Isolation and ectopic expression of a bamboo MADS-box gene

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Chinese Science Bulletin Vol. 50 No. 3 February 2005 217

Chinese Science Bulletin 2005 Vol. 50 No. 3 217—224

Isolation and ectopic expression of a bamboo MADS-box gene TIAN Bo1,2, CHEN Yongyan1,3, YAN Yuanxin1

& LI Dezhu1

1. Laboratory of Plant Biodiversity and Biogeography, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650204, China;

2. Graduate School, Chinese Academy of Sciences, Beijing 100039, China;

3. College of Bioengineering, Dalian University, Dalian 116622, China Correspondence should be addressed to Li Dezhu (e-mail: dzl@mail. kib.ac.cn)

Abstract A cDNA named DlMADS18 was isolated from the young spikelets of the sweet bamboo, Dendrocalamus latiflorus by RACE. DNA sequence analysis showed that DlMADS18 was composed of full ORF and 3′UTR, but without 5′UTR. The cDNA contained 1039 nucleotides and encoded a putative protein of 249 amino acid residues. The gene displayed the structure of a typical plant MADS box gene, which consisted of an MADS domain, K domain, a short I region, and the C-terminal region. Phylogenetic analysis of plant MADS box genes based on amino acid se-quences revealed that DlMADS18 was grouped into the AGAMOUS-LIKE 6 (AGL6)-like subfamily. It was most likely homologous to the OsMADS6 of rice (Oryza sativa), with 88% sequence identity for the entire amino acid sequences. The DlMADS18 also showed relatively high amino acid sequence identity (59%) to AGL6 of Arabidopsis thaliana. To study the functions of DlMADS18, DlMADS18 cDNA clone driven by the CaMV 35S promoter was transformed into Arabidopsis plants. Transgenic plants of DlMADS18 exhibited the pheno-types of curled leaves, dwarfism, and early flowering with clustered terminal flowers. These results indicated that DlMADS18 may probably be involved in controlling the flowering time of D. latiflorus.

Keywords: Dendrocalamus latiflorus, MADS-box gene, AGL6-like transgenic plants.

DOI: 10.1360/982004-304

Since the 1990s, the study of the molecular and ge-netic mechanisms of floral development has become one of the very important hotspots for plant molecular biolo-gists, and rapid progress has been made. Floral develop-ment was influenced by multiple environmental and en-dogenous genetic inputs. Studies on plant molecular biol-ogy showed that the floral development was controlled by a complicated gene network[1,2]. In Arabidopsis thaliana, there are two kinds of genes related to flowering time, i.e. the floral repressive genes and the floral promotive genes. The interaction between them regulates flowering time[1—6]. The MADS box genes that encode regulatory proteins play important roles in both the formation of

flower meristem and the determination of floral organ identity[7—16]. A typical plant MADS-box gene is com-posed of an MADS domain, K domain, a short I region between the MADS and K domain, and the C-terminal region downstream of the K domain[17,18]. In Arabidopsis, some MADS-box genes act as floral repressor, such as Flowering LocusC (FLC)[8], Short Vegetative Phase (SVP)[9], and AGL27[19]. They regulate the floral transition negatively through repression of the floral pathway inte-grators FT, SUPPRESSOR OF OVEREXPRESSION OF CO 1 (SOC1), and LEAFY (LFY)[11] or by encoding a dosage-dependent repressor of flowering which prolong all vegetative stages in wild-type plants independently of photoperiodic control and vernalization[9,19]. Except for a few floral repressors, there are so many MADS-box genes that act as floral promoter, such as SOC1[20], APETALA1 (AP1)[7], CUALIFLOWER (CAL)[21], AGL24[12] and most of AGL2-like genes[16,22—24]. Their overexpression under the CaMV 35S promoter leads to early flowering and their mutants are later flowering ones. A phylogenetic analysis of the then known plant MADS-box genes recognize 12 major gene clades[16]. AGL6-like gene clade or subfamily was known in the plant MADS-box gene family several years ago[16—18]. Despite quite a number of AGL6-like genes had been cloned from diverse seed plants, the func-tion of AGL6-like genes remained poorly-known.

