Genome analysis of medicinal Ganoderma spp. with plant-pathogenic and saprotrophic life-styles

Phytochemistry xxx (2015) xxx–xxx

Contents lists available at ScienceDirect

Phytochemistry

journal homepage: www.elsevier .com/locate /phytochem

Genome analysis of medicinal Ganoderma spp. with plant-pathogenicand saprotrophic life-styles

http://dx.doi.org/10.1016/j.phytochem.2014.11.0190031-9422/� 2014 Elsevier Ltd. All rights reserved.

⇑ Corresponding author.E-mail address: [email protected] (C. Liu).

1 Contributed equally to this work.

Please cite this article in press as: Kües, U., et al. Genome analysis of medicinal Ganoderma spp. with plant-pathogenic and saprotrophic life-styletochemistry (2015), http://dx.doi.org/10.1016/j.phytochem.2014.11.019

Ursula Kües b,1, David R. Nelson c,1, Chang Liu a,⇑,1, Guo-Jun Yu d,e,1, Jianhui Zhang a, Jianqin Li a,Xin-Cun Wang a, Hui Sun d,e

a Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences & Peking Union Medical College, 151 Malianwa North Road, Haidian District, Beijing 100193, Chinab University of Göttingen, Büsgen-Institute, Department for Molecular Wood Biotechnology and Technical Mycology, Büsgenweg 2, D-37077 Göttingen, Germanyc Department of Microbiology, Immunology and Biochemistry, University of Tennessee Health Science Center, 858 Madison Ave., Memphis, TN 38163, USAd State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, Chinae Key Laboratory of Combinatorial Biosynthesis and Drug Discovery (Ministry of Education), Wuhan University, Wuhan, China

a r t i c l e i n f o

Article history:Available online xxxx

Keywords:GanodermaWhole genome sequencingMitochondrial genomeTranscriptomeCYP450matAmatB

a b s t r a c t

Ganoderma is a fungal genus belonging to the Ganodermataceae family and Polyporales order. Plant-path-ogenic species in this genus can cause severe diseases (stem, butt, and root rot) in economically impor-tant trees and perennial crops, especially in tropical countries. Ganoderma species are white rot fungi andhave ecological importance in the breakdown of woody plants for nutrient mobilization. They possesseffective machineries of lignocellulose-decomposing enzymes useful for bioenergy production and biore-mediation. In addition, the genus contains many important species that produce pharmacologically activecompounds used in health food and medicine. With the rapid adoption of next-generation DNA sequenc-ing technologies, whole genome sequencing and systematic transcriptome analyses become affordableapproaches to identify an organism’s genes. In the last few years, numerous projects have been initiatedto identify the genetic contents of several Ganoderma species, particularly in different strains of Ganoder-ma lucidum. In November 2013, eleven whole genome sequencing projects for Ganoderma species wereregistered in international databases, three of which were already completed with genomes being assem-bled to high quality. In addition to the nuclear genome, two mitochondrial genomes for Ganoderma spe-cies have also been reported. Complementing genome analysis, four transcriptome studies on variousdevelopmental stages of Ganoderma species have been performed. Information obtained from these stud-ies has laid the foundation for the identification of genes involved in biological pathways that are criticalfor understanding the biology of Ganoderma, such as the mechanism of pathogenesis, the biosynthesis ofactive components, life cycle and cellular development, etc. With abundant genetic information becom-ing available, a few centralized resources have been established to disseminate the knowledge and inte-grate relevant data to support comparative genomic analyses of Ganoderma species. The current reviewcarries out a detailed comparison of the nuclear genomes, mitochondrial genomes and transcriptomesfrom several Ganoderma species. Genes involved in biosynthetic pathways such as CYP450 genes andin cellular development such as matA and matB genes are characterized and compared in detail, as exam-ples to demonstrate the usefulness of comparative genomic analyses for the identification of criticalgenes. Resources needed for future data integration and exploitation are also discussed.

� 2014 Elsevier Ltd. All rights reserved.

1. Introduction which are characterized by unique double-wall basidiospores with

1.1. Importance of Ganoderma

Ganoderma P. Karst. is an important fungal genus consisting of amorphologically diverse assemblage of mushroom taxa, all of

ornamented endospores. Although a mere 80 species are recog-nized by the Dictionary of Fungi (Kirk et al., 2011), there are 427name records in Index Fungorum (http://www.indexfungorum.org/). Species of Ganoderma have attracted world-wide attentionfrom three aspects: as therapeutic fungal bio-factories (Paterson,2006), as plant pathogens (Hushiarian et al., 2013) and as‘‘bio-bags’’ of ligninolytic enzymes (Zhou et al., 2013). It is evidentfrom the literature that the interests in particular aspects are fre-quently associated with particular geographic areas and climates.

s. Phy-

http://www.indexfungorum.org/

http://www.indexfungorum.org/

http://dx.doi.org/10.1016/j.phytochem.2014.11.019

mailto:[email protected]


http://www.sciencedirect.com/science/journal/00319422

http://www.elsevier.com/locate/phytochem


2 U. Kües et al. / Phytochemistry xxx (2015) xxx–xxx

For example, species of Ganoderma (represented by G. lucidum(Curtis) P. Karst. and G. sinense J.D. Zhao, L.W. Hsu & X.Q. Zhang)are used as highly valued traditional medicines in East Asian coun-tries including China, Japan and Korea. (A paper on an updatednomenclature of these species is presented in this Special Issue(in this issue) and also refer to the Editorial (in this issue).)Scientific reports produced from these areas therefore mostly focuson the active chemical components and therapeutic effects ofGanoderma. In contrast, most reports on Ganoderma as plant patho-gens come from South Asia including India and Pakistan, fromIndonesia and Malaysia in Southeast Asia, and North America butthere are also notable incidences in Africa. The reports regardingthe potential use for bio-energy production are mostly associatedwith the US and East Asian areas. One possible reason is thatGanoderma is a genetically highly diverse group of fungi andGanoderma spp. from different geographical areas might be genet-ically distinct and possess unique properties (Flood et al., 2000).

As therapeutic fungal biofactories, Ganoderma spp., representedby G. lucidum and G. sinense, were found to produce a wealth of bio-active polysaccharides, oligosaccharides, triterpenoids, peptidesand proteins alcohols and phenols. Fruiting bodies and myceliumare furthermore worthy sources of terpenoids, mineral elements,vitamins and selections of amino acids. Health enhancing proper-ties of Ganoderma spp. have variously been reported in preventionand treatment of objectionable and ill-fated conditions listed as‘‘hepatopathy, chronic hepatitis, nephritis, hypertension, hyperlip-idemia, arthritis, neurasthenia, insomnia, bronchitis, asthma, gas-tric ulcers, atherosclerosis, leucopenia, diabetes and anorexia’’(Rai et al., 2005; Xu et al., 2011). However, the anti-cancer andimmuno-modulation properties of G. lucidum are the most studiedeffects, in particular the cytotoxic and apoptotic actions of b-glu-cans and ganoderic acids as a class of terpenoid compounds spe-cific to Ganoderma mushrooms (Boh, 2013; Jin et al., 2012; Xuet al., 2012, 2011). The chemical components, biological activitiesand mechanisms of action are topics of intensive research andinterested readers may consult several recently published excel-lent reviews (Habijanic et al., 2013; Lee et al., 2012; Popovicet al., 2013; Soares et al., 2013; Wu et al., 2013).

As plant pathogens, Ganoderma spp. have been observed togrow on various dead or dying trees and shrubs in the NorthernTerritories of Australia, including ornamental palms (such as thegolden cane palm and Carpentaria acuminata) and broadleavedtrees (amongst others Brachychiton populneus, Delonix regia, andCasuarina equisetifolia), crop perennials (e.g. Citrus species, guava,custard apple), and several other woody species valued for theirproducts (wood, seeds, gum, fragrances, bioactive compounds,etc.) and their ecological importance (e.g. Ficus and Acacia species,white cedar), (Hennessy and Daly, 2007). In the USA, there aremany different species of Ganoderma, of which G. applanatum (Pers.ex Wallr.) Pat. is reported to kill aspen in the western states (USForest Service, 2011) and G. zonatum Murrill is considered to be apathogen of mature palms growing in southern regions (Elliottand Broschat, 2000). In southeastern countries, such as Malaysiaand Indonesia, G. boninense Pat. is a major pathogen on oil palmsthat eventually kills the trees. The economic loss caused by thispathogen is estimated to be up to 500 million USD a year(Hushiarian et al., 2013). In Mexico, G. oerstedii (Fr.) Murrill wasidentified to be a tree parasite species (Mendoza et al., 2011). Adetailed evaluation of records on Ganoderma disease of perennialcrops in India concludes that members of the G. lucidum and G.applanatum species complexes are widespread pathogens in thesubcontinent (Sankaran et al., 2005). Likewise, attacks of G. applan-atum (Pers. ex Wallr.) Pat. and G. lucidum have been documentedfor Pakistan on species of Pinus, Dalbergia, Artocarpus, Morus,Cedrus, Melia, Quercus, Populus and other trees (Nasir, 2005).Incidences of Ganoderma spp. infestations of oil palms causing

Please cite this article in press as: Kües, U., et al. Genome analysis of medicinatochemistry (2015), http://dx.doi.org/10.1016/j.phytochem.2014.11.019

basal stem rots are also known from many tropical African coun-tries (Miller et al., 2000). Stem, butt and root rot diseases are typ-ical results from Ganoderma attacks (Nasir, 2005; Elliott andBroschat, 2000). Loss of foliage of the stressed trees and die-backof individual branches can be symptoms in stages of infectionsprior to tree death (Hennessy and Daly, 2007; Paterson, 2007;Hushiarian et al., 2013). Main routes of infections may be throughroots in the soil by vegetative spread (Irianto et al., 2006;Hushiarian et al., 2013) but entry by spores through wounds is alsoconsidered (Paterson, 2007; Rees et al., 2012). To manage the dis-ease, different techniques have been attempted with varying suc-cess, such as soil mounding, surgery and removal of diseasedmaterial, isolation trenching, ploughing and harrowing, fallowing,chemical treatment, application of fertilizers, biological controland selection of resistant planting materials (Hushiarian et al.,2013). Unfortunately, measures to cure the disease or to at leasteffectively stop its spread have not been found. Current diseasemanagement relies on preventive measures reducing the frequencyof devastating infections (Paterson, 2007; Hushiarian et al., 2013).

The molecular basis of Ganoderma spp. being plant pathogens istheir ability, as white-rot fungi, to degrade lignocellulose (Paterson,2007). In nature, Ganoderma spp. as saprotrophs have their impor-tant ecological position in nutrient mobilization from dead wood byenzymatic decomposition (Clinton et al., 2009). The same abilitycan also be employed to applications where degradation of ligno-cellulose is favorable, such as bio-energy production, treatment ofwastewater and bioremediation (da Coelho-Moreira et al., 2013).The ability of Ganoderma species to degrade lignocellulose hasgained significant attention starting from the 1980s. Several studieshave shown that Ganoderma spp. have strong abilities to enzymat-ically degrade lignocellulose (Adaskaveg et al., 1990; Maeda et al.,2001; Silveira Carneiro et al., 2009; Martínez et al., 2011; Sonet al., 2010; de Andrade et al., 2012). The enzymes that participatein lignin degradation include lignin peroxidase (LiP), manganese-dependent peroxidase (MnP), laccase (Lac) and oxidases producinghydrogen peroxide, such as glyoxal oxidase (GLOX) and aryl-alcoholoxidases. A large number of studies have been conducted to identifyenzymes and their genes that are potentially involved in lignin deg-radation from Ganoderma spp. (Zhou et al., 2013). Screening forGanoderma spp. with stronger LME (lignin-modifying enzyme)activities, good stability and ability to endure extreme conditionsare active areas of research.

1.2. Benefits of whole genome sequencing projects

The research studies in the above mentioned areas have runinto several problems. Foremost is the confusion over the identityof Ganoderma spp. For example, the Ganoderma spp. used astraditional medicines are called ‘‘lingzhi’’ in China; Munnertake,Sachitake and Reishi in Japan, and Youngzhi in Korea. It has beendifficult to determine whether or not these represent thesame species. Actually, the Chinese pharmacopeia (ChinesePharmacopoeia Commission, 2010 Edition) specifically explainsthat ‘‘lingzhi’’ refers to both G. lucidum and G. sinensis. Withoutan unambiguous determination of the species identity, comparisonof chemical components and therapeutic effects obtained from var-ious studies would be difficult. Similarly, a major problem in con-trolling Ganoderma caused disease is confusion over the identity ofthe species causing the disease and the lack of effective tools forearly disease detection under field conditions (Flood et al., 2000;Paterson, 2007; Hushiarian et al., 2013; Naher et al., 2013). Whileseveral markers such as the intergenic spacer regions (ITS) for theribosomal genes have been used for DNA barcoding analysis (Smithand Sivasithamparam, 2000; Moncalvo and Buchanan, 2008), theyare not always suitable for the determination of Ganoderma spp.(Glen et al., 2009). The inter-specific and intra-specific variations

l Ganoderma spp. with plant-pathogenic and saprotrophic life-styles. Phy-


U. Kües et al. / Phytochemistry xxx (2015) xxx–xxx 3

of Ganoderma are largely unknown (Pilotti et al., 2003; Pilotti,2005; Rakib et al., 2014), thus a particular marker might not haveenough distinguishing power to discriminate closely relatedspecies. Furthermore, it is unknown whether or not such molecularmarkers are associated with particular host ranges (Nusaibah et al.,2011) or, for example, with particular biochemical properties ofinterest.

The second difficulty lies in the overall lack of informationregarding the genes responsible for secondary metabolite produc-tion, ligninolytic activities and pathogenesis. Without the knowl-edge of the complete genetic makeup of Ganoderma spp., largescale, brute-force screening or breeding of Ganoderma spp. orstrains with preferred characteristics would be tedious andcost-ineffective. A metabolic engineering approach would not beapplicable for the production of active compounds and ligninolyticenzymes. Without a gene catalog, specific drugs cannot easily bedeveloped to target control of the spread of pathogenic Ganodermaspp.

There are several benefits that whole genome sequencing pro-jects can bring, such as (species specific) marker identification,gene discovery and pathway elucidation. First and foremost, theentire genome would provide ultimate information for theselection of markers capable for species identification. Second,whole genome sequencing would be the most-cost effectivemethod for the discovery of critical genes and the elucidation ofbiosynthetic pathways for chemical compounds. Although,approaches like transcriptome profiling can also achieve the goalof gene discovery, it might not be able to detect genes expressedat lower levels and might not provide full-length information ofgenes. Furthermore, it does not provide the information regardinglocation of genes on chromosomes. Many chemical compounds infungi are synthesized by genes clustered together on the chromo-somes. As a result, co-localization of genes would be an effectivemethod to discover genes involved in the biosynthesis of particularchemical compounds (Andersen et al., 2013) and to develop strat-egies for the eventual concerted manipulation of their expression(Brakhage and Schroeckh, 2011). Third, comparative analysis ofthe genomes from multiple species can help elucidate the evolu-tionary history of critical genes (Floudas et al., 2012), such as forthe ligninolytic enzymes LiP, MnP, Lac, etc. This information canprovide guidance in screening of new species for favorable charac-teristics or for developing better strategies to combat pathogens.With rapid development of DNA sequencing technologies, wholegenome sequencing of Ganoderma spp. became affordable and adozen research projects are already on the way. This reviewintends to survey the recent progress in this area, the preliminaryresults from which would usher the research and application ofGanoderma spp. into a new era.

Table 1List of whole genome sequencing projects in NCBI’s BioProject section.

