多枝赖草谷胱甘肽还原酶cDNA片段的克隆与分析

从盐胁迫处理的多枝赖草(Leymus multicaulis)新鲜叶片中提取分离出RNA，然后根据报道的多种植物谷胱甘肽还原酶氨基酸序列上两个保守区设计简并引物。RT-PCR获得了1条大小约400bp的条带，回收该条带并进行TA克隆，蓝白斑筛选，得到阳性克隆。经过质粒大小比较和PCR验址．进行序列测定和分析，发现该序列属于GR基因片段，其Genbank注册号为AY781786．编码的氨基酸序列与Oryza sativa、Zea mays、Arabidopsis thaliana和Nicotiana tabacum的GR相应区段的氨基酸序列一致性分别为91％、89％、86％和83％。蛋白质序列分析发现该序列含有一个吡啶二硫酸核苷酸氧化还原酶(pyridine nucleotide-disulphide oxideoreductase)保守结构域。进化树分析表明，该多枝赖草cDNA片段编码的氨基酸序列在进化上与水稻和玉米较近。

Cloning and Analysis of cDNA Segment of Glutathione Reductase in Leymus multicaulis

RNA was extracted and separated from the fresh leaves of Leymus multicaulis plants under salt stress and then primers were designed according to two conserved regions of glutathione reductase reported in many species.And a ban of about 400 bp was obtained by RT-PCR.The recovery as well as TA cloning,white/blue screening was conducted to obtained a positive clone.The plasmid-size comparison and PCR identification and sequence determination and analysis revealed that the sequence belonged to GR segment and its registry code in the Genbank was AY781786.The amino acid sequence under the registry code was consistent with the sequence of the correspondent GR segments in Oryza sativa,Zea mays,Arabidopsis thaliana and Nicotiana tabacum at the uniformity rates of 91%,89%,86% and 83%,respectively.The protein sequence analysis found that the sequence contained a conserved structural region of pyridine nucleotide-disulphide oxidoreductase.The phylogenetic analysis revealed that the amino acid sequence of the cDNA segment in Leymus multicaulis was relatively close to those in Oryza sativa and Zea mays.