文献类型：期刊文献

中文题名：南极衣藻(Chlamydomonas sp.ICE-L)谷胱甘肽还原酶基因的原核表达及其条件优化

英文题名：PROKARYOTIC EXPRESSION AND ITS CONDITIONAL OPTIMIZATION OF GLUTATHIONE REDUCTASE GENE OF ANTARCTIC CHLAMYDOMONAS SP.ICE-L

作者：刘莹[1];丁燏[1];简纪常[1];吴灶和[1];缪锦来[2]

机构：[1]广东海洋大学水产学院广东省水产经济动物病原生物学及流行病学重点实验室,湛江524025;[2]国家海洋局第一海洋研究所海洋生物活性物质重点实验室,青岛266061

年份：2011

卷号：42

期号：6

起止页码：817

中文期刊名：海洋与湖沼

外文期刊名：Oceanologia Et Limnologia Sinica

收录：CSTPCD、、Scopus、CSCD2011_2012、北大核心2008、北大核心、CSCD

基金：国家自然科学基金资助项目;40876102号;40876107号

语种：中文

中文关键词：南极衣藻Chlamydomonas;sp.ICE-L;谷胱甘肽还原酶;原核表达;诱导条件;冰藻

外文关键词：Antarctic Chlamydomonas sp. ICE-L, Glutathione reductase （GR）, Prokaryotic expression, Induced condition, Ice algae

中文摘要：采用RT-PCR技术克隆南极衣藻GR基因ORF全长cDNA,然后经酶切、连接等步骤构建其原核表达载体;并对其表达的诱导时间、IPTG浓度、温度进行了优化,以期获得较大量的酶蛋白。结果表明,将构建的原核表达载体pET-GR导入大肠杆菌BL21,可以高效表达融合蛋白;且表达的蛋白均以包涵体的形式存在;经SDS-PAGE电泳结果显示,获得的目的蛋白分子量为52.2kDa,符合预期分子量;GR蛋白在大肠杆菌中诱导表达优化条件为,1.0mmol/L的IPTG,28℃诱导3h。这些结果为进一步深入研究该基因的特性与功能奠定了基础。

外文摘要：Glutathione reductase （GR） is an important antioxidative enzyme in organisms. In order to construct prokaryotic expression vector and to study the recombinant gene expression in Escherichia coli, complete gene of Chlamydomonas sp. ICE-L GR ORF was cloned by the RT-PCR method followed by restriction enzymes cutting, connection. The induced time, IPTG concentration and temperature of prokaryotic expression were optimized using SDS-PAGE. The results showed that a recombinant of prokaryotic expression vector pET-GR was constructed successfully, and ICE GR protein in inclusion body can express effectively after the expression vector pET-GR was transformed to E. coli BL21. The SDS-PAGE results indicated that the molecular weight （52.2kDa） of the target protein was similar to the theoretic molecular weight. The optimal expression condition was set as 1.0mmol/L IPTG, induced temperature 28℃, and induced time 3 hours. These results, are expected to lay a foundation for further studies on the properties and function of this gene.