One of the satisfying things about the field of flower development was the applicability of the floral patterning mechanisms to a wide range of plant species[4]. Moreover, careful analysis in multiple distantly related species may help us better understand the floral patterning mechanisms. Bamboo is a group of very important economical plants. Although bamboo belongs to the grass family, the Poaceae, which includes rice and wheat, it owns several unique characteristics in blossom. The woody bamboo is peren-nial. Many of them have 30—60 years, even 120 years juvenile phase, and then engage in a suicidal bout of sex-ual reproduction[25]. More remarkably, the onset of flow-ering is synchronous among ramets, genets and stands in a range of spatial scales, while only a few species are annual flowering or periodic flowering[26—29]. All these special characters are the main obstacles to the study and utiliza-tion of bamboo. Based on analyses of different environ-mental cues, some hypotheses were made to explain the bamboo flowering [25,29—31], while other ones referred to the endogenous mechanism such as a biological clock[25,30,31]. However, the mechanism of bamboo flow-ering is still unclear, and furthermore, there are no reports on bamboo flowering development at molecular level. In this paper, we report the isolation and ectopic expression of a bamboo MADS-box gene belonging to AGL6-like subfamily. 1 Materials and methods

(ⅰ) Plant materials and growth conditions. Young

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218 Chinese Science Bulletin Vol. 50 No. 3 February 2005

spikelets were taken from in vitro flowering cultures of the sweet bamboo, Dendrocalamus latiflorus. The Columbia ecotype of Arabidopsis thaliana was used in this study. Escherichia coli DH5α was used as the recipients for rou-tine cloning experiments. Agrobacterium tumefaciens GV3101 was used for transformation of Arabidopsis plants. The plasmid pGEM-T and the binary vectors pWM 101 and pCambia 2301 were used to form the plant ex-pression vector. Plants of Arabidopsis were grown in growth chambers under long-day conditions (16-h light/8-h dark) at 22 . The light intensity of the growth ℃

chambers was 150 μE·m−2·s−1. (ⅱ) Inducing in vitro flowering. In vitro seedlings

of D. latiflorus were produced by using the methods of Zhang et al.[32]. The culture medium which induced preco-cious flowering was modified MS[33] medium with 5 mg/L VB1, 1 mg/L VB6, 1 mg/L Niacine (nicotinic acid), and 3/4 times of normal MS medium the concentrations of NH4NO3, KNO3, MgSO4·H2O, KH2PO4 and CaCl2·2H2O. Hypoplastic spikelets were induced when the modified MS medium contained 6 mg/L BA (6-Benzylaminopurine). Then these cultures were transferred to new modified MS medium containing 3 mg/L BA, 0.5 mg/L KT (Kinetin) and 10% coconut water to improve the quality of spikelets.

(ⅲ) Cloning of DlMADS18 cDNA from D. latiflorus and sequence analysis. mRNA was isolated from young spikelets of D. latiflorus. cDNA was synthesized from 500 ng of mRNA using SMARTTM RACE cDNA Amplifica-tion Kit (Clonetech, USA). According to the conservative sequence of MADS box genes in monocots, 3′-RACE (rapid amplification of cDNA ends) degenerate primer was designed at two bases before the initiation code, and 5′-RACE primer was unnecessary. The 3′-RACE primer was 5′-GCATGGGNAGSGGNAASGTNGASCTSAA-3′. Touch down PCR amplification was performed with 3′-RACE primer and UPM (universal primer a mix). PCR products were cloned into pGEM-T vectors (Promaga) and sequenced. Sequence analysis was carried out with the BLAST search and MegAlign (DNASTAR) program of the DNASTAR software package. The PAUP 4.0b10 pro-gram[34] was used to construct the phylogenetic tree with conceptual amino acid sequence of MADS box genes.