Accession Strain Status

PRJNA182009 Ganoderma tornatum strain NPG1 In progressPRJNA182007 Ganoderma zonatum strain POR69 In progressPRJNA182006 Ganoderma miniatocinctum strain 337035 In progressPRJNA182005 Ganoderma boninense strain PER71 In progressPRJDA61381 Ganoderma lucidum strain BCRC 37177 Whole genomPRJDA61379 Ganoderma lucidum strain BCRC 37180 In progressPRJNA77007 Ganoderma lucidum strain Xiangnong No. 1 High qualityPRJNA71455 Ganoderma lucidum strain G.260125-1 High qualityPRJNA42807 Ganoderma sinense strain gasi0214-1 In progressPRJNA42873 Ganoderma lucidum strain Hu-nongke No. 1 In progressPRJNA68313 Ganoderma sp. strain 10597 SS1 High quality

* BROAD: BROAD Institute of Massachusetts Institute of Technology (MIT) and Harvardof Medical Science, Beijing, China; IEF: Institute of Edible Fungi, Shanghai Academy of AgCalifornia USA; GLRC/NYMU: Ganoderma lucidum Research Consortium, National Yang-University of Science and Technology, Wuhan, China; NA: Not available.


2. Sequencing of Ganoderma lucidum nuclear genomes

By November 2013, eleven whole genome sequencing projectsfor Ganoderma species have been registered in the BioProject sec-tion of the National Center of Biotechnology Information (NCBI),which collects biological data related to initiatives provided byinstitutions or consortiums (http://www.ncbi.nlm.nih.gov/biopro-ject/). The details of each project and the organization involvedare shown in Table 1. Among these projects, four were conductedby research groups from institutes in China, which include TheInstitute of Medicinal Plant Development (IMPLAD), HuazhongUniversity of Science and Technology (HUST), and The Instituteof Edible Fungi (IEF). Two projects were conducted by the researchgroups of The National Yang-Ming University (NYMU) from Tai-wan. The five remaining projects were conducted by researchgroups from institutes in the United States, which include (a) TheBroad Institute of Massachusetts Institute of Technology (MIT)and Harvard, and (b) The Joint Genome Institute (JGI). Interest-ingly, five of these projects focus on the genome of Ganoderma luci-dum, demonstrating the high level of interest on thisrepresentative species for the Ganoderma genus. So far, threeassembled genomes have been published, two from strains of G.lucidum and one from a strain of an unidentified Ganoderma spe-cies (Binder et al., 2013; Chen et al., 2012; Liu et al., 2012). Onewhole genome shotgun project with unassembled sequences of athird G. lucidum strain (GenBank BACH00000000) has also beensubmitted to NCBI (Huang et al., 2013). The rest of the projectsare still in progress.

The genome assembly from IMPLAD was obtained from G. luci-dum strain G.260125-1 using next-generation sequencing andoptical mapping technologies. The assembly is 43.3 Mb long andencodes 16,113 predicted genes. Due to the better assembly of thisgenome compared to those of others (see below), the richest sets ofwood degradation enzymes among all of the sequenced Basidio-mycetes were identified from this assembly. Furthermore, 24 phys-ical CYP gene clusters have been identified, which are potentiallyinvolved in the biosynthetic pathways of various biologically activecompounds. The IMPLAD researchers claimed that the genome ofG. lucidum should be investigated as a potential model system forsecondary metabolic pathways and regulation in medicinal fungi(Chen et al., 2012).

The HNAU assembly is from G. lucidum strain Xiangnong No. 1using Illumina sequencing technology alone. It has a 39.9 Mb longdraft genome, which encodes 12,080 protein-coding genes.Approximately 83% of these genes are similar to sequences foundin international databases. The rest were novel. The HNAU groupannotated their G. lucidum assembly and compared the annotatedgene models with those of other fungal genomes. The genes

Organization* References

BROAD NABROAD NABROAD NABROAD NA

e shotgun sequences available GLRC/NYMU Huang et al. (2013)GLRC/NYMU NA

genome assembly available HNAU/HUST Liu et al. (2012)genome assembly available IMPLAD Chen et al. (2012)

IMPLAD NAIEF NA

genome assembly available JGI Binder et al. (2013)

, Boston, USA; IMPLAD: Institute of Medicinal Plant Development, Chinese Academyricultural Sciences, Shanghai, China; JGI: DOE Joint Genome Institute, Walnut Creek,Ming University, Taiwan; HuNan Agriculture University (HNAU); HUST: HuaZhong


http://www.ncbi.nlm.nih.gov/bioproject/

http://www.ncbi.nlm.nih.gov/bioproject/



involved in the biosynthesis of the main biologically active compo-nents of G. lucidum, specifically ganoderic acids, were characterizedin detail. Particularly, the HNAU group identified a fusion gene thatwas only found in Basidiomycetes. Consistent with G. lucidumbeing a white rot fungus with wood degradation ability, abundantcarbohydrate-active enzymes and ligninolytic enzymes were iden-tified in the HNAU assembly (Liu et al., 2012).

The genome assembly of JGI is obtained from Ganoderma sp.strain 19597 SS1 using a combination of ABI3730 (fosmids), 454-Titanium and Illumina GAII sequencing platforms. The assemblyis 39.52 Mb long and encodes 12,910 predicted genes. The mixedsequencing reads were assembled using a JGI specific assemblyprocess and Newbler (2.3-PreRelease-6/30/2009, Roche) withdefault parameters. The draft assembly was then annotated usingthe JGI annotation pipeline, which takes multiple inputs (scaffolds,ESTs, known genes) and runs several analytical tools for gene pre-diction and annotation. Single-copy genes were identified andcompared with those from other Polyporales. Three phylogenomicdatasets, which contain 25, 71 and 356 genes respectively wereconstructed to evaluate their potential for phylogenetic systemat-ics of the Polyporales (Binder et al., 2013).

The overall comparison of the three genome assemblies is listedin Table 2. The statistical characteristics of the three assemblieswere extracted from the corresponding papers. The sizes of thethree genome assemblies are similar, ranging from 39.5 Mb forthe HNAU assembly to 43.3 Mb for the IMPLAD assembly. The IMP-LAD assembly is probably of the highest quality, in which 82 scaf-folds were mapped to 13 chromosomes thanks to a physical mapconstructed with the optical mapping technology. The HNAUassembly was obtained using a typical shotgun approach and con-tains 634 scaffolds with a total length of 39.9 Mb. Similar to theIMPLAD project, the JGI project used both 454 and Illumina datato produce an assembly, which contains 156 scaffolds. The totalGC content of the genome and protein-coding genes were compa-rable for the three genome assemblies. The numbers of predictedgenes were also similar. 16,113, 12,080 and 12,910 gene modelswere found in the IMPLAD, HNAU and JGI assemblies, respectively.The average gene length, number of exons per gene, average exon

Table 2Characteristics of three sequenced Ganoderma genomes.

Strain name G. lucidumstrainG.260125-1(Chen et al.,2012)

G. lucidumstrainXiangnongNo. 1 (Liuet al., 2012)

Ganodermasp. 10597SS1 (Binderet al., 2013)

Research group IMPLAD HNAU JGINumber of chromosomes 13 NA NALength of genome assembly

(Mb)43.3 39.9 39.52

Scaffold total number 82 634 156Scaffold maximum length

(bp)4,834,011 1,953,398 4,606,855

Scaffold minimum length(bp)

2064 1004 2002

GC content (%) 55.9 55.6 55.59Number of protein-coding

genes16,113 12,080 12,910

Average gene length (bp) 1556 1959 1541GC content of protein-

coding genes (%)59.3 58.9 59.0

Average number of exonsper gene

4.7 6.3 13.8

Average exon size (bp) 268 230 148Average coding sequence

size (bp)1188 1435 1413

Average intron size (bp) 87 100 62Repeat sequences (%) 8.15 5.07 2.53


size, average coding sequence size, and average intron size for genemodels predicted from the three assemblies showed some level ofvariation. These variations might result from different methods ofgenome assembly and gene prediction used in these projects andmight not reflect actual differences in the gene structures in thesegenomes. Validation of these predicted models are needed toexplain the observed variations.

We compared the three genome assemblies in more detail byplotting the number of scaffolds along with their total accumulatedlength (Fig. 1a). For the HNAU assembly (dashed line), the accumu-lated total length increased rather slowly, suggesting that thelength of the scaffolds was similar. By contrast, the accumulated

Fig. 1. Comparisons of the three different genome assemblies of G. lucidum strainsG.260125-1 and Xiangnong No. 1 and Ganoderma sp. strain 10597 SS1. (A)Accumulated total assembly length vs. numbers of scaffolds; (B) dot plot comparingthe genome structures between all IMPLAD and JGI scaffolds having lengths greaterthan 1 Mb.




length of the IMPLAD assembly and the JGI assembly increasedremarkably; this result suggested the presence of a few very longscaffolds along with many shorter ones. For example, the five lon-gest scaffolds of the IMPLAD assembly were 4,834,011, 2,306,357,1,960,581, 1,936,600, and 1,698,588 base pairs (bp) long,respectively. The five longest scaffolds of the JGI assembly were4,606,855, 3,722,621, 3,223,173, 2,972,551, and 2,861,455 bp long.However, the JGI scaffolds were not ordered and mapped to chro-mosomes because of the lack of a corresponding physical map.Since the IMPLAD and JGI assemblies appear to be of higher quality,these two assemblies were compared further using a dot plot. Allscaffolds with a length >1 Mb from the IMPLAD and JGI assemblieswere extracted to construct the dot plot (Fig. 1b). The North-Amer-ican Ganoderma sp. strain 19597 SS1 sequenced by JGI belongs to aGanoderma species complex (Binder et al., 2013) and consequentlyit might be a species different from that sequenced by IMPLAD.Although more work needs to be done to accurately determinethe species identity from which the materials used for sequencingwere derived, this comparison of genome assemblies suggests that,if there are different species background, the genomes of variousGanoderma species may share a high degree of overall similarity.Comparing the largest scaffolds from the IMPLAD and JGI assem-blies showed that the similarities between high scoring pairs ran-ged from 77.8% to 100%, with an average of 89.2%.

In summary, significant variations were identified among thereported genome assemblies. These might result from several rea-sons. First, the species identity of the materials used for genomesequencing has not been determined or agreed upon. As a result,it is unclear whether or not the materials used are of the same ori-gin or not. Second, it suggests that the level of completion of a gen-ome assembly affects the results significantly. The shotgunsequencing strategy alone is insufficient to obtain high-qualityassemblies even with high coverage produced by next generationDNA sequencing technologies. A physical map constructed usingthe optical mapping technology is invaluable to establish a chro-mosomal-level assembly. In spite of these variations, these assem-blies allow us to appreciate the diversity and complexity ofGanoderma’s genetic makeup.

3. Sequencing of Ganoderma mitochondrial genomes

The Ganoderma genus comprises approximately 80 species withsimilar morphological characteristics. The lack of powerful molec-ular markers to determine these species has caused widespreadmislabeling and misuse of Ganoderma products, which threatentheir safe usage (Wang et al., 2012). The mitochondrion, containingDNA referred to as the second genome, is an organelle found inmany eukaryotic cells (Henze and Martin, 2003), and its DNAevolves faster than the corresponding nuclear DNA. Consequently,it is more suitable to differentiate closely related organisms thanthe nuclear genome (Bullerwell and Lang, 2005; Burger et al.,2003; Ghikas et al., 2010). Hence, the analysis of mitochondrialDNA (mtDNA) is useful for evolution and population studies onGanoderma species.

By mining the sequence reads generated by the Illumina plat-form, Li et al. (2013) successfully assembled the mitochondrialgenome (mtDNA) for G. lucidum strain G.260125-1. This genomeis a typical circular DNA molecule of 60,630 bp with a GC contentof 26.67% (EMBL Accession Number: HF570115). By contrast, themitochondrial genome for G. sinense has also been assembled(GenBank Accession Number: KF673550). The total length of thisgenome is 86,451 bp with a GC content of 26.8%. The alignmentof these two complete genomes is shown in Fig. 2a. A total of 15pairs of highly similar fragments were present. Almost all of thesefragments represent the conserved regions encoding proteins, as


well as small and large ribosomal RNAs and tRNAs. The orders ofthe conserved genes in the two genomes are similar.

Annotation of G. lucidum mtDNA identified genes that encode15 conserved proteins (atp6, atp8, atp9, cob, cox1, cox2, cox3,nad1, nad2, nad3, nad4, nad4L, nad5, nad6, and rps3), small andlarge rRNAs (rns and rnl), 27 tRNAs, four homing endonucleases(ip1, ip2, ip3, and ip4), and two hypothetical proteins (orf1 andorf2; Fig. 2b). All genes except those encoding trnW and two hypo-thetical proteins are located on the positive strand. Annotation ofG. sinense mtDNA identified the same 15 conserved proteins, aswell as small and large rRNAs. In contrast to G. lucidum mtDNA,28 tRNA genes were detected. No genes for homing endonucleaseswere reported in G. sinense mtDNA. However, 33 hypothetical pro-teins were identified. Minor differences were observed in the spe-cies of tRNAs encoded in the two genomes. For example, trnU-UCAis present in G. lucidum only, whereas trnE-UUC occurs in G. sinenseonly. Both genomes contain three copies of trnM-CAU. G. lucidumcontains two trnR genes (trnR-UCG and trn-UCU), with each genecontaining only one copy. By contrast, G. sinense contains five trnRgenes, including three copies of trnR-UCG and two copies of trnR-UCU (Li et al., 2013).

In-depth studies have been performed for G. lucidum mtDNA,including gene expression levels across different developmentalstages, the possible presence of long non-coding RNAs (lncRNAs),potential transfer of DNA fragments between the mtDNA and thenuclear genome, and genome rearrangement comparing to thosefrom other fungal species (Li et al., 2013). Unfortunately, no otheranalyses other than the complete sequences and general annota-tions have been reported for G. sinense mtDNA. With the progres-sion of the various whole genome sequencing projects (Table 1),more mitochondrial genomes will be available and comprehensivecomparative analyses can be performed.

4. Transcriptome analysis of Ganoderma lucidum

To date, several transcriptome studies have been conducted forvarious strains of G. lucidum (Table 3), although none have per-formed a detailed molecular species identification of the materials.These studies differ in terms of the technologies employed and thesamples used. The samples were collected at various developmen-tal stages, including mycelium, primordium, and fruiting body, atvarious points in time. Therefore, the results cannot be easilyintegrated.

One of the earliest studies was conducted in 2010, in which RNAwas extracted from 50-day-old fruiting bodies and then used toconstruct a cDNA library for Sanger sequencing (Luo et al., 2010).A total of 879 high-quality ESTs were obtained and subsequentlyassembled to 600 unique sequences (Table 1). These were thencompared with those in the international nucleotide standarddatabase, such as Nr and Nt, to identify their possible functions.A total of 173 unique sequences showed similarities to knowngenes involved in the biosynthesis of secondary metabolites anddevelopmental regulation (p < 1e�10). Two unique sequencesshowed high similarity to genes encoding squalene epoxidase(SE) and farnesyl pyrophosphate synthase (FPS), respectively,which are two rate-limiting enzymes in the catalysis of triterpe-noid biosynthesis in G. lucidum (Shi et al., 2010). Furthermore, atotal of 13 simple sequence repeat (SSR) motifs were identifiedfrom these EST sequences and the candidate genes involved inthe regulation of G. lucidum development were also studied. Thisdata set exhibits only a limited coverage of the transcriptomic con-tents of G. lucidum due to the low throughput sequencing technol-ogies used.

