kc beginsto increase from the initial Kc value, Kc ini, at the beginning of rapid plantdevelopment and reaches a maximum value, Kc mid, at the time of maximumor near maximum plant development. During the late season period, as leavesbegin to age and senesce due to natural or cultural practices, the Kc begins todecrease until it reaches a lower value at the end of the growing period equalto Kc end.

Dual Crop Coefficient

The single ‘time-averaged’ Kc curve, incorporates averaged wetting effectsinto the Kc factor. The value for Kc midis relatively constant for most growingand cultural conditions. However, the values for Kc ini and Kc end can varyconsiderably on a daily basis, depending on the frequency of wetting byirrigation and rainfall. The dual crop coefficient approach calculates the actualincreases in Kc for each day as a function of plant development and the wetnessof the soil surface.

As the single Kc coefficient includes averaged effects of evaporation fromthe soil, the basal crop coefficient, Kcb describing only plant transpiration,lies below the Kc value. The largest difference between Kc and Kcb is found inthe initial growth stage where evapotranspiration is predominantly in the formof soil evaporation and crop transpiration is still small.

Because crop canopies are near or at full ground cover during the mid-season stage, soil evaporation beneath the canopy has less effect on cropevapotranspiration and the value for Kcb in the mid-season stage will be nearlythe same as Kc. Depending on the ground cover, the basal crop coefficientduring the mid-season may be only 0.05-0.10 lower than the Kcvalue.Depending on the frequency with which the crop is irrigated during the lateseason stage, Kcb will be similar to (if infrequently irrigated) or less than theKc value.

POTENTIAL FOR DECIDUOUS FRUITS

Area Expansion: Both North-West and North-Eastern regions of Indiaoffer large areas ideally suitable for cultivation of pome and stone fruits. Inthe North-West India, stone fruits like peach, plum and apricot come up wellat elevations between 900 to 1500m with an annual rainfall of 90-100cm. Apple,cherry and pear are commercially successful at elevations between 1500-2700

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m above msl. The cold arid regions between 1550-3650m with annual rainfallof 25-40 cm are again highly suitable for dry nuts and drying type of apricots.There are vast tracts of land still available for further expansion of these crops.

In the North-Eastern Hills, excepting the rain shadow belts of ArunachalPradesh and high altitude Lachung area of Sikkim, apple may not be successfulcommercially. Other crops like pear, peach, plum and apricot offer good scopefor further expansion.

Widening the Cultivar Base: The low productivity and poor quality ofapple are often linked with monoculture of a few old cultivars and theirdegeneration over the years. The U.P hills, particularly Kumaon division, hasthe unique advantage of harvesting apple fruits for early market. Similarly,the rainshadow belts in North-Eastern Hills can offer good quality apple forthe eastern Indian markets, thus reducing the cost of long distancetransportation from North-Western Hills. Part of the markets of Bangladeshcan be captured by the fruits of the North-East.

There is good scope for introduction of new promising cultivars, replacingthe Delicious group. Similarly, use of clonal rootstocks of Malling and Malling-Merton series and even their indexed material ‘EMLA’ selections will greatlychange the productivity and quality of fruits. High density planting with spurtype cultivars offer good scope.

The identification of low chilling peach, plum and pear cultivars offergood possibilities for their cultivation in the low hills and in sub-tropicalplains. Some of the new hybrids, including scab resistant apple cultivars needverification trials on a commercial scale.

Management Practices: Scientific water management and practisingproper training and pruning of trees including introduction of renewalpruning techniques, will make significant impact on increased production evenin the existing orchards. Drip irrigation, in-situ water harvesting and correctingmacro and micro-nutrient deficiencies will go a long way in bringing notableimprovement in productivity as well as fruit quality. By adopting IPM strategyand organic farming practices, selected export markets can be targeted well.

Processed Products: There is immense scope for increasing variousprocessed products of pome and stone fruits, for which technologies areavailable. The existing capacity of the most organized processing unit ofHPMC is only marginal as compared to the volume of fruit available forprocessing. The HPMC utilized only 1 to 1.5 percent of total cull fruit available.The present combined capacity of two units of HPMC is 30,000 tons annually,which can be easily raised to 50 to 75 thousand tons.

The expansion of grading and packing stations, their furthermodernization with mechanical grading equipment, use of CFB boxes andmore number of pre-cooling and cold storage units will improve the marketingsystem and enhance marketability of the produce. Apple has been identifiedas one of 6 most promising fruits for fresh fruit exports.

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FRUIT CULTURE

Introduction: The Cashewnut (Anacardium occidentale L.) is one of theimportant nut tree grown in the tropical world. This fruit tree was introductedinto Malbar coast of India in 16th century by Portuguese. It is the native ofBrazil. It is grown mostly on West Coast area of Maharashtra, Goa, Karnataka,Kerala and Tamil Nadu.

Importance: The dry nuts - Kernels are of high nutritive value and alsorich in protein, carbohydrates, minerals like calcium, phosphorus and iron.This has a great export value and is considered as one of the importanthorticultural crop in India.

Climate: The Cashew requires a minimum rainfall of 60 cm per annum,but can stand extremes of rainfall from 20 cm to 400 cm. If there issufficient water supply, it can withstand a long period of dry spell and lowhumidity.

The Cashew is a sun loving tree and does not tolerate excessive shade,also it does not favour very high temperatures above 45 deg cent. During thefruit set and development. Heavy rains and cloudy weather during floweringadversely affect the yield.

Soil: The Cashew is cultivated on a wide variety of soils. It is considereda crop of marginal land and is recommended for slopy and light soils. Thebest soils for better production are deep, friable, well drained and without ahard pan upto 2/3 m in depth. The Cashew is mainly grown on Laterite, redand coastal sands in India.

Varieties: The following are the important varieties of Cashew grown indifferent parts of the country.

i. Vengurla 1;ii. Vengurla 2;iii. Vengurla 3;iv. Vengurla 4;v. Vengurla 5;vi. Vengurla 6;vii. Bhubaneshwar;viii. Kanaka;ix. Dhana;x. Selection.Propagation: The cashew is grown by the following methods:i. Seed Propagation: It is the oldest and cheapest method of propagation.

It is also used to raise the plants for the purpose of grafting;ii. Layering this method is more successful in coastal and more humid

areas;iii. Soft Wood Grafting: This is followed both in the nursery as well as

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iv. Top-working: This method is used not for propagation but forconverting inferior free into desirable one.

Planting and Season: Normally cashew grafts are planted at the spacingof 7.0 x 7.0 or 7.5 x 7.5 or 8.0 x 8.0 m on plain land or on desirable contours onslopy lands. After marking, the pits of 60x60x60 cm are dug and filled with acompost,top soil, single super phosphate and karnaj cake. This is to be wellbefore the monsoon starts. Planting should be done during June-July i.e. inthe beginning of monsoon.

Interculturing:i. Removal of weeds is done once twice;ii. Intercrops are planted in interspace;iii. Some perenial forest plants and/or some seasonal crops can be taken

as per the local need and situation.Care of young orchard:i. Gap filling for the missing plants;ii. Removal of outgrowth on stocks in case of grafted plants;iii. Staking with bamboos;iv. Cover cropping;v. Providing protective irrigation during first few summers. These are

some of the points for young plantations.Special Horticultural practices:i. Pruning of dead and dried shoots alongwith crisscross branches

and water shoots;ii. Pruning of leader shoots in June followed by 2% KNO spray;3iii. Spraying with 10 ppm NAA twice during flowering for increasing

fruit set and minimizing flower and fruit drop.Irrigation: The cashew is mostly grown as the rainfed crop and requires

no irrigation in the high rainfall areas. However, if the rainfall is low, thecashew responses well to irrigation water at the time of fruit set upto fulldevelopment stage of nuts. Irrigation should not be given before or at thetime of flowering, as it is likely to promote vegetative growth.

Nutrition: To improve the growth and yield the cashew trees should bemanured and fertilized with bio-fertilizers like biomeal and NpK 500,200, 200gm per tree per year.

Spraying with ultrazyme and 8:12:24:4 NPKmg twice, during fruitdevelopment is useful to increase size and yield of nuts.

Plant Protection: The crop particularly blossom and fruits should beprotected against Tea Mosquito, Stem and root borer is also an important pest.Leaf minor, Leaf and blossom webber, Flower thrips are the common pests inneglected orchards. Inflorescence blight, pink disease, and anthrac nose aresome diseases found on cashew. Clean cultivation, pruning, better aerationand preventive measures should be adopted besides occassional sprays ofselected pesticides and fungicides.

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Harvesting and Yield: The Cashew comes to its full bearing stage at theage of 10th year and continuous to give yield further for 30-40 years. Usuallythe nuts are collected after they fall off from the tree. They are separated fromapples and dried for 2/3 days to reduce the moisture. Regarding yield thereis a wide variation and differences of opinions. However, the desirable varietyat good management yields 50 kg of raw nuts.

Post Harvest Handling: The processing of raw nuts involves roasting,shelling, drying, peeling, grading and packing `A' grade nuts are always indemand from the outside markets and fetch a good price. A Fenny like drinksare prepared from the apples.

GERMPLASM OF MANGO

Mango is one of the most important fruit crops in India. It has been incultivation for almost last five thousand years. Apart from India, mango isalso an important crop in the Philippines, Indonesia, Thailand, Myanmar,Malaysia, Sri Lanka, U.A.E., South-East Africa, South Africa, U.S.A., Mexico,West Indies, Brazil and tropical Australia. All the cultivated Indian mangoesbelong to the single species Mangifera indica L. The geographical distribution,phytogenetic trend, pollen morphology, chromosome number, cytogeneticaland breeding behaviour indicate that the highest concentration of species ofMangifera is found in Malayan Peninsula followed by Sunda Islands and theEastern Peninsula comprising Myanmar, Thailand and Indo-China.

Occurrence of wild species like M. sylvatica and M. caloneura inMyanmar, Assam and northeastern India and evidence of fossil leaf impressionof M. pentandra which looks similar to M. indica having edible fruits indicatethat mango originated in this region. In India, more than thousand varietiesof mango are available and these have arisen mainly as a result of openpollination. It is to be admitted that most of the varieties lack in one or theother character. As opined by Naik et al. (1958), this very diversity has tendedto limit commercial production of fruits of standard quality in India. In therecent past, efforts were made to exploit natural variability and breed newvarieties for different purposes. So there has been a need to prevent the lossof varieties, and to conserve and use them in the breeding programme. Apartfrom intervarietal breeding programmes, progress has been made with regardto identifying the location of wild species of Mangifera and their utilizationin breeding.

M ANGIFERA SPECIES—PRESENT STAT US

It was thought earlier that the genus Mangifera comprises 41 species. DingHon (1978) opined that there are only 32 species. Tardieu (1962) added 4species to this list from Indo-China. Ling (1983) added one species from Chinaand in Malaysia two more were added by Kochummen (1983). However,

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Mukherjee (1985) reorganized the classification and recognized 39 species inthe genus Mangifera. Subsequent surveys carried out by various workers invarious parts have shown the existence of different races.

Yadav (1996) reported the occurrence of races of wild species of Mangiferain the northeastern region of India. Many of the seedling types, growing inthe forest area of northeast region still have many primitive characters suchas polyembryony and dwarf growth habit. Studies of wild mango trees inManipur showed that primitive characters still persist and their mixing withthe present day commercial varieties has not taken place.

Some wild forms recorded in Manipur are similar to M. indica, but closerlook at the flower and leaf characters showed that flower staminodes areabsent and the disc is thick and tuberculate. This wild type resembles morewith M. caloneura (a species occurring in Myanmar) in having smooth ovaryand stone not exactly kidney shaped. This also resembles M. siamensis becauseof the absence of staminodes. Another wild type with small fruits and bigleaves was observed on Silchar-Imphal road which resembles the specimenreported by Sharma and Sen Chaudhary (1976) from Tripura, except that thebase of the petiole in this type was swollen.

Mukherjee (1948) has reported five species, M. indica, M. andamanica,M. camptosperma (treated under M. gedebe), M. khasiana (now treated underM. sylvatica) and M. sylvatica to be indigenous to India. Another species M.nicobarica is said to occur in Nicobar Island. Areas of concentrated populationof the trees of species have been identified for M. andamanica, M.camptosperma and M. sylvatica. M. indica is found all over the country as allthe varieties in existence in the country belong to this species.

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6

Commercial Horticulture

Consider any undesirable plant in turf to be a weed. Weeds areopportunistic and are virtually impossible to eradicate from turf. A properweed control programme limits weed infestations rather than attempting toeliminate them entirely. Weak stands of turf that lack density will soon beinfested by weeds. In some situations, extremely competitive weeds can infestdense turfgrass. For either situation, combine chemical weed control with amanagement programme directed at improving turfgrass density and vigour.Proper management includes mowing, watering, fertilizing and cultivation.Once weeds have been suppressed by an effective chemical and cultural weedcontrol programme, eliminate your subsequent use of herbicides or reducethem to spot treatments where problem weeds reappear.

CHEMICAL WEED CONTROL

The proper application of herbicides can dramatically reduce turfgrassweed populations in a short period of time. A combination of their cost,effectiveness and simplicity of use often makes herbicides the primary meansof weed control -- with little attention given to other weed reduction measures.Remember, herbicides are only one facet of the total weed control puzzle.Effective long-term weed control and a reduction of pesticides in theenvironment will be achieved only when an effective cultural weed controlprogramme has been implemented.

CULT URAL WEED CONT ROL

The basic principle involved in cultural weed control is to grow a standof grass that is dense and competitive enough to prevent weed encroachment.Weeds are not the cause of poor turf, but rather the result.

Weeds require light, water and nutrients to grow. As turf loses density,light will penetrate the canopy and cause weed seeds at the thatch and soilsurface to germinate. Once germinated, weeds can develop rapidly, especiallyif turf continues to decline.

Weeds probably will receive sufficient light to develop if you can see baresoil or thatch when you look straight down on a stand of grass. Managementpractices that discourage turf weeds include proper mowing, watering,fertilizing, thatch control and cultivation. The chance for weed encroachmentcan be reduced by using locally adapted turfgrasses and establishing themduring the correct season of the year.

• Mowing: To prevent weed germination, mow frequently at the tallestrecommended mowing height. Weeds germinate rapidly when turfis scalped by mowing too short or when it is not mowed frequentlyenough. Both mistakes decrease turf density and cause an opencanopy that favours weeds. Experts recommend a range of mowingheights to meet specific turf activities. Lower mowing heightsrequire more frequent mowing. Each mowing should not removemore than one-third of the total leaf height. Annual grassy weeds -- such as crabgrass -- are especially a problem on turfs that lackdensity as the result of poor mowing.

• Fertility: Apply nitrogen at least once a year. Areas that receivechemical weed control but no nitrogen are probably beingmismanaged. Even in low- maintenance situations, the first defenceagainst weeds is dense turf. For cool-season grasses, apply nitrogenin the fall. For warm-season grasses, apply it in the summer.Additional nitrogen should be supplied during the growing seasonto provide adequate growth and density with less attention givento turfgrass colour. On cool-season grasses avoid heavy applicationsof nitrogen in the spring. That will encourage excessive vertical leafgrowth, bringing with it a greater chance for scalped turf and aneventual decrease in turf density. Don't apply nitrogen to dormantwarm-season grasses, since it will encourage winter annual weeds,such as chickweed, henbit and speedwell.

• Establishment: Weeds can prevent the establishment of grasses atcertain times of the year. Spring seeding of cool-season grasses isseldom successful -- because the fertility requirements and frequentwatering necessary for turf establishment will also create intenseweed pressure from crabgrass and summer annual broadleaf weeds.Consequently, late summer and early fall are preferred forestablishment of cool-season grasses. Cool-season turfgrasses thatare germinating by early September will not be affected by summerweeds and will usually establish a competitive stand of grass beforewinter annuals, such as chickweed and henbit, become a problem.

• Thatch and Cultivation: These have a dual effect in terms of weedcontrol. Weed seeds below a thatch layer lack the necessary lightfor germination, so weeds that germinate in the thatch layer usuallydie because of the poor environment for seedling development in

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thatch. In this respect, thatch provides some degree of weedsuppression. On the other hand, thatch can cause turf decline byharbouring disease-causing pathogens, reducing water infiltrationand tying up pesticides and fertilizers. Power-raking andverticutting are typically used to remove thatch.

• Cultivation: Soil compaction from traffic and tight soils will oftendecrease turf competition and favour some weeds, especiallygoosegrass and knotweed. Two ways to fix the problem are coringand slicing. These procedures improve the soil surface and thegrowing environment for turfgrass.

DAM AGED CORN FOR SILAGE

Corn that has been damaged by drought, high temperatures, blight, frostor hail can be salvaged for silage. Quality will not be as high as where cornhas reached the dent stage. Feeding value will depend upon the state ofdevelopment and how it is handled after the damage occurred. Silage fromimmature corn usually is higher in moisture, does not ferment in the samemanner, frequently has a sour odour and is more laxative when fed in largequantities.

Frosted corn has a low carotene content and should be cut as soon aspossible. It will dry out quickly and lose leaves. It may be necessary to addwater to corn that has frosted and become too dry to pack well. Drought cornalso may need added water. When the corn forage is dry, keep the chopperknives sharp and chop as fine as possible.

Immature corn that has been damaged by extremely high temperaturesshould not be immediately ensiled. Although these plants may never producean ear, some additional stalk growth and consequently some additional feedmay be produced by delaying harvest.

If the plants are harvested for silage soon after they have beenextensively damaged by heat, the stalk will have so much moisture that avery low quality silage will result. Nutrients also will decline significantlythrough seepage.

Corn that has been damaged by leaf diseases such as the southern cornleaf blight often are made into silage. The blight organism is not believed tobe toxic to ruminants. It also has been shown that it does not survive theensiling process. In severe cases, a secondary infection of molds on thedamaged areas of the plant may produce a harmful toxin. However, limitedresearch indicates that this is unlikely.

The greatest problem with this type of silage stems from its lack of energydue to reduced grain formation and improper fermentation due to theexcessive dryness of the damaged plant. In severe cases where large areas ofthe corn plant are dead, fermentation problems caused by the lack of plantmoisture could arise.

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Nitrates

Under certain soil and environmental conditions, the corn plant may storean excessive amount of nitrogen compounds.

Nitrate poisoning and drought conditions often are associated, but thecondition is difficult to define. The degree of drought and nitrogen availabilitygenerally confuses the issue. As the soil moisture level becomes acute, nitratesmove towards the soil surface above the corn roots. Some drought-strickencorn may be short of nitrate instead of oversupplied.

If heavy rains occur at this point, excess nitrogen is leached downwardand may be taken up by the corn root system. This could cause abnormallyhigh nitrate levels in the plant. Under these conditions, avoid harvesting silagefrom the stricken fields for a few days.

One measure that may be helpful in reducing the amount of nitrate inthe silage is, to allow the corn plant to grow beyond the period of droughtdamage. If the plant is capable of making some regrowth following a drought,it may be able to use much of its excessive stores of nitrogen for additionalregrowth.

The highest concentrations of excessive nitrogen usually are in the lowerportion of the stalk. Raising the chopper cutter blade so that the lower 18 or20 inches of the stalk remain in the field also may reduce the concentration inthe silage. Testing for nitrates before silage harvest is frequently unreliable asa feeding guide. Nitrate level in the plant changes rapidly from day to dayand usually is reduced about 1/3 in the ensiling process.

If silage is suspected to contain excessive nitrates, it can be detected bytest prior to feeding. This analysis should be taken as near to the time whenthe silage will be fed as practical.

Silo Gases

Lethal gases may occur at any time during silo filling. The greatest dangeris 12 to 72 hours after filling, but gas may occur up to 10 days after the lastsilage is put in the silo.

Silo gases may appear in any ensiled material, grown on any type of soiland under any level of fertilization.

If the gases are present in higher concentrations, two of them, nitrogendioxide (NO2) and nitrogen tetroxide (N2O4) may be recognized by theirirritating odour and colour. Nitrogen dioxide is reddish brown and nitrogentetroxide is yellow. Nitric oxide (NO) is a colorless gas that may be present indeadly concentrations without being visible.

The gases are heavier than air and remain beneath the air mass over thesilage. They often will layer on top of the silage just below the upper edge ofthe top door. They may settle down the chute to accumulate at the bottom ofthe silo, in unloading chutes, in adjacent feed rooms and may even move intothe barn, loading areas or milk houses. They usually leave a yellow stain on

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silage, wood or other materials they contact. The presence of dead birds andsmall animals around these areas is one indication that the gases are atdangerous levels.

A few simple rules will prevent tragedy and injury:• Run the blower 15 to 20 minutes before going into a partly filled

silo. Keep the blower running while anyone is inside.• Stay out of the silo for at least a week or preferably two after the

silo is filled.• If you experience the slightest throat irritation or coughing, get into

fresh air quickly. Immediate treatment by a doctor is an absolutemust.

• Ventilate the silo room for at least two weeks after filling by openingoutside doors and windows to carry away fumes. Removing thechute doors on the silo down to the level of the settled silage willpermit natural ventilation where gas tends to be concentrated.

• Keep the doors between the silo room and the barn closed to protectlivestock.

Length of Cut

Corn silage should be cut into particles 1/2 to 3/4 inches in length. Particlesof this size will pack more firmly in the silo and are more palatable to cattle.Very finely cut silage may be made with a recutter. This will increase theamount of dry matter that can be stored in a silo, but very finely cut silage isless palatable and has resulted in lower butterfat tests when this feed was theprimary source of roughage for dairy cattle.

Adding Water to Dry Silage

If silage is too dry, it may be necessary to add water in order to establishairtight conditions. As a rule of thumb, add four gallons of water per ton ofsilage for each 1 percent desired rise in moisture content. Add this water asthe silo is being filled. If water is added after the silo is filled, it tends to seepdown the silo walls and does not permeate the silage mass. This may causeleaching of silage nutrients, seepage that may break the air seal and improperfermentation.

Frozen silage: Frozen silage is sometimes a problem, especially withtrench or bunker silos. Freezing does not impair the keeping quality of silageso long as the silage is not disturbed, but frozen silage may cause digestivedisturbances when eaten by cattle. It is best to thaw silage before feeding it.

Silage additives: Top quality silage can be made without the addition ofany additives or preservatives. There is no reliable evidence that addingenzymes, yeast cultures, antibiotics or acid-forming bacteria will increase thefeeding value of corn silage. The two additives most often used in corn silageare limestone and non-protein nitrogen compounds.

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Molasses and grain may be added to corn forage at the time of ensiling.However, this practice only tends to enrich the resulting corn silage as a feedrather than to improve the quality of the corn forage itself. There is also somefermentation loss with the added molasses and grains.

Molds: Molds are common in silage, especially around silo doors andedges of bunker silos. White and gray molds, which are caused by exposingsilage to air, are seldom toxic to livestock, but intake is sometimes reduced.A mold known as monascus causes silage to form lumps or small clumps,which are white on the outside but have a reddish centre. Cattle usually willeat silage containing this mold. There have been no reported cases of toxicity.

In the spring as warm temperatures occur, a rapid-forming red mold mayappear on the face of the silage between morning and evening feedings. Thisis commonly known as bakery mold, monilla sitophilia. The mold spores areactivated by the heat produced in the silage process and grow quickly whenexposed to air. It has not produced any known cases of toxicity.

Silo Capacities and Filling

Tables giving estimated silo capacities often vary due to the crop ensiled,length of cut and effectiveness of distribution and packing at filling time. Mosttables are based on silage harvested at 30 to 35 percent dry matter.

Silo capacities actually vary little in terms of dry matter storage, despitedifferences in moisture content at filling time. An exception to this is materialthat is 50 percent dry matter or more at the time it is placed in the silo. Withthis type of material, dry matter capacity may be reduced as much as 10percent for estimates based on 30 to 35 percent dry matter.

With large-capacity silos and high-speed filling methods, the distributionand packing of silage in silos has been sacrificed. Improper distribution andpacking may cause excessive seepage, poor fermentation and losses in storagecapacity. Half the capacity of the silo, which is 14 feet in diameter, is in theoutside 2 feet. If an overabundance of fluffy, light material is in this outsidearea, silo capacity may be reduced as much as 20 percent. Most large towersilos should be equipped with a distributor to aid in proper silage distributionand packing. Length of cut of corn should be 1/2 to 3/4 inches to ensure properplacing.

Covering Filled Silos: A loss of nutrients occurs in all silage during theensiling process. Microorganisms that carry out the fermentation processrequire energy. This loss depends on the effort made to exclude air at time offilling and to prevent loss of carbon dioxide, which is necessary to arrestrespiration of the ensiled plant cells, as well as prevent seepage losses andundesirable fermentation or spoilage due to surface exposure. There isprobably a greater range of losses within a particular type or design of silothan occurs in different types of silos where equally good ensiling practicesare followed. So, regardless of silo structure, good ensiling practices save feed.

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A plastic cover on a trench or bunker silo or a large-diameter tower silocan materially cut feed losses. For best results, the cover must be appliedimmediately after the last load is packed in the silo.

On trench or bunker silos, it is important to mound or crown the forageso that the rain water will drain off the silo. Plastic covers that are not weightedto hold them firmly on the surface of the silage will be only partly effective.Covering the plastic with old tires plus limestone, sawdust or similar materialcan be of great benefit.

HARVESTING

Corn should be harvested for silage after the ear is well dented but beforethe leaves turn brown and dry. The quantity and quality of corn silage are attheir peak in this stage of development. The ear has accumulated most of itspotential feeding value, but there has been little loss from the leaves and stalks.After the dent stage, feeding value of corn stalks and leaves decreases whilefield losses increase.

Fall temperatures influence the maturity rate of the grain. Maturityusually refers to the time when the ear has accumulated 100 percent of its drymatter production potential. In many years this potential is not achievedbecause of cool temperatures and cloudy weather. Values listed may be usedas a guide to determine when maximum dry matter production has occurred,but variety and weather interactions will exhibit some influence on the result.Ears usually will be well dented somewhere between the 32- to 35-percentmoisture stage.

Table : Relationship of Kernel Moisture to Yield Potential

Water in kernels Yield of grain as percent of maximum

40 percent 93.538 percent 94.836 percent 96.334 percent 98.026 percent 100.021 percent 98.0

Corn harvested for silage in the milk or dough stage will yield less feednutrients per acre than if harvested later. Corn also may ferment improperlyin the silo if harvested too soon. Corn silage often is made too early because itis believed that feed is lost if undigested corn kernels appear in the manure.This is not true. Digestibility is as high for well-dented kernels as for immaturecorn grain.

Corn silage that is cut late and has brown and dead leaves and stalksusually will make fair- to good-quality silage, but total production per acremay be sharply reduced. Field losses as high as 30 percent have been foundwhen silage is made late into the fall or early winter. A 10 percent reduction

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in the amount of dry matter stored in the silo also has been noted with late-cut silage.

Results of feeding trials with late-cut silage tend to vary, but in most casesthe quality of late-cut corn silage has been slightly lower than silage madefrom corn cut soon after the dent stage of the ear.

Present research does not support recommending late-cut or mature cornsilage as a standard farm practice. However, it does indicate that in emergencyconditions, corn may be harvested over a wide period of time and still makea satisfactory feed.

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7

Greenhouse Crops System inHorticulture

GREENHOUSE CROPS

The primary crops grown in greenhouses include: pepper, tomato,cucumber, lettuce, herbs, and strawberry. The industry in Florida has changedfrom primarily either tomato or cucumber in the early 1990s to the diversityof today. A variety of structure types are used, as well. Structures includeboth fan and pad or naturally ventilated systems. Both are successfully usedin the state depending on cropping intentions.

SUBIRRIGAT ION FOR GREENHOUSE CROPS

Subirrigation is becoming an increasingly common way of watering andfertilizing greenhouse crops in Massachusetts. This article is for growersconsidering a subirrigation system or just starting out with a new system.

Advantages to Subirrigation

There are three major economic advantages to subirrigation. The mostcommonly cited advantage is the savings in labor needed for watering theplants: a single person can water thousands of plants by operating the floodingsystem manually or with the help of a computer. Additionally, there is apotential savings in water and fertilizer with subirrigation since both arerecirculated and not lost by leaching or runoff. Also, depending on the systemand how it is installed, a grower can expect an increase greenhouse spaceefficiency (percentage of total floor area in use for growing plants).

Many growers report more uniform plant growth and less foliar diseasewith subirrigation. The increase in plant uniformity may be the result of moreeven and complete moistening of the growth medium and better distributionof nutrients absorbed by capillary flow. The absence of water on the leaveswith subirrigation probably results in less foliar disease.

The elimination of fertilizer and pesticide leaching and runoff from thegreenhouse is a very important reason for using subirrigation. In order toachieve the goal of reduced leaching and runoff the system must be maintainedas a truly closed system. The immediate practical value of preventing irrigationeffluent from escaping the greenhouse is not always apparent, but protectionof water used for drinking and recreation from contamination is probablythe most important long-term benefit of subirrigation.

Challenges to Using Subirrigation

Like any other new way of growing greenhouse crops there are a numberof challenges to overcome to use subirrigation successfully. The two greatestchallenges for most growers is the initial cost of the system and the ability toretrofit the system in an existing greenhouse. A conservative estimate ofpayback time is 5-10 years, but the period could be as short as 2-3 yearsdepending on the system chosen, whether existing bench frames can beretrofitted, and whether productivity of the system is maintained at a highlevel. An excellent economic analysis of subirrigation systems was recentlypublished by Wen-fei Uva and her colleagues of Cornell University. Somereaders may have heard Wen-fei speak on her work at the New EnglandGreenhouse Conference in October 2000. Her article is very detailed, butconcise, and would help growers in choosing a subirrigation system.

Fig. Greenhouse Crops.

A grower beginning to use subirrigation will have to learn some newways of irrigating and fertilizing to use the system successfully. Growthmedium and irrigation solution testing for pH and EC is one important skillto acquire. Since the growth medium tends to accumulate salts withsubirrigation it is critical to be able to test for EC on a regular basis without

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having to wait for results from a commercial lab. Also, growers who maintainnutrient and pH levels in the irrigation solution by adding fertilizer or waterto stock tanks manually rather than with automatic equipment need tocarefully monitor EC and pH to maintain the proper ranges. Successful useof subirrigation requires extra attention to cleanliness to avoid disease andinsect problems. The use of pesticides and other chemicals, particularly asdrenches, can be problematic with subirrigation so adoption of IPMtechniques, especially pest population monitoring, is very important.Cleanliness will be discussed a little more later in the article.

Subirrigation Systems

There are three basic closed, recirculating subirrigation systems currentlyin use in New England: ebb-and-flow benches, trough benches, and floodedfloor systems. There are some variants on these, for example, the "Dutchmovable tray system" is very similar to ebb-and-flow, but a complete systemis highly mechanized for a number of tasks. Capillary mats and collectiontrays are also a form of subirrigation, but they are not normally closed systems.