(ⅳ) Contruction of expression vector. A BamHⅠ- SalⅠ fragment containing cDNA ORF of DlMADS18 was amplified by PCR. This fragment was introduced into bi-nary vector pWM 101 under the control of CaMV 35S promoter, and then 35S:DlMADS18 fragment was cloned into binary vector pCambia 2301. Primers used to gener-ate this BamHⅠ-SalⅠ fragment were DlMB (5′-TTAAG- GATCCATGGGTAGGGGGAACGTGGA-3′) and DlMS (5′-AAATGTCGACTCAAAGAACCCACCCCAGCA-3′). These two primers contained the generated BamHⅠ (5′-GGATCC-3′, underlined) or SalⅠ recognition site

(5′-GTCGAC-3′, underlined) that facilitated the cloning of the cDNA.

(ⅴ) Plant transformation. The resulting plasmids were transformed into Agrobacterium tumefaciens GV3101 via the freeze-thaw method, then the Agrobacte-rium containing 35S:DlMADS18 were used to transform Arabidopsis plants using floral dip method[35]. Transfor-mants were selected on agar plates containing 1/2 times MS medium and 50 μg/mL kanamycin.

(ⅵ) Northern-blot analysis. Total RNA was iso-lated from leaves of 35S:DlMADS18 transgenic Arabi-dopsis plants. 20 μg of total RNA were fractionated on 1.5% formaldehyde-agarose gels[36] and transferred to Hybond N+ membranes. The membranes were prehybrid-ized for 20 min and hybridized with a 32P-labeled DNA probes overnight at 68℃ in the same solution (Perfect Hyb TM Plus Hybridization buffer, Sigma). After hy-bridization, the blot was washed one time with a solution containing 2×SSC and 0.5% SDS for 10 min at 68 then ℃washed twice with 0.5×SSC and 0.1% SDS for 20 min at 68 , ℃ respectively, followed by a wash with 0.1×SSC and 0.1% SDS for 20 min at the same temperature. The blots were then covered with plastic wrap and autoradiographed. The DNA probes specific for DlMADS18 was partial cDNA fragments (without MADS domain) amplified from cDNA clone through PCR. 2 Results

(ⅰ) Isolation of DlMADS18 gene and sequence analysis. We identified a cDNA clone named DlMADS18 from the young spikelets of D. latiflorus by using 3′- RACE method. The DlMADS18 cDNA was 1039 bp long and encoded a putative protein of 249 amino acid residues (Fig. 1). It was composed of full ORF and 3′UTR, but without 5′UTR. The gene contained the structure of a typical plant MADS box gene, which consisted of an MADS domain, K domain, a short I region, and the C- terminal region. Based on the amino acid sequence align-ment, DlMADS18 showed high identity to some MADS- box genes of other monocotyledons, 88%, 84%, 82%, 86%, 86%, 87%, 87%, 63% identity to OsMADS6 of rice, ZAG3 and ZAG5 of maize, HvAGL6 of barley, TaMADS12 of wheat, PaMADS1 of Poa annua, LpMADS4 of darnel, OMADS1 of orchid, respectively. Furthermore, DlMADS18 showed 59% identity to AGL6 of Arabidopsis (Fig. 2). Phylogenetic tree constructed based on the de-duced amino acid sequences of the MADS-box genes showed that DlMADS18 belonged to AGL6-like subfamily (GenBank Accession Number of DlMADS18 was AY599755) (Fig. 3).

(ⅱ) Construction of expression vector. The vector pCambia 2301 used in this paper contained neomycin phosphotransferase (NPTⅡ) gene and β-Glucuronidase (GUS) gene and was kanamycin resistance. First, the ORF of DlMADS18 was introduced into the vector pWM101

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Fig. 1. Nucleotide and deduced amino acid sequences of the DlMADS18 cDNA. The double underline indicated the MADS domain, with the single underline being the K domain. The positions of nucleotides and amino acids are shown on the left and right, respectively.