In the second study, next generation DNA sequencing technol-ogy was used instead of Sanger technology (Chu et al., 2012). The



Fig. 2. Comparison of mitochondrial genomes from G. lucidum strain G.260125-1 (EMBL: HF570115) and G. sinense (GenBank: KF673550). (A) Dot plot analysis of the twogenomes. (B) Venn diagram showing the genes shared by the two genomes.


study used 16 day-old mycelia and 9 week-old fruiting bodies of G.lucidum strain yw-1. In total, 6,439,690 and 6,416,670 high-quality90 bp long reads were obtained from the mycelium and fruitingbody, respectively, and these reads were assembled into 18,892unigenes for the mycelia and 27,408 unigenes for the fruitingbodies (Table 1). The mean size of the mycelium unigenes was498 bp and the fruiting body unigenes mean size was 514 bp, withlengths ranging from 100 bp to over 3000 bp. Unigenes from eachsample were assembled together by the sequence clustering soft-ware TGICL, resulting in a total of 28,210 unigenes. A similaritysearch was performed against the NCBI non-redundant nucleotide


database and a customized database composed of the establishedgenomes of five other Basidiomycetes (Laccaria bicolor, Coprinopsiscinerea, Schizophyllum commune, Phanerochaete chrysosporium, andPostia placenta). A total of 11,098 and 8775 unigenes were foundsimilar to entries in these databases, respectively. All unigeneswere subjected to annotation by the Gene Ontology (GO), theEukaryotic Orthologous Group terms (KOG), and the KyotoEncyclopedia of Genes and Genomes (KEGG).

The unigenes were examined in detail to identify the specificgenes involved in the biosynthetic pathway for ganoderic acids,which represent the major bioactive compounds in the medicinal



Table 3Comparisons of the G. lucidum transcriptome contents described in four studies.

Study No./references(Year)

Material/strain(s) Outline Mycelium Primordium Fruitingbody

Total

1. Luo et al. (2010) 50 days old fruiting bodies of anon-specified G. lucidum strain

Numbers of ESTs –⁄ – 879 879Total number of unique genes – – 600 600Average unique gene length (bp) – – 288 288

2. Yu et al. (2012) 16-day-old mycelium and 9-week-oldfruiting body of G. lucidum strain yw-1

Number of reads 6,439,690 – 6,416,670 12,856,360Average read length (bp) 90 – 90 90Number of unigenes 18,892 – 27,408 28,210Average unigene length (bp) 498 – 514 507Number of genes 13,332 13,055 13,144 13,731

3. Huang et al. (2013) Mono- and dikaryotic G. lucidum strainsBCRC 36123 and BCRC 37180

ESTs from 5 days dikaryon 1001 – – 47,285ESTs from 14 days dikaryon 6,848 – –ESTs from 18 days dikaryon 21,547 – –ESTs from 18 days monokaryon 11059 – –ESTs from 30 days dikaryon 6830 – –

4. Ren et al. (2013) MeJA-induced mycelium ofG. lucidum strain HG

Number of transcript-derivedfragments

3910 – – 3910

* ‘‘–’’ indicates that no data is available.


mushroom G. lucidum (Shi et al., 2010). A total of 13 identifiedexpressed unigenes were involved in the terpenoid backbone bio-synthetic pathway. Among these unigenes, 7 were up-regulated inthe mycelium and six did not show significant expression changesin the two developmental stages (Table 4). The products of theseunigenes show high similarity to hydroxymethylglutaryl-CoA syn-thase (78%), mevalonate kinase (55%), phosphomevalonate kinase(52%), mevalonate disphosphate decarboxylase (63%), farnesylpyrophosphate synthase (FPPS, 66%), geranylgeranyl pyrophos-phate synthetase (66%), squalene synthase (SQS, 54%), squalenemonooxygenase (62%), and lanosterol synthase (LS, 61%), respec-tively. The expression levels of these unigenes were confirmed byquantitative PCR (qPCR). Five genes involved in G. lucidum develop-ment were also identified: ATP synthase F1; citrate synthase;EXG1; hydrophobin 2; and a cytochrome P450. EXG1(AAO32563.1) is a glycosyl hydrolase. The transcriptome dataand qPCR results showed that ATP synthase F1, citrate synthaseand EXG1 might play potential roles in carbohydrate transportand metabolism. By contrast, the cytochrome P450 was highlyexpressed in the fruiting bodies. The expression of the hydropho-bin gene was not confirmed by qPCR (Chu et al., 2012).

A third transcriptome profiling project with the mono- anddikaryotic G. lucidum strains BCRC 36123 and BCRC 37180 was per-formed to study the development of the mycelia in the first 30 days(Huang et al., 2013). Five mycelium samples (5 days dikaryon,14 days dikaryon, 18 days dikaryon, 30 days dikaryon, and 18 daysmonokaryon) were collected for the analyses. A total of 47,285ESTs were obtained from these five mycelial samples and theirlengths ranged from 50 bases to 949 bases (Table 3). Almost allof the ESTs (47,283 out of 47,285) can be mapped to the genomesequence of G. lucidum (Yu et al., 2012). Approximately 83% ofthe EST-genome alignments (38,395) contained gaps that werepresumably introns of the multi-exon genes. Most (approximately99.9%, 38,347/38,395) putative introns were <2 kb. ESTs aligned tothe reference genome were further clustered and merged to gener-ate 7774 non-redundant EST genes. Alternative splice forms in G.lucidum were also analyzed in this study. The results showed that262 of 7774 EST genes may have alternative splice forms, including50 and/or 30 alternative splice junctions, cassette exons, andretained introns.

Genes that were differentially expressed after methyl jasmo-nate (MeJA) treatment were studied in the dikaryotic G. lucidumstrain HG to identify genes involved in the regulation of ganodericacid production (Table 3) (Ren et al., 2013). In that study,cDNA-amplified fragment length polymorphism (cDNA-AFLP)


was performed to identify genes differentially expressed inresponse to MeJA in G. lucidum, which can regulate production ofGA. A total of 64 primer combinations were used to identify morethan 3910 transcriptionally derived fragments (TDFs). Reliablesequence data were obtained for 390 of 458 selected TDFs. A totalof 90 TDFs were annotated with known functions by performing aBLASTX search of the GenBank database. Of these, 12 annotatedTDFs were assigned to secondary metabolic pathways by searchingthe KEGG PATHWAY database. 25 TDFs were also selected for qRT-PCR analysis to confirm the expression patterns observed withcDNA-AFLP. The qRT-PCR results were consistent with the alteredpatterns of gene expression as revealed by the cDNA-AFLPtechnique. Furthermore, the transcript levels of 10 genes weredetermined at the mycelium, primordium, and fruiting body devel-opmental stages of G. lucidum. The greatest expression levels werereached during the primordial stage for all genes except the cyto-chrome b2 gene, which reached the highest expression level in themycelium stage.

Since ganoderic acids are the major active components of G.lucidum, enzymes related to their biosynthesis are of great interest.Sequences found similar to those enzymes involved in the biosyn-thetic pathway (MVA pathway) for ganoderic acids are shown inTable 4. In terms of their expression profiles, Chen et al. (2012)found that the genes involved in the MVA pathway were down-regulated from the mycelia to the fruiting bodies (Table 4) (Yuet al., 2012). By contrast, most genes were expressed at a higherextent in the fruiting body stage than in the mycelial stage (Chuet al., 2012). Huang et al. (2013) observed that the highest expres-sion of the genes involved in the MVA pathway are observed at18 d in the dikaryon. In addition to genes involved in the MVApathway, other genes likely to be involved in the biosynthesis ofganoderic acids have also been identified. For example, 919 tran-script-derived fragments exhibited altered expression patternsafter MeJA induction (Ren et al., 2013). However, since MeJA mayfunction as a global regulator, not all of the differentially expressedgenes after treatment are related to differential biosynthesis. Fur-ther studies should be conducted to identify factors specificallyregulating the biosynthesis of ganoderic acids in G. lucidum andother Ganoderma species. The JGI study likely used a species otherthan G. lucidum; as such, their transcriptomic data were not com-pared in the present study.

There are two challenges facing G. lucidum transcriptome anal-yses. First, the materials used in various studies were usuallyinconsistent. Materials from various strains were collected forthe studies at different time points in cellular growth and



Table 4Expression of genes involved in the biosynthetic pathway of ganoderic acids.

Gene description (product) Chen et al. (2012) Yu et al. (2012) Huang et al. (2013)

Gene_ID Log2(M/F)

Gene_ID Log2(M/F)

Gene_ID M_5thdaya

M_14thdaya

M_18thdaya

M_30thdaya

Acetyl-CoA acetyltransferase GL23502 1.09 Unigene9161_All 0.8 YMGLESTG49648 0 5 33 12GL26574 �0.90 Unigene27186_All 2.3

3-Hydroxy-3-methyl glutaryl-CoAsynthase

GL24922 �1.40 Unigene27186_All 2.3 YMGLESTG52067 0 2 68 12Unigene6833_All 2.3Unigene6227_All 0.3

3-Hydroxy-3-methyl glutaryl-CoAreductase

GL24088 0.77 Unigene2279_All 0.0 YMGLESTG53675 0 0 16 4

Mevalonate kinase GL17879 0.58 Unigene25033_All 2.1 YMGLESTG52428 0 0 1 1Unigene14395_All 1.2

Phosphomevalonate kinase GL17808 �0.05 Unigene6822_All �0.5 YMGLESTG50562 0 0 2 0Mevalonate pyrophosphate

decarboxylaseGL25304 �0.69 Unigene6187_All 0.6 YMGLESTG50973 0 1 5 1

Isopentenyl diphosphate isomerase GL29704 0.71 Unigene9721_All 1.6 YMGLESTG47454 0 4 3 6Farnesyl diphosphate synthase GL22068 �1.66 Unigene803_All 0.3 YMGLESTG55240 0 0 8 2

GL25499 �1.73 YMGLESTG52116 0 0 11 8Squalene synthase GL21690 �0.98 Unigene111_All 0.3 YMGLESTG51888 0 2 3 4Squalene monooxygenase GL23376 4.78 Unigene2082_All 2.2 – – – – –

Unigene5606_All 1.8 – – – – –2,3-Oxidosqualenelanosterol cyclase GL18675 �0.74 – – – – – – –Geranylgeranyl pyrophosphate

synthetase– – Unigene688_All �0.6 – – – – –

Lanosterol synthase (LS) GL18675 �0.74 Unigene4718_All 1.5 – – – – –

More than one gene may be found in a particular gene description.M, mycelium; P, primordium; F, fruiting body.–: No gene was reported in the particular study.

a The number in columns of M_5th day, M_14th day, M_18th day, and M_30th day from Huang et al. (2013) study represent the EST counts for each corresponding gene.


development, or from different cultural media. A previous study onthe production of ganoderic acids has already shown that theabove factors affected gene expression levels and bioactive com-pound contents (Liang et al., 2010). As a result, it is uncertainhow many of the differences observed in the aforementioned stud-ies actually resulted from genetic diversity rather than the envi-ronmental factors. Second, the variations caused by thetechnologies should also be considered. In these studies, Sangersequencing and next-generation sequencing technologies, such as454 and Illumina, were employed. These technologies differ inthroughput, sequence length, and error rate, among others(Shendure and Ji, 2008). Furthermore, added additional possiblevariations to the data, make it more difficult for: (a) identificationof the complete transcriptomes of Ganoderma species and (b) com-parison of the expression levels of various genes during cellulardevelopment and production of secondary metabolites.

The use of diverse samples from different strains of the samespecies at various developmental stages under different culturalconditions is thus not helpful to address particular biological ques-tions. Future transcriptome studies should consider the followingpoints from the very beginning. For example, studies should theo-retically use materials from one particular, completely sequencedstrain, which has been cultured under standardized conditions,such as medium, temperature, and length of growth, among others(Berovic et al., 2003, 2012; Habijanic et al., 2013; Wang et al., 2014;Wen et al., 2010). The sequencing strategy and bioinformatic dataprocessing steps should also be standardized. For example, thecomplete genome sequences should be used as the reference toassemble the transcriptome whenever possible.

5. Genes belonging to the cytochrome P450 family

The number of cytochrome P450 genes in fungi is highly vari-able. The obligate intracellular parasites Encephalitozoon cuniculiand Nosema apis (microsporidia) have no P450s. The taphrinomy-cetes Schizosaccharomyces pombe, Schizosaccharomyces japonicus,


and Schizosaccharomyces octosporus each contain CYP51 andCYP61 for ergosterol biosynthesis but no other P450s (Weeteet al., 2010). The taphrinomycetes are the earliest diverging Asco-mycetes, so it is assumed that their ancestors had only these twoP450s. This is probably true for the ancestor of Dikarya and forthe Basidiomycetes descended from that common ancestor. Fromhere on, it is assumed that all other Basidiomycete P450s are addi-tions to this P450 baseline.

Cytochrome P450 analysis was performed on two high-qualityfinished Ganoderma genomes: G. lucidum strain G.260125-1(Chen et al., 2012) and the Ganoderma sp. strain 10597 SS1sequenced at JGI (Binder et al., 2013; Grigoriev et al., 2012). P450genes are abundant in these genomes. G. lucidum has 197 P450genes and 19 pseudogenes. Ganoderma sp. strain 10597 SS1 has181 P450 genes and 29 pseudogenes (Syed et al., 2013). Theseare among the largest fungal CYPomes, though they are smallerthan some plant CYPomes. The Ganoderma CYPs are distributedin 42 CYP families and 14 CYP clans. Clans are clades of genes thatshare a common ancestor and are the deepest reliable clusters ofP450s on phylogenetic trees. The clan system of fungi is still cur-rently being defined, but it seems that there are approximately25 P450 clans for fungi as a whole (from more than 5000 namedP450 sequences). Early recognized fungal clans in P. chrysosporium(CYP51, CYP61, CYP52, CYP64, and CYP505) have been discussedpreviously (Yadav et al., 2006).

A total of 377 P450 sequences from the two Ganoderma CYPo-mes are included in the two phylogenetic trees presented inFig. 3 and Fig. 4, respectively. Pseudogenes, short sequences, orotherwise problematic sequences were omitted. Fig. 3 shows 12of the 14 CYP clans including 194 sequences. The CYP547 clan isthe largest with 119 sequences in eight families. The CYP54 clanhas only one family, while the CYP512 family has 41 sequences.Surprisingly, 10 of the 12 clans shown have only 1 CYP family inGanoderma. Orthologs are adjacent red–blue pairs. The alternationof red and blue around the tree indicates there are no species-spe-cific gene blooms. Many of the genes have orthologs, with a fewhaving small expansions or contractions compared to the other



Fig. 3. Neighbor-joining tree of 194 Ganoderma P450s from 12 CYP clans. Clans are labeled on the spokes and each clan is shown in a separate color. G. lucidum strainG.260125-1 labels are red and Ganoderma sp. strain 10597 SS1 labels are blue. Sequence alignments were computed using CLUSTAL W and checked manually for consistentalignment of known CYP motifs. Neighbor-joining trees were generated with the Phylip package (Ropelewski et al., 2010) using ProtDist (a program in Phylip) to computedifference matrices. Trees were drawn and colored with FigTree ver. 1.3.1 using midpoint rooting (http://tree.bio.ed.ac.uk/software/figtree/) and labeled in Adobe IllustratorCS ver. 11.0.0 (Adobe Systems Incorporated). (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)


species. The largest expansion is only 4:1 from the CYP512U sub-family in the CYP54 clan.

