陆地棉小GTP结合蛋白基因GhRop4的克隆及其表达分析

Molecular Cloning and Expression Analysis of a small GTP-binding Protein Gene Gh Rop4 in Gossypium hirsutum L.

【目的】对陆地棉小GTP结合蛋白基因(Small GTP-binding protein)GhRop4(AY965614.1)在多种逆境胁迫下的应答表达模式进行研究分析,为棉花抗逆相关基因克隆及棉花抗逆分子机制研究奠定基础。【方法】利用生物信息学方法分析了GhRop4基因的结构特征和进化树关系,并通过荧光定量PCR(Real-time quantitative polymerase chain reaction,qRT-PCR)的方法分析Gh Rop4基因的组织表达特异性和不同逆境诱导条件下的表达模式。【结果】以陆地棉c DNA为模板克隆得到GhRop4基因,其开放阅读框为588 bp,编码包含195个氨基酸残基的Ⅰ型Rop蛋白。氨基酸多重序列比对表明,GhRop4与其它植物Rop蛋白高度同源,符合Rop蛋白结构特点。qRT-PCR分析表明,GhRop4基因在棉花幼苗根、茎、叶、子叶和下胚轴中均有表达,且在子叶和茎中表达水平较高。GhRop4基因对高盐、干旱、低温和棉花黄萎病菌处理都有一定程度的响应。【结论】Gh Rop4基因在陆地棉的逆境胁迫适应过程中可能具有重要的作用。

[Objective]This study is to dissect upland cotton small GTP-binding protein gene GhRop4(AY965614.1)expression alteration responsive to different abiotic stresses,and provide a foundation for stress-related genes clone,and elucidate the molecular mechanism of cotton stress-resistance.[Method]In this study,the protein structural characteristics and phylogenetic analysis of GhRop4 gene were analyzed by bioinformatics methods.Tissue-specific expression of GhRop4 under various induction conditions were conducted using quantitative real-time polymerase chain reaction(q RT-PCR).[Result]The GhRop4 gene was cloned from the upland cotton cDNA,sequence analysis showed that it contained a 588 bp open reading frame,putatively encoding a typeⅠRop protein with 195 amino acid residues.Multiple sequence alignment showed that GhRop4 shared high similarity with Rop proteins of other plants and was in accordance with the structural characteristics of Rop proteins.qRT-PCR analysis showed that GhRop4 gene expressed in roots,stems,leaves,cotyledons,and hypocotyl of cotton seedlings,and especially expressed at a relatively higher level in cotyledons and stems.GhRop4 gene expression was regulated with dehydration,high salinity,low temperature and Verticillium wilt treatments.[Conclusion]The results suggested that GhRop4 gene may play important roles in the adaptation of upland cotton to adversity stress.