融合表达形式可实现外源蛋白的高水平表达,尤其适用于小分子多肽类物质的表达。本课题中,利拉鲁肽前体分子Arg34GLP-1(7-37)属于短肽分子(含31AA),选用该质粒作为重组表达载体,并在Arg34GLP-1(7-37)的N端前设计肠激酶识别位点与之紧邻,C端连接终止密码子,三者构成目的基因,然后插入多克隆位点AIwNI、XhoI之间,以融合蛋白KSI-Arg34GLP-1的包涵体形式表达,从而实现重组融合蛋白的高效表达,后期再用高特异性的肠激酶切割融合蛋白,就可获得大小正确的利拉鲁肽前体分子Arg34GLP-1,表达流程如图5。万方数据王雪娜遵义医学院硕士学位论文7图1111KSI-Arg34GLP-1KSI-Arg34GLP-1KSI-Arg34GLP-1KSI-Arg34GLP-binantvectorvectorvectorvectorandandandandexpressionexpressionexpressionexpressionofofofofKSI-Arg34GLP-1KSI-Arg34GLP-1KSI-Arg34GLP-1KSI-Arg34GLP-1Note:ThesologenesegmentofArg34GLP-1isnamedasLG;ThedesiredtagetgeneisgainedbyPCRandnamedLG1;LG1andprimarypET-31b(+)vectorarefirstlydigestedrespectivelywithAIwNIandXhoI,binantpET-31b(+)-LG1vector;binantvectorwasfinallytransformedintoE.coliBL21(DE3)andinducedwithIPTGtoexpressfusionproteinKSI-Arg34GLP-1.万方数据
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