The remaining 183 P450 sequences in the large CYP53 andCYP64 clans are compared in Fig. 4. Similarly, there are manyortholog pairs and little evidence of gene blooms. In this figure,the individual families are colored differently since there are only2 clans in this tree. The CYP5359 family occupies half of the tree.The CYP5035 family is also quite large. This shows the dramaticexpansion of these two families (and CYP512, 5136, 5139, and5150 in Fig. 3). The family expansion happened before speciationas evidenced by the large number of ortholog pairs between the2 Ganoderma species analyzed. The large increase in intact (pre-sumably functional) P450s in these families implies some evolu-tionary advantage gained by these oxidative enzymes. Some maybe involved in secondary metabolite synthesis. Others may beinvolved in the oxidative degradation of the wood components,such as lignin, once these degradation products are imported intothe fungus after pre-digestion outside the cells with peroxidasesand laccases. This seems plausible since 1063 transporters werefound in G. lucidum strain G.260125-1 (Chen et al., 2012). The fruit-ing bodies of these bracket fungi are hard and have cell structuresthat should maintain their growth patterns. Lignin evolved in tra-cheophyte land plants to provide rigidity for water transportingvessels. The woody context of bracket fungi would necessarily becreated by an independently evolved polymer analogous to lignin


in plants. Some of the Ganoderma P450s may contribute to the for-mation of a rigid fungal superstructure.

Chen et al. (2012) pointed out that G. lucidum strain G.260125-1has 24 gene clusters with three or more CYP genes (suppl. Data 6 oftheir paper). Some of these clusters are quite large. Three clustersare over 200 kb in size and have more than 70 genes, larger thanmany known secondary metabolite clusters. Therefore, these mayneed to be broken down into 2 or more smaller clusters. The auto-mated gene cluster finding software Antismash identified 17 geneclusters (suppl. Data 9 of their paper), but only P450 cluster 10overlapped with these predictions, including one of the 5 polyke-tide synthase (PKS) genes found in the genome. A second genecluster predictor, SMURF, found 5 gene clusters, but only one over-lapped P450 cluster 23 (suppl. Data 10 of their paper). Comparisonof the 2 software predictors favors Antismash, since it found clus-ters that included all 5 PKS genes, one NRPS gene, and 11 of the 12terpene synthase genes. SMURF found the NRPS and 2 of the PKSgenes in its clusters, but none of the terpene synthases. Therequirement for at least 3 P450s to define a cluster may be toostringent. There are characterized secondary metabolite gene clus-ters that have fewer than 3 P450s. A more successful approach maybe to look for P450s within 50 kb of a cluster anchor gene like theNRPS, PKS, terpene synthase, and dimethylallyl tryptophan syn-thase (DMAT) genes. The product of DMAT is dimethylallyl trypto-phan, which is the precursor of indole alkaloids in Ganoderma.


http://tree.bio.ed.ac.uk/software/figtree/


Fig. 4. Neighbor-joining tree of 183 Ganoderma P450s from the CYP53 and CYP64 clans. Each P450 family is shown in a separate color. G. lucidum strain G.260125-1 labels arered and Ganoderma sp. strain 10597 SS1 labels are blue. The tree was computed and drawn in the same way as Fig. 3. (For interpretation of the references to color in this figurelegend, the reader is referred to the web version of this article.)


The P450 families in the 2 analyzed Ganoderma species and 2other polypores P. placenta and P. chrysosporium are shown sortedinto their clans in Suppl. Table A. This set of 4 genomes has 15clans, except for Ganoderma, which is missing CYP5153. Two ofthe clans are CYP51 and CYP61 in the ergosterol pathway. Theseclans frequently cluster together and may form a larger clan. TheCYP53 clan has 5 families in Ganoderma, 3 of which (CYP53,CYP537, CYP5140) have only 1 sequence each. Singletons like theseare probably dedicated to 1 specific function. In Aspergillus niger,CYP53A1 is encoded by the benzoate-para-hydroxylase gene(bphA) (Faber et al., 2001). Phenolics like benzoate are toxic tofungi and CYP53 acts as an early step in the b-ketoadipate detoxi-fication pathway (Podobnik et al., 2008). The enzyme is highly con-served and may be a new antifungal drug target (Berne et al.,2012). The functions of CYP537 and CYP5140 are not known.

The CYP52 clan primarily includes the CYP63 family in theGanoderma species. This is an expanded family for both Ganodermastrains under comparison in this present review. CYP63 is alsomoderately sized in P. chrysosporium and P. placenta. The CYP63genes of P. chrysosporium have been studied for induction by lignincomponents, with the verification that some were induced, sug-gesting these genes may encode lignin-degrading P450s(Subramanian and Yadav, 2008).

The CYP54 clan has only 1 family in Ganoderma, which isCYP512, with 41 members (Fig. 3). The CYP512A1 gene was clonedfrom Trametes (Coriolus) versicolor, a lignin-degrading Basidiomy-cete. This gene is inducible by dibenzothiophene, and was the first


chemical stress-induced P450 gene reported in Basidiomycetes(Ichinose et al., 2002). The substrates for the CYP512 family havebeen investigated recently (Chigu et al., 2010; da Coelho-Moreiraet al., 2013; Hirosue et al., 2011). The CYP512 members were foundto metabolize plant resins used as defense barriers to pathogens.The CYP512 family was also expanded in wood degrading Basidio-mycetes, suggesting a role in colonization of wood. Additional dis-cussion on the roles of CYP512 family can be found elsewhere(Syed et al., 2014).

The CYP505 clan has 7 sequences (Fig. 3), 3 ortholog pairsCYP505D10, CYP505D12 and CYP505D13. CYP505D11 clusterswith CYP505D12 that may represent a co-ortholog. These P450sare self-sufficient because they contain a reductase fused to theP450 part of the sequence. The first CYP505A1 or P450foxy wascloned from Fusarium oxysporum (Nakayama et al., 1996), whichhydroxylates fatty acids, and forms omega-1 and omega-3 prod-ucts. There are 93 named CYP505 fungal sequences obtained frommany species. Two other P450s in Ganoderma, CYP6005A1 andCYP6005B1, are also fusion proteins and belong to the CYP6001clan. This clan has 4 families: CYP6001, CYP6002, CYP6004, andCYP6005, among which Ganoderma only has CYP6005. CYP6003is in another clan by itself. These proteins are also called Psi-factorproducing oxygenases PpoA (CYP6001A1), PpoB (CYP6002A1), andPpoC (CYP6001C1). CYP6001A1 (PpoA) from Emericella (Aspergillus)nidulans is fused with a diooxygenase. The first part adds 2 atomsof oxygen, then the P450 (C-terminus) part acts like a hydroperox-ide isomerase. They act in the synthesis of oxylipins (Koch et al.,



Fig. 5. The matA locus and its gene products. (a) The gene structure of the chromosomal regions containing the matA locus of four different Ganoderma strains. The followinggenes are shown: HD1 gene a1 and HD2 gene a2 (allele numbers are indicated by the second number in the names), mip1 that encodes a mitochondrial intermediatepeptidase, b-fg that encodes an unknown conserved fungal protein and glgen that encodes a glycosyltransferase family 8 protein. The genes are indicated by the arrow-shapedboxes (black: conserved genes; hatched: mating type genes with little conserved allele sequences; grey shaded: genes from retro-transposons; white: potential(pseudo)genes originating possibly from transposons or retro-transposons). The arrows indicate their transcriptional direction. Numbers above the maps indicate positions inkb on the corresponding scaffolds and contigs of the strains. The ends of the two contigs of G. lucidum strain G.260125-1 (at 0 kb and 87.4 kb, respectively) are positionedrelative to each other in their likely natural order. (b and c) Sequence alignment of HD1 and HD2 transcription factors identified from the four Ganoderma strains. N-terminaldiscrimination and dimerization domains (dashed lines), stretches of K/R-rich motifs (filled boxes) as parts of potential bipartite NLSs (nuclear localization signals;underlined), HD1- and HD2-specific peptides (open boxes) and the localization of the 60 aa-long HD2 homeodomain (underlined) with its three helices I to III (filled boxes)and the conserved DNA-binding motif WFXNXR (WFQNRR in all Ganoderma HD2 proteins) are indicated.


2013). E. nidulans has all three. In Aspergillus fumigatus PpoA is alinoleate 5,8-diol synthase (Jerneren and Oliw, 2012). The P450part of these enzymes does not need molecular oxygen, since itis supplied by the substrate of the fusion protein. This leaves theI-helix region (traditionally the oxygen binding motif in P450s)free to diverge, so these CYP6001 clan sequences are typically deepbranches on P450 trees. This makes them similar to the oxylipinproducing CYP74 P450s of plants and some marine animals. Someoxylipins are developmentally significant (Tsitsigiannis et al.,2005). The CYP6005A and CYP6005B sequences may be makingsignaling molecules in Ganoderma that are related to the products


of the Psi factor producing oxygenases. Those molecules have beenshown to be involved in fungal colonization of plants (Koch et al.,2012).

The CYP7 clan is an evolutionary curiosity since it includesvertebrate CYP7 and CYP8 families (not shown in Fig. 3). It isimprobable that the common ancestor of opisthokonta (fungi andanimals) had a CYP7 clan sequence. The 11 CYP clans in animals,including CYP7, all appear to have evolved from CYP51 throughtandem duplication at 1 original locus (Nelson et al., 2013). Analternative explanation is that they are the results of lateral trans-fer from animals. The functions of CYP7 clan sequences and other




single family Ganoderma clans CYP534, CYP608, and CYP609 arenot known.

The CYP547 clan has 113 sequences out of the 194 CYP450genes identified in G. lucidum (Fig. 3). The expansion, especiallyin the CYP5150 family, suggests a function in the metabolism ofmultiple related compounds. The multiplicity of closely relatedsequences does not fit easily with biosynthetic pathways that tendto be relatively short linear chains of enzymes. However, it may notbe true that all members of a family work on closely related sub-strates. Experience from insecticide resistance shows that newfunctions can be obtained from almost any P450 family (Sezutsuet al., 2013). Such a large collection of P450s will almost certainlyhave some novel functions that have developed based on opportu-nity or need. Similar remarks apply to CYP5359 in the CYP64 clanand CYP5035 in the CYP53 clan (Fig. 4). One issue to consider is if 1clan is disproportionately represented in secondary metabolitegene clusters. A casual examination of several fungal toxins (afla-toxin, trichothecene, sirodesmin, gliotoxin, and some others) forP450s found 21 P450s in 16 families. A total of 10 were in theCYP53 clan, 3 were in the CYP613 clan, and 2 of each were in theCYP54 and CYP64 clans. The rest were in 4 different clans. Theremay be favoritism for the CYP53 clan of P450s for use in fungal sec-ondary metabolite biosynthesis. This has relevance for Ganodermaas a source of medicinal compounds, as many of these will be madein fungal gene clusters.

6. Mating type loci in Ganoderma lucidum

G. lucidum is a tetrapolar species (Adaskaveg and Gilbertson,1986) which means that 2 distinct mating type loci (matA andmatB) control development of a dikaryotic mycelium after fusionof 2 compatible monokaryons, as well as fruiting and sexual repro-duction via formation of basidiospores on the established dikaryon.For monokaryons to be compatible, it is essential that they differ inalleles at both loci. Molecular analysis of the Agaricomycetesmodel species, C. cinerea and S. commune, revealed that the A mat-ing type locus in Agaricomycetes encodes 2 types of homeodomaintranscription factors (HD1 and HD2) in one or more divergentlytranscribed HD1–HD2 gene pairs (alphabetically named a pair, bpair, etc.), and the B mating type locus pheromone precursorsand G-protein-coupled 7-transmembrane-domain pheromonereceptors (Ste3-type receptors). Alleles of the 2 loci can differ muchin sequence, up to a level at which one cannot easily recognizetheir direct relationship. The resulting different amino acidsequences of allelic protein products determine their distinct mat-ing type specificities. In compatible matings, HD1 proteins fromone strain have to interact with HD2 proteins of the other strainto form active heterodimeric transcription factor complexes forthe regulation of the expression of dikaryon-specific genes. Like-wise, pheromones from one mating partner have to interact withthe pheromone receptors of the other mating partner to inducethe pheromone-response pathway required for dikaryon develop-ment and maintenance. Importantly in this system, all monokar-yons can possess all types of genes. However, the HD1 and HD2products encoded in the same allele of the A locus do not formfunctional transcription factor heterodimers, and the pheromonesand pheromone receptors encoded in the same allele of the B locusdo not switch on the pheromone response pathway (Kües et al.,2011). Below, the identification of matA and matB loci from 4 Gano-derma genomes is described in detail.

Sequences from the 3 published genome assemblies (Binderet al., 2013; Chen et al., 2012; Liu et al., 2012) and the publishedwhole genome shotgun project (GenBank BACH00000000)(Huang et al., 2013) were analyzed together for mating type loci(James et al., 2013; this review). Candidate genes from the


G. lucidum strains BCRC 37177, G.260125-1, and Xiangnong No. 1were identified using the BLAST module at GenBank with the pro-gram TBLASTN and the databank ‘‘Whole genome shotgun contigs(wgs)’’. By contrast, candidate genes from Ganoderma sp. strain10597 SS1 were identified using the TBLASTN option on the gen-ome portal of JGI (Grigoriev et al., 2012) and data presented byJames et al. (2013). Due to poor sequence conservation, matingtype genes are generally difficult to find based on sequence simi-larity. As a result, defining conserved flanking genes (such as genesmip1 and b-fg found in most Agaricomycetes next to matA) is fre-quently used to identify their positions within the genome (Küeset al., 2011; James et al., 2013).

6.1. matA locus

HD1 protein Pda1 (GenBank AAS46746) and HD2 protein Pda2(AAS46747) of Pleurotus djamor, as well as Mip1 (EKV51093), andb-fg (EKV51091) from Agaricus bisporus var. bisporus were usedas the query sequences in database searches for matA. In the gen-omes of all 4 Ganoderma strains, the alleles of the A locus are about3.6 kb in length, although they differ significantly in sequence. InGanoderma sp. strain 10597 SS1, the A genes reside on scaffold 1at position 2,506,908–2,503,303 (James et al., 2013), in strain Xian-gnong No. 1, the genes reside on contig_850 (AHGX01000850) atposition 241,855–238,254, in BCRC37177, the genes reside oncontig03243 (BACH01002782) at position 91,031–94,647, and instrain G.260125-1, they reside on scaffold GaKu96scf_1_ctg_9(AGAX01000044) at position 3839–7483, respectively.

In all 4 strains, the matA region contains only a single pair ofdivergently transcribed HD1 and HD2 genes, which were subse-quently specified as a1 and a2 genes (Fig. 5a). As typical for Polypo-rales (James et al., 2013), the HD2 gene is flanked by the conservedmip1 gene in all 4 strains. In contrast, the conserved b-fg gene isonly found to be next to the HD1 gene in G. lucidum strainsBCRC 37177 and G.260125-1. It is found to have been translocatedto another place in G. lucidum strain Xiangnong No. 1(AHGX01000411; contig_411 at position 33,961–32,989) andGanoderma sp. strain 10597 SS1 (scaffold 8 at position 544,298–545,271). The sites of b-fg integration in the 2 strains are identical.As seen in the genome assembly of Ganoderma sp. strain 10597SS1, the gene has been moved from the locus near matA to scaffold8, which carries the matB locus (Fig. 6a). Translocation of b-fg hasbrought the conserved gene glgen in G. lucidum strain XiangnongNo. 1 directly next to matA. In other strains, there are transposonrelicts in variable composition and order located in between glgenand matA, respectively b-fg, indicating that the flanking region ofmatA is a place of frequent rearrangements (Fig. 5a).