Fig. 2. Alignment of DlMADS18 with other plant MADS box proteins. (a) Sequence alignment of the MADS domains. The asterisks represented amino acid residues identical to the corresponding DlMADS18. The numbers on the right represented the percentage of amino acid residues identical to the MADS domain (M) of DlMADS18. (b) Alignment of the K domains. The asterisks were the same as repre-sented in (a). The numbers on the right represented the percentage of identical amino acid residues with the K domain (K) and the overall region (O) of DlMADS18.


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Fig. 3. Phylogenetic analysis of plant MADS box genes based on amino acid sequences. DlMADS18 was grouped into AGL6-like clade. The name of the DlMADS18 protein was underlined. Names of the plant species for each MADS box gene were listed behind the protein names. Numbers on major branches indicated bootstrap estimates for 1000 replicate analyses.

Fig. 4. Construction of the plant expression vector containing DlMADS18.

between BamHⅠ and SalⅠ, so the ORF was controlled by CaMV35S promoter and 35SpolyA. Then the 35Spro- ORF-35SpolyA segment was cloned into pCambia 2301 and generated an expression vector with GUS report gene

(Fig. 4). (ⅲ) Ectopic expression of DlMADS18 in transgenic

Arabidopsis plants. To investigate the function of DlMADS18, ectopic expression of DlMADS18 in trans-


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genic plants was applied. ORF of DlMADS18 driven by the CaMV35S promoter was transformed into Arabidopsis plants for functional analysis. Twenty-one independent 35S:DLMADS18 transgenic Arabidopsis T1 plants were obtained. Three plants were phenotypically indistinguish-able from wild-type plants, whereas the other 18 plants showed the identical novel phenotypes described below. These eighteen plants were small and produced stalkless and small curled leaves after germination (Fig. 5(a)), while wild-type plants produced more round leaves with petioles after germination (Fig. 5(b)). Later, these 35S:DlMADS18 plants flowered significantly earlier than wildtype plants by producing only 6—8 small curled

rosette leaves (Fig. 5(c)). At the same age, wild-type plants had about 11—15 round rosette leaves with petioles and remained in vegetative stage (Fig. 5(d)). These 35S:DlMADS18 plants had 2—3 small flowers clustered at the end of either main inflorescence or branches (Fig. 5(e)). The curled cauline leaves were sepal-like (Fig. 5(e)). No obvious alteration of floral organs was observed in 35S:DlMADS18 transgenic Arabidopsis plants. To explore if the severe phenotype observed in 35S:DlMADS18 transgenic plants was correlated with the level of DlMADS18 expression, Northern blot analysis was performed. The results indicated that high DlMADS18 expression was observed in the severe phenotype of the transgenic plants

Fig. 5. Phenotypic analysis of transgenic Arabidopsis plants ectopically expressing DlMADS18. (a) A 12-day-old 35S:DlMADS18 Arabidopsis plant grown on MS agar medium with curled rosette leaves (cr). (b) A 12-day-old wild-type Arabidopsis plant grown on MS agar medium with round rosette leaves (r). (c) A flowered 18-day-old 35S:DlMADS18 Arabidopsis plant grown on soil. fb, flower bud; cr, curled rosette leaves. (d) An 18-day-old wild-type Arabidopsis plant remained in vegetative development; (e) A 35S:DlMADS18 Arabi-dopsis plant grown on soil. tf, terminal flower; cc, curled cauline leaves.


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(Fig. 6). The DlMADS18 expression was nearly undetect-able in wild-type plants and those three 35S:DlMADS18 transgenic plants which were indistinguishable from wild-type plants (Fig. 6). These results indicated that the alteration of phenotypes observed in 35S:DlMADS18 transgenic Arabidopsis plants was due to the ectopic ex-pression of the bamboo DlMADS18 gene.

Fig. 6. The detection of DlMADS18 expression in 35S:DlMADS18 transgenic Arabidopsis by Northern blot analysis. 1, wild-type untrans-formed plants; 2, wild-type like 35S:DlMADS18 transgenic plants; 3, severe phenotype transgenic plants.

