滨麦HMW-GS启动子和编码区基因的分离与序列分析

Isolation and Sequence Analysis of the HMW-GS Promoter and ORF Genes from Leymus mollis

为了深入了解滨麦(Leymus mollis)HMW-GS基因的结构特征,采用PCR方法,从滨麦基因组中分离出HMW-GS基因启动子序列(Genbank登录号:FJ600498)和编码区序列(Genbank登录号:GQ169797)。启动子序列(FJ600498)从5′至3′方向依次有E-box、N-box、G-box、HMW谷蛋白特异增强子和TATA-box等麦谷蛋白特异调控元件,推断其为滨麦HMW-GS启动子基因。编码区序列(GQ169797)具有单一完整的开放阅读框(ORF),编码396个氨基酸残基,依次包含信号肽、N-末端区、中部重复区和C-末端区等HMW-GS的典型结构区域;中部重复区主要重复单元为6肽(PQQGQQ)和9肽(GYYPTSPQQ);有6个半胱氨酸残基(Cys),分布在N-末端区(5个)和C-末端区(1个),第3、4个相邻。推断其为滨麦的y-型HMW-GS编码区基因。系统进化分析表明,启动子序列(FJ600498)与异形花草(He.piliferum)和冰草(Ag.cristatum)的HMW-GS启动子基因序列具有相对较近的同源关系;编码区序列(GQ169797)与纤毛鹅观草(Elymus ciliaris)和披碱草(E.glaucus)的HMW-GS编码区基因序列具有相对较近的同源关系。

HMW-GS promoter sequence（FJ600498） and coding region sequence（GQ169797） were isolated from the genome of L.mollis by the method of PCR.The HMW-GS promoter sequence of L.mollis from 5′ to 3′ flanking in turn contained E-box,N-box,G-box,38bp HMW glutenin specific enhancer and TATA-box,which were the promoter regulatory elements of typical HMW-GS gene.The HMW-GS coding region sequence had a single complete open reading frame（ORF）,which encoding 396 amino acids.The deduced amino acid sequence contained the signal peptides,N-terminal region,the central repeat region,and C-terminal region.In repeat domains,hexapeptide（PGQGQQ） and nonapeptide（GYYPTSPQQ） were the main repeating unit.The sequence had six cysteine residues,five within N-terminal and one within C-terminal domains,and the 3rd and the 4th cysteine neighboring.These structure characteristics suggested that GQ169797 is a y-type HMW-GS gene of L.mollis.Phylogenetic analysis showed that the promoter（FJ600498） was closely related with the HMW-GS promoter sequence of Heteranthelium piliferum and Agropyron cristatum and the ORF sequence has a relatively close homology with y-type HMW-GS gene of Elymus ciliaris and Elymus glaucus.