Ebb-and-flow. The ebb-and-flow system is very common and is quitefamiliar to most growers. The system consists of a shallow, molded plasticbench top which is flooded to water and fertilize the plants; when irrigationis complete the remaining solution drains from the bench and is pumped backto a storage tank.

Ebb-and-flow is very versatile because the bench tops can accommodateall sizes of pots and bedding plant flats (although not on the same bench orirrigation zone at the same time because of the differences in water absorptionrates between container sizes). The bench tops can be installed on existingframes and, with the rolling feature, ebb-and-flow benching can be 80-90%space efficient. Ebb-and-flow benches are easy to retrofit in clearspangreenhouses, but not in greenhouses with many internal supports. This systemhas the highest initial cost, $4 to 6/ft2, installed on existing bench frames andincluding tanks, delivery and return pumps, plumbing, and installation. Amajor portion of the cost comes from the specially molded plastic bench topswhich cost about $2.50/ft2.

Troughs. This system works by running a film of irrigation solution downa slightly inclined, shallow metal trough holding the plants. The troughs emptyin a return channel for recirculation. The pots or flats in the trough have plentyof opportunity to absorb solution as it runs past.

The trough system is very easy to retrofit on existing bench frames. Thetroughs can be obtained in various lengths and widths from a commercialmanufacturer or they can be fabricated by a local metalworking firm to thegrowers specs. A trough system is about 70-80% space efficient, less than ebb-and-flow, because normally spaces are left between the troughs. Most growersuse this system mainly for potted crops, but it is possible to do bedding plant

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flats if the open mesh style of tray is used to hold the paks. However, becauseof the trough spacing, it isn’t possible to space flat-to-flat except in anindividual trough. The initial cost of the trough system is about 2-6/ft2. Thecost of this system can be fairly low if the troughs are made locally or if theyare installed on existing benches. Most of the plumbing is simple to puttogether and inexpensive.

Flooded floor. In this system the entire floor of the greenhouse is coveredwith a concrete carefully designed and installed to pitch toward openings inthe floor. Through these openings the irrigation solution enters to flood thefloor and, following flooding, the excess drains back to the storage tank. Thefloors can be installed with bottom heating and divided into zones for separateflooding and bottom heating. Flooded floors can be used to grow plants in allcontainer types and sizes as long as separate irrigation zones are providedfor each type. Space efficiency is about 85-95%. Most greenhouses with floodedfloors were built with them rather than retrofitted later. The bottom heatingoption an efficient way of providing the proper growing temperature for theplants because the air close to the plants is heated and the larger volume ofthe greenhouse does not have to be heated so much. Some growers complainthat in a flood floor plants close to the flood/drain openings tend to beoverwatered, especially bedding plants. Also, as in the case of any floorgrowing system, all the bending and squatting needed to work with the plantscan be tiring for workers. Initial cost for a flooded floor is $3-5/ft2, but costscan vary significantly depending on the amount of excavation required forthe storage tanks and piping, whether or not bottom heat is installed, andwhether the floor is divided into zones for separate irrigation. A very skilledconcrete contractor is needed to get the pitch of the floor right to encourageproper drainage and to prevent puddling.

FERT ILIZING SUBIRRIGAT ED PLANT S

Since there is little or no nutrient leaching with subirrigation less fertilizeris needed compared to traditional overhead watering. The general rule forfertilizing subirrigated plants is to use one-half the rate (ppm) of fertilizernormally applied by overhead irrigation. Several years ago I subirrigatedpoinsettias with solutions of 100, 175, 250, or 325 ppm N from peat-lite 20-10-20 fertilizer. The plants finished about the same size with nearly as large bractsas plants watered from overhead. Leaf analysis revealed normal levels of mostnutrients at all fertilizer rates and no evidence of a serious nutrient deficiencyor excess. EC (soluble salts) levels were higher with subirrigation thanoverhead watering. EC was highest near the top of the growth mediumbecause of surface evaporation and deposition of nutrient residues. None ofthe treatments developed had excess EC.

The results of this study demonstrated that poinsettias grow well over awide range of fertilizer concentrations in subirrigation including levels applied

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by traditional overhead watering. In fact, most growers I’ve visited in NewEngland who subirrigate poinsettias on a large scale use fertilizer rates in therange of 200-250 ppm N. Use of fertilizer rates above 250 for subirrigatedpoinsettias increases the risk of excess EC leading to growth inhibition andplant injury. Learning to use an EC meter to monitor soluble salts on a regularbasis is very important with subirrigation.

Chemicals and Subirrigation

Many insect and disease problems can be prevented by adopting a newstandard of greenhouse cleanliness and through the use of simple IPMpractices to prevent infestations and infections from getting out of control.This means that for now growers must apply pesticides as they would tooverhead watered plants only more carefully. Heavy or frequent foliarspraying, or use of growth medium drench treatments, are risky practicesbecause enough chemical may enter the irrigation solution to causeundesirable effects to the plants in the long term. To avoid this problem, somegrowers divert irrigation water from their subirrigation system forconventional disposal following a pesticide application rather than letting itreturn it to the tank for recirculation. In the absence of definitive informationon the extent of buildup and effects of recirculated chemicals, growers shouldtry to limit pesticide treatments as much as possible especially growth mediumdrenches. Zero Tolerance™ disinfectant is one chemical that can berecirculated in subirrigation with beneficial effects. Zero Tolerance™ cancontrol algae and a wide variety of root disease organisms. The product labelhas specific directions on its use in subirrigation systems.

Interestingly, there is some interest in applying plant growth regulators(PGRs) through subirrigation. Currently A-Rest™ and Bonzi® are labeled foruse in "chemigation" systems including subirrigation by ebb-and-flow andfrom saucers. Labels for both PGRs have detailed instructions on how to applythe chemical so as not to cause plant injury and to protect water supplies. Inthe author’s opinion, it too early to draw conclusions about the efficacy andsafety of PGR application this way but it is being studied in Florida (Barrett,1999) and I will have some preliminary results to report soon.

Finally, cleanliness is very important. As a routine practice dead plantmaterial and other large "stuff" should be removed from growing areas, insidetanks, and plumbing after each crop. Then the system should be disinfectedwith Zero Tolerance™ or Green-Shield™.

STRUCTURE OF GREENHOUSE

A greenhouse is a structure with different types of covering materials,such as a glass or plastic roof and frequently glass or plastic walls; it heats upbecause incoming visible solar radiation (for which the glass is transparent)

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from the sun is absorbed by plants, soil, and other things inside the building.Air warmed by the heat from hot interior surfaces is retained in the buildingby the roof and wall.

In addition, the warmed structures and plants inside the greenhouse re-radiate some of their thermal energy in the infrared, to which glass is partlyopaque, so some of this energy is also trapped inside the glasshouse. However,this latter process is a minor player compared with the former (convective)process. Thus, the primary heating mechanism of a greenhouse is convection.This can be demonstrated by opening a small window near the roof of agreenhouse: the temperature drops considerably.

This principle is the basis of the autovent automatic cooling system. Thus,the glass used for a greenhouse works as a barrier to air flow, and its effect isto trap energy within the greenhouse.

The air that is warmed near the ground is prevented from risingindefinitely and flowing away. Although there is some heat loss due to thermalconduction through the glass and other building materials, there is a netincrease in energy (and therefore temperature) inside the greenhouse.Greenhouses can be divided into glass greenhouses and plastic greenhouses.Plastics mostly used are PEfilm and multiwall sheet in PC or PMMA.Commercial glass greenhouses are often high tech production facilities forvegetables or flowers. The glass greenhouses are filled with equipment likescreening installations, heating, cooling, lighting and may be automaticallycontrolled by a computer.

USES

Greenhouses protect crops from too much heat or cold, shield plants fromdust storms and blizzards, and help to keep out pests. Light and temperaturecontrol allows greenhouses to turn inarable land into arable land, therebyimproving food production in marginal environments. Because greenhousesallow certain crops to be grown throughout the year, greenhouses areincreasingly important in the food supply of high latitude countries. One ofthe largest greenhouse complexes in the world is in Almeria, Spain, wheregreenhouses cover almost 50,000 acres (200 km2).

Sometimes called the sea of plastics. Greenhouses are often used forgrowing flowers, vegetables, fruits, and tobacco plants. Bumblebees are thepollinators of choice for most greenhouse pollination, although other typesof bees have been used, as well as artificial pollination. Hydroponics can beused in greenhouses as well to make the most use of the interior space.

Besides tobacco, many vegetables and flowers are grown in greenhousesin late winter and early spring, and then transplanted outside as the weatherwarms. Started plants are usually available for gardeners in farmers’ marketsat transplanting time. Special greenhouse varieties of certain crops such astomatoes are generally used for commercial production. The closed

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environment of a greenhouse has its own unique requirements, compared withoutdoor production. Pests and diseases, and extremes of heat and humidity,have to be controlled, and irrigation is necessary to provide water. Significantinputs of heat and light may be required, particularly with winter productionof warm-weather vegetables. Because the temperature and humidity ofgreenhouses must be constantly monitored to ensure optimal conditions, awireless sensor network can be used to gather data remotely. The data istransmitted to a control location and used to control heating, cooling, andirrigation systems.

WATER DISINFECTION AND REUSE INGREENHOUSE HORTICULTURE

The production in European greenhouse horticulture is not yet as waterefficient as it could be. Soil-less growing systems are becoming common inhorticultural practice in most of the European countries, although not in eachcountry on a large scale yet. The advantages of soil-less growing systemscompared to soil grown crops are as follows:

• Growth and yield are independent of the soil type of the cultivatedarea;

• Better control of growth by use of improved water quality and abetter fertilization;

• Increased quality of products;• Pathogen-free start by use of substrates other than soil and/or easier

control of soil-borne pathogens.The disadvantages of these systems are:• The required high quality of water;• High investments and high costs for fertilizers;• Low quantity of the water.In most cases open or run-to-waste systems are adopted. In such open

systems, superfluous nutrient solution freely leaches to ground and surfacewater. Because of economical motives and environmental concerns closed soilless systems can be applied. These closed systems are more efficient with theuse of water and fertilizers, and cause less damage to the environment. Thedisadvantage of the closed systems is the risk of a rapid dispersal of soil-bornepathogens by the recirculating nutrient solution. To eliminate these pathogens,several disinfection methods can be used.

OZONE T REAT M ENT

Ozone treatment can be used to disinfect the drain water. Ozone is thesecond most powerful sterilant in the world and its function is to destroybacteria, viruses and odours. An ozone supply of 10 g/h/m3 water with anexposure time of 1 h is sufficient to kill all pathogens.

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UV Disinfection

Another way to disinfect the drain water is the use of UV-radiation. Ultra-violet radiation (or UV) is a proven process for disinfecting water, air or solidsurfaces that are microbiologically contaminated. For eliminating bacteria andfungi an energy dose is recommended of 100mJ/cm2. For viruses a dose of250 mJ/cm2 is recommended.

Heat Treatment

When heat treatment is applied, a solution is heated for about 30 secondsto a temperature of 95ÚC. At this temperature all pathogens are killed. Adisadvantage of heat treatment is the consumption of gas. Also warm drainwater contains less oxygen.

Slow sand Filtration

For several years commercial growers have used a slow sand filtrationinstallation to eliminate pathogens.

Sand filtration is frequently used and a very robust method to removesuspended solids from water. The filtration medium consists of a multiplelayer of sand with a variety in size and specific gravity. Sand filters can besupplied in different sizes and materials both hand operated or fullyautomatically.

SAND CULTURE

This term may be used in a general way to refer to cultures in sand, finegravel or cinders, but here it applies specifically to cultures of plants in sand,using nutrient solutions to supply the mineral elements required for plantgrowth. Sand is adapted ibr such use where free drainage is possible, as inthe constant-drip and flush or “slop” techniques. In small installations, coarsergrades of sand may be used with sub-irrigation, which see. In largerinstallations, it is not suitable for the subirrigation method because of slowdrainage, but it can be used in a greenhouse bench with free drainage andthe surface application of the nutrient solution.

The best type of sand is a quartz sand of the grade used for makingconcrete. It should be washed clean. This can be done by placing the sand ina pail or tub and forcing water up through it by delivering the water througha hose at the bottom of the receptacle so the flooding will carry out clay andsilt particles. Washing is especially important if the sand is fine. If the sand istoo coarse, it will require more nutrient solution to maintain the propermoisture relation.

If the only source of sand available is a limestone sand, it may beadvisable’ to use cinders, for the lime sand contains much calcium which, inexcess, will cause plant foliage to turn yellow. However, by using a more acid

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nutrient solution, or by running the nutrient solution daily through thelimestone sand for two weeks before placing the plants in the medium, thesand grains can be given a coating of phosphates and be made safe for growingplants.

GROWING SEEDLINGS IN SAND

Seedlings can be raised by sand culture either to be transferred to culturesfor chemical gardening or for use in the outdoor garden. By this methodseedlings with very fine root systems can be produced; if handled properlythey will transplant into soil readily and, given proper care, start to growpromptly. In addition, losses from damping-off are minimized, while anotheradvantage is that if the seedlings are not too crowded, they can remain in thesand if supplied with nutrient solution in the proper concentration.

A good way to handle them is to construct a box not less than 4 in. Deepon the inside with openings in the bottom so excess solution can drain out.These spaces between the bottom boards or small drainage holes can becovered with cheesecloth, burlap, or glass wool to prevent the sand from siftingthrough. Fill the box with clean sand and flush it well with water to settle thesand thoroughly. Then flush with nutrient solution diluted with five times asmuch water.

The seeds may be sown in drills or rows or broadcast; if in rows, makethe furrows barely deep enough so that the seeds can be covered after beingsown thinly. If the broadcast method is used, spread the seeds uniformly andnot too thickly over the surface, and cover with about one-fourth inch of sand.In either case, firm or compact the sand, then apply the dilute nutrient solutionuntil the sand is saturated.

Cover the box with newspaper to reduce evaporation and place in theproper temperature for the kind of seed sown. Since it would be fatal to theembryo of the seed if the surface should become dry, the sand must be keptmoist. The dilute nutrient solutions should be applied as a fine spray, toprevent disturbing the surface; a rubber bulb syringe with a fine rose nozzleis excellent for this purpose.

As soon as the seeds have germinated and the shoots appeared abovethe surface, remove the newspaper covering. When the seedlings have grownto suitable size, they may be transplanted to a permanent location or to othervessels where they can be more widely spaced.

If it is desired to leave them in the flat, thin them out to a proper spacing.They are now able to manufacture food in their leaves, and the nutrientsolution should be applied in a stronger concentration – one part of nutrientsolution to one part of water. Should it be desired or necessary to remove theseedlings from the sand medium, flood it with nutrient solution and gentlypull the plants out one by one, assisting the removal with sonic instrumentthrust under the root. A label, spatula, or case knife will be suitable.

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“Sand culture is one of the most efficient and cost-effective methods ofsoilless culture, and is widely used in the dry arid regions of the Middle East.Although it is not used in Australia on a commercial scale, it has provedpopular among some growers for crop trials. Its simplicity and low capitalcost makes it an attractive alternative to existing growing methods.”

Sand culture is one of the most efficient and cost-effective methods ofsoilless culture, and is widely used in the dry arid regions of the Middle East.Although it is not used in Australia on a commercial scale, it has provedpopular among some growers for crop trials. Its simplicity and low capitalcost makes it an attractive alternative to existing growing methods.” by LeoWright History records that Aztec Indians were among the first to use sandculture techniques.

During the 12th century the Aztecs developed extraordinary irrigationsystems in the Mexican basin, with swamp reclamation their most significantachievement, even including the colonisation of surrounding lakes. The Aztecsgrew beans and squash on primitive rafts covered with sand removed fromshallow lake beds. The plant roots grew through the sand down into thenutrient-rich water of the lake. A few of these so-called ‘floating gardens’ canstill be seen on the lakes surrounding Mexico City today.

In modern times, sand culture re-emerged during the 1960s, ironically inPuerto Penasco, Mexico, where successful trials led to commercial operationsin Mexico, the U.S. Southwest and in the Middle East. Using coarse beachsand, leached free of excess salts, vegetables were either sown directly in thesand or planted as seedlings.

The Advantages

The sand culture system has many advantages over traditionalhydroponic techniques. The fact that it is an ‘open’ (‘run-to-waste’) system,whereby the nutrient solution is not recycled, greatly reduces the likelihoodof diseases such as Fusarium and Verticillium spreading in the medium. Italso means there is no nutrient imbalance since plants are fed with freshnutrient solution at each irrigation cycle. Another advantage is the excellentcapillary action of sand, which results in lateral movement of nutrients sothat there is an even distribution of nutrients throughout the root zone.Additionally, water retention is high owing to the smallness of the sandparticles, allowing fewer irrigation cycles during the course of the day. Unlikeother systems, particularly NFT (Nutrient Film Technique, in which there isno medium), in the event of a fractured pipe, power or mechanical failure,there is more time available to repair the system before plants consumeexisting water in the medium and begin to experience stress due todehydration.

Practical advantages of sand culture include lower construction costs,simplicity of operation, and easy maintenance and service.

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The Disadvantages

The disadvantages of sand culture are few, and in some cases are incommon with other hydroponic techniques. A major disadvantage is the needto use chemical or steam sterilisation between crops in order to destroy media-borne pathogens. Such methods are thorough, although somewhat time-consuming.

Like all drip irrigation systems, feed lines can become blocked with finenutrient particles, grit or sand. This can be overcome by using in-line meshfilters which can be easily cleaned. Perhaps the overriding factor which makessand culture unattractive to end-users is the seemingly high consumption rateof nutrients because of the need to run to waste.

However, with careful management, the waste should account for nomore than 8% to 10% of the total nutrient solution added. Salt build-up isanother common problem, but this can be corrected by flushing the mediumperiodically with fresh water. Again, regular and careful monitoring of thedrainage water for evidence of salt accumulation is important to prevent excesssalt problems.

Sand Culture Systems There are basically two proven methods of utilisingsand as a growing medium. The first is to use plastic-lined beds in aboveground troughs; the other is to spread sand over the entire floor of the growingarea. Both methods use a sand depth of between 300 and 400 mm. Sand bedscan be easily constructed with wooden sides, or with concrete reinforcementwire, cut and bent to form an above ground trough and lined with thick, blackplastic film.

The bed must be watertight as leaks mean wasted nutrients. Polyethylenefilm (6-20micron), or swimming pool liner are ideal. If using black polyethylenefilm, it should be doubled over for maximum strength – a single layer ofpolyethylene will stretch and tend to mould around sharp objects once sandis shovelled into the bed, which may cause it to rupture.

The bottom of the trough should have a 1:400 incline (150mm drop per60 metres), so that it can be drained or leached when required. A drain pipeshould be set along the entire length of each sand bed, which in turn shouldbe connected to a main pipe at one end to collect waste water from all bedsand to conduct it away from the greenhouse or growing area.

For sand beds deeper than 400mm, 50mm agricultural pipe, covered withblue metal, can be used to channel excess nutrients away. For a standard sandbed, 70mm pressure pipe can be used and this should be seated on a shallowlayer of blue metal. The pipe should have drainage holes, cut across the pipewith a saw every 450mm, with the holes positioned on the undersurface ofthe pipe to discourage plant roots entering the pipe. For greenhouseoperations, the floor should have a gradient of 150mm per 30 metres, (1:200incline), and should be covered with 6 micron black plastic film. Generally,two layers of black film are used to overlap and cover the entire floor.

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A 30mm-50mm diameter drainage pipe, cut in the manner describedearlier, is placed on top of the plastic at a uniform spacing of 1-2 metresbetween pipes, depending upon the grade of sand in use. The finer the sand,the closer the pipes should be spaced.

Drain pipes must run parallel with the slope of the floor. At the low endof the greenhouse a connecting pipe is installed to conduct excess nutrientsaway from the greenhouse where it can be used for outside irrigation. Oncethe pipes are in place, the entire greenhouse floor is covered with sand to adepth of 300mm.

A simple home system can be designed along the same lines ascommercial units, but on a much smaller scale. It should consist of a bed orgrowing tray, nutrient reservoir and a trickle feeding system operated by apump that is controlled by a timer. The growing tray can have small holes inthe bottom of the plastic liner or a perforated plastic pipe can be used fordrainage.

One of the oldest and still popular hydroponic methods is to use a wicksystem. This consists of a double pot, one containing the sand and plant, andthe other the nutrient solution. A fibrous wick is set into the growing pot aboutone-third of the way with the other end suspended in the nutrient solution.As the sand dries out, capillary action draws more solution through the wickto the plant root zone. Ideal Sand Aggregate Research shows that beach sandis usually too fine and causes puddling, indicated by water coming to thesurface upon vibration of the sand.

Examples of puddling can often be seen in footsteps while walking alongwet beach sand. It is caused by the high percentage of silt and fine sand. Theideal sand aggregate is river sand, washed free of fine silt and clay.

The sand particle size should be between 0.6mm and 2mm in diameter,which allows the aggregate to drain freely and not puddle after an applicationof water. Cuming (1986) recommends a mixture of grades (30-40% of 0.5mm,40-60% of 0.2mm to 0.5mm, and 5-15% of 0.2mm). Drip irrigation System Adrip irrigation system must be used with sand culture. In a greenhousesituation piping should be capable of delivering 6-10 litres per minute foreach 100 square metres, or 30-45 litres per minute for each 500 square metresof growing area. However, the rate and length of irrigation cycle will dependupon the crop, its maturity, weather conditions and time of day.

The volume of water can be regulated using a flow control valve for eachsand bed. The valve is adjusted for each bed according to plant requirements.In this way several plant types can be grown. The flow valve should bepositioned upstream from the solenoid valve which automatically controlsthe irrigation cycle.

Where several beds are cultivated, there is no need to water themsimultaneously. The water arrangement can be set so that individual bedsare watered separately, thus ensuring mains pressure is not reduced. For small

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sand beds, a single 13mm black polyethylene pipe can be run down the lengthof the bed with spaghetti lines inserted every 300mm. Larger beds will require13mm poly pipe run along the inside of each plant row.

Emitters can be used to deliver nutrients to plants, adjusted to deliver 4-6 litres per hour. Alternatively, spaghetti tubing can be inserted into the main13mm feed line. A short length (50mm) of 13mm poly tube can be attached tothe end of each spaghetti line to channel nutrients out to each side of theirrigation point. Emitters, above ground pipes and fittings should be black toprevent algae growth inside the piping system.

Watering

If a timer is used, it should be programmed to deliver nutrients two tofive times daily, depending on the maturity of plants, weather and seasonalfactors. The water is added to each cycle to allow 8%-10% drain off. Twice aweek a sample of the drain off should be taken and tested for total dissolvedsalts. If the dissolved salts reach 2000 parts per million (ppm), then the entiresand bed should be leached free of salts using fresh water.

The Nutrient Tank

The nutrient tank should be large enough to meet feeding requirementsfor at least one week. And if several crops are grown which have differentnutrient requirements, then two tanks should be used, each with its ownspecific nutrient formulation. In this event, independent irrigation systemsmust be connected to each nutrient tank. Since sand culture is an open system,there is no need to change the nutrient solution regularly. However, the tankshould be drained and cleaned periodically of any sludge or sediment whichmay accumulate owing to inert carriers in the fertilizer salts.

Sand culture beds can be constructed on any surface, including hardbedrock or stony ground with a minimum of fuss. That, combined with lowinstallation costs, simplicity of operation and retention of moisture, even inhot weather, makes it an ideal system for Australian conditions. It isparticularly well-suited to rain shadow areas. Perhaps the only significantdisadvantage is that it is likely to become waterlogged during heavy rainperiods in those systems exposed to the elements.

GREENHOUSES WORK

Greenhouses are often seen as romantic structures. Originally theexclusive property of the wealthy and well-born, the first greenhouses wereprobably built in Roman times to cultivate exotic fruits and vegetables. In thefirst century, Pliny the Elder made a reference to the Emperor Tiberius havinghad a portable greenhouse that was protected with a covering made of

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transparent stone. This unusual and rare greenhouse was devised to cultivatethe emperor’s favourite vegetable, the cucumber.

Produce that we can find today in most local grocery stores was at onetime considered priceless in many parts of the world. In the 17th century,entire buildings were erected to house and propagate oranges and pineapples.Before they were called greenhouses, the names for these structures were asexotic as the fruits they contained. They were called specularia, orangeriesand pineries.

Reproducing plants out of season gave man a measure of control overnature. The allure of it sparked the imagination and inspired new methodsfor building structures devoted to plants. Precious glass began to be used moreand more in greenhouse construction.

Harnessing the plant world and exploring the possibilities of cultivatinguseful, exotic species led to building larger and more elaborate greenhouses,some of which are still in existence today. This fascination with propagatingplants in a controlled and protected environment has never dimmed.Greenhouses have grown from a novelty to an essential component in theway we feed hungry populations around the world, cultivate plants formedical research and preserve plants for future generations to enjoy. In thischapter, we’ll see how greenhouses work to make all of this happen and whyyou may just want a greenhouse in your backyard.

USES FOR GREENHOUSES

A greenhouse can extend a plant’s growing season by a few weeks, or itcan create a complete microclimate that’s a successful substitute for the plant’snative environment. From its origins as an indulgence for the wealthy andprivileged, greenhouses are now used in many important and unexpectedways to help man understand and make use of the natural world.

Since World War II, there’s been a huge increase in the commercial useof greenhouses to feed the world’s population. In temperate climates,greenhouses extend the growing season and protect plants from harsh weatherconditions. In higher latitudes, greenhouses increase plant production byoptimizing the available light. Even in hot, arid regions, specializedgreenhouses have been developed that can help lower temperatures andmanage water loss in plants due to transpiration.

Cultivating attractive and hardy plants for commercial nurseries andflorists is a thriving industry that relies on greenhouses to create perfect, pestfree plants.

In many areas, controlling the environment inside a greenhouse is easierand more reliable than trying to manipulate all the variables of growingdelicate plants outdoors.

Universities and research facilities around the world, like the Eden Projectin St. Austell, England, are using the controlled environments in greenhouses

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to recreate specialized ecosystems in order to understand plants better.Greenhouse technology is making it easier to study the potential value ofmedicinal plants and explore ways to increase plant yields and make plantsmore disease resistant. Greenhouses are even being used to preserve plantspecies whose natural habitat is threatened. So let’s take a look at how theywork.

HEAT ING A GREENHOUSE

Greenhouses create a sheltered environment for plants by using solarradiation to trap heat. This system of heating and circulating air helps to createan artificial environment in a greenhouse that can sustain plants when theoutdoor temperature is too cool or variable. Heat enters the greenhousethrough its covering of glass or plastic and starts to warm the objects, soiland plants inside. The warmed air near the soil begins to rise and isimmediately replaced with cooler surrounding air that starts to heat up. Thiscycle raises the temperature inside the greenhouse more rapidly than the airoutside, creating a sheltered, warmer microclimate.

In temperate climates, the sun might do all the heating in the greenhouse,but where the temperatures plummet, artificial heat may be necessary tomaintain temperatures above freezing. Where some greenhouses have accessto central heat from the main building, others have to rely on natural or bottledgas, heating coils or heating fans. These will usually work in conjunction witha thermostat. Because heat is one of the biggest expenses of keeping agreenhouse, other sources of energy are always being explored, like the useof solar batteries or animals as heat sources.

There are also other processes acting on the air inside a greenhouse. Thesun’s energy can travel through greenhouse glass easily, but the radiationemitted by the plants and soil that have absorbed the heat doesn’t get out aseasily, helping to trap heat inside.

This makes it possible to keep a greenhouse warm, but it also can causeproblems with overheating. In order to keep plants from getting too hot, somemethod of heat control is necessary. Vents that allow the lighter, hotter air toexit the greenhouse near the roof and cooler air to enter closer to ground levelact as air conditioning. Proper ventilation keeps the air in a greenhousecirculating. This helps maintain a stable temperature and also cycles the carbondioxide (CO2) that plants need for photosynthesis. Generally, greenhouseshave at least two vents, one on or near the roof and one on the lower half ofthe structure. Mechanical ventilators can also help maintain good airflow andheat control by opening and closing the vents automatically when thetemperature in the greenhouse changes.

And of course, all of the plants inside a greenhouse need some sort ofwater. Whether you use a garden hose, watering can or a sophisticatedautomated system with water sensors, water is essential in a greenhouse.

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Because watering is the most time-consuming greenhouse chore, the use ofsome type of automated system, like wicking, capillary matting or dripirrigation can make the process more consistent and reliable. Even if feedingwater directly to the greenhouse via an underground pipe isn’t possible,placing a greenhouse near water is a practical necessity.

T YPES OF GREENHOUSES

There are many types of greenhouses, and this can include plasticgreenhouses, rion greenhouses, those made of glass, temporary structures,permanent structures, and many others. The type you choose will depend ona number of factors. One important factor is the types of plants you intend toput in the building, because this will determine the temperatures required. Ifyou are just looking for types of greenhouses to start your yard plants earlyin then a cold house or cold frame will work. These structures do not haveany heating supplied at all, and they will only stay slightly above thetemperature which is outside. These buildings can not be used all year longin colder areas where snow and freezing temperatures are common.

One of the most popular types of greenhouses are rion greenhouses, whichare a form of plastic greenhouses. These are known for their incrediblestrength, and they can stand up to strong winds and heavy snows that woulddamage most other types of plastics. Portable or temporary types ofgreenhouses are usually very light, and they can be moved when the locationis not the best possible for the plants being grown. These structures are usuallyvery easy to put together and install, and they can be conveniently storedaway when you are not using them. Greenhouses come in many materials,sizes, and types, and the ones that you choose should reflect your individualneeds and requirements. This is a matter of personal preferences as well asthe pros and cons, and each person may choose differently.

Since greenhouses use the sun to create a warm environment for plants,when the temperature drops, there’s less sun-generated heat in the greenhouse.For plants that need more heat than a greenhouse can provide naturally,heating systems are necessary to make up the difference. Greenhouses aredivided into categories based on how much supplemental heat they need toproduce in order to keep plants at a certain temperature.

Cold Houses and Cold Frames: Cold houses provide protection for plants,but the temperatures inside can still dip below freezing during the winterbecause they have no supplemental heat source. Cold houses can help startspring crops a few weeks early and extend the growing season in fall, butthey’re limited by the weather.

Cool Houses: Warmer than cold houses, cool houses keep plants abovefreezing and in a temperature range of between 45 to 50 degrees Fahrenheit(7 to 10 degrees Celsius). Keeping the temperature above freezing will protectfrost sensitive plants, like geraniums and hibiscus, which would be impossible

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to keep year round in areas that experience freezing temperatures.Warm Houses: A warm house will allow a broader range of plants, but

requires slightly warmer temperatures too, around 55 degrees Fahrenheit (13degrees Celsius). Although the temperature range doesn’t support manytropical plants, some varieties of orchids and ferns can over-winter in a warmhouse environment.