3 Dicussion Bamboo is of important economic value. In order to

investigate the role of MADS box genes in regulating bamboo flower development, an MADS box gene, DlMADS18 was cloned from D. latiflorus and character-ized in this study. We compared the DlMADS18 cDNA with many of the MADS-box genes identified in Arabi-dopsis and other plants. The results indicated that DlMADS18 showed high identity to AGL6-like genes from other plants (Fig. 2) and was grouped into the AGL6-like subfamily (Fig. 3) which formed a clade together with AGL2- and SQUA-like genes[16,37], implying that they could have similar functions in plant development. Many AGL6-like genes had been cloned from angiosperms and gymnosperms. Their expression patterns mainly in floral organs indicated that their functions were possibly related to floral development. For example, AGL6 was expressed in all four whorls of floral organs[38], while AGL13 was restricted to ovules[39], ZAG3 in both male and female in-florescences of maize, also in carpels, but not in sta-mens[40], GGM9 and GGM11 in male as well as female reproductive cones of Gnetum[41], OMADS1 in inflores-cence meristem, flower buds and pedicle in an orchid[42]. However, no single loss-of-function mutant has been re-ported, implying that the function of AGL6-like genes has remained enigmatic[16]. Recently, two AGL6-like genes, OMADS1 of an orchid(Oncidium ‘Gower Ramsey’)[42] and DAL1 of a spruce (Picea abies)[43], were characterized. Transgenic plants exhibited phenotypes with early flow-ering, extremely reduced size, curled leaves and terminal flowers. The phenotype of the ectopic expression of

DlMADS18 in Arabidopsis shared very similar features with them (Fig. 5). Similar phenotypes were also observed in a range of AGL2- and SQUA-like genes transgenic plants as a result of high level expression[7,21,44—49]. The available data showed that these MADS-box genes could have similar functions in plant development.

Transgenic rice overexpressing the C-terminal dele-tion mutants of OsMADS1 had slightly early-flowering and dwarf phenotypes, but without obvious alteration of floral organs. On the other hand, ectopic expression of the OSMADS1 mutant lacking the MADS domain showed no phenotypic alteration. Therefore, the MADS domain of an MADS box protein was essential for inducing the early-flowering and dwarf phenotypes, while the C-terminal region was not needed. However, the C terminus was important for specific functions of these proteins[50]. Hsu et al.[42] proved that the reason OMADS1 inducing the early-flowering and dwarf phenotypes was due to the in-teraction between the OMADS1 and flowering time genes. The expression of flowering time genes FT, SOC1 and flower meristem identity genes LFY, AP1 was signifi-cantly upregulated in 35S:OMADS1 transgenic Arabidop-sis plants. Similar results were observed for two AGL2- like genes of lily, LMADS3 and LMADS4[21]. It suggested that DlMADS18 might act in similar mechanisms with other AGL6-/AGL2-/SQUA-like genes in flowering initia-tion. This could explain why the 35S:DlMADS18 trans-genic Arabidopsis plants produce similar phenotypes to the other plants ectopically expressing AGL6-/AGL2-/ SQUA-like genes. In summary, DlMADS18 may probably be involved in controlling the flowering time of D. lati-florus. Usually, AGL2-like genes were diverse in one plant species. The high sequence similarity and parallel expres-sion patterns among the Arabidopsis AGL2-like genes suggested that they might encode proteins of redundant functions[51]. Indeed, single gene mutations generated by reverse genetics produce only subtle phenotypes[52]. Like AGL2-like genes, AGL6-like genes are not restricted to a single member in one plant species[16]. Also there are probably redundant functions among AGL6-like genes proteins, even between AGL6-like genes and AGL2- like genes. whether there were any more AGL6-like genes in D latiflorus, was under research. Similar to the case with some AGL6-/AGL2-/SQUA-like genes[44-46], no obvi-ous alteration of floral organs was observed in 35S:DlMADS18 transgenic Arabidopsis plants.

