Identification,clone And Function Analysis Of SAUR And DELLA Genes In Moso Bamboo

Moso bamboo(Phyllostachys edulis)is one of the most important economic bamboo species in China.Bamboo shoot could grow from 0 to 20 meters in three months under optimum conditions.Cell division and elongation were both involved in rapid growth after anatomy research.Many studies on Arabidopsis thaliana and other model plants indicated that gibberellic acid(GA)and indole-3-acetic acid(IAA)have great influence on cell division and elongation respectively.Small auxin-up RNA(SAUR),one of the early auxin responsive gene families,was demonstrated to promote cell elongation.DELLA protein,one regulatory factor in GA signal pathway,was demonstrated to regulate downstream genes involved in cell division and elongation.In this study,moso bamboo shoots were used for transmission electron microscope(TEM)analysis.SAUR and DELLA genes were identified based on moso bamboo genome database with analysis of bioinformatics.Fluorescent quantitative real-time PCR(qRT-PCR),gene clone,genetic transformation,subcellular localization and prokaryotic expression were used for biological function analysis.Results were as follows.1.TEM was used to observe the cells in moso bamboo shoots of three different heights.Winter bamboo shoots cells were smaller than that in 3 and 9 meters shoots and kept division ability.Cells became mature gradually in later period.38 SAUR genes(PheSAUR)were identified based on moso bamboo genome database and the length of coding sequences,number of amino acid residue,promoter element,isoelectric point(Pl)and some other bioinformatics of which were analyzed.Majority PheSAUR genes contain no introns at all.Several sister pairs were identified by phylogenetic analysis of total PheSAUR proteins.Most of the PheSAUR proteins were closely related to OsSAUR proteins after phylogenetic analysis of all the PheSAUR,Os SAUR and AtSAUR proteins.Three highly conservative motifs were identified in all the PheSAUR,OsSAUR and AtSAUR proteins.In addition,one DELLA gene with no introns was identified.However,some conserved domains of protein were lost.2.PheSAUR2,PheSAUR6,PheSAUR18,PheSAUR20,PheSAUR26,PheSAUR35 were tissue specific expressed in moso bamboo seedlings by qRT-PCR.Majority PheSAUR genes were up regulated after IAA treatment.Six PheSAUR genes were cloned.Subcellular localization showed that PheSAUR4 was localized to cell membrane,PheSAUR28 was localized to cytoplasm,PheSAUR29 and PheSAUR34 were localized to cell membrane and nucleus simultaneously.Those localizations indicated that PheSAUR proteins may be involved in IAA signal transporting.35S:PheSAUR29-GFP lines showed faster growing speed than controls in the 1/2 MS medium containing GA3.PheSAUR29 may be involved in GA signal pathway.PheSAUR4 and PheSAUR29 were both up and down regulated after GA3 and S3307(uniconazole)treatments in seedlings respectively.PheSAUR7,PheSAUR13,PheSAUR16,PheSAUR17,PheSAUR18 were up regulated after GA3 treatment in seedlings.Moreover,prokaryotic expression vectors of PheSAUR4 and PheSAUR29 were constructed and prokaryotic expression conditions of which were obtained.3.DELLA gene(1 857 bp)was cloned with no introns and the protein of which was consist of 618 amono acids.GC content was 69.04%.All the DELLA conserved domains were involved in protein.This protein was most closely related to SLR1 protein in rice by phylogenetic analysis with other DELLA proteins.Hence,this DELLA gene was named PheSLR1.A 1 933 bp promoter of PheSLR1 was cloned.Light and GA responsive elements were identified in the promoter.PheSLR1 was up regulated during shoots growth and could be up regulated by GA3 and down regulated by S3307 treatment respectively.Moso bamboo seeds treated with S3307 showed slower growing speed than with DMSO and GA3.PheSLR1 expressed less in seeds treated with S3307 than with DMSO and GA3.PheSLR1 protein was localized to nucleus by subcellular localization analysis.The expression levels of putative downstream genes of PheSLR1 protein,such as PheGA20 ox,PheCes A1,PheCesA2 and PheGASA6,were analyzed.Most of those genes were up-regulated in bamboo shoot development.Moreover,prokaryotic expression condition and purified protein of PheSLR1 were obtained.In this study,SAUR and DELLA genes were identified based on moso bamboo genome database.Biological functions of SAUR and DELLA genes were also explored.Those results provided some information for moso bamboo shoot development mechanism research.