Hot Houses: These greenhouses are designed to house tropical plants likecaladium, dieffenbachia and gardenia, which need a temperature range of 60degrees Fahrenheit (15.5 degrees Celsius) and higher. They require the mostsupplemental heat and insulation and can be expensive to maintain.

Conservatories: Conservatories are designed to display plants, not justmaintain and propagate them. They often have finished floors, ornate windowtreatments and space for furniture. Window greenhouses and small tabletopgreenhouses are also considered conservatories because they’re used primarilyfor display.

DIFFERENT T YPES OF GREENHOUSE

Greenhouses add growing time for plants in areas with extremetemperatures and weather conditions. The structures are built to houseseedlings and mature plants for personal or commercial use. There aredifferent types of greenhouse structures that either attach to a building or arefreestanding units. The structures have a variety of shapes and coveringmaterials from which to choose.

Attached Structures

A lean-to structure butts up against a building. Window mount and lean-to greenhouses are attached to a building, usually a house. The window mountis a small unit that is attached to the window frame in a building.

The lean-to structure has two types of roofing. The curved roof will allowthe one-story greenhouse to be placed against a taller building. A straight-eave structure will fit up under the eaves and roofing system of a single-storyhouse.

Even-span Greenhouse

The even-span greenhouse can be located at the end of an existingbuilding. The open end of the full-size greenhouse is attached to the outerwall of the building. An entrance can be made into the building where thetwo structures connect. The even-span has a large area for growing and betterair circulation than a lean-to.

Freestanding Structures

Freestanding greenhouse structures are built as self-supporting frames.They are made in styles such as Quonset, rigid frame, Gothic, A-frame, post

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and rafter. These different structures are built to stand on the ground or afoundation. Freestanding greenhouses are built in a variety of sizes.

Coverings

Greenhouses have a variety of coverings from which to choose. The homegardener may choose to build a greenhouse from recycled windows for a low-budget project. Glass has been traditionally used for years and can be a long-lasting cover. Tempered glass is also an option for the builder. Clear ortransparent, quality-grade fibreglass is a good option for a greenhousecovering as it is lightweight and can last 15 to 20 years. Plastic is used indouble-wall and film forms. These are lightweight coverings with varyingdegrees of quality.

SELECTION OF COMMERCIAL GREENHOUSE FRAMES

A good selection of commercial greenhouse frames and framing materialsis available. The frames are made of wood, galvanized steel, or aluminum.Build-it-yourself greenhouse plans are usually for structures with wood ormetal pipe frames. Plastic pipe materials generally are inadequate to meetsnow and wind load requirements. Frames can be covered with glass, rigidfibreglass, rigid double-wall plastics, or plastic film. All have advantages anddisadvantages. Each of these materials should be considered—it pays to shoparound for ideas.

FRAM ES

Greenhouse frames range from simple to complex, depending on theimagination of the designer and engineering requirements. The following areseveral common frames.

Quonset: The Quonset is a simple and efficient construction with anelectrical conduit or galvanized steel pipe frame. The frame is circular andusually covered with plastic sheeting. Quonset sidewall height is low, whichrestricts storage space and headroom.

Gothic: The gothic frame construction is similar to that of the Quonsetbut it has a gothic shape. Wooden arches may be used and joined at the ridge.The gothic shape allows more headroom at the sidewall than does the Quonset.

Rigid-frame: The rigid-frame structure has vertical sidewalls and raftersfor a clear-span construction. There are no columns or trusses to support theroof. Glued or nailed plywood gussets connect the sidewall supports to therafters to make one rigid frame. The conventional gable roof and sidewallsallow maximum interior space and air circulation. A good foundation isrequired to support the lateral load on the sidewalls.

Post and rafter and A-frame: The post and rafter is a simple constructionof an embedded post and rafters, but it requires more wood or metal than

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some other designs. Strong sidewall posts and deep post embedment arerequired to withstand outward rafter forces and wind pressures. Like the rigidframe, the post and rafter design allows more space along the sidewalls andefficient air circulation. The A-frame is similar to the post and rafterconstruction except that a collar beam ties the upper parts of the rafterstogether.

Coverings

Greenhouse coverings include long-life glass, fibreglass, rigid double-wallplastics, and film plastics with 1- to 3-year lifespans. The type of frame andcover must be matched correctly.

Glass: Glass is the traditional covering. It has a pleasing appearance, isinexpensive to maintain, and has a high degree of permanency. An aluminumframe with a glass covering provides a maintenance-free, weather-tightstructure that minimizes heat costs and retains humidity. Glass is availablein many forms that would be suitable with almost any style or architecture.Tempered glass is frequently used because it is two or three times strongerthan regular glass. Small prefabricated glass greenhouses are available fordo-it-yourself installation, but most should be built by the manufacturerbecause they can be difficult to construct.The disadvantages of glass are thatit is easily broken, is initially expensive to build, and requires must betterframe construction than fibreglass or plastic. A good foundation is required,and the frames must be strong and must fit well together to support heavy,rigid glass.

Fibreglass: Fibreglass is lightweight, strong, and practically hailproof. Agood grade of fibreglass should be used because poor grades discolour andreduce light penetration. Use only clear, transparent, or translucent gradesfor greenhouse construction. Tedlar-coated fibreglass lasts 15 to 20 years. Theresin covering the glass fibres will eventually wear off, allowing dirt to beretained by exposed fibres. A new coat of resin is needed after 10 to 15 years.Light penetration is initially as good as glass but can drop off considerablyover time with poor grades of fibreglass.

Double-wall plastic: Rigid double-layer plastic sheets of acrylic orpolycarbonate are available to give long-life, heat-saving covers. These covershave two layers of rigid plastic separated by webs. The double-layer materialretains more heat, so energy savings of 30 percent are common. The acrylic isa long-life, nonyellowing material; the polycarbonate normally yellows faster,but usually is protected by a UV-inhibitor coating on the exposed surface.Both materials carry warranties for 10 years on their light transmissionqualities. Both can be used on curved surfaces; the polycarbonate materialcan be curved the most. As a general rule, each layer reduces light by about10 percent. About 80 percent of the light filters through double-layer plastic,compared with 90 percent for glass.

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Film plastic: Film-plastic coverings are available in several grades ofquality and several different materials. Generally, these are replaced morefrequently than other covers. Structural costs are very low because the framecan be lighter and plastic film is inexpensive. Light transmission of these film-plastic coverings is comparable to glass. The films are made of polyethylene(PE), polyvinyl chloride (PVC), copolymers, and other materials. A utilitygrade of PE that will last about a year is available at local hardware stores.Commercial greenhouse grade PE has ultraviolet inhibitors in it to protectagainst ultraviolet rays; it lasts 12 to 18 months. Copolymers last 2 to 3 years.New additives have allowed the manufacture of film plastics that block andreflect radiated heat back into the greenhouse, as does glass which helps reduceheating costs. PVC or vinyl film costs two to five times as much as PE butlasts as long as five years. However, it is available only in sheets four to sixfeet wide. It attracts dust from the air, so it must be washed occasionally.

GREENHOUSE HIGH PRESSURE HEATING SYSTEM

In this system it makes no difference whether the floor of your boiler-house is on the same level as the greenhouses or two feet below or two feetabove. It will make no difference in the working of the system. In my case theground at the north-east end of the range happened to be two feet lower thanthe average level of the greenhouse site. I mention this because I think thenorth-east is where a building such as a boiler-house necessitates will castthe least shade and do the least harm. Being almost on a level with thesurrounding ground the fuel can be wheeled to the furnace doors, but betterthan that, the ashes can be wheeled out. There is no iron ladder to climb downand break your neck on Christmas eve or other occasions when a falteringstep is excusable, no water or stopped up sewer to fret and annoy.

Now all there is about this system besides saving labor and expense ininstallation is that you carry from forty to sixty pounds of steam on the boilerall the time. One practical friend said he carried sixty-five pounds and believedit was more economical of fuel than forty pounds, and by the use of acontrolling valve the pressure is reduced one, two, or five pounds on yoursystem of heating. The controlling valve recommended to us was the Mason,made in Boston, Mass., and not once in three years’ use has there been theslightest hitch or trouble with it. The slightest turn of the brass key on top ofthe valve will increase or diminish the quantity of steam passing through thevalve. A small steam clock connected with the main steam pipe a few feetbeyond the controlling valve will tell you how much steam you have on yourheating system. In our case the main steam pipe leaving the boiler is only a 3-inch, carrying of course the fifty pounds of steam till it reaches the controller,which in moderately cold weather is set at, three, in severe weather at five orsix, and this is enough to carry the steam in some of the houses for full 500

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feet. It is just worth mentioning here that while you have fifty pounds of steamin your boiler, if a sudden storm comes up or a sudden fall of temperature,all of which often happens, there is no great commotion at the fire door butsimply a turn of that little brass key on the controller and the opening of afew valves. You have the reserve steam at all times and it costs no more tokeep it there than it does to keep up two pounds of steam on your gravitysystem.

Now, whatever arrangement of heating pipes you have there will be onepipe you call your main return, which in the gravity system empties thecondensed steam into the boiler. In the pressure system it empties into anautomatic steam pump or a trap. It is my duty to be honest, so I must confessI have no experience with a steam trap of any make, but I have watched themworking and have consulted practical men who have used both the trap andautomatic pump and in every case they gave a strong preference for the pump.I understand that a trap does not work unless you have five or six poundspressure on your heating system. In the case of the pump you can have butone on your radiating pipes. That will make no difference because your pumpis supplied by a small pipe (1-inch will do) from top of your boiler - where itgets whatever pressure is in the boiler.

The return, pipe will most likely leave the greenhouse about the level ofthe floor and run as directly as convenient to the pump, where it falls into adrum or cylinder which in a small pump is about two feet long and fifteeninches in diameter. In this drum there is a float and as soon as there is a gallonof water returned the float rises and lets in a jet of steam, which starts thelittle pump working, and by a 1 1/4-inch pipe throws the water into the boilersmoothly and almost silently. This pump keeps your system clear of anycondensation. Our pump is in a small brick pit eighteen inches below the floorof the boiler-house. This can always be arranged, but it appears that is notnecessary, and if the pump were set on the floor it would work perfectly,because the pressure behind the condensed steam would raise the water twofeet into the drum. Wherever convenient, however, I would prefer to havethe drum a few inches below the return pipe so that there would be decidedfall into the drum.

In the chapter on gravity I said about all I could on the distribution ofthe radiating pipes, so what more I can say on the high pressure system willbe devoted to remarks on its advantages. Although reduced in pressure bythe controller the steam is still hotter and drier than steam only raised slightlyabove the boiling point. When occasion arrives, it will travel to the farthestpoint of your system much quicker than by increasing the supply of steamwith a stronger fire. In many places the draining of a deep pit is almostimpossible without great expense. There is no need of it in the least.

It is money wasted. You will find that after this steam under the pressuresystem has been through your heating pipes it will return to the pump in the

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shape of steam, showing that you have got the utmost benefit from it, but thechief claim of the advocates of the high pressure system is that it is a decidedsaving of fuel, and I quite agree with them. Why this should be I am not ablescientifically to argue or demonstrate, yet it seems that after you have oncegot the forty to sixty pounds on your boiler, very little additional fuel willhold it there. It is certain that any condensation returning to the boiler is muchhotter and nearer steam than under the gravity system, hence less fuel isrequired to convert into steam.

In conclusion, without an elaborate drawing it would be difficult todemonstrate the arrangements of pipes, position of the controller, pump, by-path and other points, but any one thinking of installing this up-to-date systemwould surely visit one of the many establishments where this system is inuse. He could take quite a journey for the price of digging ten feet into theearth.

There is one point more. Whatever steam system you use, don’t dependon one boiler. If you need eighty horsepower, buy two forties, if a hundredget two fifties, and it is a saving in fuel to have plenty of boiler power. As it isin hot water heaters, a little recent real experience may be worth quoting. Fortwo years we heated about 22,000 feet of glass with one boiler rated at fiftyhorsepower. Then we built on another 12,000 feet of glass situated 250 feetfrom the boiler-house, and put in another boiler rated at seventy-five horse-power. Now on occasions either one of these boilers will heat and has heatedthe whole place, but both night and day firemen are positive that one-thirdless coal is burned when they are using both boilers to do the work.

What wonderful stuff this asbestos covering is and how it enables you tocarry steam long distances in the open an! You have heard of the rival safemakers, one of whom, to demonstrate the imperviousness of his safe to heat,locked a mouse in the safe and then subjected it to great heat. On unlockingthe safe the poor mouse was found frozen to death. A more truthful experience,and one nearer home, has astonished us; in running a 2-inch steam pipe fromthe boiler house to some carnation houses 250 feet distant, we have to crossan orchard ten feet above the ground. The 2-ineh pipe is covered with asbestosan inch thick and then a thin covering of tar paper. We have noticed frequentlythat while five pounds of steam was coursing through the 2-inch pipe, icicleswere hanging from the tar paper covering.

GREENHOUSE HEATING BY HOT WATER

I must preface this very important subject by a confession. Some fifteenyears ago I made the acquaintance of a steam-fitter, a clever workman, whohad been quite successful in heating dwelling houses and thought he had ideason greenhouse heating, but on that he was away off. A weakness of the writeris to be anxious to learn from any one who he thinks knows more on any

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subject than he does. So I learned from this man several fallacies on greenhouseheating which I have many times regretted putting into print. It does me goodto confess this and I must ask the reader of the article on hot water in the firstedition of this book to believe that where my present views conflict with whatI wrote six or seven years ago, it is the result of later experience and practice.One of the fallacies was the use of small pipe, that is, less than 2-inch. Lookingat this question superficially, the small pipe, 1-inch or 1 1/4-inch has theadvantage over larger because you get more radiating surface in proportionto volume of water, but in practice this does not hold good. Friction is sogreat that the circulation is retarded, radiation is rapid and the small amountof water in the small pipe soon cools. While 11/4-inch pipe may work verywell in a house not over fifty feet, for any longer distance use nothing lessthan 2-inch.

The next and most costly mistake for me was running the pipe straightup from the boiler perhaps to within a foot of the greenhouse ridge, using a 21/2-inch or 3-ineh pipe, and at the farther end then dropping into a numberof returns containing eight or ten times the volume of water of the flow pipe.Here is a dismal specimen of one of the houses, and I have reason to believethe same mistake has been made in hundreds of cases.

He had a 4-inch pipe rising from the boiler to the height of the greenhouseridge, then branched off with a 3-inch and suspended the 3-inch about a footfrom the ridge to the farther end. The house was used for soft-wooded potplants, and was 20x100. At the farther end the 3-inch branched into two 2-inch pipes and dropped to the level of the 4-inch cast iron pipe under thebenches, with which the house was formerly heated, four under each bench.By a complicated lot of fittings one of the 2-inch pipes was actually expectedto heat the four 4-inch. Could anything be more absurd? Roughly estimated,the four 4-inch contained sixteen times as much water as the one 2-inch. Nowthis was an extremely absurd case, yet I have seen many others, where thesame mistake exists differing only in degree. What were the principal featuresin this failure? The pipe near the roof at the boiler end of the house was veryhot for perhaps twenty feet; then it gradually cooled and for more than halfthe length of the house there was not heat enough for any to be diffused amongthe plants on the benches. When the 2-inch pipe emptied into the four 4-inchreturns, 16 slightly warmed them for a few feet, and then for seventy-fivefeet there was no-heat under the benches.

I must digress a moment here to say that although growers generallywant no steam or hot water pipes beneath their benches of roses or carnations,they are of undoubted benefit to our soft-wooded plants like geraniums andbegonias. Every one who grows bedding plants must have noticed this, butunder the arrangement described we had little but cold, damp benches. Thereis a time of year, fall and spring, when this pipe near the ridge is useless.Days or nights when it may be 50 degrees outside you need a little fire heat

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to keep out dampness and keep up a healthy circulation of the atmosphere.While firing gently you will also want a little ventilation, and the heat fromoverhead pipe will entirely be wasted and your house left chill and damp.

There was an absurd controversy in the florists’ papers some fifteen yearsago on this subject of overhead heating. Some said that as the natural sourceof the heat was from above, meaning the sun, that our pipes should be nearthe roof, a great fallacy. Another piece of information which appeared someyears ago in answer to a query was, “ If you use hot water you need notexcavate for heater, if you intend to use steam then you must keep the boilerwell down below the level of the greenhouse.” Nowadays we know that,broadly speaking, the very reverse of this is the truth, for it is the return, orcool, water pressing on the warmer and lighter water in the heater that is thepower or cause of circulation, therefore the greater the height of the descendingcolumn of water the faster the circulation. In truth there has been nothingnew discovered of late about the circulation. There may have been in itsapplication.

Some forty-five years ago, perhaps before, there was published in Londonby Hood a volume on hot water. There has never been a better work on thesame subject since. We may have found out better and cheaper modes ofapplying it than prevailed in his day, but all the laws of circulation which hedemonstrates so finely are just the same today and always will be, for theyare natural laws, and can never be altered. Hood says that the circulation ofhot water was well known by the Romans, and used for heating their baths,so this wonderfully useful method of warming our houses did not originatein London, New York or Kalamazoo.

The cause of circulation is finely illustrated by Hood by having twocolumns of water, say two lengths of 4-inch pipe, each three feet high andconnected at the bottom with a piece of pipe of the same size. Half-way inthis connecting pipe there is a valve. Fill one pipe with water at thetemperature of 45 degrees, the other with hot water at a temperature of 180degrees. Then open the valve connecting the hot and cold columns. The hotwater will rise or overflow. Now this is all there is to the circulation of waterin your system. The water in the cold column or your return pipe is denserand of greater specific gravity, in simpler words, heavier and forces the lighterand warmer water out through the flow.

Now this illustrates the motive power that first starts the circulation ofhot water. It is the difference between the weight of the water in the returnpipe and that in the boiler. The water in the boiler being made lighter by thefire, the colder and heavier water forces it up and it is replaced with coldwater, so it must follow that the higher, and consequently heavier, the columnof water in the return pipe the faster will be your circulation. And it followsagain that the faster the circulation the hotter will your pipes be, for the waterreturning quickly to the fire has not time to get cool. “When your return pipe

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near the boiler is nearly as hot as the flow where it leaves the boiler yourcirculation is perfect. All of which goes to prove that the lower the boiler thebetter the apparatus will work.

Reserve all your drop till you get near the boiler and then dropperpendicularly down. This talk about giving the pipes a rise of one foot in ahundred, or the same drop, is all bosh. If the pipes were a dead level in thehouse it Mould be perfect, but it is better to have a rise or fall of two inches ina hundred feet because you want when emptying the pipes to have a drainout. Providing your boiler is well down, and that is the very essence of thewhole job, it makes no difference whether you have a slight rise in the flowpipes in the house or a slight fall.

As admitted at the start of this chapter, I was the victim of false theoriesand it took a few years to break away and return to principles I knew of manyyears ago. Now here is what we did some fourteen years ago and it was wrong.The top of the heater was some two feet six inches below the joists of theboiler shed. It would have been lower could drainage have been secured. Asit was we had to lay a 4-inch socket tile 650 feet. The two 3-inch flow pipesrose straight up through the floor and eight feet higher. This was done toavoid the doorway leading from the shed into the greenhouses. Please imaginethese two houses to be 20x125, running east and west and connected bywooden partitions and wooden gutter. The flow pipe in the houses was a 2-inch on each side of the houses, running a foot below the gutter and on thesides, on the posts a foot below the plate from which the bars spring; that is agood place for them, but it is by no means overhead heating, it is resisting thecold at a very important spot.

As we had carried the flow pipe from the boiler so high in the shed, wehad to drop again to the level of the flow pipes in the houses, which wasdone with two elbows and a short piece of 2-inch pipe. Although it worked,this was against correct principles. The hot water had no natural tendency todescend, it would rather flow out on a level. At the farther end of the 125 feetthe 2-inch flow went into a manifold with five 1 1/4-inch openings and theywere the returns.

This was not anything like as wrong as the case described earlier, yetthere was not hot water enough in the 2-inch to supply the five 114-inch. Thatand other causes, friction and rapid radiation, all combined made it a poorjob. There was no need of air pipes or petcocks in this system because a 3/4-inch pipe was tapped into the highest point of pipes in the shed and led up toa large tank which supplied us with water for the houses.

This tank held 150 barrels of water and from it a 1 1/4-inch pipe wasconnected with the return pipe close to the boiler. It may have given us aconstant pressure of fifteen pounds. Please don’t attach the slightestimportance to the fact that this feed pipe was from a vessel holding 150 barrelsof water. Pressure in a liquid depends entirely on height, not bulk. A funnel

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thirty feet high and three feet in diameter at the top, tapering to one inch atbottom, would give no more pressure at the one inch opening than a columnof water one inch in diameter and the same height as the spreading funnel.Now we ran along several years with the arrangement above described,thought it perfection, and read that others in the east had adopted the sameprinciple. Yet now we feel sure that many tons of coal were wasted in tryingto get those return pipes hot. With the small amount of radiation we wantedthem hot, not lukewarm.

So two years ago last summer we made a great change, not with theslightest feeling of doubt or uncertainty, because we were only going on thesound laws and principles of hot water circulation. Needed a new heater,wrongly called boiler, and purchased the Burnham. Let me say here that Ihave reason to deeply regret recommending boilers in the past withoutsufficient experience with them.

Still, after two years’ trial I must say that the Burnham boiler has provedmost satisfactory. It is of heavy castings, simply put together and easily andthoroughly cleaned. In place of one 2-inch flow pipe we hung two 2-inch andat the farther end ran each flow pipe into a manifold with three 1 1/4-inchopenings. The 2-inch could amply supply the three 11/4-inch and they werehot back to the shed. I should have started at the boiler. Will say now that theBurnham boiler, or the size we have, has a 5-inch opening for flow, same forreturn. We ran up as near as possible to the joist with a 5-inch pipe, thenbranched with a tee into a horizontal 4-inch pipe. I now remember there wereeight 2-inch pipes to be supplied from this 4-inch. On reaching the south sideof the north house we branched with a tee into a 3-inch and rose to the levelof the flow pipes. Remember, the 4x3x3 tee came out of a 4-inch horizontal,not an upright, pipe. After supplying that run the flow was reduced to 3-inchand went on to supply the north run of the north house. From the shed end tothe farther end the 2-inch flow pipes have a rise of two inches. On reachingthe manifolds provision had to be made to let air escape, so we tapped intoeach manifold a small petcock. For the first ten days after we begin to firewith a new supply of water there is a considerable accumulation of air. Aftera week or two there seems no more in the water, and if these petcocks areneglected for weeks no harm results. This I have found repeatedly to be thecase. You may not have a tank of water raised up thirty feet, and there is noneed of it. A barrel or box holding twenty gallons of water, or a metal boiler,such as is used in our kitchens, will do equany as well. Let it be seven oreight feet above the bottom of your heater, and when there is a gallon of waterin the barrel your system is full. In this case you do not need petcocks, but inplace of them simply tap in a 1/2-inch pipe and let it run up a rafter oranywhere out of the way; that, of course, will constantly and with-, outwatching vent the heating pipes of any air. See that the top of this little pipeis a foot or two above the highest point of the water in your feeding cistern.

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This rearrangement of pipes was a great satisfaction and unqualified success,so much so that in seven or eight other houses where the overhead and riseand fall delusion had existed it was all pulled down and common sense anda smooth, quick circulation exists.

If an amateur came to me and asked for advice about heating a house20x100, or 20x75. and wanted it so arranged that he could leave it at 11 p.m.till 6 a.m., I should without hesitation tell him to get plenty of boiler powerand use the 4-inch pipe for radiation. Use the heavy cast iron pipe used byhorticultural builders, 9-foot lengths and four inches outside diameter, notwhat many greenhouse men have used, Mott’s soil pipe, 5-foot lengths, fourinches inside diameter; this is a thin, brittle metal, unsatisfactory in many ways.

I have had no experience with water under pressure, unless the pressureof our city mains constitutes that system. Our city water has a pressure ofabout thirty-five pounds to the square inch, and in my experience it is a verycheap heating system for a store or office. Wishing to heat a flower store inthis city, which is something like 19x80 feet in area, I put a small heater in thecellar. It is simply three lengths of 3-inch pipe, each about three feet long, andrun into a manifold at both ends. The coil is resting on two 4-inch brick wallsabout two feet from the floor and is bricked over top, sides and ends. Oneend of this coil is raised about three inches and from it rising to the floor isthe 11/4-inch flow, which leads off, and by the help of some tees connectswith three radiators on the floor of the store, and from the other end of theradiators the returns drop to the lower end of the coil. There are two naturalgas burners under this very simple heater, which in the coldest weather hasnever been turned on more than one-third its force.

A 1-inch pipe from the city water is connected with the lowest part of thecoil and the valve is never closed, so there is always a pressure of thirty-fivepounds on the pipe and radiators. The highest part of the system is the top ofthe radiators, and in them is a petcock which should be opened every day tolet out air, but often is not for weeks, and in a radiator it is not of so muchconsequence.

There is nothing more about it, only the radiators can be made very hot;a great success. If a strong fire should expand the water in the heater it has tofind room by driving the water back into the mains. The whole thing cost lessthan $50, and $5 worth of gas was consumed in the coldest month. Now thissystem could be used with great success wherever you have a boiler thatwould stand the pressure. You could use it on either the uphill or downhillsystems, but you could not have any open air vents, and unless you trustedto the automatic air valves you would have to daily open the petcocks at thehighest point.

The architecture of many greenhouse establishments, the smaller andolder places more especially, is so complicated and houses are on so manydifferent levels, that sound advice as to how to arrange pipes is impossible

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unless you are on the spot. Three rules can be accepted as sound. Provideplenty of capacity in both flow and return pipes. Don’t use less than 2-inchpipes, and reserve all the drop on your return pipes till just before you enterthe boiler. Watch these points and you will save coal.

On a cold, wet, windy, winter’s day there is nothing more therapeuticthan pottering about in a greenhouse. A cup of tea, packet of biscuits, theradio and some seeds to sow or cuttings to take; life doesn’t get much better.The trouble is such simple horticultural pleasures don’t come cheap.Depending on the temperature which has to be maintained and the severityof the weather greenhouse heating bills can quite literally go through thepoorly insulated glass roof and sides.

Due in part to the high cost of heating many greenhouse gardeners onlyturn their heating systems on towards the end of winter or early spring. Forthose without any form of heating now is the time to decide which type offuel to use and the best kind of heater to suit their plants and budget.

FOOD PRODUCED IN GREENHOUSES

Although greenhouses have been in existence since 1800 (or earlier), andgreenhouse food production started to develop as an industry in the secondhalf of the nineteenth century, the largest growth and expansion of thegreenhouse industry occurred throughout the world following World WarII. Today, food production in greenhouses can be found in all continents. Mostpopular food crops grown in greenhouses are tomato (beefsteak, cluster, Italian, cherry), cucumber, and sweet pepper. Other greenhouse grown vegetables include watermelon, muskmelon, summer squash, zucchini, lettuce, eggplant, snap beans, celery, cabbage, radish, Welsh onion, and asparagus. Fruits such as grapes, strawberry, banana, pineapple, papaya, orange, mandarin, cherry, and figure, as well as culinary and medicinal herbs, are also grown in greenhouses.

T ODAY’S GREENHOUSES

The main greenhouse covering materials are glass and polyethylene (PE).Glass has been used since the early days of greenhouses. The introduction ofPE film after World War II was the main reason for the expansion ofgreenhouse production around the world, and it is now the most widely usedcovering material in the world. Glass-covered greenhouses are concentratedmainly in northern Europe and North America. The low cost of the PEgreenhouse is the main reason for its high popularity, especially in developingcountries. In recent years, the use of PE-greenhouses has even spread tonorthern regions. Research has shown that, under Canadian climaticconditions, heating costs of a double-layer PE-greenhouse are 20 to 30 percentlower than for a glass-covered greenhouse. Most of the greenhouses built now

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in Canada are covered with PE. Standard PE film blocks the ultraviolet, butnot the infrared radiation, and has a short durability. However, improved PEfilms retain the infrared, but allow the ultraviolet, radiation (necessary forthe bees, used for pollination of plants, to orient themselves) in the greenhouse,and are more durable. Polyvinyl chloride (PVC), another plastic film used tocover greenhouses, is used mostly in Japan. Other covering materials forgreenhouses include rigid plastic acrylic, fibreglass, polycarbonate, and PVCpanels, but their use is generally limited because of their high cost, comparedto PE. Beside glass and PE, polycarbonate is often used on the sidewalls ofpolyethylene greenhouses in northern regions because of its good insulation,durability, and reasonable cost.

Greenhouses come in many styles and sizes, from the original houseswith minimal climate control (furnace and vents) to the modern 10-ha (25-acre) or more, multispan greenhouses with high-tech climate controls(sophisticated and powerful heating system, CO 2 enrichment, evaporativecooling pads, exhaust fans, roof vents, thermal/shade curtain, computercontrols, light sensors). Most sophisticated greenhouses are generally foundin the developed, northern countries. Phytotrons are highly sophisticatedstructures that allow for accurate control of environmental conditionsincluding light, and are generally used for scientific research in universitiesand research institutes. However, phytotrons cannot be consideredgreenhouses since they are not covered with a transparent material. The degreeof environment control needed depends on various factors. The first factor isthe location of the greenhouse (local climatic conditions). Northern regionsare characterized by cold winters and warm summers. If the objective is togrow plants all year long, then such large differences in climatic conditionsbetween winter and summer require a high-tech greenhouse. In regions suchas the Mediterranean (Spain, Italy, Morocco, Greece), the mild winter climatedoes not require the use of powerful heating systems, and low-techgreenhouses are sufficient for winter production. However, these regions havevery hot summers, and the use of a low-tech greenhouse may not providesatisfactory temperature control to grow plants during summertime.

The production schedule also affects the level of environment control andthus the level of technology. A greenhouse in northern regions may require ahigh level of climate control if the objective is to grow crops all year long (orlong-season crops). If the objective is only to extend the production season(e.g., one early crop in spring), then a less sophisticated greenhouse could besatisfactory. Optimal growing conditions differ from one species to another.For example, lettuce prefers cooler temperatures than cucumber. Thus, thecrop grown in the greenhouse may influence the level of environment controlneeded or desired. A low-tech greenhouse may provide sufficient climatecontrol for lettuce but not for cucumber, depending on the location of thegreenhouse and the production schedule.

