小麦冷休克蛋白基因TaCSP的克隆及表达分析

为深入研究小麦冷休克蛋白(cold shock proteins,CSPs)基因,根据大肠杆菌(E.coli)CSPs蛋白保守氨基酸序列,借助生物信息学技术检索小麦EST序列,采用同源序列法拼接了小麦3个冷休克蛋白基因,并以小麦叶片总RNA为模板,采用RT-PCR对其进行扩增;同时,采用荧光定量PCR对3个基因的组织表达特性以及ABA、低温、高温、干旱和盐胁迫条件下的表达模式进行了研究。结果表明,小麦3个冷休克蛋白基因TaCSP1、TaCSP2和TaCSP3全长分别为290、374和377bp,各编码69、69、80个氨基酸残基。序列分析表明,TaCSP1、TaCSP2和TaCSP3之间氨基酸相似性为72.9%~84.3%,与大肠杆菌来源CSPs的相似性为40.0%~79.7%。荧光定量PCR表达谱分析显示,TaCSPs在抽穗期小麦的根、茎、叶和幼穗中均能表达。胁迫分析表明,3个冷休克蛋白基因在小麦幼根中的表达,在低温和ABA处理下,表现为早期诱导、后期抑制,在高温、干旱和盐胁迫下,则表现为早期抑制、后期有所恢复。克隆获得的小麦3个冷休克蛋白基因在根组织中强烈表达,并受ABA和冷胁迫诱导,推测其在小麦抵御逆境胁迫过程中发挥重要作用。

Cloning and Expression Analysis of TaCSP genes Encoding Cold Shock Proteins from Wheat

In this study,we reported that the cloning and expression pattern analysis of three full length cDNA encoding cold shock proteins from wheat ( Triticum aestivum L. Chinese Spring). Based on the conserved amino acid sequence of the cold shock protein, three pair of primers were designed based on simulation of gene splicing and RT-PCR, and then were used to clone cold shock protein genes TaCSP1, TaCSP2 and TaCSP3, respectively. FQ-PCR assay were performed to examine the tissue-specific expression of TaCSPs , and their expressing patterns during low temperature, high temperature, drought, salt and ABA. Sequencing and structural analysis showed that TaCSP1, TaCSP2 and TaCSP3 were 290 bp, 374 bp and 377 bp in full length, and encoding 69, 69 and 80 amino acids, respectively. Homology analysis showed that the deduced amino acid sequence of TaCSP1, TaCSP2 and TaCSP3 shared 72.9%-84.3% identity with each other, and 40.0%-79.7% identity with CSPs from E.coli. Tissue-specific expression analysis showed that TaCSP1, TaCSP2 and TaCSP3 were constitutively expressed in various wheat tissues including roots, stems, leaves and spikes. The expression profiling also showed that the three wheat CSPs genes were strongly induced at 1 h after exposure to cold and ABA stresses in wheat seedlings roots, and then reduced to the baseline. The expression profiling also showed that wheat CSPs were down regulated by high temperature, drought and salt stresses. However, the expressing levels of these CSPs increased after long time treatment. Three cold shock protein genes were isolated from wheat, and the expression pattern suggested the close relationship between the expression of this gene and various abiotic stresses.