文献类型：期刊文献

中文题名：哈氏弧菌谷胱甘肽还原酶基因克隆及原核表达

英文题名：Cloning and Prokaryotic Expression of Glutathione Reductase Gene in Vibrio harveyi

作者：朱帆[1,2,3];丁燏[1,2,3];鲁义善[1,2,3];简纪常[1,2,3];吴灶和[2,3,4]

机构：[1]广东海洋大学水产学院;[2]广东省水产经济动物病原生物学及流行病学重点实验室;[3]广东省教育厅水产经济动物病害控制重点实验室;[4]仲恺农业工程学院

年份：2015

卷号：31

期号：6

起止页码：183

中文期刊名：生物技术通报

外文期刊名：Biotechnology Bulletin

收录：CSTPCD、、北大核心2014、北大核心、CSCD_E2015_2016、CSCD

基金：广东省教育厅高等学校高层次人才项目

语种：中文

中文关键词：哈氏弧菌;谷胱甘肽还原酶;基因克隆;原核表达

外文关键词：Vibrio harveyi;glutathione reductase;gene cloning;prokaryotic expression

中文摘要：经克隆哈氏弧菌谷胱甘肽还原酶(GR)基因,并构建其原核表达载体,以获得相应的表达蛋白。将GR和p ET-32a(+)通过Bam H I和Xho I双酶切后,体外用T4连接酶连接,构建重组质粒p ET-GR;然后转化至大肠杆菌BL21(DE3)中,利用异丙基-β-D-硫代半乳糖苷(IPTG)诱导表达,应用SDS-PAGE分析表达情况和表达条件。SDS-PAGE电泳获得分子量约为68.9 k D融合蛋白条带。在E.coli BL21(DE3)中重组质粒p ET-GR的表达条件为28℃,0.7 mmol/L的IPTG浓度诱导4 h表达量最高,且主要以包涵体形式表达。哈氏弧菌谷胱甘肽还原酶基因在大肠杆菌中获得了高效表达。

外文摘要：The purpose of this study is to clone glutathione reductase（GR）gene fromVibrio harveyi,construct the prokaryotic expression vector of it, and obtain the corresponding expressed protein. Digested GR and pET-32a（＋）were cut with the double enzymesBamH I and Xho I, then ligated with T4 ligase to construct the recombinant plasmid pET-GR. Then pET-GR was transformed into Escherichia coli BL21 （DE3）, which was induced by IPTG, and their expressions were analyzed by SDS-PAGE. An approximately 68.9 kD exogenous protein was observed on the SDS-PAGE. The optimal expression condition for the recombinant plasmid pET-GR was that the recombinantE. coli BL21 （DE3）was induced for 4 h at 28℃ by 0. 7 mmol/L of IPTG and it expressed in E. coli as the inclusion bodies. The conclusion of the study is that GR gene fromV. harveyican efficiently express inE. coli.