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Economic development also plays a role in the level of technology usedin the greenhouse. In developing countries, growers may not be able to affordthe most sophisticated equipment, and may lack technical expertise andtechnical support. Although greenhouses were developed in northern regionsas a means of protecting crops against cold temperatures, and are thereforegenerally associated with cold climates, they are also used in arid regions suchas Saudi Arabia. In such regions, the objective of the greenhouse is to protectplants from the excessive solar radiation and temperature, and to preventexcessive water loss by plants (especially since water resources are generallylimited in those regions). Therefore, technology in greenhouses in these regionsis directed towards cooling. In northern countries, high-tech greenhouses canprovide optimal growing conditions (temperature, humidity, carbon dioxide)for vegetable crops even during the coldest winter months. However, evenwith excellent climate control, yield and quality of crops grown during thesemonths are low due to the low light level available. Research has shown thatit is possible to produce good yield of high-quality produce during the wintermonths by using artificial light to supplement the natural radiation. The mostcommon artificial lighting is the high-pressure sodium lamp. The high costof electric energy in many regions is the most important factor preventing anincreased use of artificial light.

PRODUCTION SYSTEMS

Since the early days of greenhouses, plants have been grown in soil or insoil-filled containers. The first technique for fertilizing plants, which is still inuse today in organic production, was the use of manure. Today, fertilizationof plants can also be accomplished by incorporating chemical fertilizers inthe soil, or by distributing fertilizers dissolved in water (so-called fertigation)to plants with a drip (trickle) irrigation system.

Intensive and repetitive cultivation of crops on the same soil generallyresults in a degradation of soil properties and fertility. Salt accumulation maybe another problem in soil cultivation. Incorporation of manure, compost, andother organic materials into soil can be used to improve its structure andreplenish its fertility. However, ensuring perfect fertilization of plants grownin soil is still a difficult task. Furthermore, intensive and repetitive cultivationof crops on the same soil can also result in insect or disease infestation. Soilreplacement and soil fumigation are two solutions, but the first technique isexpensive and the second is not always successful. Greenhouse productionin soil is still used widely.

GROWING WIT HOUT SOIL

In order to better control fertilization for optimizing plant growth andyield, and also to avoid the problems occurring in soil, growing systems that

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do not use soil (soilless) were developed for the cultivation of greenhousecrops. These soilless systems can be classified in two groups: liquid (water)and solid (artificial substrates that are either inorganic or organic). Systemsusing water as a growing medium are the nutrient film technique (NFT), deepflow technique (DFT), and aeroponics.

Common inorganic media are rockwool, vermiculite, perlite, and claypellets. Organic substrates are peat, coconut coir, sawdust, and straw.Inorganic and organic substrates are usually contained in bags, and plantsare irrigated with a complete nutrient solution distributed by a drip irrigationsystem. The excess of nutrient solution can either be allowed to leak into theground or is recuperated and recirculated (after treatment) to plants. In liquidsystems, plant roots are continuously exposed to nutrient solution, which isnot leaked into the ground.

Growing methods using artificial substrates or water are known as soillessculture or hydroponics. Hydroponics is literally defined as the growing ofplants in water, but the plants are actually grown in a complete nutrientsolution. Ideally, the term hydroponics should be reserved for water culture,and the term soilless culture for plant cultivation on artificial substrates. Inpractice, the terms hydroponics and soilless culture are used indiscriminately todescribe water and substrate-based systems.

Although official statistics are unavailable, hydroponic systems are knownto be used extensively for food production in greenhouses. The most popularsoilless medium for hydroponic vegetable production is rockwool. Thenutrient film technique is also often used, but to a much lesser extend thanrockwool. In some regions, the availability of low-cost materials may providealternative substrates. For example, in British Columbia, sawdust, a residueof the large forestry industry, is commonly used as a substrate. Both aeroponicsand DFT remain in little use today.

INSECT AND DISEASE CONT ROL IN GREENHOUSES

One objective of hydroponics is to avoid insects and diseases that mayoccur in soil. In a soilless culture system, such as rockwool, it is easy to removeinfected plants. However, spread of diseases can occur very quickly in systemswhere nutrient solution is recirculated. Methods such as filtration of thenutrient solution, and disinfection with ozone or ultraviolet light, have beendeveloped to eliminate pathogens that may be present in the nutrient solution.However, these methods are often expensive and not completely effective.

Greenhouses are used to create and maintain an environment ideal forplants. However, this environment is often favorable for insects and pathogenstoo. In the past, the control of insects and diseases in greenhouses wasaccomplished with the use of pesticides, but over time both insects anddiseases have developed resistance to such pesticides, while consumers havebegun to demand pesticide-free produce. Biological agents are now used to

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control whitefly, thrips, aphids, and two-spotted spider mite in greenhouses;few reliable biological agents are currently available for the control of diseases.

RESEARCH ON GREENHOUSE FOOD CROPS

In countries or regions where greenhouse production is an importantindustry, government and universities are generally involved in research ongreenhouse production. The general objective of the research is to improveyield and quality of produce and profitability of production, by investigatingall aspects of greenhouse production: greenhouse design and coveringmaterials, growing methods, environment controls, substrates, plant nutrition,plant pathology, and insect control. Grower associations may also be involvedin the development of research priorities, and may contribute financially tothe expenses of research.

Due to the presence of a large and technologically advanced greenhouseindustry in the Netherlands, the most notable research institutions are foundthere. The Research Station for Floriculture and Glasshouse Vegetables (underthe Ministry of Agriculture, Nature Conservancy and Fisheries) has five sites.The other important Dutch institution is the University of Wageningen.

In the United Kingdom, Horticulture Research International (HRI), thelargest horticultural research establishment in the world, maintains an activeresearch programme on greenhouse crops and provides its services (fromfundamental research to technology transfer) to research councils, governmentdepartments, growers, and commercial industries, in the EuropeanCommunity (EC) and other countries.

In the Americas, the Greenhouse and Processing Crops Research Centre(GPCRC; Agriculture and Agri-Food Canada) is the largest research facilityspecializing in greenhouse vegetables. The GPCRC is a leading member ofthe Canadian Network for Greenhouse Vegetable Research.

Japan, Spain, and Israel are some of the other countries with importantresearch programmes in horticulture, including greenhouse food production.

The International Society for Horticultural Science (ISHS) is aninternational organization of horticultural scientists, which aims at promotingresearch in all branches of horticulture, including greenhouse food production.Within the ISHS, there are various commissions and working groups relatedto greenhouse production.

FUT URE OF GREENHOUSE FOOD PRODUCT ION

As the world population continues to increase, and more agricultural landis lost to urban development, intensive food production in greenhouses mayplay a more important role in food production. Furthermore, improvingeconomic conditions in developing countries and an increasing preoccupationwith health and nutrition will increase demand for high-quality food products.

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Through controlled climate and reduced pesticide use, greenhouses can meetthis consumer demand. Foods with improved health characteristics orcontaining nutraceuticals (substances with pharmaceutical or health-beneficialproperties that can be extracted or purified from plants) can be grownpesticide-free in greenhouses.

ORGANIC GROWING MEDIA ANDFERTILIZERS FOR GREENHOUSES

Organic methods of plant production are of increasing interest to manygrowers of horticulture crops including greenhouse operators. Somegreenhouses are seeking “organic certification” in the same way as farmersof edible crops and livestock. The purpose of this chapter is to review thecurrent status of “organic” growing media and fertilizers for greenhouse cropproduction. It is said that the true organic grower is seeking to produce cropsin “balance” or “harmony” with their environment so that finding chemicalpesticide and fertilizer substitutes will be unnecessary. I believe this mighttrue in outdoor crop production, but in the rather unique greenhouseenvironment this might not be readily achieved.

The greenhouse environment is very effective at promoting rapid plantgrowth, as well as insects and diseases in a short time period. Greenhousecrops are grown in small volume containers and they are irrigated frequentlywith large volumes of water. The combination of rapid plant growth, limitedroot volume, and frequent leaching make fertility management a challengewithout the use of water-soluble chemical fertilizers.

Right now, because of the special challenges of growing in the greenhouse,it may be difficult for a traditional greenhouse grower to quickly transforminto an “organic grower” without first looking for substitutes.

Personally I’ve got no problem with looking for organically acceptablesubstitutes for the traditional soilless growing media and chemical fertilizers.I believe that finding substitutes may be about the only way for mostgreenhouse growers have in immediate future to get into organic growingand that’s what this piece is about. Probably the first thing that comes to mindwhen we talk “organic” is composting and using the compost as a substitutefor traditional soilless media.

T HE BASIC COM POST ING PROCESS

The general steps in the biological process which creates compost are thesame regardless of the raw materials being composted or the size andcomplexity of the production facility. A compost must pass through all of thesteps outlined here in order for it to be considered of high enough quality foruse in organic potting mixes. The progress of organic matter decompositionduring composting can be followed by monitoring the temperature of the

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compost pile. During the initial phase of composting the temperature of thepile increases rapidly as the population and activity of decay microorganismsincreases in response to the readily decomposable carbon in the raw materials.The goals are to reach a temperature between 130°F or more and to maintainthis temperature range until the microorganisms begin to exhaust the readilyavailable carbon. Heating to these temperatures is critical for high qualitygreenhouse compost because it is in this range that weed seeds and most plantand human pathogens that might come from animal manure are killed. Duringcomposting the pile is turned and remixed several times to ensure completeheating and decomposition.

Following the high temperature phase there is an extended period ofgradual temperature decline until the pile reaches ambient air temperature.Now, if the pile is turned, reheating will not occur. At this point the compostis said to be “near maturity”, but to ensure that the compost is stable andready to use, most producers allow some extra time for the compost to “cure”.How long composting lasts varies with the method. It could take about 1-2years in a static unturned pile, 6-9 months if the pile is turned occasionally,or only 1-4 months the pile is turned frequently.

To comply with the National Organic Programme standards compostpiles must maintain 131-170ºF for at least 3 days (static pile) or at least 15days (windrow, turned at least 5 times). High temperatures are necessary tokill any human pathogens especially if farm manure is a component. Also,weed seeds and plant diseases are most successfully killed at hightemperatures.

Many types of raw materials can be used for making compost; somecommon materials are listed in the following table. Pay close attention to thecomments in the table.

Table : Some Raw Materials for Making Compost

Material Comments

Farm animal manure Must be compostedStraw and bedding Crop residues Must be pesticide freeFruit & vegetable wastes Must be pesticide freeFood processing wastes Seafood processing waste Grass clippings Must be pesticide freeSawdust & other wood wastes Use in moderation, low nutrient value.Newspaper Black ink only, shredded, <25%Leaves Shredded

It is important that the raw materials be properly prepared prior to mixingand the start of composting. Most organic materials must be shredded orground to reduce particle size and help make them less resistant to decay.

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During composting, oxygen and moisture levels are critical factors indetermining the degree of decomposition which takes place and the length oftime it takes to reach a stable product. Oxygen levels below 5% and moisturelevels above or below the range of 40-65% inhibit the composting process.Most composting operations aerate the piles and irrigate them if conditionsfavour excessive drying. The volume of the finished compost is smaller thanthe volume of raw materials because of the breakdown of organic matter andthe evaporation of water.

When is the compost ready for use? Currently there is no single widelyaccepted criterion to determine when compost is “done”. Measurements oftemperature, respiration, ammonia production, pH, and carbon:nitrogen (C:N)ratio are among the potential indicators of compost maturity, but no one factoris completely reliable. Generally, at the end of active composting (heatingperiod) producers allow a “curing” period of about 1-2 months to make surethe compost is stable before it’s used.

IRRIGATION WATER QUALITY FOR GREENHOUSE PRODUCTION

Most greenhouse media contain 30 to 60 percent peat moss alone or incombination with composted pine bark. Other materials are added fordrainage and aeration. In terms of air/water relations in the root zone, thequality of the peat used is very important. Peat that has been milled too muchhas a smaller fibre size. Media settling may result in loss of plant-rootingvolume. Also, aggregates such as vermiculite may or may not improvedrainage and air space, depending on the size and shape of the particle.

Fine-grade vermiculite particles may actually decrease media aeration.Only a portion of the water added to media is available for root uptake.

Available water-holding capacity is the amount of water held in the root zoneand available to plants between irrigating and when the plant wilts. In a 6-inch pot, approximately 65 percent of the pore space is filled with water afterthe pot has been saturated and allowed to drain.

Generally only about 70 percent of that water is available; the rest is calledunavailable water. The amount of available water depends on how tightlythe water is held to the particles of materials that make up the media (matrictension).

For example, peat has relatively higher unavailable water contents at agiven matric tension compared to rock wool. This variability in the availabilityof water in different types of media components means no two media areexactly alike in terms of providing water to plants. This makes knowing whento water difficult. Another important characteristic of media components thatinfluences watering practices is wettability, i.e., the ability of dry media torapidly absorb water when moistened.

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The choice of media should be influenced by irrigation systems andpractices.

M EDIA COLUM N HEIGHT /CONTAINERS

Another factor relating media to air/water relations in the root zone isthe size of the growing container. With media in containers, the amount ofair and water held in a given media is a function of the height of the columnof media. The taller the column, the smaller the ratio of water to air spaces.

This is most important in plug production where the small cells drainvery poorly or not at all, resulting in poor root zone aeration.

The dramatic effect of container height on air space is evident, whichshows the change in percent volume air space of a 1:1 peat:vermiculite mediain various growing containers.

In all containers, there will be a certain amount of saturated media at thebottom of the container after drainage. This is due to what is called a >perchedwater table. The saturation zone is a larger part of the total volume of thegrowing media in a very short container, such as a plug cell.

A good way to illustrate the effect of container height is to use a sponge.A sponge of the dimensions 2@ x 4.25@ x 8.5@ (72.25 cubic inches or 1,184milliliters) represents the media in a container. When fully saturated, thesponge holds 950 ml; that is, the total porosity is 80 percent. Holding thesponge so it is 2 inches high results in about 50 ml water draining out, resultingin a volume air space of 4.2 percent. If it then is held so it is 4.25 inches high,another 125 ml drains out, resulting in a volume air space of 14.8 percent. Ifthe sponge is then held so it is 8.5 inches high, another 375 ml drains out,resulting in a volume air space of 46.5 percent. Starting with the same volumeof media (sponge), the effect of container height (sponge height) on media airspace is dramatic. We can conclude that the choice of containers is importantin managing water/air relations in the root zone, especially of plugs.

Another issue is whether square or round plug cells are better. In general,square cells with their greater volume make crop management a little easier.As long as the height is the same, however, there is no difference in air space.

Media Handling

How media are handled can greatly influence their air and watercharacteristics. The major concern is to avoid compaction. Containers,including plug trays, should be lightly filled and the excess brushed off thetop. Air space can be drastically reduced by compaction. At no time shouldany growing containers be stacked. The moisture content of the media priorto filling containers may also be important. Adding water to peat based mixesbefore filling plug trays causes the media to swell and helps create moreaeration. Water added to about 100 percent by weight of the media is sufficientfor cell packs. Plug mixes should have about 200 percent by weight water

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added before filling plug trays. Moistening of the media before filling largercontainers does not have much benefit.

GREENHOUSE TECHNOLOGY AND DESIGN

The Dutch and pseudo Dutch (particularly Britain and Sweden) industriesare founded on glass. Two key greenhouse designs are used – the narrowglasshouse, often called the Dutch Venlo is the dominant design used forvegetable production, while the wide span is favoured in the production ofcutflowers.

The height of greenhouses has a significant impact of the capacity tocontrol the growing environment. In these industries, the older structures thathave not been replaced have gutter heights in the range of 3 – 4 metres. Thereare few of these structures remaining. More recently constructed greenhouseshave higher structural designs. Common heights are 4.5 metres, with somebeing built to 5.5 metres. Some greenhouse manufacturers expect futuredesigns will have higher gutters.

Thermal screens are used in a lot of this industry as a method of savingenergy (as much as 19%) in heating the greenhouse, however, though theycan have a dual purpose by preventing excess energy from entering thegreenhouse in summer, they are seldom used for this second purpose. Growershave found that when drawn, the adverse impact on ventilation, coupled withlower light levels is not acceptable. Subsequently, greenhouses arewhitewashed for the summer period instead. Many operators apply thewhitewash through sprinklers installed on the roof of the greenhouse usingproducts that either readily wash off or becomes clear in rain. This ensuresmaximum light reaches the crop in overcast conditions.

The Dutch industry produces approximately 60% of the cutflowers soldin the world market and 50% of potted plants. The production of potted plantsis increasing at a faster rate than cutflowers. In vegetables, tomatoes have thereputation as the largest crop area, but new developments have tendedtowards capsicums. Overseas, growers build greenhouses for particular crops;they specialise and become expert in a single crop. Very few growers producemore than one type of crop.

Automation of the greenhouse is standard practice in these industries.As in Australia, labour is expensive and often difficult to recruit. Computersmonitor and control the growing environment as well as the nutrient dosingand irrigation of the hydroponics system. The pinnacle of this drive forautomation is found in some of the nursery operations in which a computeris able to control not only the growing environment of the plant, but also moveand sort plants within the greenhouse. In one 20 hectare nursery, a total ofjust six employees, are involved in little more than supplying inputs such aspotting mix to the potting machine and nutrients to the dosing system, and

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plastic sleeving the plants that are ready for shipping –everything else isundertaken by the computer.

T HE CANADIAN INDUST RY

The greenhouses used in Canada are similar to the Dutch industry andthe Dutch influence is quite strong. Growers travel to The Netherlands on afrequent basis. The primary difference is seen in the use of plastic cladding,which is rarely used in Holland. Plastic offers the Canadians a less expensiveoption and because of the higher light levels experienced in Canada,particularly Ontario, compared with Holland, there are not the samedisadvantages. As a result, plastic greenhouses outnumber glass by roughly2:1. A notable innovation, however, is the development of curved glass sheetsenabling the construction of glasshouses with curved roof designs. A curvedroof permits a higher average level of light transmission over the course ofthe day and season.

Structures in this industry have high gutter heights, which enableimproved management of the growing environment.

Greenhouses tend to have gutters 4 – 5 metres above the ground level.Older sheds with smaller air volumes (lower rooves) are generally used forcucumber production because they are less suitable for solanaceous (tomato,capsicum and eggplant) crops.

The floricultural industry makes greater use of glasshouses. The largestflower producing area in the country is the Niagara Peninsula. In BritishColumbia floriculture has been growing at a rate of around five percent perannum. This is similar to most other countries, including Australia, whichare experiencing a strong demand for cutflower crops. Fully automatedfloriculture greenhouse operations may cost as much as $350/m2 to build.

The Spanish Industry

There are over 40000 hectares of greenhouse structures in Spain,concentrated along the coast from Alicante on the east to Almeria in the south.The majority of the structures are of the traditional Spanish greenhouse, butnew developments are mostly of modern European designs. Almost half thisarea is used to produce tomatoes, though flower exports from Spain aregrowing annually.

The structures in the traditional Spanish industry are basic.Typically a tent structure, the greenhouses are used for production of

vegetable crops sold into the European supermarkets. The supermarket chainstend to source Spanish product during winter.

This traditional greenhouse evolved in Spain as poor land holders beganto realise the benefits of covering crops with plastic. These structures aregenerally unheated and poorly ventilated. The greenhouses are used toproduce crops from autumn to spring. Very few are used during the hot

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summer months. The structures are essentially two layers of wire nettingsuspended by posts. The older greenhouses have wooden posts, while themore recently built structures make use of stainless steel. Plastic film andoccasionally woven plastic sheeting is inserted between the netting. Productionmay be in either hydroponics or soil. Though the move towards soillesssystems is occurring, the vast scale of the industry and the relaxed attitude ofthe growers will probably mean that soil production will continue for manyyears to come.

The newer greenhouse industry takes after the more typical European,that is Dutch, designs. Structures have a minimum gutter height of 4 metres,several extending to 5.5 metres to attain better environmental managementduring the intensely hot summers. Plastic and glass cladding are used, thoughwhere the investment directly involves a Dutch connection, glass is usedwithout question – a habit more than a requirement, as the Spanish industryis quite south compared with other greenhouse production areas in Europeand consequently receives a lot more sunlight. Polycarbonate is increasinglypopular and is used on some or all the walls as this material provides extrastructural integrity in the face of moderate sea breezes. Gable and curved roofdesigns are most common. Hydronic heating is standard and often involvesco-generation of carbon dioxide for use in the crop. Insect screens are used inmany of the modern structures.

SUCCESSFUL CROP PRODUCTION INCOMMERCIAL GREENHOUSES

Successful crop production requires that crop pests and diseases bemanaged so that the effects of diseases and pests on the plants are minimized.The management of crop diseases is directed at preventing the establishmentof diseases and minimizing the development and spread of any diseases thatbecome established in the crop. Managing pest problems is directed atpreventing the pest populations from becoming too large and uncontrollable(Portree 1996). The presence of pests and diseases are a fact of crop productionand growers must use all available options and strategies to avoid seriouspest and disease problems. Integrated pest management (IPM) is a term usedto describe an evolving process where cultural, biological, and chemicalcontrols are included in a holistic approach of pest and disease control. Keycomponents of effective pest and disease control programmes include: cropmonitoring, cultural control, resistant cultivars, biological control and chemicalcontrol.

CROP M ONIT ORING

Crop monitoring is the continually on-going surveillance to detect thepresence of a pest or disease at the very early stages of development of the

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disease or pest population, before economic damage has occurred. Everyoneinvolved in working the crop should be made aware of the common pest anddisease problems and what to look for to detect the presence of problems inthe crop. In addition this general surveillance of the crop, dedicated monitoringof the crop should be included in the weekly work schedule. Blue sticky cards,placed throughout the crop, are a useful monitoring tool to help trap and detectpest problems before they become a problem. Yellow sticky cards are knownto attract and catch some biological control agents e.g. Aphidius sp. (Don Elliott,pers comm) Biological control agents can be released well in advance of anypest population explosion thus allowing for the establishment of the controlagents and prevention of a serious pest problem.

Crop monitoring should begin when the crop is still on the seedling tableor at the transplant stage (especially when transplants are obtained by anothergreenhouse i.e. Purchased from a propagator). If transplants are beingpurchased from a propagator it is important to be in contact with thepropagator regarding any pest problems encountered during the productionof the transplants. It is also important to know what pest control measureswere used, if any, to control the problems. It is advisable to establish inadvance, what control measures you are willing to have applied to thetransplants at the propagator’s prior to receiving them into the greenhouse.The concern is that any pesticides that are applied are compatible with thepest control programmes i.e. biological control programmes that will be usedfor the duration of the crop. Some growers may insist that only biologicalcontrols be used during the production of transplants, and/or that biologicalcontrol agents be introduced preventively to the transplants before they arereceived.

Cultural Control

Cultural control involves providing the conditions that favour the growth,development and health of the crop, and where ever possible, providingconditions that work against pest and disease. Many disease causing fungiand bacteria require the presence of free water or condensation on the plantsin order to cause disease. High relative humidity promotes the developmentof disease, and maintaining the environment below 85% relative humiditywill help to escape or avoid disease problems. Ensuring proper ventilationand air movement within the crop canopy, as well as maintaining optimumplant spacing and a relatively open canopy, will ensure good air circulationand minimize the establishment of micro-climates that favour diseasedevelopment. Proper contouring of the greenhouse floor will avoid the poolingof water which contributes to localized high relative humidity. Optimizingthe greenhouse environment to favour the development of the plant willensure a strong, healthy plant which is not only a prerequisite for high yieldsbut also results in plants that are better able to resist diseases and insect pests.

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Good crop sanitation is another important component of successfulcultural control. The plants must be pruned and maintained on schedule, allcrop debris should be promptly removed from the vicinity of the greenhouse.Any weeds that happen to gain a foot hold through gaps in the floor plasticshould be removed immediately upon discovery and the floor repaired.Personal plants “pet” plants should not be grown in the greenhouse. Bothweeds and “pet” plants can be as source and “haven” for pest and diseaseproblems.

Pruning tools and other equipment should be cleaned and disinfectedon a regular basis. Aprons or other clothing worn by the workers should bewashed frequently. When a disease or pest problem area exists in thegreenhouse, that area of the greenhouse should be worked last, to avoid thespread of the disease or pests by the workers. In this situation, special caremust be taken to disinfect tools and to clean clothing.

Maintain a 6 to 10 meter wide buffer zone around the outside of thegreenhouse by regularly mowing any weeds that try to grow in this zone.The presence of plants in close proximity to the greenhouse can serve as areservoirs for continual introduction of pests and diseases into the greenhouse.Screening of the intake vents can also play an important role in excludingpests from the greenhouse. It is not enough just to screen-off the intake ventsas the screening restricts the air flow into the greenhouse, it is important toensure that the surface area of the screening used is large enough so that itdoes not restrict the flow of air into the intake vents. This may require that ascreen ‘chamber’ be constructed over the vent.

Resistant Cultivars

Plant breeders have had considerable success in developing cultivars thatcontain genetic resistance or tolerance to diseases. When selecting the cultivarsto be grown, it is important to consider the genetic resistance of the cultivarsto the prevalent disease problems in the region. The development of cultivarspossessing genetic resistance to pests has been relatively unsuccessful,however, the techniques of genetic engineering have made inroads inconferring pest resistance in plants. Genetically modified, pest resistant plantsmay become available to greenhouse growers in future. The development useof genetically modified plants or genetically modified organisms (GMOs) iscurrently a contentious issue, and may not be accepted by growers orconsumers.

Biological Control

Biological control uses beneficial organisms, primarily predators andparasites, to control pest populations below economically important levels.The goal is to establish a balance between the pest population and its parasitesand predators to keep the pest population under control. The complete

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eradication of the pest population is not the goal of biological controlprogrammes, as some pest organisms are required so the parasites andpredators can reproduce.

The greenhouse industry has a well established reputation for usingbiological pest control agents more than any other crop production industry.The reason for this is, in part, due to the ability of greenhouse growers tomanage the environment to favour the biological control agents. Another factoris the relatively limited number of pest species in greenhouses, as well as ageneral tolerance of greenhouse crops to leaf damage caused by these pests.The high value of greenhouse produce is another reason why the use ofbiological controls is economical in greenhouse crops.

The increased use of biological controls has led to a reduction in pesticideapplications as the industry leads in environmentally responsible, intensivecrop production.

Effective biological control of diseases is a more difficult goal and to-date, has rarely been achieved. However, research in developing biologicalcontrols for greenhouse crop diseases is ongoing and it is likely that biologicalcontrol products for greenhouse diseases will be available in Canada in thenear future. The primary strategy of biological control for greenhouse plantdiseases is to introduce fungal parasites to control populations of diseasecausing fungi in the greenhouse environment so that they are unable, or havea reduced ability to infect the plants. Some of the promising biological controlagents, for example, fungi in the Genus Trichoderma are also strong competitorsof the disease causing fungi such as Botrytis cinerea, and can be used to protectwound sites to prevent Botrytis from colonizing the wound site.

Chemical Control

Pesticides are valuable tools when used as a component of an integratedpest management programme. Insecticides should be applied only in supportof biological control programmes, dealing with localized pest outbreaks inthe crop that have escaped the biological control agents. When insecticidesare used, care must be taken to ensure that they are compatible with biologicalcontrol agents, that there will be minimal long term adverse residual effectson biological control programmes. Fungicides are used only when a diseaseproblem is detected.

Pesticides are regarded as the controls of last resort because their misusescreates high-profile environmental and food safety problems. Also, theapplication of some pesticides to a crop can cause stresses that reduce theproductive life of the crop and can make the plants susceptible to other pestsand diseases. If the use of biological control agents is to obtain a balancebetween pests and predators that does not threaten the productive yield ofthe crop, the indiscriminate use of pesticides creates imbalance and uncertaintyin the crop.

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GREENHOUSE ENVIRONMENT TO CONTROL PLANT DISEASES

Numerous plant disease problems can arise in greenhouse situations.These diseases can cause extensive damage of allowed to develop unchecked.Since plant disease are strongly affected by temperature and humidity, thebest way to combat disease is to manipulate the greenhouse environment.Unlike the weather outdoors, we can control the greenhouse environment.

Plant disease control in the greenhouse is generally more effective if thefollowing aspects of the greenhouse environment are managed properly.

HUM IDIT Y

High humidity levels encourage the development of many plant diseases.The relative humidity is usually 25%-70% during the day in greenhouses andgenerally no problem. However, humidity levels are generally 90-100% duringthe night. During periods of rainy weather in winter, the relative humiditymay stay near 100% for a number of days and nights.

Maintain adequate plant spacing. When plants are crowded together,disease development is encouraged by the high humidity in the canopy. Plantshung overhead reduce normal water evaporation and contribute to highhumidity in the crop canopy.

Maintain air circulation during periods of high humidity. Mostgreenhouses are equipped with air circulation, fan-jet or horizontal air flow,systems. These systems should operate continuously when high humidityoccurs in the greenhouse, i.e., every night of the year and during rainy overcastdays. Ventilate the greenhouse to reduce internal relative humidity. Mostwinter evenings are cool enough to raise the humidity to 100% and causeconsiderable condensation in the greenhouse. This condensation can bereduced if the greenhouse is ventilated at dusk each day.

In the late afternoon, turn on the ventilation fans to exhaust the warmmoist air from the greenhouse. The warm moist greenhouse air is replacedwith cool/cold, moist air from outdoors. When this outside air is heated inthe greenhouse it becomes much drier than the previous greenhouse air. Thismanagement practice greatly reduces the relative humidity in the greenhouseand reduces potential disease problems.

Watering

The risk of plant diseases is reduced when the foliage and flowers arekept dry. Most disease organisms need water on the plant surface for normalgrowth. Additionally, splashing water is the primary method of spreadingdisease from plant to plant.

Apply water only to the growing medium surface, when possible, ratherthan “showering” the whole plant. Water early in the day; don’t water after 4p.m., except in the summer.

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Watch plants closely and water judiciously. Remember that withvariations in light levels, temperature and humidity the plants need for waterwill change. Watering practices should be attentive to these changes. Plantsbeneath overhead plants will not dry as quickly as neighbouring plants infull sun.

Water thoroughly and do not water again until the growing medium isdry. Excess water in the growing medium weakens plants and makes themmore susceptible to damping-off and root rotting diseases. Good drainageand the related good aeration of the growing medium is important to preventroot rot problems. Separate weaker plants that do not use water as fast as thestrong plants; They will not be over-watered as easily. Be sure to use a goodgrowing medium. Professionally prepared media are generally the mosteffective and are recommened to commercial growers. When using mineralsoil in a mix, be sure it is adequately sterilized. Never use more than 15%mineral soil in a growing media that will be used in containers less than 4inches tall.

