Identification Of MADS-box Genes And Their Function In Floral Organ Development Of Phyllostachys Edulis

Bamboo is one of the most important non-timber forest products in the world. They flower at the end of very long vegetative growth phases simultaneously, often following a large-area death, which seriously threatens the bamboo management. So far, most researchers have focused on flowering habit, physiologic and biochemical index, flower organ dissection and structure, while them neglected molecular mechanisms involved in floral organ development in Phyllostachys edulis. Therefore, studying on the key regulatory genes involved in floral formation and discussing the bamboo flowering mechanism are of important theoretical and practical significances. Leaf, pistil, stamen, young embryo, glume, palea, flower bud and bract of P. edulis were collected. The flowers organs of P. edulis are mainly studied through the transcriptome sequencing and in situ hybridization. In order to find the key MADS-box genes in flower regulation which provides guidance for studying mechanism of bamboo floral organ development. The main results are as followed:We performed a whole-genome survey of P. edulis and identified 34 MADS-box genes, which were classified into four groups. We provided detailed analyses of the gene structure, motif, phylogenetic classification, comparison of the gene divergence and duplication. The number of introns in most MADS-box genes which belonged to MIKCC and MIKC* type ranged from 1 to 8. Comparative analysis of MADS-box genes among P. edulis, rice, and Arabidopsis suggests that there were more gene duplication events in P. edulis, leading to a faster birth and death rate after bamboo diverged from other grasses.The differences in ABCDE genes expression during four flower development stages were extensively analyzed. The A- and B-class genes were highly expressed in flower bud formation. With inflorescence maturing, expression level of them decreased gradually. Instead, C-, D- and E-class genes were lowly expressed in flower bud formation, but mainly expressed in bloom and young embryo formation. The above results reveal important roles of the P. edulis ABCDE genes during floral organ development.Transcriptome sequencing for leaf of non-flowering plant, pistil, stamen, young embryo, glume, palea, flower bud and bract was performed by Illumina HiSeqTM 2000. By comparing the eight libraries at a statistically significant value(FDR ≤ 0.001 and log2 Ratio ≥ 1), 13,051 genes were detected by changed-expression, including 7,831 genes up-regulated and 5,220 genes down-regulated. Differentially expressed genes presented a dynamic development among different flower organs, which showed a largest number of differential expression genes in flower bud and pistil but smallest in glum.By applying Illumina HiSeqTM 2000 sequsncing technology on studying transcriptom and differentially expressed genes profiles among different flower organs of P. edulis, we found the A-class genes, PheMADS14, PheMADS15, Phe MADS18-1 and PheMADS18-2 were mainly expressed in flower bud, while PheMADS29 and PheMADS31 were preferentially expressed in bract and young embryo. Three B-class genes, PheMADS2, PheMADS4-1 and PheMADS4-2, had high mRNA levels in the stamen and pistil development. PheMADS3 and PheMADS21 belonging to the C- and D-class, respectively, were highly expressed in stamen and pistils. The E-class gene PheMADS1 was preferentially expressed in stamens, pistil and young embryo of P. edulis. The above results show that these MADS-box genes were mostly similar to those of Arabidopsis A-, B-, C-, D-, E-class genes, supporting the notion that the ABCDE model might be extended to P. edulis.In situ hybridization showed that PheMADS15 mRNA were obviously accumulated in early spikelet meristem, the primordia of flower organs, and the reproductive organs. PheMADS3 and PheMADS21 mRNA were highly expressed in the glumes, lemma, palea, pistil and developing embryo, but hardly detected in lodicules. The spatial and temporal expression patterns of PheMADS1 and PheMADS5 were detected from the early floral bud to the mature floral organ by in situ hybridization in P. edulis. This finding further suggests that ABCDE model can be applied to P. edulis.PheMADS15 was located in the nucleus as its function as a transcription factor. PheMADS15 could activate the expression of the His-3 and β-Gal reporter genes, indicating that it is a transcriptional activator.Moreover, we overexpressed PheMADS15, an AP1-like gene, in Arabidopsis, and found that the transgenic plants had early flowering phenotype. Phe MADS15 gene might accelerate expression of TFL1 and SOC1 repress transcription of LFY, respectively, which caused early flowering in transgenic Arabidopsis. The results suggest that PheMADS15 may be a regulator of flower development and flowering transition in P. edulis.