For most plants, cytokinins promoted flowering. For example, the vegetative stage of a Malus species could be weakened by intrinsic cytokinins and promoted to flower-ing one[53]. High concentrations of BA (10—100 mg/L), a cytokinin, induced the formation of dwarf stems and the abundant appearance of stem axillary buds as well as flower bud formation. All young leaves had some mor-phological abnormalities, being wrinkled and not very green. The cessation of BA application resulted in the


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cessation of flower bud transition to flowers[54]. DlMADS18 was isolated from the young spikelets of D. latiflorus tissue cultures induced by high concentrations of BA. Similar phenotype suggested that BA induced the expression of the DlMADS18 genes and promoted the D. latiflorus culture flowering.

We studied the bamboo floral development using molecular biological method. The result is conducive to a better understanding of the mechanism of bamboo floral development and the functions of AGL6-like genes. Flow-ering is a very complicated process controlled by multiple genes. The mechanisms of bamboo flowering at molecular level should be revealed in more detail. Because of the unique property of bamboo , to stabilize the tissue culture systems of bamboo is essential for the future research. Acknowledgements We thank Prof. Guangchu Zhang (Guangdong Forest Research Institute) for presenting the in vitro flowering cultures, Dr. Diqiu Yu (Xishuangbanna Tropical Botanical Garden of the Chinese Academy of Sciences) for critical reading of this manuscript. This work was supported by the National Natural Science Foundation of China (Grant No. 30200015) and the Natural Science Foundation of Yunnan Province (Grant No. 2002C0056M).

References 1. Levy, Y. Y., Dean, C., The transition to flowering, Plant Cell, 1998,

10: 1973—1989. 2. Blázquez, M. A., Flower development pathways, J. Cell Sci., 2000,

113: 3547—3548. 3. Mouradov, A., Cremer, F., Coupland, G., Control of flowering time:

Interacting pathways as a basis for diversity, Plant Cell, 2002: S111—S130.

4. Jack, T., Molecular and genetic mechanisms of floral control, Plant Cell, 2004, 16: S1—S17.

5. Boss, P. K., Bastow, R. M., Mylne, J. S. et al., Multiple pathways in the decision to flower: Enabling, promoting, and resetting, Plant Cell, 2004, 16: S18—S31.

6. Simpson, G. G., Dean, C., Arabidopsis, the rosetta stone of flower-ing time? Science, 2002, 296: 285—289.

7. Mandel, M. A., Yanofsky, M. F., A gene triggering flower formation in Arabidopsis, Nature, 1995, 337: 522—524.

8. Michaels, S.D., Amasino, R. M., FLOWERING LOCUS C encodes a novel MADS domain protein that acts as a repressor of flowering, Plant Cell, 1999, 11: 949—956.

9. Hartmann, U., Hohmann, S., Nettesheim, K. et al., Molecular clon-ing of SVP: A negative regulator of the floral transition in Arabi-dopsis, Plant J., 2000, 21: 351—360.

10. Lee, H., Suh, S. S., Park, E. et al., The AGAMOUS-LIKE 20 MADS domain protein integrates floral inductive pathways in Arabidopsis, Genes Dev., 2000, 14: 2366—2376.

11. Sheldon, C. C., Rouse, D. T., Finnegan, E. J. et al., The molecular basis of vernalization: The central role of FLOWERING LOCUS C (FLC), Proc. Natl. Acad. Sci. USA, 2000, 97: 3753—3758.

12. Yu, H., Xu, Y., Tan, E.L. et al., AGAMOUS-LIKE 24, a dosage-dependent mediator of the flowering signals, Proc. Natl. Acad. Sci. USA, 2002, 99: 16336—16341.

13. Borner, R., Kampmann G., A MADS domain gene involved in the transition to flowering in Arabidopsis, Plant J, 2000, 24(5): 591—599.