Sanitation

Keep the greenhouse clean and free of plant debris and outside soil.Remove dead leaves, flowers, plant refuse and weeds from the greenhouse.Debris should be gathered regularly and discarded as soon as possible aftercollecting. Immediate removal is important because certain fungal pathogenscan develop and produce spores on the plant debris. Weeds along walkwaysand under benches or even weeds growing just outside the greenhouse canharbour diseases that can be transmitted to greenhouse crops. Insects on weedsmay vector plant virus diseases.

Non-sterile soil from outdoors should not be allowed into the greenhouse.Since many soilborne pathogens can be a serious problem if introduced intothe greenhouse, the best control method is to exclude these pathogens fromthe greenhouse. This can be accomplished by using a soilless medium whichis free of plant pathogens. Surface sterilize all work surfaces and tools regularlyto prevent accidental disease spread. Diseases are often transferred in thegreenhouse through used pots, dirty tools, messy work surfaces, unsweptfloors, etc. Precautions should be taken to prevent this. Surface sterilizationof tools and work surfaces in and around the greenhouse can be accomplishedby washing with a dilute solution of household bleach (9 parts water to 1part beach) or the use of commercial disinfectants such as Greenshield andPhysan.

GREENHOUSE PLANT DISEASES AND THEIR CONTROL

The construction of greenhouses must foster a number of criticalconditions for the optimal health of plants. These include good soil drainage,

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ample sunlight, optimum water and air quality, a good exchange of air withthe use of adequate-sized exhaust fans, continuous fresh air and ample airmovement between the plants. Given the fact that there are a large number ofplants confined in a relatively small space it is common for greenhouse plantsto develop viral diseases, leaf spots, root rot and other diseases very quickly.

DAM PING OFF

Damping off is caused in greenhouses by the pathogens Pythium spp.,Fusarium spp. and Rhizoctonia solani. The pathogens are likely to infest theseedlings of nearly all plants. The disease is most common due to poorlydrained soil in spring; it causes the death of the younger seedlings, and rootrot and stunted growth in older seedlings.

Recommended control includes maintaining moderate soil moisture andensuring proper soil drainage. Seeds should also be germinated at highertemperatures. Chemical fungicides for treating seeds include captan,mefenoxam and metalaxyl.

Black Root Rot

Black root rot is a common disease in floral greenhouse plants and iscaused by the fungus Thielaviopsis basicola. Greenhouse plants mostsusceptible to root rot include geranium, poinsettia, pansy, fuchsia, petuniaand begonia. The fungus is found in soil and can live in the soil withoutcausing disease for many years in thick walled spores. The disease causesstunting, yellowing, wilting and eventual death of the entire plant.Management of the disease includes avoiding plant stress by unnecessarilycool or warm temperatures, ensuring well drained soils and correcting anynutritional imbalance in soil.

Gardening procedures should be sanitary, such as avoiding reusing traysand containers and using good quality potting soils. Chemical controls includesoil drenching with Domain, Banner and Terragard.

Powdery Mildew

Powdery mildew is a frequently occurring disease in greenhouse plantsand is caused by various fungi like Erysiphe, Leveillula, Microsphaera andSpaerotheca. The fungi survive on greenhouse weeds and crops and attackthe healthy, vigorous plants. Powdery mildew appears on floral plants likeAfrican violet, dahlia, begonia and hydrangea. It is also common onherbaceous perennials like aster, phlox and sedum. Herbs like rosemary, sage,mint and greenhouse cucumbers and tomatoes are also susceptible.

Prevention includes proper spacing of plants to reduce humidity levels,keeping humidity level below 93 percent, and keeping greenhouse clean ofweeds. Since powdery mildew only colonizes the upper cell layers, there areno recommended chemical control methods for the disease.

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NUTRIENT AND FEEDING GUIDELINES IN HYDROPONICS

All plants need nutrients to supply them with the elements needed forvital biochemical processes. Nitrogen (N), phosphorus (P) and potassium (K)are the top three generally listed, but there are more than a dozen others.Magnesium (Mg), iron (Fe), calcium (Ca) and several more perform essentialroles in the life of hydroponic plants, just as they do in soil-based gardens.

Nitrogen is used by growing leaves. But despite the fact that the air isabout 79% nitrogen, plants need it in the form of a supplement. The N2molecule in air is very stable and plants don’t break it apart to use singlenitrogen atoms. Phosphorus is essential to root growth. Potassium aids indisease resistance owing to its role in enzyme formation.

The other elements perform a variety of functions. Calcium, for example,is a large component of cell walls and also helps deliver ions to various partsof the plant. Chlorine (Cl) is a component of chlorophyll, an importantparticipant in photosynthesis. Iron is essential to the hemoglobin molecule,which is formed by plants as well as animals, where it helps transport oxygenneeded for cellular respiration.

Pre-made solutions are the easiest to work with to supply all the neededelements. As with any compound, dosage is important. For very young plants,such as small cuttings or those that are just germinating, 1/3 teaspoon ofcalcium nitrate dissolved in a gallon of water is about right, for example. Plantsthat are flowering will require more, about 3/4 teaspoon.

Water and temperature conditions are important factors, as well, whenfeeding your plants. Any solution should be fed at room temperature, whichshould also be the temperature of any water used in hydroponic gardens.

Dry plants should not be fed nutrients. Nitrogen burning is possible.That’s rarely a problem with hydroponics, but one ‘branch’ known asaeroponics, where the plants are grown in air, can suffer that problem.

Allowing any water to stand overnight will help evaporate any excesschlorine from home water sources. Mineralized water is preferable to distilledsince it will contain calcium and other useful elements.

Regulate the pH to keep it as near neutral as possible. As plants take upnutrients they’ll tend to make the water alkaline. Add tiny small amounts ofsulfuric acid to move it back to neutral. Sodium hydroxide will help shiftexcessively acidic water back to a neutral pH. Testing kits are available toaccurately measure the pH of your hydroponic water.

Hydroponically grown plants are more sensitive to nutrient levels andless able to self-regulate than those in soil-based gardens.

In soil, for example, they can take up or shed compounds. Releasingcompounds into the water medium doesn’t move them away from the plant.The hydroponic gardener will need to exercise more care to keep plantshealthy.

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PACIFIC HYDROPONICS

Every day of the year, lettuces from Pacific Hydroponics fly all over theworld, not as packaged exports, but dished up as salads and garnishes inthousands of airline meals. As supplier to Qantas and Ansett airlines, PacificHydroponics has carved out a secure, but highly particular, market nichewhich demands year round supply and an assurance of top quality.

Looking at the streamlined growing operation at Pacific Hydroponicstoday, it’s hard to imagine the problems which plagued its creation. But inreality, a period of “no crops and no income” was a part of the difficult birthof today’s prosperous enterprise. For owner Garry Cahill, unraveling themystery, and sometimes secrecy, surrounding commercial hydroponics hasbeen one of the biggest challenges to turning the operation around.Fortunately, persistence has paid off.

Like many of today’s hydroponic operators, Garry came into the industrylooking for a new business venture.

In 1989 he bought 25 acres of land at Lake Munmorah on the CentralCoast of NSW, did a TAFE course in hydroponics, and engaged a companyin Melbourne to install a hydroponic lettuce growing operation for him. Thecompany offered a complete package which included the installation of areliable growing system and an established marketing arrangement with alarge supermarket chain. It sounded ideal, but unfortunately the manypromises proved hollow. Having paid for the system, Garry then found thatthe manufacturers were unable to get it going. At the same time, the so called‘established market” failed to materialise. There followed a year with nothinggrowing in the system and a high interest debt to service.

“In other words,” Garry says, “we couldn’t get the thing going and wehad no market.”

Leaving his wife and four children in Sydney, Garry moved up to thefarm and spent two difficult years turning the system around and finding amarket for his product. His task was assisted by the support of farm manager,Phil Ritchie, as well as significant contributions from Geoff Creswell of theNSW Department of Agriculture, among others. Between them, they wereable to overcome the inherent problems in the system and start producinggood lettuce crops. Simultaneously, Garry tackled the question of marketing,convinced of the need for a planned approach.

“I knew we had to get out and market it professionally,” he says.And market it professionally he did, establishing not only an airline

market (Qantas also supply meals to all the other airlines), but also a localrun to quality restaurants on the central coast.

Today, Pacific Hydroponics harvest lettuce every day of the year,delivering them direct to the airlines in Sydney each morning. The farm’sdistance from Sydney (about 110km) makes this system practical, but Garrycomments that he would not go any further away from a capital city market.

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The farm also benefits from a benign coastal climate, where frosts are rare,and an immediate environment which is conducive to good production. Thereis good natural windbreak protection, to the west, north and east, and duringsummer the prevailing north easterly winds pass over the large water area ofnear by Lake Munmorah, creating a cooling effect. Garry is convinced that afarm’s surrounding environment can play a significant role.

“We didn’t know that when we built it,” he says, “it’s a bonus. But nowI realise how important environment is to the operation.”

At any one time, upwards of 90,000 lettuces are growing in the system atPacific Hydroponics. Varieties include mignonette, butter, red and green coral,red and green oak, mini cos, green cos and radicchio. Plants are harvestedevery morning, between 6 and 9am, and around 1000 boxes a week are soldfresh. “Boxes’ is actually a misnomer, since Garry has managed to avoid theexpense of buying and printing cardboard boxes for packing and transport.Instead, lettuce are packed in red plastic crates which are subsequentlyreturned to him by Qantas for re-use.

Despite the problems Garry experienced in getting to production stage,the system as it stands now is impres sive in its efficiency and ease of operation.It is based on a highly interesting concept, using plastic covered, 14 metrelong tables. Seedlings are planted through slits in the plastic cover, whichalso supports them, and fed by the Nutrient Film Technique (M). The plasticis reflective white on the outside and black underneath, to create a darkenedchamber which is favourable to the roots, but not to the growth of algae. Thereare 410 lettuces in each table, and harvesting involves simply pulling out theplants, roots and all. Hosing down the tops of the tables after harvest is thesimple clean up procedure.

The tables are arranged in six sheds’, roofiess structures with sides Ofmesh fabric that can be rolled up or down as required. Each table, however,has its own plastic cover which can be rolled over to create a mini green house,or to provide hail protection. The sheds measure 110m by 17m and eachcontains 40 tables. The tables themselves are custom made from fibreglasswith slight corrugations, creating small channels down which the nutrientsolution flows. The plant roots are free to wander into the nearest channel,and there is no ponding effect. The nutrient is recirculated and the tables havea slope of 300mm over their 14 metre length which, Garry comments, is higherthan the norm.

“It’s far greater than farms I’ve seen, “ he says, “ but I’ve found out thatthe quicker you turn the water around the better.”

Indeed speed is one of the keys to the way Garry runs his operation.“We’re in the game of fast growing produce – that’s what it is,” Garry

explains. “The faster you grow it, the more money you make because the morecrops you get per year. So it is all speed and in fact they’re easier to managethe faster they’re grown.”

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The farm uses town water, which is stored in two 45,000 litre (10,000 galIon) tanks. It requires pH stabilization before using, since according to Garry,the pH can vary widely – anywhere from pH 5 to 9. The nutrient solution ismixed in smaller 8000 litre (1758 gallons) tanks, which can be heated if -necessary, and a standard lettuce mix nutrient is used, manufactured bySydney company Simple Grow. Brian Heame, Simple Grow’s owner andmanager, has been involved with the operation from the start, helping Garryto get his nutrient formulation right.

The growing system is a re-circulating one with a flow rate of 1.25 litresper minute and the nutrient solution is topped up when necessary, notablyafter periods of rain. Each of the six sheds operates quite independently ofthe others, to minimize production loss should a problem arise. Duringsummer, the solution is dumped about every two months, but dumping israrely found to be necessary in winter. Interestingly, all monitoring of ECand pH is done manually, using standard meters, and automation is kept toa minimum. This, Garry believes, ensures a more hands-on approach to theoperation.

“I really don’t think you can fully automate a commerdal set-up. Onceyou do that, you don’t have people here and if you don’t have people here,you’ll never find out if some thing’s wrong. It can’t be a 9 to 5 thing,” he says.With select clients such as airline companies, high priority has to be given toproducing crops that are dean and free from insects. Pests and diseases areminimised by keeping the area around the farm clean, and apart from an attackof Rutherglen Bug two years ago, there have been few problems. Garrysubscribes to a preventative approach to such problems.

“We know now, during certain times of the year, you’ve got to watchout for certain things,” he says. “So before it happens we take precautions”.

A precautionary approach also extends to the prevention of pythium,which, during the last summer season, didn’t manifest itself. Farm managerPhil Ritchie believes management practices have an important influence onpythium prevention and that by not stressing the plants, you can avoid theproblem appearing. To this end, he shaded the plants lightly last summer, byletting the plastic covers hang down a little over the tables and believes thishad a positive effect. As a result, he is now considering trialing shadecloth asa roof over some of the sheds.

Another pythium preventative measure at Pacific Hydroponics is theaddition of Agral, a non-ionic wetting agent, to the nutrient tank. Agral wasoriginally developed in the UK to con trol Big Vein virus in lettuce and has aspecial registration for use in hydroponic nutrient solutions. It works bydissolving away the tail of the pythium spores, where the virus is alive, andalso dissolving the vessicle from which the pythium spores are released. Agralwas recommended to Phil by the NSW Department of Agriculture and is nowrun in the nutrient all summer. No seedling propagation is done on the farm.

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All the lettuce seedlings come from Leppington Seedlings, professionalgrowers south west of Sydney, as part of a standard weekly order. Accordingto farm manager Phil Ritchie, the relationship with a seedling grower is animportant one. “I’ve explained to him what we’ve been after in a crop,” Philsays. “I can say to him that this type of lettuce is having certain difficultiesand then we get a replacement. And he has been able to supply us with lettucesthat suit this district.”

When Garry Cahill bought the farm, it was being used as a small flowergrowing enterprise and with it came plantings of Kangaroo Paw(Anigozanthos sp.) and Proteas. These he has maintained as a sidelineenterprise, selling the cut flowers on the export market. While Garry admitsthat he would not have chosen to plant them, he believes in taking advantageof all possible sources of income, particularly since he has a permanent labourforce of six people. The flowers are drip irrigated, and the spent nutrient fromthe lettuce system is also run onto them, rather than dumping it into apaddock. “If we were going to build another of these, we’d make some changesand do it a lot cheaper.”

While Garry would consider diversifying into other crops, he maintainsthat it would have to be under certain terms. “We’d want a market first andwe’d say to the client, OK we’ll grow that crop for you and this is how we cangrow.”

The best market, he adds, is a reliable year-round one, rather than one Ithat fluctuates seasonally. With the benefit of hindsight, Garry can also seeways of improving on his current growing system. “If we were going to buildanother of these, we’d make some changes and we’d do it a lot cheaper,” hesays. “So consequently, we’d get it running as well as this is running, but thecapital cost would be lower, so we’d be more profitable.”

Location is something Garry would not change significantly. He believesthere are advantages in being close to the coast, because of the mild climaticinfluences and consequently would not consider going any further inland,where the frost belt begins.

Out of his adversity, Garry has managed not only to survive, but to turnthe enterprise around to profit. His problems, which started with an ineptcompany who were unable to get their own system design up and running,were not alleviated by what he felt at the time was a degree of secrecy withinthe industry. While things are improving now, Garry feels that a moreprofessional attitude to hydroponic farming operations needs to be adopted,whereby they are run along strict business guidelines.

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8

Biotechnology andGenetic Transformation

APPROACHES FOR WHEAT TRANSFORMATION

The last two decades have witnessed the widespread use of variedapproaches for introduction of exogenous DNA into wheat. Wheatimprovement by genetic engineering requires the delivery, integration andexpression of defined foreign genes into suitable regenerable explants. Initialattempts at introducing transgenes into wheat employed protoplasts asexplants due to the absence of cell walls. The introduction of marker geneconstructs into protoplasts provided valuable information regarding theexpression pattern and tissue specificity of various promoters and regulatoryelements in the transformed tissue. However, the difficulties associated withplantlet regeneration from protoplasts have compelled researchers to look foralternate target cells/tissues with better regeneration capabilities.

Therefore, attention shifted to embryogenic suspension cells andembryogenic callus cultures derived from scutellar tissue of mature andimmature embryos. A detailed and in depth account of the in vitro cultureresponse of wheat from different explants is available in numerous reviewsand so has not been d ealt w ith in detail here (Bajaj, 1990; Maheshwari et al.1995; Vasil and Vasil, 1999). In recent years, with the development of suitableregeneration protocols, microspore embryos and immature inflorescences areemerging as suitable target tissues for genetic transformation experiments.Till date, the biolistics approach has been most successful in delivering foreigngenes into wheat.

SCORABLE M ARKERS

Initial steps for genetic transformation involves delivery of a gene cassetteinto recipient cells followed by analysis of the expression of delivered gene.The results of the above events can be detected by assaying the expression of

a reporter gene introduced into plant cell cultures or intact tissues. The reportergenes produce a visible effect, directly or indirectly, due to their activity inthe transformed cells. Analysis of reporter gene expression does not requirethe integration of the transgene into the host genome and is commonly usedto test promoter and gene functions. Initial studies on the introduction offoreign DNA to wheat have relied on the use of the E. coli gene for the enzymechloramphenicol acetyl transferase (cat).

However, the detection of transgene by enzymatic and immunochemicalmethods for measurement of CAT activity and amount of CAT protein,respectively, was tedious; and moreover, presence of inhibitors of CAT activityand endogenous CAT activity hampered its use as a reporter gene in wheat.With the availability of a protocol for the rapid assay of-glucuronidase (gus)gene from E. coli, this gene has emerged as the most widely used scorablemarker in wheat transformation. GUS enzyme hydrolyzes b glucuronidecompounds and gives reaction products that can be quantifiedspectrophotometric or spectrofluorometrically. The gus reporter gene systemis extremely useful for optimisation of parameters for genetic transformation,due to the availability of a simple histochemical detection procedure.

One of the major limitations of gus reporter gene system, however, is thedestructive nature of its assay. Thus, to study the fate of introduced transgenesin living cells, vital reporter genes encoding for anthocyanin biosynthesis,green fluorescent protein, and firefly luciferase have been used successfully.The R genes of Zea mays which stimulate endogenous anthocyaninaccumulation in the vacuoles of plant tissues is useful as a scorable marker inmature and differentiated cells. Due to high sensitivity and ease ofvisualization, the use of R genes in wheat transformation have been reportedby different groups. A synthetic, spectrally modified, version of greenfluorescent protein (gfp) from the jellyfish, Aqueorea victoria has also beenused as a vital marker in wheat transformation. Recently Jordan (2000)reported the use of a modified gfp as a visible marker for the detection oftransgenic wheat plants on the basis of gfp expression alone.

Amongst the other scorable markers, luciferase gene from the fire fly,Photinus pyranus, has been successfully used in stable transformation ofwheat. Luciferase and modified versions of gfp permit non-destructiveanalysis of transgene activity, ease to follow the fate of introduced transgenesin individual tissue samples, facilitate rapid assessment and comparison ofdifferent transformation procedures, and provide valuable insight into theconditions influencing the efficiency of DNA integration and stable expression.

SELECTABLE M ARKERS

The varied frequency of DNA delivery in cells of different explants hasnecessitated the development of methods for efficient selection of cells thatcarry and express the introduced gene sequences. The selection regimes for

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transformed cells are based on the expression of a gene termed as the selectablemarker producing an enzyme that confers resistance to a cytotoxic substanceoften an antibiotic or a herbicide. The most commonly used selection markerin wheat transformation is the bar gene (bialaphos resistance gene) encodingfor phosphinothricin acetyl transferase (pat). Both bar and pat genes isolatedfrom different Streptomyces species, encode for phosphinothricin acetyltransferase. Amongst the antibiotic resistance markers, the bacterial neomycinphosphotransferase II (nptII) gene providing resistance to aminoglycosideantibiotics is commonly used in wheat transformation.

Herbicide resistance genes offer an alternative to antibiotic-resistantmarkers. Genes coding for 5-enolpyruvyl-shikimate-3-phosphate synthase(EPSPS), a critical enzyme for aromatic amino acid biosynthesis, andphosphinothricin acetyl transferase (PAT) provide tolerance to glyphosate andglufosinate ammonium herbicides, respectively.

The enolpyruvylshikimate-phosphate synthase (CP4) gene isolated fromAgrobacterium strain CP4 and the glyphosate oxidoreductase (GOX) alsoprovide tolerance to glyphosate by degrading glyphosate into aminomethylphosphoric acid, and have been added to the list of selectable markersavailable for wheat. The hygromycin phosphotransferase (hpt) gene, widelyused in rice transformation, has also been reported as an efficient selectablemarker for achieving stable genetic transformation in wheat. The efficiencyof hpt as a selectable marker is comparable to the other widely used selectablemarkers encoding resistance for antibiotics and herbicides.

TRANSFORMATION OF WHEAT PROTOPLASTS

Protoplasts are regarded as competent targets for gene transfer due tothe absence of cell walls. During the early 80's protoplasts were the chosenexplants for wheat transformation and their use as explants for direct genetransfer was facilitated by the development of suitable techniques for isolationof protoplasts from different tissues. However, due to limited success withculturing protoplasts from organized tissues, cell suspension/calli derivedprotoplasts have been used in wheat. To overcome the hindrance exercisedby the plasma membrane, protoplasts are subjected to either chemicaltreatment [polyethylene glycol (PEG)] or subjected to physical forces likeelectric pulses (electroporation), either alone or in combination with PEG, forthe introduction of foreign DNA. For wheat, isolation of protoplasts fromembryogenic callus cultures have been the most popular tissues of choice.

The first report of direct gene transfer in wheat protoplasts was by Lorzet al. (1985) from cultured cells Triticum monococcum by PEG-mediateduptake. The transformed cells were selected on a kanamycin-containingmedium and identified by detection of nptII activity. Subsequently, Werr andLorz (1986) reported the expression of the introduced genes but the majority

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of foreign DNA introduced was found to remain extra-chromosomal andtransient gene activity was found to be dependent on the presence of thepromoter fragment of the Shrunken gene of maize. This observation alsoindicated for the first time, the efficacy of a cereal promoter.

Lee et al. (1989) introduced chimeric gene constructs having the catreporter gene under the control of CaMV 35S promoter in aleurone protoplastsisolated from developing caryopses. The resultant gene activity detected inprotoplasts from developing aleurone layers raised the possibility ofinvestigating tissue-and development-specific control of genes for cereal seedproteins. By and large, the physiological age of the developing grain and thephysiological state of the isolated protoplasts affected the level of transientgene expression. The significant factors affecting the transient activity of thereporter gene constructs were the presence of the divalent cation Mg2+ withPEG, time lapse after DNA uptake, pre-culture medium and the regulatoryelements of the vector construct. Nonetheless, these reports yielded lowtransformation efficiencies in the order of 0.005% and the low regeneratingability of the explant hampered work in this area.

Introduction of the E. coli plasmid, pCGN1055, containing the hpt geneinto wheat protoplasts, by cationic liposome-mediated transformation resultedin the production of transgenic albino plantlets. The presence and activity ofthe transgenes was confirmed by assay of enzymatic activity and southernhybridisation and the transformation efficiencies reported were in the orderof 6%.

ELECT ROPORAT ION OF PROT OPLAST S

Electroporation is a technique that utilizes a high intensity electric pulseto create transient pores in the cell membrane thereby facilitating the uptakeof macromolecules like DNA. Ou-Lee et al. (1986) reported the expression ofthe bacterial chloramphenicol acetyl transferase (cat) gene in three importantgramineous plants, i.e. rice, sorghum and wheat. The survival percentage ofprotoplasts after electroporation depended largely on the tissue of origin. Thetransient activity of cat gene was equal when it was fused to either CaMV 35Spromoter or the copia long terminal repeat promoter of drosophila.

This study also demonstrated, interestingly, that the Drosophila promoter(copia) was capable of directing the synthesis of functional CAT enzyme inplant cells as efficiently as one of the strongest constitutive promoters knownto function in plant cells. Besides the promoter influencing the foreign geneexpression levels, presence of an intron was also found to be helpful as shownby Oard et al. (1989) with a maize alcohol dehydrogenase (Adh) intron inchimeric constructs. This was evidenced by ribonuclease protection assaysthat demonstrated increased mRNA levels for the intron-containing constructsversus those without the intron. This report highlighted the use of regulatoryelements for enhancing the transgene expression.

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Zaghmout and Trolinder (1993) optimised other conditions for asignificant increase in gus activity which included the protoplast source, thepromoter, the PEG concentration, as well as, electroporation parametersespecially the electric field strength, preincubation with the plasmid, andrecovery period on ice after the electric pulse.

Stable transformation by electroporation was reported by the use of theplasmid pBARGUS into protoplasts isolated from cell suspension initiatedfrom an anther-derived callus. This was the first report of stable transformationof wheat protoplasts and also confirmed by Southern hybridization. Stabletransformation of protoplasts was also reported by He et al. (1994) byelectroporation employing the bar gene as the selectable marker, which alsofacilitated the selection of phosphinothricin-resistant colonies. Southernanalysis and PAT assay revealed the presence of the bar gene and its product,respectively, in the regenerated colonies and plants. He et al. (1994) obtainedtransient expression of gus gene at a frequency of 1x 10-5 which was more ascompared to that observed by Lee et al. (1989). Electroporation of protoplaststhus considerably improved the gene transfer efficiency as compared to thatachieved with PEG mediated approach.

TRANSGENE SILENCING IN WHEAT

Success at developing improved wheat cultivars through geneticengineering depends on stable and predictable expression of the inserted gene.However, gene silencing is a common phenomenon in the production oftransgenic plants and needs to be effectively controlled for the desired results.This is all the more important because gene silencing is an importantphenomenon involved during natural plant defence. Gene silencing is acomplicated phenomenon as it includes both transcriptional gene inactivationand post transcriptional gene inactivation. The complex and the large genomesize of wheat is expected to be prone to silencing by introduced transgenes.

With the development of transformation methodologies for wheat, moreinformation has started to accumulate regarding the inheritance andexpression of the introduced transgenes. Information regarding the long termstability of transgenes is thus of immense significance for the use of geneticmanipulation as a tool in wheat crop improvement and is necessary for theintroduction and subsequent expression of desirable agronomic trait.

DNA methylation plays a significant role in establishing and maintainingan inactive state of the gene by rendering the chromatin structure inaccessibleto the transcription machinery. Methylation of DNA is expected to result inreduced gene expression. Muller et al. (1996) studied the variability oftransgene expression in clonal cell lines of wheat and found a negativecorrelation between the degree of methylation and marker gene expression.PEG-mediated approach was used to transform protoplasts isolated from

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suspension cultures of T-aestivum. The integration and expression of theselectable marker gene nptII fused with different promoters, namely, alcoholdehydrogenase, shrunken from maize, and actin1 from rice, was confirmedby Southern analysis and enzyme activity test.

Transgenic cell lines under selection were 'protoplasted' and clonal calluslines were cultivated from genetically identical single cells without selectionpressure. A reduction/loss of marker gene expression was observed due to areduction in the nptII transcript level and was seen to be associated withhypermethylation of the integrated DNA. The silencing effect was reversedby a 4-week culture phase on media supplemented with demethylation agent,5-azacytidine, thus confirming the role of methylation in transgene silencingDemeke et al. (1999) studied the inheritance and stability of an Act1D-uidA:nptII expression cassette in T4 and T5 transgenic plants. Based on thehistochemical localization of GUS activity, this study demonstrated the lackof any cytoplasmic effect on the inheritance of the transgene. The transgenesmaintained a multiple integration pattern similar to that observed in the T1generation. The transgenic plants which produced low gus and nptII activityin seeds had an intact expression cassette. Southern blot analysis of genomicDNA performed with the methylation sensitive enzyme, HpaII, showed thetransgene in GUS negative plants to be highly methylated relative to thetransgene in GUS positive plants. Some of the studies on gene silencing alsoindicate that multiple integration pattern and copy number is also associatedwith DNA methylation.

Cannell and coworkers studied the inheritance of gus and bar markergenes over three generations. The integration, inheritance and expression ofthese marker genes in the population studied were stable and predictable witha few exceptions. The inheritance of integration patterns was stable, andtransmission/ inheritance of the transgenes followed Mendelian ratios in amajority of lines. From this study, the authors claim that the transformationprocedure, transgene integration, and marker gene expression had little effecton the transmission of transgenes to the subsequent progenies. However, overthe three generations studied, a uniform, 'progressive' transgene silencing,specific to the gus gene, was observed. This gradual loss of the gus geneexpression was not accompanied by a reduction in bar expression. Since inthis case, the gus and the bar genes are located on the same plasmid,observations indicate a post-transcriptional gene silencing.

The silencing of HMW glutenins was also observed in transgenic wheatexpressing extra HMW subunits. Characterization of six independent eventsinvolving the transformation of wheat HMW-GS genes, 1Ax1 and 1Dx5, in acultivar expressing five subunits by particle bombardment resulted in partialor complete gene silencing. Silencing of all the HMW glutenin subunits wasobserved in two different events of transgenic wheat expressing the 1Ax1subunit transgene and overexpressing the 1Dx5 subunit. Control of gene

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silencing thus remains a challenge for the immediate future. Meyer, (1995)advocated that the problem of gene silencing in wheat can be minimized byoptimising methods for simple integration patterns, use of promoters and genesequences isolated from cereals, use of matrix associated regions (MARs) orscaffold attachment regions (SARs), which insulate transgenes fromsurrounding chromatin, might help in reducing the gene silencing problem.

TRANSPOSON TAGGING IN WHEAT

Transposon mutagenesis has been widely exploited in various organismsto isolate genes that encode unidentified products. The maize activator (Ac)and Dissociation (Ds) elements are the best-studied transposable elements inheterologous host plants. Initial studies have reported the introduction andactivation of the Ac/Ds elements into cultured wheat cells by using a wheatdwarf virus by particle bombardment. Takumi and coworkers has developeda transposon tagging system in wheat by introducing the Ac transposase geneunder the CaMV 35S promoter into cultured wheat embryos by particlebombardment. For the development of a transposon tagging system, embryosisolated from a stable Ac line were bombarded with a plasmid containing themaize dissociator (Ds) element located between the rice Act1 and the gusgenes. The transient expression of the gus gene was observed after the excisionof Ds elements. Southern and northern analysis of the T0 and T1 plants hasexhibited the stable expression and inheritance of the Ac transposase gene.These results have demonstrated the precise processing of the maize Actransposase gene and the synthesis of the transposase protein in transgenicAc lines. The Ac transposase gene causes the transactivation and excision ofthe Ds in the Ac transgenic lines transformed with the maize Ds elements.Thus in near future we can expect traits of commercial importance to be taggedfor a wider utility in important crop plants.