The products of the HD1 and HD2 genes for the 4 strains arevery dissimilar in sequence (Fig. 5 b and c). Therefore, they shouldrepresent 4 different mating type specificities (Kües et al., 2011)and allele numbers 1–4 have been assigned to these different matAalleles and their respective a1 (HD1) and a2 (HD2) genes (Fig. 5a),starting with the already published matA allele of Ganoderma sp.strain 10597 SS1 (James et al., 2013). Sequence alignments of the4 HD1 and the 4 HD2 proteins are shown in Fig. 5b and c, respec-tively. HD1 proteins of Ganoderma are about 480 amino acids (aa)long. Strikingly, a typical HD1 homeodomain for DNA binding,which is the most conserved region in HD1 proteins, is not recog-nizable in the translated sequences of Ganoderma HD1 genes(Fig. 5b). The HD1 genes were thus defined in the alleles of thematA locus based on their position relative to that of the diver-gently transcribed HD2 genes (James et al., 2013) and based onthe low level conservation of the N-terminal parts of their productscompared to those from some other wood-decay fungi. Using theBLASTP method and the NCBI database, the first 120–140 aa ofthe Ganoderma HD1 proteins were aligned with the N-termini of



Fig. 6. The matB locus and its gene products. (a) The gene structure of the chromosomal regions containing the matB locus of four different Ganoderma strains. The likelyexpansion of matB is underlined. The following genes are shown: pheromone receptor genes (prefixed with ste3; the first number afterward indicates a specific gene, a letter aor b afterward indicates duplications of a specific gene, the second number indicates an allele of a gene) and pheromone precursor genes (prefixed with ph; the first numberafterward indicates a specific gene, the second number indicates an allele of a gene, letters a and b alleles result in the same product). The genes are indicated by the arrow-shaped boxes (black: genes with highly conserved allele sequences, likely non-mating type genes; grey shaded: genes with poorly conserved allele sequences and possible Bmating-type activity; white: pheromone receptor genes inactivated by stop codon mutations and/or deletions of parts of genes; note that the ste3.1 alleles in two of thestrains cannot produce a functional protein). The arrows indicate the transcriptional direction. Blocks of matB genes closer related to each other are indicated by black, greyand hatched boxes, respectively. Numbers above the maps indicate positions in kb on the respective scaffolds and contigs of the strains. The four contigs of G. lucidum strainBCR 37177 have been positioned using sequence alignments of the complete contigs, so that its genes are arranged in the most similar order as those of other strains. (b and c)Phylogenetic trees of pheromone receptors, precursors of pheromones and pheromone-like peptides (see compilation in Table 5) from four different Ganoderma strains,calculated by the neighbor-joining method (500 bootstrap repeats). In the analysis of pheromone receptors, the sequences of seven receptors encoded in the genome of C.cinerea strain Okayama 7 (#130) (Kües et al., 2011) were included (shaded in grey) for recognition of possible mating type and non-mating type receptors. Pheromoneprecursors whose mature pheromones and pheromone-like peptides likely have identical sequences are also shaded in grey. The groupings of proteins (indicated by brackets)base on the locations of their genes in the genomes (within matB: Mating type; outside matB: Non-mating type) and on the sequences of the 2 or 3 aa that are directly linkedto the C-terminal CAAX motif of pheromone precursors, respectively.


several HD1 mating type proteins from Dichotomus and Heterobas-idion (data not shown). As shown in other species, the N-termini ofboth types of mating type proteins mediate individual mating typespecificity. The N-termini are crucial for discrimination of HD1 andHD2 proteins encoded in the same matA allele and are needed as


interfaces for heterodimerization with suitable partner proteinsencoded in other matA alleles (Kües et al., 2011). In accordance,the N-terminal sequences of the 4 Ganoderma HD1 proteins showonly 42–54% amino acid identity and 63–75% similarity to eachother (Fig. 5b). Furthermore to the N-terminal dimerization and




specificity domains, some KR-rich protein stretches are sharedwith several HD1 mating type proteins of other species (data notshown). In other species, these regions localize in sequences C-ter-minal to the HD1 DNA-binding motif (pfam12737: Mating_Csuperfamily), whereas in Ganoderma, they follow directly the N-terminal discrimination and dimerization domains (Fig. 5b). Thisfurther suggests that the regions encoding the HD1 homeodomainhave been deleted from the Ganoderma a1 genes. Regardless of theinternal gene deletions, the a1 genes still contain an open readingframe for a protein consisting of all the remaining N- and C-termi-nal regions known from HD1 proteins in other species. Neverthe-less, even in the apparent absence of the HD1 homeodomain, theGanoderma a1 proteins might still perform their necessary functionas partners of HD2 proteins in formation of active heterodimerichomeodomain transcription factor complexes. This idea is sup-ported by data from C. cinerea, which showed that HD1 proteinscan be functional even without an operational homeodomain(Asante-Owusu et al., 1996). However, the C-terminal K/R-richstretches in the HD1 proteins are essential as part of bipartite NLSs(nuclear localization signals) to transport the HD1–HD2 heterodi-mers into the nucleus (Spit et al., 1998). From sequence alignmentsmade with the Ganoderma HD1 proteins and those from other spe-cies, a highly conserved 11-aa-long peptide (GPRLHAVSDSF/Y)linked to the K/R stretches (Fig. 5b) was newly observed that isspecific to numerous Agaricomycetes HD1 proteins and whosemolecular function remains to be elucidated.

The Ganoderma HD2 genes encode proteins that are approxi-mately 550–560 aa-long. The proteins have the typical highly con-served HD2 DNA binding motif (70–80% identity, 81–92%similarity), which is a classical 60 aa-long homeodomain with 3a-helices (Kües et al., 2011). The conserved DNA-binding motifWFQNRR can be found in the third helix (Fig. 5c). The 148–153 aa-long N-termini are less conserved, exhibiting only 34–43%identity and 48–59% similarity to each other. As indicated above,this high degree of dissimilarity is important for their functionsas recognition and dimerization domains with the N-termini ofHD1 proteins (Kües et al., 2011). Since all 4 HD2 proteins differ sig-nificantly from each other in the N-termini, each should representa distinct mating type allele specificity. The regions C-terminal tothe HD2 motifs are about 350 aa-long. They are the most variableregions in the entire protein sequences and contain only a fewshort stretches of sequences with higher similarity (Fig. 5b). Nota-ble is a newly observed 20-aa-long conserved peptide (V/SPXHAYPXP/TYPPL/SCXY/HX,P/TFP/S) of yet unknown functionthat is found in the Ganoderma HD2 proteins (Fig. 5b) and also inseveral other Agaricomycetes HD2 proteins. However, data fromprotein truncation experiments showed in C. cinerea that largerparts of the poorly conserved C-termini of HD2 proteins are dis-pensable for function (Kües et al., 1994) and the low level of sim-ilarity between C-termini of Ganoderma HD2 proteins mightsuggest this also for species of this genus.

6.2. matB locus

To identify B locus genes, pheromone receptor Rcb3 of C. cinerea(AAO17256) was used to search the 4 genome sequences. In total, 8to 9 different pheromone receptor genes and also parts of formerpheromone receptor genes were detected to reside in the 4 strainswithin a 60–100 kb long sequence region. These genes are on scaf-fold 8 of the genome of Ganoderma sp. strain 10597 SS1, on con-tig_857 (AHGX01000857) of the genome of G. lucidum strainXiangnong No. 1, on contig00760 (BACH01000746), contig00761(BACH01000747), and contig00762 (BACH01000748) of the gen-ome of G. lucidum strain BCRC 37177, and on scaffoldGaLu96scf_14_ctg_1 (AGAX010000010) of the genome of G. luci-dum strain G.260125-1 (for details on positions, see (Fig. 6a)).


These regions show restricted sequence similarities among thegenomes, supporting the notion that the matB genes reside here,and that encoded genes in all 4 strains have different B mating typespecificities. Hence, numbers 1–4 were assigned to the matB allelesfrom the different strains following the same order as that for thematA alleles (Fig. 6a).

Consistent with B mating type function, 5 genes for pheromoneprecursors per strain were also found in these regions and 3 addi-tional genes were found 20–30 kb apart (Fig. 6a). Moreover, thereare 2 other genes elsewhere in each genome, summing up to a totalof 10 pheromone precursor genes per strain (Table 5). The firstpheromone precursor genes were spotted in low stringencyTBLASTN searches using pheromone precursor sequences of thepolypore P. placenta (Kües et al., 2011), which detected short con-served N-terminal MDAFF motifs combined with the essentialC-terminal CAAX (C = cysteine; A = aliphatic aa, X = any aa) motifs(compare sequences in Table 5). Additional genes were thenobtained by cross-checking the TBLASTN search results with theidentified Ganoderma pheromone precursors. The marking ofgenomic positions by PF08015 (pfam08015: fungal mating-typepheromone) on the JGI portal helped additionally in someinstances for Ganoderma sp. strain 10597 SS1. In general however,due to their short length and the high sequence variation of theencoded proteins, pheromone precursor genes are difficult to findthrough the TBLASTN method alone, even under conditions of low-est stringency and with inactivated filters for low complexityregions. All 6 frame-translations of this entire DNA region weremanually inspected to search for N-terminal MDAFF-like motifsand C-terminal CAAX-motifs, which are indicative of pheromoneprotein sequences. No new pheromone precursor genes werefound.

From other species, it is observed that matB pheromone recep-tor genes and matB pheromone precursor genes localize together.Other genes for pheromone receptors and for pheromone-like pep-tides can be present in addition but apparently have no function inmating. Such non-mating-type genes do not necessarily cometogether in the genome (Kües et al., 2011). Some of pheromonereceptor genes in Ganoderma have neighboring pheromone precur-sor genes (Fig. 6a), hinting to B mating type function. Starting froma pheromone receptor gene with a neighboring pheromone precur-sor gene, the receptor genes in Ganoderma sp. strain 10597 SS1were named ste3.1–ste3.8 and the linked pheromone precursorgenes ph1–ph8. All were given the allele number 1 (Fig. 6a). Next,a phylogenetic analysis was done with receptor sequences of all4 Ganoderma strains and in addition the sequences for B-mating-type and non-mating-type receptors of C. cinerea (Fig. 6b), usingClustalX and the neighbor-joining function in the program MEGA(Tamura et al., 2011). In the C. cinerea matB locus, there are 3 para-logous groups of pheromone receptor and pheromone precursorgenes, all of which have mating type function but act indepen-dently from each other (Kües et al., 2011; Riquelme et al., 2005).In the phylogenetic tree of pheromone receptors, the mating-type-active receptors of C. cinerea can be found in clusters withGanoderma receptors coming from genes that have neighboringpheromone precursor genes (compare Fig. 6a and b). It is thuslikely that at least 3 paralogous groups of matB genes and productsexist in Ganoderma, similar to the Subgroups 1–3 seen in C. cinereaand other Agaricales (Kües et al., 2011; Riquelme et al., 2005).Other receptors in Ganoderma appear to have diverged from thosewith putative mating type function (Fig. 6b) and might be non-mating type receptors similar to extra receptors found in otherAgaricomycetes (Kües et al., 2011). Analysis with the TMpred pro-gram at the ExPASy portal (Swiss Institute of Bioinformatics, Lau-sanne, Switzerland) predicted that all of the Ganoderma receptorshave strong 7-transmembrane helices as to be expected for func-tional receptors.



Table 5Sequences of pheromone precursors as deduced from the Ganoderma genomes.

Strain/position in genome Namea Sequence of pheromone precursor

Ganoderma sp. strain 10597 SS1 (JGI scaffold 8)1,522,224–1,522,346 Ph1-1 MDEFENLFGSIETKWDAPMTLEVPVNADDPSHAGVYCVIA1,520,557–1,520,388 Ph2-1 MDAFSFTLDELLAPQHPPAADPLPSPSSSDDEPKGMPVDFEYINNNYSHSWCTVA1,509,586–1,509,425 Ph3-1 MDYFESLHASFLKSETSSAQSLDIVDISLPADIPVPVDEEHQPSTSHFWCVIV1,507,928–1,508,116 Ph4-1 MDEFATITVSLSEPTTSFSPSVPPLGLQLPAHERAAVLARLSSSLPVNEDNNDQYTAYCIII1,506,415–1,506,549 Ph5-1 MDSFFVISEPLYDSPGQAVELSDDLPNDSEHYGGGQTSFYCVIA1,454,925–1,455,062 Ph6-1 MDAFFVIASPIQADEPTSSSVEEIEEIFVDYENLNNSWHSGCVIA1,448,245–1,448,126 Ph7-1a MDAFFTISSPIPVDEPAVEEILIDSEGSSTSEHSGCIIA1,443,942–1,444,061 Ph8-1 MDAFFTIAPAVPVEEPAADIVFINCDSGTGDNHSGCIIA

Ganoderma sp. strain 10597 SS1 (JGI scaffold 16)200,939–201,082 Phl1-1 MDAFFVISAPIVASPESATSSSSELDETFVDQDSFGYVGWHAGCVVA202,641–202,781 Phl2-1 MDAFFHLAAPIPSQEPACSSSSEGTEVFVDEDTFGYVGVHSGCIIA

G. lucidum strain Xiangnong No.1 contig_857 (GenBank Accession No. AHGX01000857)90,217–90,035 Ph2-2 MDSFDTLSLPLVTLEDTHTLSSSSAFGETCAPSGVPSLLDPPVNQEVIDGNGTYSWCTIA92,109–92,231 Ph1-2 MDEFDIFDISLGISLQEPLSGDIPVNEDDPSRAGVYCIIA100,547–100,753 Ph4-2 MDEFSIITTLPSDHSETHLNPISPPYGPSRPFANISPHQRRALLAHIASSVTVDEDSNDQYQAYCIIA103,674–103,823 Ph5-2 MDIFEPIFIAPPIPKLPSPPEPSDIDESDVLADFEHSNPASPKFYCVIS108,751–108,554b Ph3-2 MDDFAFTVIAPPIPDAGVAVDVGDIVLIDEDRPWHFASACTIA151,870–151,733 Ph6-1 MDAFFVIASPIQADEPTSSSVEEIEEIFVDYENLNNSWHSGCVIA158,563–158,682 Ph7-1b MDAFFTISSPIPVDEPAVEEILIDSEGSSTSEHSGCIIA162,882–162,763 Ph8-1 MDAFFTIAPAVPVEEPAADIVFINCDSGTGDNHSGCIIA

G. lucidum strain Xiangnong No. 1 contig_841 (GenBank Accession No. AHGX01000841)14,304–14,438 Phl1-2 MDAFFVISAPIVASPESATSSSSEYDETFVDQDSFGYVGWHAGCVVA16,003–16,143 Phl2-2 MDAFFHLAAPIPSPEPASSSSSEGTEVFLDEDTFGYVGVHSGCIIA

G. lucidum strain BCRC 37177 contig00761 (GenBank Accession No. BACH01000747)4053–4175 Ph1-3 MDAFTTIVSIATTGETTAEITNDQREVTYGSSSYSWCTVA297–467 Ph2-3 MDDFTTFSAFLETEASLKDRPSVDGPPAIMSPTDGVPVDMEYITGNSSSYSWCVVA

G. lucidum strain BCRC 37177 contig00760 (GenBank Accession No. BACH01000746)37,323–37,117 Ph4-3 MDEFCIIASLSSEHSETHLNSIDSRYSPSRSFGGISSQERRALLAHIASSVAIDEDSNDQYQAYCVIA33,995–33,852 Ph5-3 MDTFELLFIAPPIPELPSSLESSDAAESDVLADYEHNNPASPKFYCVIS30,680–30,874b Ph3-3 MDEFIFTVIAAPVPDTGATADVEEIVLIDEDRPWHFGMGCTIA

G. lucidum strain BCRC 37177 contig00759 (GenBank Accession No. BACH01000745)22,403–22,266 Ph6-2 MDAFFVIASPIQAEEPANSSVEEIEETFVDHDSLNNNWHSGCVIA29,260–29,379 Ph7-2 MDAFFTISSPVPVDEPAVEEILVGSDSSSTSEHSGCVIA33,626–33,507 Ph8-2 MDAFFTIAPPVPVDEPVADVEFINYDDGTSDNHSGCTIA

G. lucidum strain BCRC 37177 contig00934 (GenBank Accession No. BACH01000906)26,505–26,362 Phl1-3 MDAFFVISAPVIASPECAPSSFSEVDETFVDQETFGYVGWHAGCVVA24,841–24,698 Phl2-3 MDAFFYIADPIPSQEPAPSSSSPAAPETFFDHDTFGYIGVHSGCIVA

G. lucidum strain G.260125-1 GaLu96scf_14_ctg_1 (GenBank Accession No. AGAX01000010)141,651–141,529 Ph1-4 MDAFVTIVSIVTTGETTAEITNDQREVTYGSSSYSWCTIA145,527–145,357 Ph2-4 MDDFATFSAFLETEASLKDQPSVDGPPAIMPPTDGVPVDMEYITGNSSSYSWCVVA163,198–163,052 Ph5-4 MDEFFVLAEPIPVDQPSDTGSSDVPAIDYPSEFEQYGSGQNTFFCVVA164,566–164,396 Ph3-4 MDLFELTHDLFLEPEAEVAVSSSPLDTEDPSLVIDLPVPVDSEHQPSSSHFWCIVA168,957–169,169 Ph4-4 MDTFDITFAISDGSPQGAPSAALSPVSSPTGSPVAPVVMDAADTPDLDFEQVPADFDHRSSGFGAYCVIA204,460–204,579 Ph8-3 MDAFFTIAPPVPVDEPTADVEFINYDDGSGDNHSGCTIA209,131–209,012 Ph7-3 MDAFFTISSPVPVDEPAVEEILVSSDSSNSSEHSGCVIA215,985–216,122 Ph6-3 MDAFFVIASPIQTDEPASSSVEEIEEVFVDQDSLSHNYHSGCVIA

G. lucidum strain G.260125-1 GaLu96scf_22_ctg_3 (GenBank Accession No. AGAX01000061)122,680–122,826 Phl1-4 MDAFFVISAPLVTSSESAATSSSSEVDEPFVDQDSFGYVGWHAGCVIA124,338–124,484 Phl2-4 MDAFFYLAAPIPSQEPASEPSSPSENPEVFVDHDTFGYIGVHSGCIIA

a Ph stands for pheromones encoded by genes within the chromosomal region containing the matB locus, Phl for pheromone-like peptides encoded by genes unlinked tomatB.

b Intron between last sense codon and stop codon.


