水稻低温响应基因OsCR1的克隆与表达分析

Cloning and Expression Profile of Cold-Responsive Gene OsCR1 in Rice

低温、高温、干旱等非生物胁迫是影响水稻产量与品质的重要非生物逆境因子。为了探索水稻耐逆的分子机理并挖掘新的水稻耐逆基因,采用Affymetrix 60K水稻基因表达芯片分析了培矮64S全基因组在上述逆境下的表达谱变化,筛选出一个受低温诱导表达水平显著升高的基因OsCR1(Oryza sativa L.cold responsive gene1,GenBank登录号:AK066066)。实时定量PCR进一步证实该基因受低温诱导高表达。利用RT-PCR方法扩增获得其cDNA序列。生物信息学分析表明,OsCR1位于水稻第10号染色体上,包含两个内含子并存在选择性剪接现象,可编码含195个氨基酸残基的蛋白质,其蛋白等电点为5.78,分子量为21.9kD。拓扑结构预测显示OsCR1蛋白上含有2个Ser/Thr蛋白激酶磷酸化位点;疏水性分析显示该基因为亲水性蛋白;启动子区域分析发现5类逆境相关的顺式作用元件。氨基酸序列比对分析显示OsCR1为未知功能基因。

Many biotic stimuli, such as cold, heat and drought adversely affect plant development. To understand mechanisms underlying stress tolerance and discover novel stress-tolerant genes in rice（ Oryza sativa L. ） , the Affymetrix 60 K GeneChip Rice Genome Array was used to analyze the whole genome expression of rice Pei＇ai 64s under cold, drough and heat stress conditions. We identified a gene OsCRl.（ Oryza sativa L. cold responsive gene l, GenBank accession ： AK066066 ） , which was highly induced by cold in leaf and panicle at all the developmental stages test. The expression profile of OsCR1 was further confirmed by quantitative real-time PCR analysis. The cDNA of OsCR1 was cloned by RT-PCR and sequenced. The cDNA is 831 bp in length and contains an open reading frame of 585 bp encoding a protein of 195 amino acid residues with a calculated molecular mass of 21.9 kD and a PI of 5.78. Search in GenBank found that OsCR1 is located in chromosome lO of rice. The OsCR1 gene sequence was found to have two introns and might have alterative splicing phenomenon. Topology prediction showed that the OsCR1 protein contains two Ser/Thr protein kinase phosphorylation sites. Hydrophobic analysis revealed that the protein is a hydrophilic protein. Five different stress-responsive cis-acting elements were found in the putative promoter region. Alignment with other proteins revealed that OsCR1 encodes a protein of unknown function.