M ARKER ASSIST ED SELECT ION IN WHEAT BREEDING

Conventionally, plant breeding depends upon morphological/phenotypicmarkers for the identification of agronomic traits. With the development ofmethodologies for the analysis of plant gene structure and function, molecularmarkers have been utilized for identification of traits. Molecular markers actas DNA signposts to locate the gene(s) for a trait of interest on a plantchromosome, and are widely used to study the organization of plant genomesand for the construction of genetic linkage maps. Molecular markers areindependent from environmental variables and can be scored at any stage inthe life cycle of a plant.

Over the last several years, there has thus been marked increase in theapplication of molecular markers in the breeding programmes of various cropplants. Molecular markers not only facilitate the development of new varieties

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by reducing the time required for the detection of specific traits in progenyplants, but also fasten the identification of resistance genes and theircorresponding molecular markers, thus accelerating efficient breeding ofresistance traits into wheat cultivars by marker assisted selection (MAS). Theavailability of back-cross derived near isogenic lines (NILs) have alsofacilitated the analysis of various lines by using different marker systems. Theintroduction of alien genetic variation into wheat is a valuable and proventechnique for wheat improvement. Wild relatives of wheat provide anenormous resource of new genes for wheat improvement, particularly diseaseand stress tolerance. These genes can be of use if recombined into lines adaptedto the conditions of a particular region.

Initial studies on the application of molecular markers in wheat reliedon the hybridisation based restriction fragment length polymorphism (RFLP)system-RFLP maps provided a more direct method for selecting desirablegenes via their linkage to easily detectable markers thereby expediting themovement of desirable genes among varieties (Tanksley et al. 1989). The factorsthat had been instrumental for the use of RFLP in wheat was the limitednumber of polymorphisms observed among wheat lines and more significantlythe availability of aneuploid stocks for the determination of chromosomallocation of genes.

In wheat, RFLP's have been used to map seed storage protein loci, lociassociated with flour colour, cultivar identification, vernalization (Vrn1) andfrost resistance gene on chromosome 5A, intrachromosomal mapping of genesfor dwarfing (Rht12) and vernalization (Vrn1), resistance to preharvestsprouting, quantitative trait loci (QTL's) controlling tissue culture response(tcr), nematode resistance, milling yield, resistance to chlorosis induction byPyrenophora tritici-repentis.

RFLP markers are also useful in selection programmes for resistanceagainst pests and pathogens, which is otherwise labor and time consuming,and to detect homozygous individuals and have been used for resistance tobarley yellow dwarf virus, resistance to wheat spindle streak mosaic virus,resistance against powdery mildew, resistance against leaf rust, resistanceagainst cereal cyst nematode. The use of RFLP analysis in wheat has, however,been of limited use in the intervarietal analysis due to low level ofpolymorphism and the high cost for screening in breeding situations.

With the development of polymerase chain reaction (PCR) methodologies,Random Amplified Polymorphic DNA (RAPD) emerged as a convenient andeffective technique for tracing alien chromosome segments in translocationlines. RAPD markers provide a useful alternative to RFLP analysis forscreening markers linked to a single trait within near isogenic lines and bulkedsegregants. He et al. (1992) reported the development of a DNA polymorphismdetection method by combining RAPD with DGGE (denaturing gradient gelelecrophoresis) for pedegree analysis and fingerprinting of wheat cultivars.

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RAPD markers can be converted to more user-friendly Sequence CharacterizedAmplified Region (SCAR) markers, that display a less complex bandingpattern.

SCAR markers linked to resistance genes against fungal pathogens havebeen characterized in combination with RAPD and RFLP. In recent years,RAPD and other PCR based markers like Sequence Characterized AmplifiedRegions (SCAR), Sequence Tagged Sites (STS) and Differential Display ReverseTranscriptase PCR (DDRT-PCR) are increasingly being used for identificationof desirable traits in wheat and related genera. These markers have been usedin particular for disease resistance against viral and fungal pathogens andalso for insect and nematode pests and have the potential of pyramiding ofresistance genes for effective breeding programmes.

PCR based markers have been extensively characterized for genes ofresistance against common bunt, Tilletia tritici,; powdery mildew, Erysiphegraminis, leaf rust, Puccinia recondita, resistance against Hessian fly,Mayetiola destructor and Russian wheat aphid, Diuraphis noxia.

Simple sequence repeats or microsatellites are more promising molecularmarkers for the identification and differentiation of genotypes within a species.The high level of polymorphism and easy handling has made microsatellitesextremely useful for different applications in wheat breeding.

Microsatellites have also been used to identify resistance genes like Pm6from Triticum timopheevii and Yr15 from breadwheat. In near future,molecular markers can provide simultaneous and sequential selection ofagronomically important genes in wheat breeding programmes allowingscreening for several agronomically important traits at early stages andeffectively replace time consuming bioassays in early generation screens.

PRODUCT DEVELOPMENT AND TECHNOLOGY TRANSFER

It is perhaps important to recall here that biotechnology may beconsidered as an enabling technique (for example, the use of genetic markersin plant breeding) or as being incorporated in a biotechnology product (forexample, a new, disease-resistant plant variety). Whether a biotechnologyproduct is imported or generated by local research effort, adaptation to localagroecological and production conditions and/or product development arenecessary.

Moving from the purely research phase, development (small-to large-scalefield testing, setting up of a pilot plant, seed multiplication, etc.) appears tobe a major constraint. This is in part because development is not alwaysincluded or is often underestimated, or both, in research budgets.

Similarly, little attention has been paid to technology diffusion ormarketing mechanisms or to the demand side aspects of biotechnology. Onthe one hand, without the incentive of strong market potential, private firms

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are unwilling to undertake the risks of production and marketing.On the other hand, public research institutions are shown to beill-equipped technically and do not generally have the financial resources toscale up from small-to large-scale testing and from pilot plant to large-scaleproduction.

Few biotechnology products are yet on the market in the countries wehave studied. In all countries, disease-free planting material produced bytissue culture and micropropagation (for vegetatively propagated crops andflowers in particular) is already available, usually marketed by commercialfirms. In some cases, demand exceeds supply.

The case of biopesticides presents a contrasting picture. A number ofgovernments are involved in trying to promote the diffusion of biopesticidesto reduce dependence on chemical products, and research on biopesticides isbeing supported in public institutions. Unfortunately, the inadequate technicalcapacity of public institutions to produce biopesticides efficiently and to ensureconsistent quality results in a lack of acceptability and effective demand onthe part of farmers, and lack of interest on the part of the private sector inproduction and commercialization.

The problem is further compounded in those situations where nationalextension services are shrinking as a result of reduced public expenditure orare being privatized. If, for reasons of environmental protection, governmentsare committed to the increased use of biopesticides, they will need to continueinvesting in or subsidizing production and technology transfer to farmers orprovide incentives to the private sector to become involved.

A number of innovative examples of efforts to stimulate involvement ofthe private sector and to facilitate the creation of markets for biotechnologyproducts have emerged from our research. These include tax exemptions andaccess to credit for local start-up firms, government procurement as a meansof assuring an initial market for local start-up firms, the provision of large-scale testing facilities or quality control services by government agencies, andefforts to seek commercial partners as soon as research results appearpromising.

A number of collaborative arrangements between public researchinstitutions in developing countries and commercial companies (both domesticand foreign) are receiving donor support. In most cases, these concernbiotechnology research and development where some elements of thetechnology, or specific research techniques, are "transferred" from developed-country laboratories to the developing-country institutions.

Sometimes, they concern research techniques, genes, or products overwhich private companies hold intellectual property rights. To date, a numberof such techniques, or elements of technology, have been "donated" todeveloping countries by multinational corporations through nonexclusive,royalty-free licences.

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POLICY AND INST IT UT IONAL ISSUES

In terms of policy implications, the research highlights, first and foremost,the lack of integration of biotechnology research in the broader nationalinstitutional and policy framework. Clearly, biotechnology policies andstrategies should be country-specific, designed as a function of the particularconditions prevailing within a country, its particular scientific andtechnological capabilities, institutions, and the range of policies affectingagriculture. Similarly, it highlights the lack of effective linkages and interactionamong the different stakeholders in biotechnology research, development, anddiffusion.

One vexing question in formulating sensible policies for biotechnologyin agriculture is that of the economic cost-benefit of biotechnologies and, moreparticularly, their economic advantage over other methods of plant protectionand production. The case of biopesticides suggests that relatively low, short-to medium-term economic costs, which would need to be met by governmentsor through development assistance, may lead to major long-term, social andenvironmental benefits. It is important that methodologies be developed forassessing the comparative cost-benefits of new biotechnology products.

Other policy and institutional issues of particular importance inbiotechnology are those of biosafety and intellectual property rights. Althougha number of countries are currently establishing biosafety guidelines, in mostcountries national procedures are not yet in place. Our research suggests thatthe lack of biosafety procedures could inhibit the transfer of biotechnology.Similarly, it is important that countries clarify their intentions with respect tointellectual property rights in plant biotechnology.

Issues of Relevance for This Workshop

Research on biotechnology at the Development Centre has beenundertaken from the broad perspective of technological change and innovationin agriculture. Essentially, the aim has been to examine what is at present inplace to stimulate or impede the development and diffusion of new technologywithin a national context, and to determine what institutional arrangementsand policies might improve the situation. This approach has the merit ofpinpointing current impediments and proposing measures that might reduceidentified constraints to technological change in the future. It also has theshortcoming of generating little quantitative, particularly economic, data.

This approach has been seen as fruitful given the formidable difficultiesof both ex-ante and ex-post assessment of the impact of biotechnology. Onedifficulty stems from problems in arriving at a satisfactory definition ofbiotechnology. In one of its early publications, the OECD, which definesbiotechnology as "the application of biological organisms, systems andprocesses based on scientific and engineering principles, to the production ofgoods and services," listed 11 definitions.

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What is important to keep in mind is that biotechnology encompasses amix in the form of an enabling tool (e.g., a genetic marker in plant breeding),a process (e.g., fermentation), or a product (e.g., a transgenic seed). The impactof each of those outputs may depend to a very large extent on interrelated,underpinning technologies and capacities. For example, a new crop varietymay be the product of a combination of biotechnology, plant breeding, andseed production techniques and the contribution of the biotechnology inputto the performance of that variety in the field may be extremely difficult topinpoint. Similarly, the impact of biopesticides will depend both on the abilityto produce products of consistent, high quality and on skill and timeliness intheir application. At the least, ex-ante assessment of a given biotechnologytechnique or product would require the following:

• Definition and description of the technology;• Specification of alternative technology options for achieving the

same objectives;• Management strategies required for the technology;• Assessment of the direct effect of the technology on yields,

production costs, productivity, input demand, and a range ofenvironmental, legal, safety, and other considerations; and

• Assessment of the indirect effects: identification of the losers/gainers,assessment of risks and uncertainties associated with the technology,and, finally, long-term impact.

In many situations, the availability of data on some of these variables, ateither the macro-or microlevel, is highly problematic. Quantitative ex-postassessment of agricultural biotechnology products is also, to date, very limited.The first wave of genetically engineered products is only now beginning toreach the market, and very little economic analysis of the earlier products oftissue culture and micropropagation has been published.

The socioeconomic assessments of bovine somatotropin (bST) now underway in the U.S. and Canada may have little relevance to the productionsystems, management, and climatic conditions of many developing countries.A large part of the work on biotechnology assessment related to developingcountries has focused on supply-side problems and constraints, with littleassessment of potential demand.

It is important that the methods used for setting national priorities inbiotechnology should seek a better balance between demand and supply-sideaspects. Effective demand will, to a large extent, determine the respective rolesof the public and private sectors in the development and dissemination ofnew biotechnology products. In the case of those technologies with a strongpublic-good aspect (such as biopesticides), but with weak short-term marketpotential, the initial costs will need to be borne by governments.

Clearly, for countries to be able to formulate sensible nationalbiotechnology strategies both quantitative and qualitative impact assessment

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are required and a number of different approaches and methods of assessmentwill be outlined by other participants. One approach that appears promisingand that the OECD has begun to explore is that of a national system ofinnovation (NSI).

Although there is as yet no consensus on a precise definition, a NationalSystem of Innovation (NSI) can be defined as the network of agents and setof policies and institutions that affect the introduction of technology that isnew to the economy and that determine the rate and direction of technologicallearning and change. It is not yet clear how relevant this concept is in adeveloping-country context.

GENE TRANSFER METHODS IN PLANTS

To achieve genetic transformation in plants, we need the construction ofa vector (genetic vehicle) which transports the genes of interest, flanked bythe necessary controlling sequences i.e. promoter and terminator, and deliverthe genes into the host plant. The two kinds of gene transfer methods in plantsare:

VECT OR-M EDIAT ED OR INDIRECT GENE T RANSFER

Among the various vectors used in plant transformation, the Ti plasmidof Agrobacterium tumefaciens has been widely used. This bacteria is knownas “natural genetic engineer” of plants because these bacteria have naturalability to transfer T-DNA of their plasmids into plant genome upon infectionof cells at the wound site and cause an unorganized growth of a cell massknown as crown gall.

Ti plasmids are used as gene vectors for delivering useful foreign genesinto target plant cells and tissues. The foreign gene is cloned in the T-DNAregion of Ti-plasmid in place of unwanted sequences.

To transform plants, leaf discs (in case of dicots) or embryogenic callus(in case of monocots) are collected and infected with Agrobacterium carryingrecombinant disarmed Ti-plasmid vector.

The infected tissue is then cultured (co-cultivation) on shoot regenerationmedium for 2-3 days during which time the transfer of T-DNA along withforeign genes takes place. After this, the transformed tissues (leaf discs/calli)are transferred onto selection cum plant regeneration medium supplementedwith usually lethal concentration of an antibiotic to selectively eliminate non-transformed tissues.

After 3-5 weeks, the regenerated shoots are transferred to root-inducingmedium, and after another 3-4 weeks, complete plants are transferred to soilfollowing the hardening (acclimatization) of regenerated plants. The moleculartechniques like PCR and southern hybridization are used to detect the presenceof foreign genes in the transgenic plants.

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VECT ORLESS OR DIRECT GENE T RANSFER

In the direct gene transfer methods, the foreign gene of interest isdelivered into the host plant cell without the help of a vector. The methodsused for direct gene transfer in plants are:

Chemical mediated gene transfer e.g. chemicals like polyethylene glycol(PEG) and dextran sulphate induce DNA uptake into plant protoplasts.Calcium phosphate is also used to transfer DNA into cultured cells.

Microinjection where the DNA is directly injected into plant protoplastsor cells (specifically into the nucleus or cytoplasm) using fine tipped (0.5 - 1.0micrometer diameter) glass needle or micropipette. This method of genetransfer is used to introduce DNA into large cells such as oocytes, eggs, andthe cells of early embryo. Electroporation involves a pulse of high voltageapplied to protoplasts/cells/tissues to make transient (temporary) pores in theplasma membrane which facilitates the uptake of foreign DNA.

The cells are placed in a solution containing DNA and subjected toelectrical shocks to cause holes in the membranes. The foreign DNA fragmentsenter through the holes into the cytoplasm and then to nucleus.

Particle gun/Particle bombardment - In this method, the foreign DNAcontaining the genes to be transferred is coated onto the surface of minutegold or tungsten particles (1-3 micrometers) and bombarded onto the targettissue or cells using a particle gun (also called as gene gun/shot gun/microprojectile gun).The microprojectile bombardment method was initiallynamed as biolistics by its inventor Sanford. Two types of plant tissue arecommonly used for particle bombardment- Primary explants and theproliferating embryonic tissues.

Transformation - This method is used for introducing foreign DNA intobacterial cells e.g. E. Coli. The transformation frequency (the fraction of cellpopulation that can be transferred) is very good in this method. E.g. the uptakeof plasmid DNA by E. coli is carried out in ice cold CaCl2 (0-50C) followed byheat shock treatment at 37-450C for about 90 sec. The transformation efficiencyrefers to the number of transformants per microgram of added DNA. TheCaCl2 breaks the cell wall at certain regions and binds the DNA to the cellsurface.

Conjuction - It is a natural microbial recombination process and is usedas a method for gene transfer. In conjuction, two live bacteria come togetherand the single stranded DNA is transferred via cytoplasmic bridges from thedonor bacteria to the recipient bacteria.

Liposome mediated gene transfer or Lipofection - Liposomes are circularlipid molecules with an aqueous interior that can carry nucleic acids.Liposomes encapsulate the DNA fragments and then adher to the cellmembranes and fuse with them to transfer DNA fragments. Thus, the DNAenters the cell and then to the nucleus. Lipofection is a very efficient techniqueused to transfer genes in bacterial, animal and plant cells.

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SELECT ION OF T RANSFORM ED CELLS FROM UNT RANSFORM ED

CELLS

The selection of transformed plant cells from untransformed cells is animportant step in the plant genetic engineering. For this, a marker gene (e.g.for antibiotic resistance) is introduced into the plant along with the transgenefollowed by the selection of an appropriate selection medium (containing theantibiotic). The segregation and stability of the transgene integration andexpression in the subsequent generations can be studied by genetic andmolecular analyses.

TRANSGENIC PLANTS WITH BENEFICIAL TRAITS

During the last decades, a tremendous progress has been made in thedevelopment of transgenic plants using the various techniques of geneticengineering. The plants, in which a functional foreign gene has beenincorporated by any biotechnological methods that generally are not presentin the plant, are called transgenic plants. As per estimates recorded in 2002,transgenic crops are cultivated world-wide on about 148 million acres (587million hectares) land by about 5.5 million farmers. Transgenic plants havemany beneficial traits like insect resistance, herbicide tolerance, delayed fruitripening, improved oil quality, weed control etc.

ST RESS TOLERANCE

Biotechnology strategies are being developed to overcome problemscaused due to biotic stresses (viral, bacterial infections, pests and weeds) andabiotic stresses (physical actors such as temperature, humidity, salinity etc).

Abiotic Stress Tolerance

The plants show their abiotic stress response reactions by the productionof stress related osmolytes like sugars (e.g. trehalose and fructans), sugaralcohols (e.g. mannitol), amino acids (e.g. proline, glycine, betaine) and certainproteins (e.g. antifreeze proteins). Transgenic plants have been produced whichover express the genes for one or more of the above mentioned compounds.Such plants show increased tolerance to environmental stresses. Resistanceto abiotic stresses includes stress induced by herbicides, temperature (heat,chilling, freezing), drought, salinity, ozone and intense light. Theseenvironmental stresses result in the destruction, deterioration of crop plantswhich leads to low crop productivity. Several strategies have been used anddeveloped to build ressitance in the plants against these stresses.

Herbicide Tolerance

Weeds are unwanted plants which decrease the crop yields and bycompeting with crop plants for light, water and nutrients. Several

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biotechnological strategies for weed control are being used e.g. the over-production of herbicide target enzyme (usually in the chloroplast) in the plantwhich makes the plant insensitive to the herbicide. This is done by theintroduction of a modified gene that encodes for a resistant form of the enzymetargeted by the herbicide in weeds and crop plants. Roundup Ready cropplants tolerant to herbicide-Roundup, is already being used commercially.

The biological manipulations using genetic engineering to develop herbicideresistant plants are:

• Over-expression of the target protein by integrating multiple copiesof the gene or by using a strong promoter.

• Enhancing the plant detoxification system which helps in reducingthe effect of herbicide.

• Detoxifying the herbicide by using a foreign gene.• Modification of the target protein by mutation.Some of the examples are:Glyphosate resistance - Glyphosate is a glycine derivative and is a

herbicide which is found to be effective against the 76 of the world’s worst 78weeds. It kills the plant by being the competitive inhibitor of the enzyme 5-enoyl-pyruvylshikimate 3- phosphate synthase (EPSPS) in the shikimic acidpathway. Due to it’s structural similarity with the substrate phosphoenolpyruvate, glyphosate binds more tightly with EPSPS and thus blocks theshikimic acid pathway.

Certain strategies were used to provide glyphosate resistance to plants:• It was found that EPSPS gene was overexpressed in Petunia due to

gene amplification. EPSPS gene was isolated from Petunia andintroduced in to the other plants. These plants could tolerateglyphosate at a dose of 2- 4 times higher than that required to killwild type plants:– By using mutant EPSPS genes: A single base substitution from

C to T resulted in the change of an amino acid from proline toserine in EPSPS. The modified enzyme cannot bind toglyphosate and thus provides resistance.

– The detoxification of glyphosate by introducing the gene(isolated from soil organism- Ochrobactrum anthropi) encodingfor glyphosate oxidase into crop plants. The enzyme glyphosateoxidase converts glyphosate to glyoxylate andaminomethylphosponic acid. The transgenic plants exhibitedvery good glyphosate ressitance in the field.

Another Example is of Phosphinothricin Resistance

Phosphinothricin is a broad spectrum herbicide and is effective againstbroad-leafed weeds. It acts as a competitive inhibitor of the enzyme glutaminesynthase which results in the inhibition of the enzyme glutamine synthase

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and accumulation of ammonia and finally the death of the plant. Thedisturbace in the glutamine synthesis also inhibits the photosynthetic activity.The enzyme phosphinothricin acetyl transferase (which was first observed inStreptomyces sp in natural detoxifying mechanism against phosphinothricin)acetylates phosphinothricin, and thus inactivates the herbicide. The geneencoding for phosphinothricin acetyl transferase (bar gene) was introducedin transgenic maize and oil seed rape to provide resistance againstphosphinothricin.

OT HER ABIOT IC ST RESSES

The abiotic stresses due to temperature, drought, and salinity arecollectively also known as water deficit stresses. The plants produce osmolytesor osmoprotectants to overcome the osmotic stress. The attempts are on touse genetic engineering strategies to increase the production ofosmoprotectants in the plants. The biosynthetic pathways for the productionof many osmoprotectants have been established and genes coding the keyenzymes have been isolated. E.g. Glycine betaine is a cellular osmolyte whichis produced by the participation of a number of key enzymes like cholinedehydrogenase, choline monooxygenase etc. The choline oxidase gene fromArthrobacter sp. was used to produce transgenic rice with high levels ofglycine betaine giving tolerance against water deficit stress.

Scientists also developed cold-tolerant genes (around 20) in Arabidopsiswhen this plant was gradually exposed to slowly declining temperature. Byintroducing the coordinating gene (it encodes a protein which acts astranscription factor for regulating the expression of cold tolerant genes),expression of cold tolerant genes was triggered giving protection to the plantsagainst the cold temperatures.

INSECT RESISTANCE

A variety of insects, mites and nematodes significantly reduce the yieldand quality of the crop plants. The conventional method is to use syntheticpesticides, which also have severe effects on human health and environment.The transgenic technology uses an innovative and eco-friendly method toimprove pest control management.About 40 genes obtained frommicroorganisms of higher plants and animals have been used to provide insectresistance in crop plants.

The first genes available for genetic engineering of crop plants for pestresistance were Cry genes (popularly known as Bt genes) from a bacteriumBacillus thuringiensis. These are specific to particular group of insect pests,and are not harmful to other useful insects like butter flies and silk worms.Transgenic crops with Bt genes (e.g. cotton, rice, maize, potato, tomato, brinjal,cauliflower, cabbage, etc.) have been developed. This has proved to be aneffective way of controlling the insect pests and has reduced the pesticide

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use. The most notable example is Bt cotton (which contains CrylAc gene) thatis resistant to a notorious insect pest Bollworm (Helicoperpa armigera). Thereare certain other insect resistant genes from other microorganisms which havebeen used for this purpose. Isopentenyl transferase gene from Agrobacteriumtumefaciens has been introduced into tobacco and tomato. The transenic plantswith this transgene were found to reduce the leaf consumption by tobaccohornworm and decrease the survival of peach potato aphid.

Certain genes from higher plants were also found to result in the synthesisof products possessing insecticidal activity. One of the examples is the Cowpeatrypsin inhibitor gene (CpTi) which was introduced into tobacco, potato, andoilseed rape for develping transgenic plants. Earlier it was observed that thewild species of cowpea plants growing in Africa were resistant to attack by awide range of insects. It was observed that the insecticidal protein was atrypsin inhibitor that was capable of destroying insects belonging to the ordersLepidoptera, Orthaptera etc. Cowpea trypsin inhibitor (CpTi) has no effecton mammalian trypsin, hence it is non-toxic to mammals.

VIRUS RESISTANCE

There are several strategies for engineering plants for viral resistance,and these utilizes the genes from virus itself (e.g. the viral coat protein gene).The virus-derived resistance has given promising results in a number of cropplants such as tobacco, tomato, potato, alfalfa, and papaya. The induction ofvirus resistance is done by employing virus-encoded genes-virus coat proteins,movement proteins, transmission proteins, satellite RNa, antisense RNAs, andribozymes. The virus coat protein-mediated approach is the most successfulone to provide virus resistance to plants. It was in 1986, transgenic tobaccoplants expressing tobacco mosaic virus (TMV) coat protein gene were firstdeveloped. These plants exhibited high levels of resistance to TMV.

The transgenic plant providing coat protein-mediated resistance to virusare rice, potato, peanut, sugar beet, alfalfa etc. The viruses that have beenused include alfalfa mosaic virus (AIMV), cucumber mosaic virus (CMV),potato virus X (PVX), potato virus Y (PVY) etc.

RESISTANCE AGAINST FUNGAL AND BACT ERIAL INFECT IONS

As a defence strategy against the invading pathogens (fungi and bacteria)the plants accumulate low molecular weight proteins which are collectivelyknown as pathogenesis-related (PR) proteins.

Several transgenic crop plants with increased resistance to fungalpathogens are being raised with genes coding for the different compounds.One of the examples is the Glucanase enzyme that degrades the cell wall ofmany fungi. The most widely used glucanase is beta-1,4-glucanase. The geneencoding for beta-1,4 glucanase has been isolated from barley, introduced,and expressed in transgenic tobacco plants. This gene provided good

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protection against soil-borne fungal pathogen Rhizoctonia solani. Lysozymedegrades chitin and peptidoglycan of cell wall, and in this way fungal infectioncan be reduced. Transgenic potato plants with lysozyme gene providingresistance to Eswinia carotovora have been developed.

DELAYED FRUIT RIPENING

The gas hormone, ethylene regulates the ripening of fruits, therefore,ripening can be slowed down by blocking or reducing ethylene production.This can be achieved by introducing ethylene forming gene(s) in a way thatwill suppress its own expression in the crop plant. Such fruits ripen very slowly(however, they can be ripen by ethylene application) and this helps inexporting the fruits to longer distances without spoilage due to longer-shelflife. The most common example is the 'Flavr Savr' transgenic tomatoes, whichwere commercialized in U.S.A in 1994. The main strategy used was theantisense RNA approach. In the normal tomato plant, the PG gene (for theenzyme polygalacturonase) encodes a normal mRNA that produces theenzyme polygalacturonase which is involved in the fruit ripening.

The complimentary DNA of PG encodes for antisense mRNA, which iscomplimentary to normal (sense) mRNA. The hybridization between the senseand antisnse mRNAs renders the sense mRNA ineffective. Consequently,polygalacturonase is not produced causing delay in the fruit ripening.Similarly strategies have been developed to block the ethylene biosynthesisthereby reducing the fruit ripening. E.g. transgenic plants with antisense geneof ACC oxidase (an enzyme involved in the biosynthetic process of ethylene)have been developed. In these plants, production of ethylene was reduced byabout 97% with a significant delay in the fruit ripening.

The bacterial gene encoding ACC deaminase (an enzyme that acts on ACCand removes amino group) has been transferred and expressed in tomatoplants which showed 90% inhibition in the ethylene biosynthesis.

M ALE ST ERILIT Y

The plants may inherit male sterility either from the nucleus or cytoplasm.It is possible to introduce male sterility through genetic manipulations whilethe female plants maintain fertility.

In tobacco plants, these are created by introducing a gene coding for anenzyme (barnase, which is a RNA hydrolyzing enzyme) that inhibits pollenformation. This gene is expressed specifically in the tapetal cells of antherusing tapetal specific promoter TA29 to restrict its activity only to the cellsinvolved in pollen production. The restoration of male fertility is done byintroducing another gene barstar that suppresses the activity of barnase atthe onset of the breeding season. By using this approach, transgenic plants oftobacco, cauliflower, cotton, tomato, corn, lettuce etc. with male sterility havebeen developed.

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9

Plant Breeding

INTRODUCTION

Plant breeding is the art and science of changing the genetics of plantsfor the benefit of humankind. Plant breeding can be accomplished throughmany different techniques ranging from simply selecting plants with desirablecharacteristics for propagation, to more complex molecular techniques. Plantbreeding has been practiced for thousands of years, since near the beginningof human civilization. It is now practiced worldwide by individuals such asgardeners and farmers, or by professional plant breeders employed byorganizations such as government institutions, universities, crop-specificindustry associations or research centers.

International development agencies believe that breeding new crops isimportant for ensuring food security by developing new varieties that arehigher-yielding, resistant to pests and diseases, drought-resistant or regionallyadapted to different environments and growing conditions.

DOM ESTICAT ION

Plant breeding in certain situations may lead the domestication of wildplants. Domestication of plants is an artificial selection process conducted byhumans to produce plants that have more desirable traits than wild plants,and which renders them dependent on artificial (usually enhanced)environments for their continued existence. The practice is estimated to dateback 9,000-11,000 years. Many crops in present day cultivation are the resultof domestication in ancient times, about 5,000 years ago in the Old Worldand 3,000 years ago in the New World. In the Neolithic period, domesticationtook a minimum of 1,000 years and a maximum of 7,000 years. Today, all ofour principal food crops come from domesticated varieties. Almost all thedomesticated plants used today for food and agriculture were domesticatedin the centers of origin. In these centers there is still a great diversity of closelyrelated wild plants, the so called crop wild relatives that can also be used for

improving modern cultivars by plant breeding. A plant whose origin orselection is due primarily to intentional human activity is called a cultigen,and a cultivated crop species that has evolved from wild populations due toselective pressures from traditional farmers is called a landrace. Landraces,which can be the result of natural forces or domestication, are plants (oranimals) that are ideally suited to a particular region or environment. Anexample are the landraces of rice, Oryza sativa subspecies indica, which wasdeveloped in South Asia, and Oryza sativa subspecies japonica, which wasdeveloped in China.

CLASSICAL PLANT BREEDING

Classical plant breeding uses deliberate interbreeding (crossing) of closelyor distantly related individuals to produce new crop varieties or lines withdesirable properties. Plants are crossbred to introduce traits/genes from onevariety or line into a new genetic background. For example, a mildew-resistantpea may be crossed with a high-yielding but susceptible pea, the goal of thecross being to introduce mildew resistance without losing the high-yieldcharacteristics. Progeny from the cross would then be crossed with the high-yielding parent to ensure that the progeny were most like the high-yieldingparent, (backcrossing). The progeny from that cross would then be tested foryield and mildew resistance and high-yielding resistant plants would befurther developed. Plants may also be crossed with themselves to produceinbred varieties for breeding.