The alleles of the likely mating-type pheromone receptor geneswithin the matB locus of the 4 strains and of likely non-mating typepheromone receptor genes positioned in the flanking chromosomalregions (Fig. 6a) were defined by the clustering of receptors in thephylogenetic tree (Fig. 6b), as well as protein length (size rangesare 414–436, 438–477, 476–494, 591–601 aa) and general genestructure. Among the strains, the genes do not simply line up inthe same order. Inversions and relocations of genes and group ofgenes are observed. This is true for pheromone receptor genes, aswell as for pheromone precursor genes (Fig. 6a). Particularly manyrearrangements occurred directly within the closer confinedregions of matB (covering about 19–30 kb; Fig. 6a), together withapparent duplications of ste3.4 genes in strains BCRC 37177 andG.260125-1, and ste3.2 genes in strain Xiangnong No. 1. Blocks of


closer related genes are recognized between the matB alleles ofthe different strains, which include both pheromone precursorand pheromone receptor genes. In summary, each combination of2 distinct matB alleles shares one closer related block of genes witheach other, and differs stronger in a second block of genes. In sucha manner, discrete mating type specificities should be granted forall 4 strains in all possible breeding combinations.

Table 5 summarizes the sequences of all pheromone precursorsidentified from the 4 Ganoderma genomes, including those whosegenes are unlinked to matB. All sequences start with an MDXFmotif, contain an internal dipeptide of charged aa for N-terminalpheromone processing (such as EH, ED, RE, DR, DD) and end withan CAAX motif required for isoprenylation and carboxymethylationof the conserved cysteine after C-terminal proteolytic processing



Fig. 7. Screen shot showing the web resources available for Ganoderma. (a) JGI’sweb portal (http://genome.JGI.doe.gov/Gansp1/Gansp1.home.html); permission touse the screen shot of the web site has been granted by Dimitrios Floudas fromClark University, Worcester, MA; (b) GaluDB (http://www.medfungi.org/galu).Permission to use the screen shot of the web site has been granted by Chang Liufrom Institute of Medicinal Plant Development, Chinese Academy of MedicalScience, Beijing, China.


(Kües et al., 2011). Some pheromone precursor genes are highlyconserved between 2 strains (such as for Ph6-1 and Ph8-1;Fig. 6a) and others encode identical precursor sequences from dis-tinct DNA sequences (such as for Ph7-1a and Ph7-1b, Table 5). Inaddition, a number of distinct precursors will result in identicalmature pheromones upon processing (marked in Fig. 6c). Maturepheromones of Ganoderma should consist of 10–13 aa. The phylo-genetic tree of all pheromone precursors shown in Fig. 6b may notsupport well the grouping by allele origin, because the underlyingClustalX alignment based mainly on the relatively conserved shortN- and C-terminal sequences of the 39–70 aa long pheromone pre-cursors of overall highly variable sequences (see Table 5). However,the clustering revealed a possibly meaningful grouping by the 2 or3 aa that are directly neighbored to the CAAX motif (Fig. 6c). Amino


acids at these positions contribute to the interaction of phero-mones with compatible pheromone receptors (Riquelme et al.,2005; Szabo et al., 2002). Pheromones should not interact withreceptors encoded by the same matB allele (Kües et al., 2011;Riquelme et al., 2005). Guesses could be made as to which of thepheromones might interact with which foreign receptors. How-ever, due to the variable positions of alleles of pheromone precur-sor and pheromone receptor genes within the alleles of the matBlocus, a clear answer of whether products of alleles of a specificpheromone precursor gene will interact with only allelic productsof a defined receptor gene can only be given by experimental work.

7. Resources for mining Ganoderma genomes

Although the INSD (International Nucleotide Sequence Data-base), such as NCBI and EMBL, are the most reliable sources of pub-lic information, databases for particular taxonomic groups are alsoneeded to help to provide rich information related to non-sequence information of the species. Furthermore, the completegenome sequences of various Ganoderma species and strains, alongwith the transcriptomes of various developmental stages fromGanoderma species or strains, are becoming more available. Thereis a need for a centralized resource that can be used to integrateand compare various genome and transcriptome data sets. Severalefforts have been taken to meet this challenge.

Two web resources are available for Ganoderma. One has beenset up by JGI (Grigoriev et al., 2012) and is shown in Fig. 7a. TheJGI portal provides various data for the JGI sequencing projects. Itprovides: (1) a genome browser, so that users can browse thegenes of the sequenced genome; (2) gene annotations based onGO, KEGG, KOG, CLUSTERS, and SYNTENY; and (3) a download pagefor the sequences of scaffolds, coding sequences, transcripts, pro-teins, EST clusters, and gene models. The portal is well designed,user friendly, and aesthetically pleasing. The data are well-orga-nized and can be easily searched and downloaded.

The second web resource is set up and maintained by IMPLAD(Chen et al., 2012) and is shown in Fig. 7b. The web server is asso-ciated with the whole genome sequencing project led by IMPLAD.Similar to the JGI’s web portal, it provides a page allowing users todownload various types of data related to IMPLAD’s G. lucidum gen-ome sequencing projects. Two categories of data are provided. Oneis the primary experimental data, which includes the genomeassemblies, other genome-sequencing data, 454 transcriptomicdata, and RNA-Seq data. These can be downloaded from GenBankunder the project accession PRJNA71455. The other category isthe secondary analysis data, which includes gene models, geneannotations, compound structures, and gene clusters. Tools canbe divided further into 4 sub-categories and support functions,such as: (1) database searches based on sequence similarity; (2)the retrieval of annotations and sequences of a list of genes, andthe IDs of genes around a particular gene of interest; (3) the cura-tion of gene models; and (4) the identification of gene clusters andthe mapping of genes to chromosomes.

One of the limitations for the above web resources is that theyfocused on particular sequencing projects. As a result, these webresources lack integration with other sources of Ganoderma infor-mation. To provide an integrated information hub, we are currentlyconstructing another web resource Gano portal at IMPLAD (http://www.medfungi.org/gano). The Gano portal aims to provide a cen-tralized resource for these datasets, because of the rapid genera-tion of genome and transcriptome data sets. The Gano portalcontains the following pages: (1) a Ganoderma diversity page,which list the reported Ganoderma species and their related infor-mation; (2) a Gano identification page, which allows users toupload a DNA barcode sequence for the Gano sample under their


http://www.medfungi.org/gano

http://www.medfungi.org/gano

http://genome.JGI.doe.gov/Gansp1/Gansp1.home.html

http://www.medfungi.org/galu



study. The sequences will then be compared to the backend data-base for species determination; (3) a Gano genome page, whichlists all the whole genome sequencing projects on Gano, their affil-iated publications, and if there are any, the affiliated datasets; (4) aGano transcriptome page, which lists all Gano transcriptome pro-jects, their affiliated publications, and the affiliated data sets; (5)a gene search page, which provides users a BLAST searchable data-base that combines all coding sequences, transcripts, and proteinsequences from various whole genome sequencing and transcrip-tome projects; (6) a download page, which keeps a reference setof genomes, coding sequences transcripts, and proteins for down-load; and (7) a page listing the Ganoderma type strains and organi-zations maintaining their culture collections. We hope thatthrough community effort, this portal could support the retrievaland comparison of genomic and transcriptomic data related toGanoderma.

8. Discussion

8.1. Major discoveries

One of the key problems cited in numerous reports is the diffi-culty in defining the species identity of the samples involved. Com-parisons of the genome sequences of 2 of the G. lucidum strains andtranscript sequences identified from the 4 transcriptomic studieshave shown high degree of discrepancies. Although, these differ-ences might be attributed to different developmental stages, differ-ent growth conditions and different sequencing technologies, thepresence of such large degree of discrepancies cast doubt on theidentical species origin of the samples claimed in these studies. For-tunately, the wealth of information provided by the nuclear andmitochondrial genomes will provide a comparative resource forthe discovery of novel molecular markers for species determination.

Similar to all other genome sequencing projects, the most valu-able output of these studies is the identification of genes that areinvolved in various biological processes. For example, 24 gene clus-ters have been discovered in the genome of G. lucidum strainG.260125-1, which are potentially responsible for the biosynthesisof biologically active compounds. Particularly, the analyses of theCYP450 genes and genes at the mating type loci can serve as goodexamples for exploiting genomic information to discover genesthat are responsible for important biochemical and developmentalprocesses. Additional studies can be conducted to compare genesthat are involved in lignin degradation and plant pathogenesis,which might result in the identification of genes that are responsi-ble for these processes. Potential drug targets can also be identifiedby comparing the genetic makeup of infective and non-infectiveGanoderma spp. This information on genes and pathways also laysthe foundation for marker-guided breeding, as well as using meta-bolic engineering technologies to overproduce active chemicalcompounds or ligninolytic enzymes of interests.

8.2. Limitations

There are several limitations in these current studies, such asunconfirmed species identity, biased species sampling, limitingsample size and less than ideal sequencing quality. First, most ofthe studies described above focused on G. lucidum, as it is thebest-studied species of Ganoderma for its pharmacological activi-ties. The research groups conducting these studies are mostly fromChina, where G. lucidum is of particular economic importance.However, the species identity in the G. lucidum complex lack con-sensus. The medicinal species has been assumed to be G. lucidum,but evidence has emerged that this is a different species(Moncalvo et al., 1995). Wang et al. (2009) divided Asian speci-


mens classified as G. lucidum into two clades; both were separatedfrom the European G. lucidum. One clade, composed of tropical col-lections, represented G. multipileum; while the identity of the otherclade was unknown. Later, Wang et al. (2012) recognized theunknown clade as G. sichuanense. Another paper by Cao et al.(2012) found that the holotype of G. sichuanense was not conspe-cific with the unknown clade and proposed the unknown cladeas a new species G. lingzhi, representing the widely cultivated spe-cies in China. Second, only 2 defined strains of G. lucidum have beensequenced so far with genomes being assembled. Considering thelarge degree of sequence variations observed between the 2 gen-omes, a larger sample size is needed to paint a complete pictureof the sequence variations at the genome level for G. lucidum,which are critical for breeding. Lastly, the quality of the availableassemblies varies significantly, as the scaffolds can be placed tochromosomes in only one of the three completed assemblies. Theless than perfect assembly would hinder the discovery of geneclusters as well as the understanding of the history of genome evo-lution, such as gene duplication, rearrangement, etc. (A paper on anupdated nomenclature of these species is presented in this SpecialIssue (in this issue) and also refer to the Editorial (in this issue).)

8.3. Future directions

The availability of genome sequences have ushered in a new eraof research on Ganoderma spp. Future work can be divided intoefforts focused on (1) genome sequencing projects alone; (2) utili-zation of genome information for functional genome studies; and(3) application of Ganoderma spp. in the areas of healthcare, bio-energy production and plant protections. For additional genomesequencing projects, better planning would be beneficial. Forexample, the list of species to be sequenced should be prioritizedbased on their economic importance and/or social impact. Thesample size for the same species should be increased to understandbetter the intra-specific and inter-specific variations. The priorityfor sequencing either the nuclear genome or the mitochondrialgenome should be considered. For example, if the goal is to developnew markers for species determination, sequencing of the orga-nelle genome probably should take precedence over the sequenc-ing of the nuclear genome, as the size of organelle genome isabout 1/1000–1/500 of that of the nuclear genome. Assemblies ofhigher quality are preferred and additional technologies such asthe construction of genetic or physical maps should be taken intoconsideration during the planning phase of any projects attempt-ing to obtain high-quality assemblies.

With the availability of sequences of more genomes, in-depthgenome information mining and the development of functionalgenomics tools in order to validate the functions of predicted genesand gene clusters will become the crucial tasks. The interest inGanoderma spp. rose from their potential applications in healthcare, bio-energy production and plant protection. As a result, gen-ome studies have to be conducted with these potential applicationsin mind. Species or strains with particular characteristics, such asthe ability to produce a larger variety of chemical compounds or lig-ninolytic enzymes with high activities should be studied first andgenomic information should be integrated with those data onchemical or enzymatic activities. A centralized resource, which willbe maintained for long term, will allow researchers working onparticular aspects of Ganoderma to access and exploit the rich infor-mation generated from these genome sequencing projects.

9. Author contributions

C.L. designed and organized the writing of the paper; C.L. andJ.L. wrote Sections 1–3, 7 and 8; G.Y. and H.S. wrote Section 4;




D.N. wrote Section 5; U.K. wrote Section 6; J.H.Z. critically reviewedthe manuscript. X.C.W. created and maintains the Gano portal. Allauthors have read and agreed to the contents of this paper.

Acknowledgements

We thank Mr. Kai Wu for helping with comparing the mito-chondrial genomes and an anonymous reviewer for constructiveadvice on the manuscript. This study was supported by a researchgrant for Returned Overseas Chinese Scholars, the Ministry ofHuman Resources of China (No. 431207) granted to C. Liu, Programfor Changjiang Scholars and Innovative Research Teams in Univer-sities of Ministry of Education of China (Grant No. IRT1150) andgrants from the National Science Foundation of China (Grant No.81373912 to C. Liu). The funders had no role in study design, datacollection and analysis, decision to publish, or preparation of themanuscript.

Appendix A. Supplementary data

Supplementary data associated with this article can be found, inthe online version, at http://dx.doi.org/10.1016/j.phytochem.2014.11.019.

References

Adaskaveg, J.E., Gilbertson, R.L., 1986. Cultural studies and genetics of sexuality ofGanoderma lucidum and G. tsugae in relation to the taxonomy of the G. lucidumcomplex. Mycologia 78, 694–705.

