黄瓜3-磷酸甘油醛脱氢酶基因CsGAPDH的克隆及其涝胁迫响应分析

Cloning,Characterization and Expression of CsGAPDH,A Waterlogging Responsive Gene in Cucumber

以黄瓜耐涝品系‘早二N’根组织总RNA为模板,运用RT-PCR结合RACE技术,获得了黄瓜3-磷酸甘油醛脱氢酶基因(CsGAPDH)的cDNA全长序列,基因编码区共1011bp,编码336个氨基酸(GenBank登录号HQ156465)。系统进化分析表明:CsGAPDH基因与胡萝卜、葡萄、大豆等双子叶植物同源基因的核苷酸序列相似性均在88%以上,与单子叶植物水稻和玉米同源基因的相似性达86.8%和86%。qRT-PCR表达分析结果表明:CsGAPDH基因快速响应涝胁迫,在涝胁迫2h,该基因在根中表达量达到对照水平的12倍,在涝胁迫12h达到表达高峰,随后表达量呈现逐渐下降趋势;CsGAPDH基因在‘早二N’的根、茎、叶中均有表达,根和叶中原始表达量高于茎,在涝胁迫4h后均诱导表达,根中诱导表达量上调幅度最大,为初始表达量的4倍。CsGAPDH属于黄瓜涝胁迫响应基因,在缓解涝胁迫伤害过程中可能具有重要的调控作用。

Glyceraldehyde-3-phosphate dehydrogenase gene（GAPDH）was cloned from root tissues of cucumber（Cucumis sativus L.）line＇Zaoer N＇with high waterlogging tolerance,based on reverse transcription polymerase chain amplification（RT-PCR）and rapid amplification of cDNA ends（RACE） methods.CsGAPDH gene contained an open reading frame（1 011 bp）and encoded a protein of 336 amino acids.Phylogenetic analysis of CsGAPDH gene showed high conserved character of GAPDH gene among different crops.CsGAPDH showed the highest homology with dicotyledonous plants such as carrot,grape and soybean,and the similarity were more than 88%.For monocot crops rice and maize,the similarities of GAPDH gene were 86.8% and 86%,respectively.The time course expressions of CsGAPDH gene in root of＇Zaoer N＇under waterlogging treatment were determined by quantitative RT-PCR.CsGAPDH gene wassignificantly induced at 2 h after waterlogging treatment,and the peak level of gene expression was at 12 h.The tissue specific expression of CsGAPDH indicated that the expression level of CsGAPDH gene in root and leaf was higher than that in stem of the control plant;However,the transcription level of CsGAPDH gene were significantly up-regulated at 4 h after waterlogging treatment,which were higher in root than that in leaf and stem.In summary,the CsGAPDH gene was involved in waterlogging response in cucumber,and might play an important role in releasing waterlogging stress.