Classical breeding relies largely on homologous recombination betweenchromosomes to generate genetic diversity. The classical plant breeder mayalso makes use of a number of in vitro techniques such as protoplast fusion,embryo rescue or mutagenesis to generate diversity and produce hybrid plantsthat would not exist in nature. Traits that breeders have tried to incorporateinto crop plants in the last 100 years include:

1. Increased quality and yield of the crop.2. Increased tolerance of environmental pressures (salinity, extreme

temperature, drought).3. Resistance to viruses, fungi and bacteria.4. Increased tolerance to insect pests.5. Increased tolerance of herbicides.

Before World War II

Intraspecific hybridization within a plant species was demonstrated byCharles Darwin and Gregor Mendel, and was further developed by geneticistsand plant breeders. In the United Kingdom in the 1880s was the pioneeringwork of Gartons Agricultural Plant Breeders. In the early 20th century, plantbreeders realized that Mendel’s findings on the non-random nature ofinheritance could be applied to seedling populations produced through

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deliberate pollinations to predict the frequencies of different types. In 1908,George Harrison Shull described heterosis, also known as hybrid vigour.Heterosis describes the tendency of the progeny of a specific cross tooutperform both parents. The detection of the usefulness of heterosis for plantbreeding has led to the development of inbred lines that reveal a heteroticyield advantage when they are crossed. Maize was the first species whereheterosis was widely used to produce hybrids.

By the 1920s, statistical methods were developed to analyse gene actionand distinguish heritable variation from variation caused by environment. In1933, another important breeding technique, cytoplasmic male sterility (CMS),developed in maize, was described by Marcus Morton Rhoades. CMS is amaternally inherited trait that makes the plant produce sterile pollen. Thisenables the production of hybrids without the need for labour intensivedetasseling. These early breeding techniques resulted in large yield increasein the United States in the early 20th century. Similar yield increases werenot produced elsewhere until after World War II, the Green Revolutionincreased crop production in the developing world in the 1960s.

After World War II

Following World War II a number of techniques were developed thatallowed plant breeders to hybridize distantly related species, and artificiallyinduce genetic diversity. When distantly related species are crossed, plantbreeders make use of a number of plant tissue culture techniques to produceprogeny from otherwise fruitless mating. Interspecific and intergeneric hybridsare produced from a cross of related species or genera that do not normallysexually reproduce with each other.

These crosses are referred to as Wide crosses. For example, the cerealtriticale is a wheat and rye hybrid. The cells in the plants derived from thefirst generation created from the cross contained an uneven number ofchromosomes and as result was sterile. The cell division inhibitor colchicinewas used to double the number of chromosomes in the cell and thus allowthe production of a fertile line. Failure to produce a hybrid may be due topre- or post-fertilization incompatibility. If fertilization is possible betweentwo species or genera, the hybrid embryo may abort before maturation. Ifthis does occur the embryo resulting from an interspecific or intergeneric crosscan sometimes be rescued and cultured to produce a whole plant. Such amethod is referred to as Embryo Rescue. This technique has been used toproduce new rice for Africa, an interspecific cross o f Asian rice (Oryza sativa)and African rice (Oryza glaberrima).

Hybrids may also be produced by a technique called protoplast fusion.In this case protoplasts are fused, usually in an electric field. Viablerecombinants can be regenerated in culture. Chemical mutagens like EMS andDMS, radiation and transposons are used to generate mutants with desirable

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traits to be bred with other cultivars- a process known as Mutation Breeding.Classical plant breeders also generate genetic diversity within a species byexploiting a process called somaclonal variation, which occurs in plantsproduced from tissue culture, particularly plants derived from callus. Inducedpolyploidy, and the addition or removal of chromosomes using a techniquecalled chromosome engineering may also be used.

When a desirable trait has been bred into a species, a number of crossesto the favoured parent are made to make the new plant as similar to thefavoured parent as possible. Returning to the example of the mildew resistantpea being crossed with a high-yielding but susceptible pea, to make themildew resistant progeny of the cross most like the high- yielding parent, theprogeny will be crossed back to that parent for several generations. Thisprocess removes most of the genetic contribution of the mildew resistantparent. Classical breeding is therefore a cyclical process.

It should be noted that with classical breeding techniques, the breederdoes not know exactly what genes have been introduced to the new cultivars.Some scientists therefore argue that plants produced by classical breedingmethods should undergo the same safety testing regime as geneticallymodified plants.

There have been instances where plants bred using classical techniqueshave been unsuitable for human consumption, for example the poison solaninewas unintentionally increased to unacceptable levels in certain varieties ofpotato through plant breeding. New potato varieties are often screened forsolanine levels before reaching the marketplace.

MODERN PLANT BREEDING

Modern plant breeding uses techniques of molecular biology to select,or in the case of genetic modification, to insert, desirable traits into plants.

ST EPS OF PLANT BREEDING

The following are the major activities of plant breeding;1. Creation variation,2. Selection,3. Evaluation,4. Release,5. Multiplication,6. Distribution of the new variety.

Marker Assisted Selection

Sometimes many different genes can influence a desirable trait in plantbreeding. The use of tools such as molecular markers or DNA fingerprintingcan map thousands of genes. This allows plant breeders to screen large

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populations of plants for those that possess the trait of interest. The screeningis based on the presence or absence of a certain gene as determined bylaboratory procedures, rather than on the visual identification of the expressedtrait in the plant.

Reverse Breeding and Doubled Haploids (DH)

A method for efficiently producing homozygous plants from aheterozygous starting plant, which has all desirable traits. This starting plantis induced to produce doubled haploid from haploid cells, and later on creatinghomozygous/doubled haploid plants from those cells. While in naturaloffspring recombination occurs and traits can be unlinked from each other,in doubled haploid cells and in the resulting DH plants recombination is nolonger an issue. There, a recombination between two correspondingchromosomes does not lead to un-linkage of alleles or traits, since it just leadsto recombination with its identical copy. Thus, traits on one chromosome staylinked. Selecting those offspring having the desired set of chromosomes andcrossing them will result in a final F1 hybrid plant, having exactly the sameset of chromosomes, genes and traits as the starting hybrid plant. Thehomozygous parental lines can reconstitute the original heterozygous plantby crossing, if desired even in a large quantity. An individual heterozygousplant can be converted into a heterozygous variety (F1 hybrid) without thenecessity of vegetative propagation but as the result of the cross of twohomozygous/doubled haploid lines derived from the originally selected plant.patent.

Genetic Modification

Genetic modification of plants is achieved by adding a specific gene orgenes to a plant, or by knocking down a gene with RNAi, to produce adesirable phenotype. The plants resulting from adding a gene are oftenreferred to as transgenic plants. If for genetic modification genes of the speciesor of a crossable plant are used under control of their native promoter, thenthey are called cisgenic plants. Genetic modification can produce a plant withthe desired trait or traits faster than classical breeding because the majorityof the plant’s genome is not altered.

To genetically modify a plant, a genetic construct must be designed sothat the gene to be added or removed will be expressed by the plant. To dothis, a promoter to drive transcription and a termination sequence to stoptranscription of the new gene, and the gene or genes of interest must beintroduced to the plant. A marker for the selection of transformed plants isalso included. In the laboratory, antibiotic resistance is a commonly usedmarker: plants that have been successfully transformed will grow on mediacontaining antibiotics; plants that have not been transformed will die. In someinstances markers for selection are removed by backcrossing with the parent

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plant prior to commercial release. The construct can be inserted in the plantgenom e by genetic recom bination using the bacteria Agrobacterium tumefaciensor A. rhizogenes, or by direct methods like the gene gun or microinjection.Using plant viruses to insert genetic constructs into plants is also a possibility,but the technique is limited by the host range of the virus. For example,Cauliflower mosaic virus (CaMV) only infects cauliflower and related species.Another limitation of viral vectors is that the virus is not usually passed onthe progeny, so every plant has to be inoculated.

The majority of commercially released transgenic plants, are currentlylimited to plants that have introduced resistance to insect pests and herbicides.Insect resistance is ach ieved through incorporation of a gene from Bacillusthuringiensis (Bt) that encodes a protein that is toxic to some insects. Forexample, the cotton bollworm, a common cotton pest, feeds on Bt cotton itwill ingest the toxin and die. Herbicides usually work by binding to certainplant enzymes and inhibiting their action. The enzymes that the herbicideinhibits are known as the herbicides target site. Herbicide resistance can beengineered into crops by expressing a version of target site protein that is notinhibited by the herbicide. This is the method used to produce glyphosateresistant crop plants.

Genetic modification of plants that can produce pharmaceuticals (andindustrial chemicals), sometimes called pharmacrops, is a rather radical newarea of plant breeding.

ISSUES AND CONCERNS

Modern plant breeding, whether classical or through genetic engineering,comes with issues of concern, particularly with regard to food crops. Thequestion of whether breeding can have a negative effect on nutritional valueis central in this respect. Although relatively little direct research in this areahas been done, there are scientific indications that, by favoring certain aspectsof a plant’s development, other aspects may be retarded. A study publishedin the Journal of the American College of Nutrition in 2004, entitled Changes inUSDA Food Composition Data for 43 Garden Crops, 1950 to 1999, comparednutritional analysis of vegetables done in 1950 and in 1999, and foundsubstantial decreases in six of 13 nutrients measured, including 6% of proteinand 38% of riboflavin. Reductions in calcium, phosphorus, iron and ascorbicacid were also found. The study, conducted at the Biochemical Institute,University of Texas at Austin, concluded in sum m ary: “We suggest that anyreal declines are generally most easily explained by changes in cultivated varietiesbetween 1950 and 1999, in which there may be trade-offs between yield and nutrientcontent. ”

The debate surrounding genetically modified food during the 1990speaked in 1999 in terms of media coverage and risk perception, and continuestod ay- for exam ple, “Germany has thrown its weight behind a growing European

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mutiny over genetically modified crops by banning the planting of a widely grownpest-resistant corn variety.”. The debate encompasses the ecological impact ofgenetically modified plants, the safety of genetically modified food andconcepts used for safety evaluation like substantial equivalence. Such concernsare not new to plant breeding. Most countries have regulatory processes inplace to help ensure that new crop varieties entering the marketplace are bothsafe and meet farmers’ needs. Examples include variety registration, seedschemes, regulatory authorizations for GM plants, etc.

Plant breeders’ rights is also a major and controversial issue. Today,production of new varieties is dominated by commercial plant breeders, whoseek to protect their work and collect royalties through national andinternational agreements based in intellectual property rights. The range ofrelated issues is complex. In the simplest terms, critics of the increasinglyrestrictive regulations argue that, through a combination of technical andeconomic pressures, commercial breeders are reducing biodiversity andsignificantly constraining individuals (such as farmers) from developing andtrading seed on a regional level. Efforts to strengthen breeders’ rights, forexample, by lengthening periods of variety protection, are ongoing.

When new plant breeds or cultivars are bred, they must be maintainedand propagated. Some plants are propagated by asexual means while othersare propagated by seeds. Seed propagated cultivars require specific controlover seed source and production procedures to maintain the integrity of theplant breeds results. Isolation is necessary to prevent cross contamination withrelated plants or the mixing of seeds after harvesting. Isolation is normallyaccomplished by planting distance but in certain crops, plants are enclosedin greenhouses or cages (most commonly used when producing F1 hybrids.)

PARTICIPATORY PLANT BREEDING

Participatory Plant Breeding is carried out, for example, in northernVietnam, where government scientists work with farmers from the Muongpeople ethnic minority to improve local rice varieties.

The development of agricultural science, with phenomenon like the GreenRevolution arising, have left millions of farmers in developing countries, mostof whom operate small farms under unstable and difficult growing conditions,in a precarious situation. The adoption of new plant varieties by this grouphas been hampered by the constraints of poverty and the international policiespromoting an industrialised model of agriculture. Their response has beenthe creation of a novel and promising set of research methods collectivelyknown as participatory plant breeding. Participatory means that farmers aremore involved in the breeding process and breeding goals are defined byfarmers instead of international seed companies with their large-scale breedingprogrammes. Farmer’s groups and NGOs, for example, may wish to affirm

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local people’s rights over genetic resources, produce seeds themselves, buildfarmers’ technical expertise, or develop new products for niche markets, likeorganically grown food.

BREEDING METHODS IN CROP PLANTS

SELF POLLINAT ED CROPS

Mass Selection

In mass selection, seeds are collected from (usually a few dozen to a fewhundred) desirable appearing individuals in a population, and the nextgeneration is sown from the stock of mixed seed. This procedure, sometimesreferred to as phenotypic selection, is based on how each individual looks.Mass selection has been used widely to improve old "land" varieties, varietiesthat have been passed down from one generation of farmers to the next overlong periods. An alternative approach that has no doubt been practiced forthousands of years is simply to eliminate undesirable types by destroyingthem in the field. The results are similar whether superior plants are saved orinferior plants are eliminated: seeds of the better plants become the plantingstock for the next season. A modern refinement of mass selection is to harvestthe best plants separately and to grow and compare their progenies. Thepoorer progenies are destroyed and the seeds of the remainder are harvested.It should be noted that selection is now based not solely on the appearance ofthe parent plants but also on the appearance and performance of their progeny.Progeny selection is usually more effective than phenotypic selection whendealing with quantitative characters of low heritability. It should be noted,however, that progeny testing requires an extra generation; hence gain percycle of selection must be double that of simple phenotypic selection to achievethe same rate of gain per unit time.

Mass selection, with or without progeny test, is perhaps the simplest andleast expensive of plant–breeding procedures. It finds wide use in the breedingof certain forage species, which are not important enough economically tojustify more detailed attention.

Pure–Line Selection

Pure–line selection generally involves three more or less distinct steps:1. Numerous superior appearing plants are selected from a genetically

variable population;2. Progenies of the individual plant selections are grown and evaluated

by simple observation, frequently over a period of several years;3. When selection can no longer be made on the basis of observation

alone, extensive trials are undertaken, involving carefulmeasurements to determine whether the remaining selections aresuperior in yielding ability and other aspects of performance.

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Any progeny superior to an existing variety is then released as a new"pure–line" variety. Much of the success of this method during the early 1900sdepended on the existence of genetically variable land varieties that werewaiting to be exploited. They provided a rich source of superior pure–linevarieties, some of which are still represented among commercial varieties. Inrecent years the pure–line method as outlined above has decreased inimportance in the breeding of major cultivated species; however, the methodis still widely used with the less important species that have not yet beenheavily selected.

A variation of the pure–line selection method that dates back centuries isthe selection of single–chance variants, mutations or "sports" in the originalvariety. A very large number of varieties that differ from the original strainin characteristics such as colour, lack of thorns or barbs, dwarfness, and diseaseresistance have originated in this fashion.

Hybridization

During the 20th century planned hybridization between carefully selectedparents has become dominant in the breeding of self–pollinated species. Theobject of hybridization is to combine desirable genes found in two or moredifferent varieties and to produce pure–breeding progeny superior in manyrespects to the parental types. Genes, however, are always in the company ofother genes in a collection called a genotype. The plant breeder's problem islargely one of efficiently managing the enormous numbers of genotypes thatoccur in the generations following hybridization.

As an example of the power of hybridization in creating variability, across between hypothetical wheat varieties differing by only 21 genes iscapable of producing more than 10,000,000,000 different genotypes in thesecond generation. At spacing normally used by farmers, more than 50,000,000acres would be required to grow a population large enough to permit everygenotype to occur in its expected frequency. While the great majority of thesesecond generation genotypes are hybrid (heterozygous) for one or more traits,it is statistically possible that 2,097,152 different pure–breeding (homozygous)genotypes can occur, each potentially a new pure–line variety. These numbersillustrate the importance of efficient techniques in managing hybridpopulations, for which purpose the pedigree procedure is most widely used.

Pedigree Breeding

Pedigree breeding starts with the crossing of two genotypes, each of whichhave one or more desirable characters lacked by the other. If the two originalparents do not provide all of the desired characters, a third parent can beincluded by crossing it to one of the hybrid progeny of the first generation(F1). In the pedigree method superior types are selected in successivegenerations, and a record is maintained of parent–progeny relationships. TheF2 generation (progeny of the crossing of two F1 individuals) affords the first

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opportunity for selection in pedigree programmes. In this generation theemphasis is on the elimination of individuals carrying undesirable majorgenes. In the succeeding generations the hybrid condition gives way to purebreeding as a result of natural self– pollination, and families derived fromdifferent F2 plants begin to display their unique character. Usually one ortwo superior plants are selected within each superior family in thesegenerations. By the F5 generation the pure–breeding condition (homozygosity)is extensive, and emphasis shifts almost entirely to selection between families.The pedigree record is useful in making these eliminations. At this stage eachselected family is usually harvested in mass to obtain the larger amounts ofseed needed to evaluate families for quantitative characters. This evaluationis usually carried out in plots grown under conditions that simulatecommercial planting practice as closely as possible. When the number offamilies has been reduced to manageable proportions by visual selection,usually by the F7 or F8 generation, precise evaluation for performance andquality begins.

The final evaluation of promising strains involves:• Observation, usually in a number of years and locations, to detect

weaknesses that may not have appeared previously;• Precise yield testing;• Quality testing. Many plant breeders test for five years at five

representative locations before releasing a new variety forcommercial production.

The Bulk–Population Method

The bulk–population method of breeding differs from the pedigreemethod primarily in the handling of generations following hybridization. TheF2 generation is sown at normal commercial planting rates in a large plot. Atmaturity the crop is harvested in mass, and the seeds are used to establishthe next generation in a similar plot. No record of ancestry is kept. Duringthe period of bulk propagation natural selection tends to eliminate plantshaving poor survival value.

Two types of artificial selection also are often applied:1. Destruction of plants that carry undesirable major genes;2. Mass techniques such as harvesting when only part of the seeds

are mature to select for early maturing plants or the use of screensto select for increased seed size.

Single plant selections are then made and evaluated in the same way asin the pedigree method of breeding. The chief advantage of the bulkpopulation method is that it allows the breeder to handle very large numbersof individuals inexpensively. Often an outstanding variety can be improvedby transferring to it some specific desirable character that it lacks. This can beaccomplished by first crossing a plant of the superior variety to a plant of thedonor variety, which carries the trait in question, and then mating the progeny

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back to a plant having the genotype of the superior parent. This process iscalled backcrossing. After five or six backcrosses the progeny will be hybridfor the character being transferred but like the superior parent for all othergenes. Selfing the last backcross generation, coupled with selection, will givesome progeny pure breeding for the genes being transferred. The advantagesof the backcross method are its rapidity, the small number of plants required,and the predictability of the outcome. A serious disadvantage is that theprocedure diminishes the occurrence of chance combinations of genes, whichsometimes leads to striking improvements in performance.

Hybrid Varieties

The development of hybrid varieties differs from hybridization. The F1hybrid of crosses between different genotypes is often much more vigorousthan its parents. This hybrid vigour, or heterosis, can be manifested in manyways, including increased rate of growth, greater uniformity, earlier flowering,and increased yield, the last being of greatest importance in agriculture.

CROSS POLLINAT ED CROPS

The most important methods of breeding cross–pollinated species are:• Mass selection;• Development of hybrid varieties;• Development of synthetic varieties. Since cross–pollinated species

are naturally hybrid (heterozygous) for many traits and lose vigouras they become purebred (homozygous), a goal of each of thesebreeding methods is to preserve or restore heterozygosity.

Mass Selection

Mass selection in cross–pollinated species takes the same form as in self–pollinated species; i.e., a large number of superior appearing plants are selectedand harvested in bulk and the seed used to produce the next generation. Massselection has proved to be very effective in improving qualitative characters, and,applied over many generations, it is also capable of improving quantitativecharacters, including yield, despite the low heritability of such characters. Massselection has long been a major method of breeding cross–pollinated species,especially in the economically less important species.

Hybrid Varieties

The outstanding example of the exploitation of hybrid vigour throughthe use of F1 hybrid varieties has been with corn (maize).

The production of a hybrid corn variety involves three steps:1. The selection of superior plants;2. Selfing for several generations to produce a series of inbred lines,

which although different from each other are each pure– breedingand highly uniform;

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3. Crossing selected inbred lines.During the inbreeding process the vigour of the lines decreases drastically,

usually to less than half that of field–pollinated varieties. Vigour is restored,however, when any two unrelated inbred lines are crossed, and in some casesthe F1 hybrids between inbred lines are much superior to open–pollinatedvarieties. An important consequence of the homozygosity of the inbred linesis that the hybrid between any two inbreds will always be the same. Once theinbreds that give the best hybrids have been identified, any desired amountof hybrid seed can be produced. Pollination in corn (maize) is by wind, whichblows pollen from the tassels to the styles (silks) that protrude from the topsof the ears.

Thus controlled cross–pollination on a field scale can be accomplishedeconomically by interplanting two or three rows of the seed parent inbredwith one row of the pollinator inbred and detasselling the former before itsheds pollen. In practice most hybrid corn is produced from "double crosses,"in which four inbred lines are first crossed in pairs (A × B and C × D) and thenthe two F1 hybrids are crossed again (A × B) × (C × D).

The double–cross procedure has the advantage that the commercial F1seed is produced on the highly productive single cross A × B rather than on apoor-yielding inbred, thus reducing seed costs. In recent years cytoplasmicmale sterility, described earlier, has been used to eliminate detasselling ofthe seed parent, thus providing further economies in producing hybrid seed.Much of the hybrid vigour exhibited by F1 hybrid varieties is lost in the nextgeneration. Consequently, seed from hybrid varieties is not used for plantingstock but the farmer purchases new seed each year from seed companies. Perhapsno other development in the biological sciences has had greater impact onincreasing the quantity of food supplies available to the world's population thanhas the development of hybrid corn (maize). Hybrid varieties in other crops, madepossible through the use of male sterility, have also been dramatically successfuland it seems likely that use of hybrid varieties will continue to expand in thefuture.

Synthetic Varieties

A synthetic variety is developed by intercrossing a number of genotypesof known superior combining ability–i.e., genotypes that are known to givesuperior hybrid performance when crossed in all combinations. (By contrast,a variety developed by mass selection is made up of genotypes bulked togetherwithout having undergone preliminary testing to determine their performancein hybrid combination.) Synthetic varieties are known for their hybrid vigourand for their ability to produce usable seed for succeeding seasons. Becauseof these advantages, synthetic varieties have become increasingly favouredin the growing of many species, such as the forage crops, in which expenseprohibits the development or use of hybrid varieties.

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BREEDING EXPERIMENTS

The following crops have been selected to serve as examples of the varioustypes of flowers encountered in plant breeding –– perfect, imperfect, andcomposite. The techniques suggested here, with slight modifications, can beused for crossing other plants having similar flowering systems.

CORN

Corn is a good plant for the amateur plant breeder to experiment withbecause it is easy to work with and often shows visible changes in the kernelsof the F1 generation. Some of the varieties that you can work with are yellowfield corn, white field corn, sweet corn, popcorn, calico corn, and strawberrycorn.

Fig. Corn has an Imperfect Flower. The Staminate Flower isthe Tassel and the Pistillate is the Ear.

Corn has an imperfect flower, with both staminate and pistillate flowerson the same plant. The staminate flower is the tassel; the pistillate flower isthe ear. The silk, which has a hairy surface throughout most of its length thatis receptive to pollen, is the corn's stigma.

Procedure:• Decide on the varieties of corn that you wish to cross and plant the

seeds in your experimental area. You can work with as manyvarieties as you wish, but if this is your first experiment it will bebest to limit yourself to three or four. Corn tassels lose their pollenin a relatively short time, so in order to have the pollen of thedifferent varieties ready at about the same time, plant a few seedsof each variety weekly over a period of weeks. Mark the rows forfuture reference. 222222


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• As soon as the plants develop ear shoots and before the silks emerge,cover the shoots with a loosely fitted bag secured at the bottomwith a paper clip or string. A clear plastic or glassine bag will allowyou to watch the silks develop. Cover several ear shoots, for themore that are pollinated the greater your chances of getting desiredresults.

Fig. When Collecting Corn Pollen, Bend the Tassel Downward toPrevent the Pollen from Spilling Out of the Bag.

• Keep a close watch on the ears. When the silks are visible the ear isready for pollination. The appropriate tassel should then be baggedfor use the next morning. The bag should be secured tightly at thebase of the tassel to keep the pollen from falling out and to keep itfrom becoming contaminated with other pollen.

Remove the bag covering the silk. Place the bag from the tassel over theear, being careful not to spill any pollen. Secure the bag to the stalk below theear and shake.

Fig. Place the Bag Containing Pollen on the Ear of the Corn, Securethe Bag to the Stalk Below the Ear, and Label the Pollinated Ear.

• Label each of the ears that have been pollinated with the name ornumber of the plants that are serving as seed and pollen parentsand enter the cross in your record book.

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• Leave the bagged ear on the plant until the leaves dry. After thepollinated ear has dried, record any changes that may have occurredin the kernels. Then harvest the kernels and save them for futureplanting.

SQUASH

The flowers of squash are large and easy to work with. The flowers areimperfect, with both the male and female flowers on the same plant.

The pistillate flower is enlarged at the base while the staminate flower isborne on a long stem. Although some varieties of squash are cross–sterile,that is, they cannot be crossed, many others can be crossed.

It is also possible to cross some varieties of pumpkin and squash.Remember that if you are crossing pure varieties, segregation won't occuruntil the F2 generation; that is, the various colours and shapes that theoffspring have inherited will not all show up until the offspring are self–pollinated and the seeds that they produce bear fruit.

Procedure:• Plant the different varieties of squash. Label the rows so that you

will know which plants are to be seed parents and which are to bepollen parents. You won't have any difficulty in having the maleand female parents ready for pollination at the same time, for squashproduces flowers over a relatively long period.

• The evening before the flowers open, place a bag over them or close theflowers with rubber bands or string. It is easy to tell in the evening whichflowers will open the following morning. The flowers will be slightly open,revealing the inner colour, which is brighter than the colour on the outsideof the petals. Both pistillate and staminate flowers should be protected, toprevent bees from entering.

• Early in the morning on the day that the flowers open, remove thestamens from the staminate flower and gently rub the anthers acrossthe stigma of the pistillate flower.

• Replace the bag on the pistillate flower.

Fig. To Pollinate Squash, Gently Rub the Anther Across theStigma of the Pistillate Flower.

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• Label the pollinated pistillate flower and, enter the cross in yourrecord book.

• You can remove the protective covering when the petals have driedon the pistillate flower or when the ovary bulges through the bag.

• Harvest the seeds when the fruit is mature. Maturity is usuallyreached about 45 days after pollination. Save the seeds until thenext year, when you will.

• Plant them.• Self–pollinate the resulting flowers.• Again harvest the seeds.The year after that, when you plant the seeds resulting from the self–

pollination of the F generation, segregation will probably occur.

Fig. Harvest the Squash Seeds and Save them for Planting Next Year.

T OM AT OES

Fig. The Reproductive Parts of the Tomato.

The tomato has a perfect flower, with male and female organs both withinthe same blossom. The sex organs are easy to tell apart and therefore thereforeit is relatively easy to cross tomatoes. For this experiment, choose tomatoeswith two different fruit colours, such as red and yellow. If the tomatoes arepure, variations in colour will show up in the F2 generation. Along with colour

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variations, there will probably be changes in fruit shape and other plantcharacteristics.

Procedure:• Plant the seed or young plants and label the rows so that you will

know which will bear red fruit and which will bear yellow fruit.The tomato produces flowers for several weeks, so there will be nodifficulty in having pollen and seed parents ready at the same time.

• As soon as the flowers that will serve as seed parents begin to open.the plant should be emasculated. The stamens are fused to thepetals, so all you need to do to emasculate the plant is to pull outthe petals.

• After you remove the stamens from the seed parents, cover theflowers securely.

• The pollen parents that you wish to use should be in the same stageof opening as the seed parents. Cover the pollen parents at the sametime that you cover the seed parents.

Fig. A Tomato Flower that has been Emasculated, Pollinated, and Labeled.

Fig. Harvest the Tomato Seeds and Save them for Planting Next Year.• As soon as the pollen–parent flower is completely open, remove

one of its stamens with scissors or tweezers. Remove the bag fromthe seed parent and gently brush the anther across the stigma.

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• Replace the bag that was removed from the seed–parent flower.• Label the flower that was pollinated and enter the cross in your

record book.• After a week remove the bag from the seed parent and leave it off.• When the fruit is ripe, harvest and store the seeds for future planting

and self–pollination.

CHRYSANT HEM UM S

Chrysanthemums are composite flowers. If they are not available forcrossing, you can use any other composite flower, such as zinnias or asters.Composite flowers are more difficult to cross than complete or incompleteflowers.

If you are crossing plants for the first time, it would probably be best ifyou gained experience by first crossing a simpler flower, such as squash, iris,or lily, before trying to cross composite flowers.

Procedure:• Plant the seeds or young plants. Mums, zinnias, and asters bloom

over a relatively long period, so there will be no difficulty in havingpollen and seed parents ready at the same time.

• Immediately after the seed–parent flowers open and before thepollen is mature, remove the disc florets with tweezers. The discflorets are in the centre of the flower. They are easy to identifybecause they are often yellow in colour and they have both pistilsand stamens, whereas the ray florets, which surround the discflorets, have only pistils. Remove some of the ray florets near thecentre of the flower just to be sure that you haven't missed any ofthe disc florets. If any disc florets are left on the flower, the floweris likely to be self–pollinated.

Fig. When Emasculating Chrysanthemums, make sure that you remove all of theDisc Florets.

• After removing the disc florets, cover the seed– parent flowers withbags.

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• As soon as the flowers of the pollen parents open, cover them withbags.

• Pollination will be more effective if you trim the ray florets on theseed parents to just above the stigmas. This step can be done atany time after the disc florets are removed and before pollinationtakes place.

Fig. Trim the Ray Florets for more Effective Pollination.

• The flower is ready for pollination about four or five days after itopens. Shortly before you pollinate the flower, dip a camel–hair brushin alcohol and allow it to dry.

Fig. Each Mature Ray Floret will have a Seed. Harvest these Seeds andPlant them Next Year.

• Remove a flower from the pollen parent. With the camel–hair brush,gently brush the disc florets of the pollen–parent flower. The pollenshould adhere to the brush. Then gently brush the trimmed rayflorets of the seed–parent flower. Each time you change pollenparents dip the brush in alcohol and let it dry.

• Replace the protective bags on the seed and pollen parents.• Label the pollinated flower and enter the cross in your record book.

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• Three or four days later, repeat step 7. Be sure to use the samepollen parent that you used the first time. Replace the bag on theseed parent.

• Remove the bag from the seed parent about a week after the finalpollination.

• After the seed head has dried, harvest and store the seed.