Adaskaveg, J.E., Gilbertson, R.L., Blanchette, R.A., 1990. Comparative studies ofdelignification caused by Ganoderma species. Appl. Environ. Microbiol. 56,1932–1943.

Andersen, M.R., Nielsen, J.B., Klitgaard, A., Petersen, L.M., Zachariasen, M., Hansen,T.J., Blicher, L.H., Gotfredsen, C.H., Larsen, T.O., Nielsen, K.F., Mortensen, U.H.,2013. Accurate prediction of secondary metabolite gene clusters in filamentousfungi. Proc. Natl. Acad. Sci. U.S.A. 110, E99–E107.

Asante-Owusu, R.N., Banham, A.H., Böhnert, H.U., Mellor, E.J., Casselton, L.A., 1996.Heterodimerization between two classes of homeodomain proteins in themushroom Coprinus cinereus brings together potential DNA-binding andactivation domains. Gene 172, 25–31.

Berne, S., Podobnik, B., Zupanec, N., Novak, M., Krasevec, N., Turk, S., Korosec, B., Lah,L., Suligoj, E., Stojan, J., Gobec, S., Komel, R., 2012. Virtual screening yieldsinhibitors of novel antifungal drug target, benzoate 4-monooxygenase. J. Chem.Inf. Model. 52, 3053–3063.

Berovic, M., Habijanic, J., Zore, I., Wraber, B., Hodzar, D., Boh, B., Pohleven, F., 2003.Submerged cultivation of Ganoderma lucidum biomass and immunostimulatoryeffects of fungal polysaccharides. J. Biotechnol. 103, 77–86.

Berovic, M., Habijanic, J., Boh, B., Wraber, B., Petravic-Tominac, V., 2012. Productionof Lingzhi or Reishi medicinal mushroom, Ganoderma lucidum (W.Curt. :Fr.) P.Karst. (higher Basidiomycetes), biomass and polysaccharides by solid statecultivation. Int. J. Med. Mushrooms 14, 513–520.

Binder, M., Justo, A., Riley, R., Salamov, A., Lopez-Giraldez, F., Sjokvist, E., Copeland,A., Foster, B., Sun, H., Larsson, E., Larsson, K.H., Townsend, J., Grigoriev, I.V.,Hibbett, D.S., 2013. Phylogenetic and phylogenomic overview of thePolyporales. Mycologia 105, 1350–1373.

Boh, B., 2013. Ganoderma lucidum: a potential for biotechnological production ofanti-cancer and immunomodulatory drugs. Recent Pat. Anticancer DrugDiscovery 8, 255–287.

Brakhage, A.A., Schroeckh, V., 2011. Fungal secondary metabolites – Strategies toactivate silent gene clusters. Fungal Genet. Biol. 48, 15–22.

Bullerwell, C.E., Lang, B.F., 2005. Fungal evolution: the case of the vanishingmitochondrion. Curr. Opin. Microbiol. 8, 362–369.

Burger, G., Gray, M.W., Lang, B.F., 2003. Mitochondrial genomes: anything goes.Trends Genet. 19, 709–716.

Cao, Y., Wu, S.H., Dai, Y.C., 2012. Species clarification of the prize medicinalGanoderma mushroom ‘‘Lingzhi’’. Fungal Divers. 56, 49–62.

Chen, S., Xu, J., Liu, C., Zhu, Y., Nelson, D.R., Zhou, S., Li, C., Wang, L., Guo, X., Sun, Y.,Luo, H., Li, Y., Song, J., Henrissat, B., Levasseur, A., Qian, J., Li, J., Luo, X., Shi, L., He,L., Xiang, L., Xu, X., Niu, Y., Li, Q., Han, M.V., Yan, H., Zhang, J., Chen, H., Lv, A.,Wang, Z., Liu, M., Schwartz, D.C., Sun, C., 2012. Genome sequence of the modelmedicinal mushroom Ganoderma lucidum. Nat. Commun. 3, 913.

Chigu, N.L., Hirosue, S., Nakamura, C., Teramoto, H., Ichinose, H., Wariishi, H., 2010.Cytochrome P450 monooxygenases involved in anthracene metabolism by thewhite-rot basidiomycete Phanerochaete chrysosporium. Appl. Microbiol.Biotechnol. 87, 1907–1916.

Chinese Pharmacopoeia Commission, 2010. Pharmacopoeia of the People’s Republicof China. Chinese Medical Science, Beijing, China.


Chu, C.L., Yu, Y.L., Kung, Y.C., Liao, P.Y., Liu, K.J., Tseng, Y.T., Lin, Y.C., Hsieh, S.S.,Chong, P.C., Yang, C.Y., 2012. The immunomodulatory activity of meningococcallipoprotein Ag473 depends on the conformation made up of the lipid andprotein moieties. PLoS ONE 7, e40873.

Clinton, P.W., Buchanan, P.K., Wilkie, J.P., Smaill, S.J., Kimberley, M.O., 2009.Decomposition of Nothofagus wood in vitro and nutrient mobilization by fungi.Can. J. For. Res. 39, 2193–2202.

da Coelho-Moreira, J.S., Bracht, A., de Souza, A.C., Oliveira, R.F., de Sa-Nakanishi, A.B.,de Souza, C.G., Peralta, R.M., 2013. Degradation of diuron by Phanerochaetechrysosporium: role of ligninolytic enzymes and cytochrome P450. Biomed. Res.Int. 2013, 251354.

de Andrade, F.A., Calonego, F.W., Severo, E.T.D., Furtado, E.L., 2012. Selection of fungifor accelerated decay in stumps of Eucalyptus spp. Bioresour. Technol. 110, 456–461.

Elliott, M.L., Broschat, T.K., 2000. Ganoderma Butt Rot in Plants. Fact Sheet PP-54.University of Florida, The Institute of Food and Agriculture Sciences (IFAS),Gainsville, FL.

Faber, B.W., van Gorcom, R.F., Duine, J.A., 2001. Purification and characterization ofbenzoate-para-hydroxylase, a cytochrome P450 (CYP53A1), from Aspergillusniger. Arch. Biochem. Biophys. 394, 245–254.

Flood, J., Bridge, P.D., Holderness, M. (Eds.), 2000. Ganoderma Diseases of PerennialCrops. CABI Publishing, Wallingford, UK.

Floudas, D., Binder, M., Riley, R., Barry, K., Blanchette, R.A., Henrissat, B., Martínez,A.T., Otillar, R., Spatafora, J.W., Yadav, J.S., Aerts, A., Benoit, I., Boyd, A., Carlson,A., Copeland, A., Coutinho, P.M., de Vries, R.P., Ferreira, P., Findley, K., Foster, B.,Gaskell, J., Glotzer, D., Górecki, P., Heitman, J., Hesse, C., Hori, C., Igarashi, K.,Jurgens, L.A., Kallen, N., Kersten, P., Kohler, A., Kües, U., Kumar, T.K., Kuo, A.,LaButti, K., Larrondo, L.F., Lindquist, E., Ling, A., Lombard, V., Lucas, S., Lundell, T.,Martin, R., MyLaughlin, D.J., Morgenstern, I., Morin, E., Murat, C., Nagy, L.G.,Nolan, M., Ohm, R.A., Patyshakuliyeva, A., Rokas, A., Ruiz-Dueñas, F.J., Sabat, G.,Salamov, A., Samejima, M., Schmutz, J., Slot, J.C., John, F.S., Stenlid, J., Sun, H.,Sun, S., Syed, K., Tsang, A., Wiebenga, A., Young, D., Pisabarro, A., Eastwood, D.,Martin, F., Cullen, D., Grigoriev, I.V., Hibbett, D.S., 2012. The paleozoic origin ofenzymatic lignin decomposition reconstructed from 31 fungal genomes.Science 336, 1715–1719.

Ghikas, D.V., Kouvelis, V.N., Typas, M.A., 2010. Phylogenetic and biogeographicimplications inferred by mitochondrial intergenic region analyses and ITS1-5.8S-ITS2 of the entomopathogenic fungi Beauveria bassiana and B. brongniartii.BMC Microbiol. 10, 174.

Glen, M., Bougher, N.L., Francis, A.A., Nigg, S.Q., Lee, S.S., Irianto, R., Barry, K.M.,Beadle, C.L., Mohammed, C.L., 2009. Ganoderma and Amauroderma speciesassociated with root-rot disease of Acacia mangium plantation tress in Indonesiaand Malaysia. Australian Plant Path. 38, 345–356.

Grigoriev, I.V., Nordberg, H., Shabalov, I., Aerts, A., Cantor, M., Goodstein, D., Kuo, A.,Minovitsky, S., Nikitin, R., Ohm, R.A., Otillar, R., Poliakov, A., Ratnere, I., Riley, R.,Smirnova, T., Rokhsar, D., Dubchak, I., 2012. The genome portal of theDepartment of Energy Joint Genome Institute. Nucleic Acids Res. 40, D26–32.

Habijanic, J., Berovic, M., Boh, B., Wraber, B., Petravic-Tominac, V., 2013. Productionof biomass and polysaccharides of Lingzhi or Reishi medicinal mushroom,Ganoderma lucidum (W.Curt.:Fr.) P. Karst. (higher Basidiomycetes), bysubmerged cultivation. Int. J. Med. Mushrooms 15, 81–90.

Hennesey, C., Daly, A., 2007. Ganoderma Diseases. NSW Government, Orange, NSW,Australia. Agnote No: 167.

Henze, K., Martin, W., 2003. Evolutionary biology: essence of mitochondria. Nature426, 127–128.

Hirosue, S., Tazaki, M., Hiratsuka, N., Yanai, S., Kabumoto, H., Shinkyo, R., Arisawa,A., Sakaki, T., Tsunekawa, H., Johdo, O., Ichinose, H., Wariishi, H., 2011. Insightinto functional diversity of cytochrome P450 in the white-rot basidiomycetePhanerochaete chrysosporium: involvement of versatile monooxygenase.Biochem. Biophys. Res. Commun. 407, 118–123.

Huang, Y.H., Wu, H.Y., Wu, K.M., Liu, T.T., Liou, R.F., Tsai, S.F., Shiao, M.S., Ho, L.T.,Tzean, S.S., Yang, U.C., 2013. Generation and analysis of the expressed sequencetags from the mycelium of Ganoderma lucidum. PLoS ONE 8, e61127.

Hushiarian, R., Yusof, N.A., Dutse, S.W., 2013. Detection and control of Ganodermaboninense: strategies and perspectives. Springerplus 2, 555.

Ichinose, H., Wariishi, H., Tanaka, H., 2002. Identification and characterization ofnovel cytochrome P450 genes from the white-rot basidiomycete, Coriolusversicolor. Appl. Microbiol. Biotechnol. 58, 97–105.

Irianto, R.S.B., Barry, K., Hidayati, N., Ito, S., Fiani, A., Rimbawanto, A., Mohammed, C.,2006. Incidence and spatial analysis of root rot of Acacia mangium in Indonesia.J. Trop. For. Sci. 18, 157–165.

James, T.Y., Sun, S., Li, W., Heitman, J., Kuo, H.C., Lee, Y.H., Asiegbu, F.O., Olson, A.,2013. Polyporales genomes reveal the genetic architecture underlyingtetrapolar and bipolar mating systems. Mycologia 105, 1374–1390.

Jerneren, F., Oliw, E.H., 2012. The fatty acid 8,11-diol synthase of Aspergillusfumigatus is inhibited by imidazole derivatives and unrelated to PpoB. Lipids 47,707–717.

Jin, X., Ruiz Beguerie, J., Sze, D.M., Chan, G.C., 2012. Ganoderma lucidum (Reishimushroom) for cancer treatment. In: Cochrane Database Syst. Rev. 6. http://dx.doi.org/10.1002/14651858.CD14007731.pub14651852, CD007731.

Kirk, P.N., Cannon, P.F., Minter, D.W., Stalpers, J.A., 2011. Dictionary of the Fungi,tenth ed. CABI Europe, Croydon, UK.

Kües, U., Asante-Owusu, R.N., Mutasa, E.S., Tymon, A.M., Pardo, E.H., O’Shea, S.F.,Göttgens, B., Casselton, L.A., 1994. Two classes of homeodomain proteins specifythe multiple A mating types of the mushroom Coprinus cinereus. Plant Cell 6,1467–1475.




http://refhub.elsevier.com/S0031-9422(14)00502-0/h0005





























































































































http://dx.doi.org/10.1002/14651858.CD14007731.pub14651852

http://dx.doi.org/10.1002/14651858.CD14007731.pub14651852









Kües, U., James, T.Y., Heitman, J., 2011. Mating type in Basidiomycetes: unipolar,bipolar, and tetrapolar patterns of sexuality. In: Pöggeler, S., Wöstemeyer, J.(Eds.), Evolution of Fungi and Fungal-like Organisms, vol. XIV. Springer Verlag,Berlin, pp. 97–160.

Koch, C., Fielding, A.J., Brodhun, F., Bennati, M., Feussner, I., 2012. Linoleic acidpositioning in psi factor producing oxygenase A, a fusion protein with anatypical cytochrome P450 activity. FEBS J. 279, 1594–1606.

Koch, C., Tria, G., Fielding, A.J., Brodhun, F., Valerius, O., Feussner, K., Braus, G.H.,Svergun, D.I., Bennati, M., Feussner, I., 2013. A structural model of PpoA derivedfrom SAXS-analysis-implications for substrate conversion. Biochim. Biophys.Acta 1831, 1449–1457.

Lee, K.H., Morris-Natschke, S.L., Yang, X., Huang, R., Zhou, T., Wu, S.F., Shi, Q.,Itokawa, H., 2012. Recent progress of research on medicinal mushrooms, foods,and other herbal products used in traditional Chinese medicine. J. Tradit.Complement. Med. 2, 84–95.

Li, J., Zhang, J., Chen, H., Chen, X., Lan, J., Liu, C., 2013. Complete mitochondrialgenome of the medicinal mushroom Ganoderma lucidum. PLoS ONE 8,e72038.

Liang, C.X., Li, Y.B., Xu, J.W., Wang, J.L., Miao, X.L., Tang, Y.J., Gu, T., Zhong, J.J., 2010.Enhanced biosynthetic gene expressions and production of ganoderic acids instatic liquid culture of Ganoderma lucidum under phenobarbital induction. Appl.Microbiol. Biotechnol. 86, 1367–1374.

Liu, D., Gong, J., Dai, W., Kang, X., Huang, Z., Zhang, H.-M., Liu, W., Liu, L., Ma, J., Xia,Z., 2012. The genome of Ganderma lucidum provide insights into triterpensebiosynthesis and wood degradation. PLoS ONE 7, e36146.

Luo, H., Sun, C., Song, J., Lan, J., Li, Y., Li, X., Chen, S., 2010. Generation and analysis ofexpressed sequence tags from a cDNA library of the fruiting body of Ganodermalucidum. Chin. Med. 5, 9.

Maeda, Y., Kajiwara, K., Ohtaguchi, K., 2001. Manganese peroxidase gene of theperennial mushroom Elfvingia applanata: cloning and evaluation of itsrelationship with lignin degradation. Biotechnol. Lett. 23, 103–109.

Martínez, A.T., Rencoret, J., Nieto, L., Jiménet-Barbero, J., Gutiérrez, A., del Río, J.C.,2011. Selective lignin and polysaccharide removal in natural fungal decay ofwood as evidenced by in situ structural analyses. Environ. Microbiol. 13, 96–107.

Mendoza, G., Guzman, G., Ramirez-Guillen, F., Luna, M., Trigos, A., 2011. Ganodermaoerstedii (Fr.) Murrill (higher Basidiomycetes), a tree parasite species in Mexico:taxonomic description, rDNA study, and review of its medical applications. Int.J. Med. Mushrooms 13, 545–552.

