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哈氏弧菌谷胱甘肽还原酶基因克隆及原核表达

DOI: 10.13560/j.cnki.biotech.bull.1985.2015.06.029, PP. 183-188

Keywords: 哈氏弧菌,谷胱甘肽还原酶,基因克隆,原核表达

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Abstract:

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pET-32a(+)通过BamHI和XhoI双酶切后,体外用T4连接酶连接,构建重组质粒pET-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68.9kD融合蛋白条带。在E.coliBL21(DE3)中重组质粒pET-GR的表达条件为28℃,0.7mmol/L的IPTG浓度诱导4h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。

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