14. Theiβen, G., Development of floral organ identity: Stories from the

MADS house, Curr. Opin. Plant Biol., 2001, 4: 75—85. 15. Ng, M., Yanofsky, M., Function and evolution of the plant

MADS-box gene family, Nat. Rev. Gen., 2001, 2: 186—195. 16. Becker, A., Theissen G., The major clades of MADS-box genes and

their role in the development and evolution of flowering plants, Mol. Phylogenet. Evol., 2003, 29(3): 464—489.

17. Ratcliffe, O. J., Nadzan, G. C., Reuber, T. L. et al., Regulation of flowering in Arabidopsis by an FLC homologue, Plant Physiol., 2001, 126: 122—132.

18. Samach, A., Onouchi, H., Gold, S. E. et al., Distinct roles of CONSTANS target genes in reproductive development of Arabi-dopsis, Science, 2000, 288: 1613—1616.

19. Kempin, S. A., Savidge, B., Yanofsky, M. F., Molecular basis of the cauliflower phenotype in Arabidopsis, Science, 1995, 267: 522—525.

20. Mandel, M. A., Yanofsky, M. F., The Arabidopsis AGL9 MADS box gene is expressed in young flower primordial, Sex Plant Re-prod., 1998, 11: 22—28.

21. Tzeng, T. Y., Hsiao, C. C., Chi, P. J. et al., Two lily SEPAL-LATA-like genes cause different effects on floral formation and flo-ral transition in Arabidopsis, Plant Physiol., 2003, 133: 1091— 1101.

22. Simon, T. M., Elizabeth, A. K., Heterogeneous expression patterns and separate roles of the SEPALLATA gene leafy hull sterile1 in grasses, Plant Cell, 2004, 16: 1692—1706.

23. Ma, H., Yanofsky, M. F., Meyerowitz, E. M., AGL1-AGL6, an Arabidopsis gene family with similarity to floral homeotic and transcription factor genes, Genes Dev., 1991, 5: 484—495.

24. Theiβen, G., Kim J., Saedler H., Classification and phylogeny of the MADS-box multigene family suggest defined roles of MADS-box gene subfamilies in the morphological evolution of eukaryotes, J. Mol. Evol., 1996, 43: 484—516.

25. Janzen, D. H., Why bamboos wait so long to flower? Ann. Rev. Ecol. Syst., 1976, 7: 347—391.

26. Sharma, M. L., The flowering of bamboo: Fallacies and facts. Bamboo in Asia and the Pacific, Thailand: Chiangmai, 1994, 68—70.

27. Ueda, k., Studies on the physiology of bamboo,with references to practical application, Tokyo: Prime Minister′s Office, 1960, 167.

28. Nelson, B. W., Natural forest disturbance and change in the Brazil-ian Amazon, Remote Sensing Reviews, 1994, 10: 105—125.

29. Franklin, D. C., Synchrony and asynchrony: Observations and hy-potheses for the flowering wave in a long-lived semelparous bam-boo, J. Biogeography, 2004, 31(5): 773—786.

30. Keeley, J. E., Bond, W. J., Mast flowering and semelparity in bam-boos: The bamboo fire cycle hypothesis, Am. Naturalist, 1999, 154: 383—391.

31. Wong, K. M., Flowering, fruiting and germination of the bamboo Schizostachyum zollingeri in Perlis, Malaysian Forester, 1981, 44: 453—463.

32. Zhang, G. C., Chen, F. S., Wang, Y. X., Study on rapid propagation in vitro of Dendrocalamus latiflous, J. Bamboo Res., 1993, 12: 7—15.

33. Murashige, T., Skoog, F., A revised medium for rapid growth and bioassays with tobacco tissue cultures, Physiol. Plant, 1962, 15: 473—479.

34. Swofford, D. L., PAUP: Phylogenetic analysis using parsimony, version 4.0b10, Sinauer Associates Massachusetts, USA, 2001.

35. Clough, S. J., Bent A. F., Floral dip: A simplified method for Agro-bacterium-mediated transformation of Arabidopsis thaliana, Plant J., 1998, 16(6): 735—43.