BREEDING TECHNIQUES

EQUIPM ENT

The equipment required for plant breeding is relatively inexpensive andeasy to use.

Here are some items that you may find useful:• Magnifying glass (10 or 15 power).• Tweezers.• Small sharp–pointed scissors.• Camel–hair brush.• Small containers or vials.• Alcohol.• Rubber bands or soft wire.• Paper or cellophane bags.• Paper clips.• Tags.• Notebook.

When to Breed

Prepollination steps generally should begin just before the flower opens.If you wait until after it opens, it may be pollinated by natural means, whichwould make the flower useless for experimentation.

Since most plants bloom over a period of time, even if some of the flowerson a plant have bloomed before you have prepared them for breeding, therewill probably be others that haven't opened yet and that you can stillexperiment with.

Extremely high temperatures or moist conditions are harmful to pollen.For best results you should pollinate plants on dry days during the cool hoursof the morning or as soon as the anthers have split open.

Selecting the Parents

Some plants have natural barriers to cross–or self–pollination. It isadvisable to check for this before you begin breeding, for although youfrequently can overcome these barriers, some plants cannot be artificiallypollinated. An example of a barrier that can be overcome is the natural cross–

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pollination prohibitor of snapdragons; snapdragon flowers are constructedso as to prevent entry of wind–borne pollen from other flowers. It is possible,however, to open the flower by hand. An example of a barrier that cannot beovercome is the self–pollination prohibitor of some orchids; the stigmas ofcertain orchids produce a substance which kills the pollen of flowers of thesame plant. The mechanism that performs this cannot be removed withoutdestroying the pistil.

The plants you select for breeding should be sturdy and healthy. It isusually easier to tell which ones are healthy after a few flowers on the planthave bloomed.

In choosing a pollen parent, select one that has a heavy yellow powderon the anther. This powder is the pollen. If you brush your fingernail againstthe anther, a trace of pollen should adhere to your nail. A fresh flower is morelikely to have healthier pollen than one that has started to wilt or dry out. Inchoosing a seed parent, examine the stigma.

It should have either a glistening substance on it that is sticky to the touchor a "hairy" surface. It is this substance or surface that retains the pollen, thusmaking fertilization possible. Once you have selected the pollen and seedparents, you are ready to begin pollination.

Prepollination Steps

The first step is to mark those flowers that are to serve as pollen parentsand those that are to serve as seed parents. This can be done with colouredthread, one colour for the male and another colour for the female. Or you canuse paper labels, covered with varnish to protect them from the weather. Someplant breeders use bands designed for marking chickens.

Fig. Emasculation is Necessary to Prevent Self–Pollination.Remove all Anthers or Stamens.

Your next step is to protect the plant from unwanted pollen. If the plantis to be cross–pollinated, the stamens will have to be removed to prevent thepossibility of selfing. The removal of the stamen is called emasculation. Itshould be performed before the anthers split open to release pollen. This mayrequire opening the flower by hand before it is ready to bloom.

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Emasculation can be accomplished by:• Pinching off the stamens or anthers with tweezers.• Snipping off the stamens or anthers with sharp–pointed scissors.• Removing the petals to which the stamens are sometimes attached.

A magnifying glass will be particularly useful in emasculation.Both the seed and pollen parents should be protected from contamination

by foreign pollen. This can be done by one of the following methods:

Closing the Flower

In many flowers, such as morning-glories, petunias, and lilies, the petalscan be closed around the floral organs with a piece of soft wire, string, orrubber band. Care should be taken not to tear the petals.

Covering the Flower

Some flowers, such as composite flowers, cannot be closed. To protectthem from unwanted pollen, you can cover the flower with a paper bag. Or,if you wish to observe the flower at all times, you can cover it with a cellophanebag. The bag should be held securely in place with a paper clip or string.

Fig. To Protect the Flower from Unwanted Pollen, close it with a String or Soft Wire(Left) or Cover it With a Bag (Right). A Cellophane or Plastic Bag will Permit you to

Observe the Flower at all Times.

Flowers that are to be self–pollinated should likewise be protected fromforeign pollen by either closing or covering the flower. If the plant is grownindoors there is little likelihood of contamination by foreign pollen and youdo not have to cover or close the flower. However, indoor as well as outdoorplants require emasculation to avoid self–pollination.

POLLINAT ION ST EPS

Crossing

There are several methods that can be used for crosspollinating flowers.Here are four of the most common methods:1. Remove the stamens from the pollen parent with tweezers. Place

the stamens in a small container. Remove the protector from theseed parent. Holding a stamen with the tweezers, gently brush theanther across the stigma. Replace the protector.

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Fig. One Way to Cross–Pollinate is to Remove the Stamens from the Pollen Parentand then, holding a Stamen with Tweezers, Brush the Anther Across the Stigma of

the Seed Parent.

2. Cut the flower that is serving as the pollen parent. Remove theprotector from the seed parent. With tweezers remove a stamenfrom the pollen parent and brush an anther gently across the stigmaof the seed parent. Replace the protector.

3. With a camel–hair brush, transfer the pollen from the anthers ofthe pollen parent into a small container. Remove the protector fromthe seed parent and brush the pollen across the stigma. Replace theprotector.

Fig. If the Stamens are too Small or Too Difficult to Grasp, you Can Cross–Pollinateby Transferring the Pollen from the Anthers with a Brush into a Container and then

Brushing the Pollen onto the Stigma.

4. Shake the bagged pollen parent so that the pollen is collected inthe bag that is covering it for protection. Remove the bag from thepollen parent, being careful not to spill the pollen. Remove the

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protector from the seed parent and place the bag containing thepollen over the seed parent and shake the bag so that pollen fallson the stigmatic surfaces. This is usually done on corn.

Each time you use different pollen, be sure to first wash with alcohol thecamel–hair brush, tweezers, and any other item which might have touchedsome pollen. This step is very important to prevent pollination of the seedparent with unwanted pollen that has adhered to the equipment. After youwash the instruments be sure that they are dry before using them again.

Selfing

Procedures for self–pollinating flowers will depend on the type of flower.For perfect flowers, your job is done once you have closed or covered theflower, although you can sometimes help the pollen land on the stigma byshaking the flower once a day for several days after the pollen develops. Onlythose composite flowers containing both disc and ray florets can be self–pollinated.

Since they have both pistils and stamens, they can be selfed in the sameway as perfect flowers. With imperfect flowers, you will be able to self onlythose flowers that are on the same plant. In selfing imperfect flowers, the pollenfrom the staminate flower must be transferred to the stigma of the pistillateflower on the same plant. To do this you can use any of the methods givenabove for cross–pollination.

OBSERVING T HE OUT COM E

Once pollination has been completed, a period must elapse during whichfertilization and subsequent development of the fertilized ovule take place.When the seed is completely developed, it should be harvested.

In plants grown for flowers, harvest the seed as soon as the seed pod isdry or when it just starts to split open. In fruits and vegetables the seed willbe fully developed and ready for harvesting when the seed–bearing parts havereached maturity. As you harvest the seed, place them in packets bearing theassigned offspring number. Be careful to keep seed from different crosses andselfs in separate packets. Air–dry the seed in a fairly warm spot for a week orso and then store them in a cool, dry place. As soon as practicable, the seedshould be planted and the results entered in your record book. The offspringcan then be used for further study.

Any seeds that are not planted should be saved in case of crop failure.An important point to remember is that if both parents are pure, segregationusually will not become apparent until the F2 generation, and you may haveto breed several generations before you can obtain a plant that will breedtrue.

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10

Forestry Horticulture System

PHARMACEUTICALS

BERBERINE

Berberine is an isoquinoline alkaloid which is distributed in roots of Coptisjaponica and cortex of Phellondendron amurense. Berberine chloride is usedfor intestinal disorders in the Orient and it takes 5 to 6 years to produce Coptisroots as the raw material.

Mitsui Petrochemical has improved the productivity, and found thataddition of 10-8 M gibberellic acid into the medium stimulated berberineproductivity up to 1.66 g per L of the medium. Using a cell sorter andprotoplasts of C. japonica, they selected many higher alkaloid-producing celllines. Thus, the Mitsui group produces berberine in a large scale at a level of1.4 g per L of their optimized medium within 2 weeks. Furthermore, theyestablished a "high-density cell culture" process to produce berberine muchmore efficiently. In order to achieve a cell mass of 70 g/L on a dry weightbasis, stirring without damaging the cells, supply of sufficient amounts ofoxygen, and that of appropriate nutrients were optimize. As a result, the yieldsreached 70 g per L of cell mass and 0.45 g/day of berberine. The continuousculture with high cell density was also conducted successfully.

Addition of a polyamine, spermidine, was found to stimulate theproduction of berberine by Thalictrum minus cell suspension cultures by Haraet al. in Tabata's laboratory although other polyamines such as cadaverine,putrescine and spermine were ineffective. They indicated that spermidineeffected an increase of ethylene generation which was associated withactivation of berberine synthesis. The maximum stimulative effect wasobtained by addition of 2 mM spermidine.

ALKALOIDS

A variety of alkaloids have been used as pharmaceuticals and most ofthem are plant metabolites. The typical tropane alkaloids, atropine,

hyoscyamine, scopolamine and cocaine, were widely used as blockers of theparasympathetic nervous system such as anodyne and antispasmodic.Research on production of these useful alkaloids by plant cell cultures hasbeen carried out for more than 25 years, however, industrial production hasnot yet succeeded because of low producing ability of the cultured cells. Plantsused for these studies are mainly Atropa belladonna, Hyoscyamus niger,Datura meteloids and others.

M ORPHINAN ALKALOIDS

Codeine is an analgesic and cough-suppressing drug and Papaversomniferum L.is a traditional commercial source of codeine, and morphinewhich can be converted to codeine. Mature capsules of P. bracteatumaccumulates up to 3.5% of thebaine which also can be converted to codeine.Although many researchers have tried to produce codeine by undifferentiatedcells of these plants, little success has been achieved.

It is indicated that morphogenetic differentiation from cultured cells ofP. bracteatum was prerequisite for producing higher levels of thebaine. Theresearchers also showed the similar results as Kamimura's report using P.bracteatum and P. somniferum. For example, it is reported that cells of P.somniferum produced norsanguinarine, sanguinarine, cryptopine and othervarious alkaloids, but not codeine and morphine.

Concerning sanguinarine, shows that fungal mycelium of Botrytis sp.elicited production of sanguinarine by P. somniferum cells. The level of thisalkaloid increased 26 times in the presence of the elicitor, 29% of the dry cellweight, compared to the medium without it. Since the alkaloid extracted fromintact plants is added to toothpastes as an anti-plaque agent, the commercialproduction of sanguinarine by cell culture technology was intensivelyinvestigated by Kurz's group at the National Research Council of Canada anda company in the U.S. several years ago, but the process has not yet beencommercialized.

The efficient production of thebaine and codeine using cell culture systemsby de novo synthesis was not successful, therefore studied thebiotransformation of codeinone to codeine using immobilized cells of P.somniferum. The conversion yield was 70.4% and about 88% of codeineconverted was excreted into the medium.

T ROPANE ALKALOIDS

Scopolamine and hyoscyamine are being used commercially as anestheticand antispasmodic drugs. These alkaloids occur in leaves of (Solanaceae)plants including D. myoporoides and D. leichhardtii. Scopolia, Atropa,Hyoscyamus and Datura also contain tropane alkaloids.

Studies on production of tropane alkaloids by plant tissue cultures havebeen actively carried out by many researchers since West et al. found tropane

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alkaloids in an Atropa belladonna callus more than 30 years ago. However,the concentrations of scopolamine and hyoscyamine in cultured cells aregenerally very low in spite of many efforts to increase the yield using variousapproaches. Therefore, the plant cell culture has not yet been employed tomanufacture these alkaloids. For example, Tabata et al. added tropic acid intoScopolia japonica suspension cultures as a precursor and could increase thelevel of alkaloids up to 15 times. Mitsuno et al. (159) selected a high tropanealkaloid producing strain of Hyoscyamus niger which produced about 7 timesmore hyoscyamine (13.9×10-3% fresh cells), than that of the parent strain.According to their results, there was no direct correlation between highproducing ability and variation of chromosomal numbers.

Cardinolides

Cardiac glycosides or cardenolides are products of Digitalis species. Someof these compounds have been employed in treatment of heart diseases.Chemical structures of the major aglycone of cardenolides and various sugarresidues link the 3-hydroxy group of aglycones. For commercial productionDigitalis plants are being cultivated in the fields of several countries includingthe Netherlands, Hungary and Argentina.

Although there are a number of papers describing production of cardiacglycosides in Digitalis tissue cultures, generally the yield was very low, andmoreover, during the successive transfers of the cultured cells the amount ofcardenolides often decreased and disappeared completely.

Instead, many researchers indicated that morphological differentiationcaused an increase in productivity. For example, it shows that organ culturesof D. lanata leaves and roots produced cardenolides and during the cultivationthe level of digoxin in the tissues rose with increasing age. Hirotani and Furuyaalso found that renewed organ differentiation from callus tissues of D.purpurea led to a new formation of cardinolides.

Nutritional factors including growth regulators, sugars, nitrogen sources,vitamins and so on in the medium affect on differentiation of shoots and otherorgans and the secondary metabolites such as cardiac glycosides are oftensynthesized. The differentiated tissue culture is prerequisite for secondarymetabolites production in some cases, however in general, the method requiresmuch longer culture time than the suspension cell culture and consequentlyit is not efficient.

Various secondary metabolites other than digitoxin and digoxin have beenfound in callus tissues of D. lanata and D. purpurea. These include cholesterol,campesterol, stigmasterol, -sitosterol, 4-hydroxy-digitolutein and others. Itis found that the levels of those secondary products tended to decrease overseveral passages in cultivation. In contrast to the de novo synthesis, thebiotransformation process with Digitalis plant cells seems to be morepromising from a commercial point of view. It is reported that D. lanata and

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D. purpurea callus cultures rapidly transformed progesterone to pregnane.Leaf and root cultures of D. lanata and shoot-forming callus tissues of D.purpurea accumulated an increased level of digoxin and/or digitoxin whenprogesterone was added to the cultures. Stohs and Staba studies on thebiotransformation of cardenolides by Digitalis cells and recognized that theglycosylation reaction occurred.

Among many studies, the biotransformation from digitoxin to digoxinusing Digitalis lanata cells investigated by Reinhard and Alfermann is themost interesting approach in terms of commercial application since digoxinhas a higher demand as a drug for heart diseases than digitoxin. It isadvantageous that Digitalis leaves contain a larger amount of digitoxin whichcan be used as a substrate. It is a hydroxylation reaction at the 12 -positionof digitoxin, and Reinhard et al. found that -methyldigitoxin was the mostsuitable substrate in this biotransformation as methyldigoxin is the majorproduct.

L-DOPA

L-DOPA, L-3,4-dihydroxyphenylalanine, is an important intermediate ofsecondary metabolism in higher plants and is known as an precursor ofalkaloids, betalain, melanine, and others. It is also a precursor ofcathecolamines in animals and is being used as a potent drug for Parkinson'sdisease.

It is found that the callus tissue of Mucana pruriens accumulated 25 mg/L DOPA in the medium containing very high concentration of 2,4-D. Wicherset al. immobilized cells of M. pruriens within alginate and found that the cellsproduced DOPA from tyrosine in up to 2% of dry cell weight. The DOPAsynthesized was secreted mostly into the medium.

Teramoto and Komamine induced callus tissues of Stizolobium hassjoo(Mucuna hassjoo), M. pruriens and M. deeringiana, and optimized the cultureconditions. The highest concentration of DOPA was obtained when S. hassjoocells were cultivated in MS medium containing 0.025 mg/l 2,4-D and 10 mg/lkinetin. The level of DOPA in the cells was about 80 nmol/g-f.w.

VALEPOT RIAT ES

Plants in the Valerianaceae have been used as folk medicines. For example,Nardostachys jatamansi, Valeriana wallichii and V. officinalis L. var.angustifolia have been used in India, and N. chinensis has been employed inChina for hundreds of years. Partrinia plants are also being used as sedativedrugs in former Soviet Union. Although the active principles in these plantshave not been identified, a group of compounds having biological activitiessuch as sedative, tranquilization, cytotoxicity and antitumor activities werenamed "valepotriates". Thies synthesized a series of valepotriate derivativesand tested for biological activities.

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It has investigated plant tissue cultures for producing valepotriatesbecause of their limited supply and uncertain availability. They induced callustissues of nine different species of Valerianaceae on MS media and found thatFedia cornucopiae and V. locusta cells produced higher levels of thecompounds than the intact plants. Isolation of cell lines resistant totrifluoroleucine and to nystatin, treatment with colchicine, cultivation of thecells in two-phase culture media and addition of several bioregulators werecarried out intensively.

As valepotriates have monoterpene skeletons, L-leucine was consideredas a precursor. In an isolated cell lines of V. wallichii resistant totrifluoroleucine, a leucine analog, but the yield of valepotriates in the cellswas not increased although the intracellular level of leucine was increased. Itwas reported that fungi resistant to nystatin, a polyene antibiotic, produced ahigh level of steroids.

One of the strains of V. wallichii to resistant to nystatin increased theamount of valepotriates produced up to 3 times, which was 88 mg/g-d.w. Thesame group treated a suspension culture of V. wallichii with colchicine whichwas expected to induce polyploid cells. As a result, they found that thecolchicine treated cells produced higher amount valepotriates than therespective untreated cultures. A two-phase culture provided by addition ofRP-8 into the culture medium was effective in inducing secretion of thelipophilic compounds from the cells, and the total yield of valepotriates wassubstantially increased.

Since the valepotriate skeleton is of the iridoid nature, they added someplant bioregulators such as dimethyl-morpholinium-bromide, dimethyl-piperidinium-bromide, dimethyl-piperidinium-chloride as well as 2-(3,4-dichloro-phenoxy)-triethylamine and 2-(3,5-diisopropylphenoxy)-triethylamine to cell suspension cultures of F. cornucopiae and V. wallichii.When they were employed in concentrations of 0.01 to 0.04 mmol during theearly exponential growth stages of the cells, the levels of valepotriates wereincreased significantly.

ANTITUMOR COMPOUNDS

The plant kingdom is one of the attractive sources of novel antitumorcompounds. The National Cancer Institute in the U.S., for example, hasconducted an intensive screening programme since 1955 and has identifiedvarious potent compounds from higher plants. These antitumor compoundsinclude maytansine, tripdiolide, homoharringtonine, bruceantin, ellipticine,thalicarpine, indicine-N-oxide, and baccharin. In addition to these compounds,some of the higher plant products such as vinblastine, vincristine,podophyllotoxin derivatives including etoposide, and camptothecin and itsderivative have already been marketed as very important anticancer drugs.

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Taxol from Taxus brevifolia and related plants, is one of the most exitingcompounds and was marketed in 1992. However, the concentrations of theseactive compounds in plants are generally low, the growth rate of the plants isslow and the accumulation pattern of these compounds is highly susceptibleto geographical or environmental conditions. Therefore, it is not an easy taskto produce economically these compounds by extraction from intact plants.

Camptothecin

Camptotheca acuminata, a native of North China, was found to producea potent antitumor alkaloid, camptothecin, by Wall et al. It is highly active inWalker 256 rat carcinosarcoma and mouse leukemia. The clinical trials inpatients with gastrointestinal cancer were at first very promising butsubsequent trials showed toxicity.

Sakato and Misawa (184) induced callus from the stem of C. acuminataon MS solid medium containing 0.2 mg/L 2,4-D and 1 mg/L kinetin. The calluswas transferred to the liquid medium. Gibberelin, L-tryptophan andconditioned medium stimulated growth of the cells. After 15 days ofcultivation in suspension, the con-centration of camptothecin in the cells was0.0025% on a dry weight basis, which was about 1/20 of the level in the intactplant. A.J. van Hengel et al. in the Netherlands established the suspensionculture system of C. acuminata and detected camptothecin in the culturedcells using TLC, HPLC and GC-MS. The highest level, 0.998 mg ofcamptothecin per litre of the medium, was accumulated in the cells cultivatedin MS medium containing 4 mg/L NAA.

Homoharringtonine

Homoharringtonine together with harringtonine and isoharringtonine,were isolated from Cephalotaxus harringtonia by Powell et al. These alkaloidsare complex esters of the inactive alcohol, cephalotaxine. These compoundsinhibit the growth of murine leukemias, L1210 and P388, and KB cells.Homoharringtonine also shows activity against colon tumors, melanoma andleukemia in mice.

Since the chemical synthesis of homoharringtonine was not efficient forcommercialization, studies of tissue culture were conducted by Delfel andhis colleagues. They detected about 5-10 mg of the alkaloids per kg dry wt. ofthe callus tissues cultivated for 3 to 6 months. The levels were approximately1 to 3% of the concentrations found in the parent plant. These products werecephalotaxin, homoharri-ngtonine, harringtonine and isoharringtonine.

MS medium containing 1 mg/L kinetin and 3 mg/L NAA was favourableto induce the callus from C. harringtonia, while the medium without growthregulators strongly promoted organogenesis. The radioimmunoassayestablished by the same group showed the levels of cephalotaxine and its esterswere only 1/300 of the intact plant.

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Taxol

Under the intensive NCl screening programme of antitumor compoundsin the U.S., Wall et al. began to isolate an active principle against KB cellsfrom a tree, Taxus brevifolia in 1965. In 1969 pure taxol was first isolated andits chemical structure was disclosed in 1971. It is a diterpene amide and hasshown against B16 mouse melanoma tumor, the MX-1 human mammaryxenograft and CX-1 colon xenografts. The mode of action of taxol is ratherunique because it stabilizes microtubles and inhibits depolymerization. Theclinical trials begun in 1983 have shown positive results in the treatment ofadvanced ovarian cancer and breast cancer as well.

The FDA in the U.S. has approved taxol (generically known as paclitaxel)at the end of 1992; Bristol-Myers Squibb produces the drug for use in thetreatment of ovarian cancer in patients who have failed to respond to otherchemotherapies. Taxol is now being manufactured by extraction from the barkof wild-grown T. brevifolia trees. The demand for taxol will be undoubtedlyincreasing since it will be applied for other cancers including breast and lungcancers in the near future, but its supply is limited. Other related plants suchas T. canadensis and T. cuspidata also contain taxol and other relatedcompounds. Generally the concentration of taxol in Taxus plants are very low.Therefore, harvesting Taxus trees for production of taxol commercially causesa serious problem in the U.S. from the environmental point of view.

As alternative ways, plant tissue and cell cultures as well as chemicaltransformation processes from baccatin extracted from needles of the plantand the total chemical syntheses have been investigated in many researchgroups. In the U.S., Phyton Catalytic Inc. and ESCAgenetics announced acouple of years ago that they were establishing plant cell culture processes tomanufacture taxol. The latter showed a photograph of a vial containing taxolpowder produced by plant cell cultures. However, both companies have neverpublished details of the plant cell culture procedure in scientific periodicalsor at any meetings, although ESCAgenetics described some of their resultsusing a couple.

A couple of patents have been published. For example, a U.S. Patent filedby A.C. Christen et al. of U.S.D.A. described that the tissue of T. brevifoliahad been successfully cultured to produce taxol, related alkaloids and alkaloidprecursors, but the patent claims are exceedingly broad and identity of whatthe actual process was remains difficult to ascertain.

Shuler et al. at Cornell University showed that a cell line of T. brevifoliain suspension provided by USDA-Agriculture Research Station and PhytonCatalytic, produced 3.9 mg/L taxol in the medium after 26 days of culture.The level of taxol was determined by reverse-phase HPLC. They found thatfresh cell weight increased by 4-fold after a lag phase of about 4 days andthat taxol was first seen at 13 days, and increased sharply after 20 days. It isof interest that all taxol produced was secreted into the medium, which was

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very unusual for plant cell cultures. Wickremesinhe and Artheca of thePennsylvania State University established callus cultures and suspensioncultures from T. brevifolia cv Repandens, T. cuspidata, T. media cvs. Hicksiiand Densiformis. Though some cell lines grew fast and their doubling timeswere 9-14 days, the levels of taxol were too low for commercial production.Induction of the hairy roots of Taxus plants has been tried.

DiCosmo and his colleagues of the University of Toronto in collaborationof Nippon Oil Co. Ltd. in Japan described that they detected and identifiedtaxol in the callus of T. cuspidata and its level was 0.02+0.005% in dry weightafter 2 months in culture. Suspension cultures of T. cuspidata were alsoestablished from the callus cultures and subsequently immobilized onto glassfibre mats. The cells were maintained as immobilized cultures for 6 months.The level of taxol in the immobilized cells was 0.012+0.007% of the extracteddry weight.

Taxol is one of the most suitable and desirable plant products to plantcell culture research because the shortage of its supply and its high-value.Bristol-Myers Squibb is therefore, financially supporting many research groupsto establish the process of cell culture production of taxol.

The root of Panax ginseng C.A. Meyer, a perennial herb, so-called"ginseng" has been widely used as a tonic and precious medicine since ancienttimes particularly in oriental countries including Korea and China. It iseffective for gastroenteric disorders, diabetes and weak circulation, and hasbeen used as an adjuvant to prevent various disorders, rather than a medicineto cure disorders. Thus, ginseng has been recognized as a miraculous medicinein preserving health and longevity. It has been known that the root containsvarious saponins and sapogenins. Among them, ginsenoside-Rb has a sedativeactivity, while Rg has stimulative activities.

Although there are several species of ginseng, the commercially importantspecies, P. ginseng grows in an area of 30-48? north latitude such as Korea.The cultivation of ginseng in the field requires four to seven years, and it isimpossible to plant consecutively for 20 to 50 years but the demand for theplant has increased dramatically in the world, and its price has soared. Theseare reasons why many researchers have tried to produce ginseng cells throughplant tissue cell cultures.

Furuya et al. at Kitasato University have studied P. ginseng callus tissuessince the early 1970's. Meiji Seika Kaisha in Japan investigated the large-scaleproduction of the cells established by Furuya using various types offermentors. According to their patent published in 1973, crown gall calli, callustissues and redifferentiated roots of P. ginseng were able to accumulatesaponins and sapogenins known from the intact plant. The callus tissues androots were cultivated on both MS solid and liquid media containing vitamins,sucrose, 2,4-D and suitable natural nutrients such as soybean powder or beefextract for several weeks at 25-28 C. The concentrations of crude saponins in

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the callus (21.1%), in the crown gall (19.3%) and in the redifferentiated root(27.4%) were much higher than those in the natural root (4.1%). The saponinswere found to contain ginsenoside-Rb and -Rg.

It is obtained cultured cells of P. ginseng containing ginsenosides. Choiof Korea Ginseng and Tobacco Research Institute has investigated in vitroculture of P. ginseng extensively. He indicated that plant growth regulatorssuch as 2,4-D and kinetin in the medium affected the levels of saponins incallus and suspension cultured cells. For example, 3.62% of total saponinswas detected in the callus cultivated in MS medium containing 5 mg/L 2,4-Dand 1 mg/L kinetin, while 8.78% was produced in 10 mg/L 2,4-D and 1 mg/Lkinetin medium.

After the Meiji Seika abandoned development of the ginseng project,another Japanese company, Nitto Denko Corporation, constructed a 20 KLfermentor to scale-up cultivation of ginseng cells in collaboration with Furuyain the middle of the 1980's. The company has optimized various environmentalconditions using 30 L jar fermentors for the cell line having partlydifferentiated tissues originally induced. Glucose in the medium promotedcell growth in the initial stages of the fermentation and sucrose fed duringthe growth cycle stimulated the productivity of saponins. Although a higherNO2/NH3 ratio was favourable to the growth, it decreased the concentrationof saponins in the cells. The growth was suppressed by moderate agitation,but the yield of saponins increased. The highest cell mass, 19 g/L on a dryweight basis was obtained using a 2 KL fermentor and the production rate ofthe cell mass was approximately 700 mg/L/day.

The chemical variation between mother plants of P. ginseng and its tissuecultured cells. Ushiyama indicated that their cultured cells contained basicallythe same constituents as those in the intact plant. Acute virulence tests, Amestests and dietary tests with livestock feed containing 12.5% of dried culturedcells for 6 months did not show any abnormalities in animals.

Podophyllotoxin

Podophyllum pelatatum, May apple, which is a common herb in easternNorth America contains an antitumor lignan, podophyllotoxin. It is active toKB cells and is used against certain virus diseases and skin cancer. A semi-synthetic derivative of podophyllotoxin, etoposide (V-16), was found to beactive against brain tumor, lymphosarcoma and Hodgkins' disease and wasapproved by the FDA in the U.S. Bristol-Myers Squibb is one of the largestmanufacturers of the drug. Production of podophyllotoxin by P. pelatum cellcultures was first attempted by Kadkade and he found that a combination of2,4-D and kinetin in the medium supported the highest amount of itsproduction. Red light stimulated the production. The Nippon Oil inducedembryogenic roots from a callus of the plant in a liquid MS mediumsupplemented 1 mg/L NAA, 0.2 mg/L kinetin and 500 mg/L casein

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hydrolysates. The roots were then transferred to the medium without growthregulators. They detected 1.6% of podophyllotoxin in the dried tissues, whichwas 6 times higher level than that in a mother plant.

To increase the yield of podophyllotoxin, Woerdenberg et al. in theNetherlands added a complex of a precursor, coniferyl alcohol, and ß-cyclodextrin to Podophyllum hexandrum cell suspension cultures. Additionof 3 mM coniferyl alcohol complex gave 0.013% podophyllotoxin of the cellson a dry weight basis but the cultures without the precursor produced only0.003%. -D-glucoside of coniferyl alcohol, coniferin, was a more potentprecursor in terms of the yield of the anticancer compound (0.055%), butunfortunately this compound is not commercially available. The same authorsreported that cell suspension cultures of Callitris drummondii (conifer) alsoaccumulated podophyllotoxin--D-glucose. In the dark, the cells producedapproximately 0.02% podophyllotoxin of the dry cell mass and 85-90% of thelignans were the -D-glucoside form, while in the light the yield ofpodophyllotoxin--D-glucose increased to 0.11%.

It is reported that callus tissues and suspension culture cells of Liliumalbum produced podophyllotoxin. One of the cell lines produced 0.3%podophyllotoxin of dried cells together with small amounts of5-methylpodophyllotoxin, lariciresinol and pinoresinol after 3 weeks ofcultivation. The callus tissue induced from P. hexandrum was reported byHeyenga to produce podophyllotoxin, 4'-demethyl-podophyllotoxin andpodophyllotoxin-4-0-glucoside when the callus was incubated in B5 mediumcontaining 2,4-D, gibberellic acid and 6-benzylaminopurine. The levels ofpodophyllotoxin and its derivatives were similar to those in the mother plant.

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Horticultural Plant Biotechnology

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