Miller, R.N.G., Holderness, M., Bridge, P.D., 2000. Molecular and morphologicalcharacterization of Ganoderma in oil-palm plantings. In: Flood, J., Bridge, P.D.,Holderness, M. (Eds.), Ganoderma Diseases of Perennial Crops. CABI Publishing,Wallingford, UK, pp. 159–182.

Moncalvo, J.M., Buchanan, P.K., 2008. Molecular evidence for long distance dispersalacross the Southern Hemisphere in the Ganoderma applanatum-australe speciescomplex (Basidiomycota). Mycol. Res. 112, 425–436.

Moncalvo, J.M., Wang, H.F., Hseu, R.S., 1995. Gene phylogeny of the Ganodermalucidum complex based on ribosomal DNA sequences. Comparison withtraditional taxonomic characters. Mycol. Res. 99, 1489–1499.

Naher, L., Yusuf, U.K., Ismail, A., Tan, S.G., Monal, M.M.A., 2013. Ecological status ofGanoderma and basal stem rot disease of oil palms (Elaeis guineensis Jacq.). AJCS7, 1723–1727.

Nakayama, N., Takemae, A., Shoun, H., 1996. Cytochrome P450foxy, a catalyticallyself-sufficient fatty acid hydroxylase of the fungus Fusarium oxysporum. J.Biochem. 119, 435–440.

Nasir, N., 2005. Diseases caused by Ganoderma spp. on perennial crops in Pakistan.Mycopathologia 159, 119–121.

Nelson, D.R., Goldstone, J.V., Stegeman, J.J., 2013. The cytochrome P450 genesislocus: the origin and evolution of animal cytochrome P450s. Philos. Trans. R.Soc. B Biol. Sci. 368, 20120474.

Nusaibah, S.A., Latiffah, Z., Hassaan, A.R., 2011. ITS-PCR-RFLP analysis of Ganodermasp. infecting industrial crop. Pertanika J. Trop. Agric. Sci. 34, 83–91.

Paterson, R.R.M., 2006. Ganoderma – a therapeutic fungal biofactory.Phytochemistry 67, 1985–2001.

Paterson, R.R.M., 2007. Ganoderma disease of oil palm – a white rot perspectivenecessary for integrated control. Crop Protect. 26, 1369–1376.

Pilotti, C.A., 2005. Stem rots of oil plam caused by Ganoderma boninense: pathogenbiology and epidemiology. Mycopathologia 159, 129–137.

Pilotti, C.A., Sanderson, F.R., Aitken, E.A.B., 2003. Genetic structure of a population ofGanoderma boninense on oil palm. Plant. Pathol. 52, 455–463.

Podobnik, B., Stojan, J., Lah, L., Krasevec, N., Seliskar, M., Rizner, T.L., Rozman, D.,Komel, R., 2008. CYP53A15 of Cochliobolus lunatus, a target for naturalantifungal compounds. J. Med. Chem. 51, 3480–3486.

Popovic, V., Zivkovic, J., Davidovic, S., Stevanovic, M., Stojkovic, D., 2013.Mycotherapy of cancer: an update on cytotoxic and antitumor activities ofmushrooms, bioactive principles and molecular mechanisms of their action.Curr. Top. Med. Chem. 13, 2791–2806.

Ren, A., Li, M.J., Shi, L., Mu, D.S., Jiang, A.L., Han, Q., Zhao, M.W., 2013. Profiling andquantifying differential gene transcription provide insights into ganoderic acidbiosynthesis in Ganoderma lucidum in response to methyl jasmonate. PLoS ONE8, e65027.

Rai, M., Tidke, G., Wasser, S.P., 2005. Therapeutic potential of mushrooms. Nat. Prod.Rad. 4, 246–257.

Rakib, M.R.M., Bong, C.-F.J., Khairulmazmi, A., Idris, A.S., 2014. Genetic andmorphological diversity of Ganoderma species isolated from infected oil palms(Elaeis guineensis). Int. J. Agric. Biol. 16, 691–699.


Rees, R.W., Flood, J., Hasan, Y., Wills, M.A., Cooper, R.M., 2012. Ganoderma boninensebasidiospores in oil palm plantations: evaluation of their possible role in stemroots of Elaeis guineensis. Plant. Pathol. 61, 567–578.

Riquelme, M., Challen, M.P., Casselton, L.A., Brown, A.J., 2005. The origin of multipleB mating specificities in Coprinus cinereus. Genetics 170, 1105–1119.

Ropelewski, A.J., Nicholas, H.B., Mendez, R.R.G., 2010. MPI-PHYLIP: Parallelizingcomputationally intensive phylogenetic analysis routines for the analysis oflarge protein families. PLOS ONE 5, e13999.

Sankaran, K.V., Bridge, P.D., Gokulapalan, C., 2005. Ganoderma diseases of perennialcrops in India – an overview. Mycopathologia 159, 143–152.

Sezutsu, H., Le Goff, G., Feyereisen, R., 2013. Origins of P450 diversity. Philos. Trans.R. Soc. B Biol. Sci. 368, 20120428.

Shendure, J., Ji, H., 2008. Next-generation DNA sequencing. Nat. Biotechnol. 26,1135–1145.

Shi, L., Ren, A., Mu, D., Zhao, M., 2010. Current progress in the study on biosynthesisand regulation of ganoderic acids. Appl. Microbiol. Biotechnol. 88, 1243–1251.

Silveira, J.S., Emmert, E., Sternadt, G.H., Mendes, J.C., Almeisa, G.F., 2009. Decaysusceptibility of Amazon wood species from Brazil against white rot and brownrot decay fungi. Holzforschung 63, 767–772.

Smith, B.J., Sivasithamparam, K., 2000. Internal transcribed spacer ribosomal DNAsequence of five species of Ganoderma from Australia. Mycol. Res. 104, 943–951.

Soares, A.A., de Sa-Nakanishi, A.B., Bracht, A., da Costa, S.M., Koehnlein, E.A., deSouza, C.G., Peralta, R.M., 2013. Hepatoprotective effects of mushrooms.Molecules 18, 7609–7630.

Son, E., Au-Yeung, T.T., Yang, C.Y.H., Breuil, C., 2010. Diversity and decay ability ofbasidiomycetes isolated from lodgepole pines killed by the mountain pinebeetle. Can. J. Microbiol. 57, 33–41.

Spit, A., Hyland, R.H., Mellor, E.J., Casselton, L.A., 1998. A role for heterodimerizationin nuclear localization of a homeodomain protein. Proc. Natl. Acad. Sci. U.S.A.95, 6228–6233.

Subramanian, V., Yadav, J.S., 2008. Regulation and heterologous expression of P450enzyme system components of the white rot fungus Phanerochaetechrysosporium. Enzyme Microbiol. Technol. 43, 205–213.

Syed, K., Nelson, D.R., Riley, R., Yadav, J.S., 2013. Genomewide annotation andcomparative genomics of cytochrome P450 monooxygenases (P450s) in thepolypore species Bjerkandera adusta, Ganoderma sp. and Phlebia brevispora.Mycologia 105, 1445–1455.

Syed, K., Shale, K., Pagadala, N.S., Tuszynski, J., 2014. Systematic identification andevolutionary analysis of catalytically versatile cytochrome p450monooxygenase families enriched in model basidiomycete fungi. PLoS ONE 9,e86683.

Szabo, Z., Tonnis, M., Kessler, H., Feldbrügge, M., 2002. Structure-function analysisof lipopeptide pheromones from the plant pathogen Ustilago maydis. Mol.Genet. Genomics 268, 362–370.

Tamura, K., Peterson, D., Peterson, N., Stecher, G., Nei, M., Kumar, S., 2011. MEGA5:molecular evolutionary genetics analysis using maximum likelihood,evolutionary distance, and maximum parsimony methods. Mol. Biol. Evol. 28,2731–2739.

Tsitsigiannis, D.I., Kowieski, T.M., Zarnowski, R., Keller, N.P., 2005. Three putativeoxylipin biosynthetic genes integrate sexual and asexual development inAspergillus nidulans. Microbiology 151, 1809–1821.

US Forest Service, 2011. White mottled rot. Root rot that topples live aspen, USDepartment of Agriculture, Washington, DC.

Wang, C.H., Hsieh, S.C., Wang, H.J., Chen, M.L., Lin, B.F., Chiang, B.H., Lu, T.J., 2014.Concentration variation and molecular characteristics of soluble (1,3;1,6)-b-D-glucans in submerged cultivation products of Ganoderma lucidum mycelium. J.Agric. Food Chem. 62, 634–641.

Wang, D.M., Wu, S.H., Su, C.H., Peng, J.T., Shih, Y.H., Chen, L.C., 2009. Ganodermamultipileum, the correct name for ‘G. lucidum’ in tropical Asia. Bot. Stud. 50, 451–458.

Wang, X.C., Xi, R.J., Li, Y., Wang, D.M., Yao, Y.J., 2012. The species identity of thewidely cultivated Ganoderma, ‘G. lucidum’ (Ling-zhi), in China. PLoS ONE 7,e40857.

Weete, J.D., Abril, M., Blackwell, M., 2010. Phylogenetic distribution of fungalsterols. PLoS ONE 5, e10899.

Wen, H., Kang, S., Song, Y., Song, Y., Sung, S.H., Park, S., 2010. Differentiation ofcultivation sources of Ganoderma lucidum by NMR-based metabolomicsapproach. Phytochem. Anal. 21, 73–79.

Wu, G.S., Guo, J.J., Bao, J.L., Li, X.W., Chen, X.P., Lu, J.J., Wang, Y.T., 2013. Anti-cancerproperties of triterpenoids isolated from Ganoderma lucidum – a review. ExpertOpin. Investig. Drugs 22, 981–992.

Xu, T., Beelman, R.B., Lambert, J.D., 2012. The cancer preventive effects of ediblemushrooms. Anticancer Agents Med. Chem. 12, 1255–1263.

Xu, Z., Chen, X., Zhong, Z., Chen, L., Wang, Y., 2011. Ganoderma lucidumpolysaccharides: immunomodulation and potential anti-tumor activities. Am.J. Chin. Med. 39, 15–27.

Yadav, J.S., Doddapaneni, H., Subramanian, V., 2006. P450ome of the white rotfungus Phanerochaete chrysosporium: structure, evolution and regulationof expression of genomic P450 clusters. Biochem. Soc. Trans. 34, 1165–1169.

Yu, G.J., Wang, N., Huang, J., Yin, Y.L., Chen, Y.J., Jiang, S., Jin, Y.X., Lan, X.Q., Wong,B.H.C., Liang, Y., Sun, H., 2012. Deep insight into the Ganoderma lucidum bycomprehensive analysis of its transcriptome. PLOS ONE 8, e44031.

Zhou, X.W., Cong, W.R., Su, K.Q., Zhang, Y.M., 2013. Ligninolytic enzymes fromGanoderma spp.: current status and potential applications. Crit. Rev. Microbiol.39, 416–426.














































































































































































Ursula Kües studied biology at the RuhruniversityBochum (Germany) and graduated with focus in botanyin 1983. After a PhD study in microbiology at theTechnical University of Berlin and postdoctoral periodsat the Free University Berlin, Queen Mary and WestfieldCollege at the University of London and the Departmentof Plant Sciences in Oxford, she was awarded a Violetteand Samuel Glasstone Fellowship of the University ofOxford. In 1994, she became a group-leader at the ETHZurich in Switzerland, where she did her habilitation aspost-doctoral degree and obtained a venia legendi inmolecular microbiology and genetics. Since 2001, she is

Full Professor at the Georg-August-University in Göttingen where she chairs theDepartment of Molecular Wood Biotechnology and Technical Mycology. Herresearch field covers litter and wood decay fungi, developmental processes of
mushrooms, and molecular genetics, genomics and proteomics of basidiomycetes.
David R. Nelson graduated from the University of TexasHealth Science Centre at San Antonio with a PhD degreein 1985. After post-doctoral periods at the University ofTexas Medical School at Houston, and the University ofNorth Carolina at Chapel Hill, he became AssistantProfessor at University of Tennessee (Memphis) in 1994,where he was promoted to Full Professor in 2014. Prof.Nelson is a world-renowned expert on P450 classifica-tion and evolution. He has named more than 26,000P450 sequences and has participated in many genomeprojects in plants, fungi and animals.

Chang Liu graduated from the University of Minnesotain 2001, Twin Cities Campus, obtaining his PhD degree.After graduation, he worked in GlaxoSmithKline Corp(Research Triangle Park, USA), Yale University and TheUniversity of Hong Kong. He has been a Professor in theInstitute of Medicinal Plant Development, ChineseAcademy of Medical Science in Beijing since 2010. Hehas published in journals such as Nature Communica-tions, Science and PNAS. Recently, his research hasfocused on the genomic aspects of medicinal fungi andhe has been a primary contributor for the sequencingand analyses of the nuclear and mitochondrial genomes

of Ganoderma lucidum.

Guo-Jun Yu graduated in 2010 from the Henan Agri-cultural University at Zhengzhou, Henan Province,obtaining his Bachelor degree. Now he is working for hisPhD degree at the Wuhan University. He is interested inlectins, transcriptomics, proteomics and glycoproteo-mics of medicinal fungi.


Jianhui Zhang joined Professor Liu’s lab in 2012 as apostdoctoral fellow. For the two last years, he wasengaged in computational identification of gene familiesfrom medicinal plants (Salvia miltiorrhiza) and mito-chondrial genome analyses of medicinal fungi (Gano-derma lucidum). His research interest is using genomicinformation, molecular biology, bioinformatics andbiochemistry to understand problems of fundamentalimportance in plant and fungal biology.

Jianqin Li graduated from the China Agricultural Uni-versity in Beijing, obtaining her PhD degree in year2010. After graduation, she worked in the Institute ofMedicinal Plant Development, Chinese Academy ofMedical Science as a postdoctoral fellow. She had par-ticipated in Ganoderma lucidum genome sequencingprojects and published in journals such as NatureCommunications and PLOS ONE. Her works focused onthe mitochondrial genome and non-coding RNAs ofGanoderma lucidum. Recently, she took on a position as ascientific editor of the Journal of Agricultural Biotech-nology.

Xin-Cun Wang has worked on taxonomy and phylog-eny of Ganodermataceae Donk since 2005 in the Insti-tute of Microbiology, Chinese Academy of Sciences,Beijing and received his PhD degree in 2013 from theUniversity of Chinese Academy of Sciences. In the sameyear, he became a postdoctoral fellow in the Institute ofMedicinal Plant Development, Chinese Academy ofMedical Sciences, Bejing, focusing on the developmentand application of DNA barcoding technologies and theunderstanding of organelle genome evolution in fungiand plants. He had published several articles aboutspecies identity and identification in journals such as

Molecular Ecology Resources, Taxon, PLOS ONE etc., and identified the widelycultivated Ganoderma species ‘G. lucidum’ in China as G. sichuanense J.D. Zhao & X.Q.Zhang with his colleagues based on microscopic and molecular evidence.

Hui Sun obtained her PhD degree in 2000 under thesupervision of academician Lianhui Xie and ProfessorQiyin Lin, in Fujian Agricultural University. After grad-uation, she has focused on research of active proteins/peptides in medicinal fungi. She has finished a series ofworks on transcriptome, proteome, and active peptidesand proteins in medicinal fungi in the past years. Cur-rently, she is interested in lectins from medicinal fungiwhich may be developed to serve as antitumor medi-cines and useful glycan identification tools in the future.



Genome analysis of medicinal Ganoderma spp. with plant-pathogenic and saprotrophic life-styles

Documents

Transcript of Genome analysis of medicinal Ganoderma spp. with plant-pathogenic and saprotrophic life-styles