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36. Sambrook, J., Russell, D. W., Molecular Cloning: A Laboratory Manual, 3rd ed, New York: Cold Harbor Laboratory Press, 2002, 540—544, 721—725.

37. Theiβen, G., Becker, A., Di Rosa, A. et al., A short history of MADS-box genes in plants, Plant Mol. Biol., 2000, 42: 115—149.

38. Mouradov, A., Glassick, T. V., Hamdorf, B. A. et al., Family of MADS-box genes expressed in early male and female reproductive structures of Monterey Pine, Plant Physiol., 1998, 117: 55—61.

39. Rounsley, S. D., Ditta, G. S., Yanofsky, M. F., Diverse roles for MADS box genes in Arabidopsis development, Plant Cell, 1995, 7: 1259—1269.

40. Mena, M., Mandel, M. A., Lerner, D. R. et al., A characterization of the MADS-box gene family in maize, Plant J., 1995, 8: 845—854.

41. Winter, K. U., Becker, A., Münster, T., et al., MADS-box genes re-veal that gnetophytes are more closely related to conifers than to flowering plants, Proc. Natl. Acad. Sci. USA, 1999, 96: 7342— 7347.

42. Hsu, H. F., Huang, C. H., Chou, L. T. et al., Ectopic expression of an orchid (Oncidium Gower Ramsey) AGL6-like gene promotes flowering by activating flowering time genes in Arabidopsis thaliana, Plant Cell Physiol., 2003, 44(8): 783—794.

43. Carlsbecker, A., Tandre, K., Johanson, U. et al., The MADS-box gene DAL1 is a potential mediator of the juvenile-to-adult transi-tion in Norway spruce (Picea abies), Plant J., 2004, 40: 546—557.

44. Chung, Y. Y., Kim, S. R., Finkel, D. et al., Early flowering and re-duced apical dominance result from ectopic expression of a rice MADS box gene, Plant. Mol. Biol., 1994, 26: 657—665.

45. Kang, H. G., An, G., Isolation and characterization of a rice MADS box gene belonging to the AGL2 gene family, Mol. Cells, 1996, 7: 45—51.

46. Kang, H. G., Jang, S., Chung, J. E. et al., Characterization of two rice MADS box genes that control flowering time, Mol. Cells, 1997, 7: 559—566.

47. Kyozuka, J., Harcourt, R., Peacock, W. J. et al., Eucalyptus has functional equivalents of the Arabidopsis AP1 gene, Plant Mol. Biol., 1997, 35, 573—584.

48. Honma, T., Goto, K., Complexes of MADS-box proteins are suffi-cient to convert leaves into floral organs, Nature, 2001, 409: 525—529.

49. Pelaz, S., Gustafson-Brown, C., Kohalmi, A. E. et al., APETALA1 and SEPALLATA3 interact to promote flower development, Plant J., 2001, 26: 385—394.

50. Jeon, J. S., Jang, S., Lee, S. et al., leafy hull sterile1 is a homeotic mutation in a rice MADS box gene affecting rice flower develop-ment, Plant Cell, 2000, 12: 987—884.

51. Savidge, B, Rounsley, S. D., Yanofsky, M. F., Temporal relation-ships between the transcription of two Arabidopsis MADS box genes and the floral organ identity genes, Plant Cell, 1995, 7: 721—733.

52. Pelaz, S, Ditta, G. S., Baumann, E. et al., B and C floral organ identity functions require SEPALLATA MADS-box genes, Nature, 2000, 405: 200—203.

53. Sedgley M., Flowering of deciduous perennial fruit crops, Hort. Rev., 1990, 12: 223—264.

54. Kochankov, V. G., Milyaeva, E. L., Zhyvukhina, E. A. et al., Effect of 6-Benzylaminopurine on stem formation and flower bud intia-tion in Rudbeckia bicolor plants of different ages under non- induc-tive conditions, Acta Horticulturae, 1989, 251.

(Received September 10, 2004; accepted December 24, 2004)

Isolation and ectopic expression of a bamboo MADS-box gene

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