| GST-SRY融合蛋白原核表达和纯化 |
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| 引用本文： | 张小玉,段君君,王江,贾霄,汪小波,何颖红. GST-SRY融合蛋白原核表达和纯化[J]. 昆明医科大学学报, 2018, 39(4): 1-4 |
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| 作者姓名： | 张小玉 段君君 王江 贾霄 汪小波 何颖红 |
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| 作者单位： | 1. 大理大学基础医学院 |
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| 基金项目： | 基金: 国家自然科学基金资助项目 (31360288); |
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| 摘 要： | 目的 构建GST-SRY融合蛋白原核表达载体, 在E.coli中诱导其表达, 并进行纯化.方法 PCR扩增SRY基因, 将SRY基因插入原核表达载体p GEX-6P-1, 经酶切和测序验证后, p GEX-6p-1-SRY转化到原核表达菌株E.coli Rosetta DE3, IPTG诱导表达GST-SRY融合蛋白, 用GST标签纯化树脂对其进行纯化.结果 经双酶切和测序证实p GEX-6P-1-SRY构建正确.重组质粒转化到Rosetta DE3经IPTG诱导后表达了分子质量单位约为49 KD的重组蛋白.结论 p GEX-6P-1-SRY原核表达载体构建成功, SRY蛋白成功诱导表达并纯化, 为今后深入研究SRY蛋白的功能奠定了实验基础.
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| 关 键 词： | SRY 基因重组 基因表达 蛋白纯化 |
| 收稿时间： | 2017-11-20 |
Prokaryotic Expression and Purification of GST-SRY Fusion Protein |
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| Abstract: | Objective To construct GST-SRY fusion protein expression vector and induce its expression in E.coli, then purificate so as to obtain GST-SRY fusion protein. Me thods SRY gene was amplified by PCR and cloned into prokaryotic expression vector p GEX-6 P-1. p GEX-6 P-1-SRY plasmid was verified by restriction enzyme digestion and DNA sequencing. After that, the recombinant plasmid was transformed into E.coli Rosetta DE3.Expression of GST-SRY fusion protein was induced by IPTG, which was purified by Glutathione Sepharose later on.Re s ults Restriction enzyme digestion and sequencing demonstrated that p GEX-6 P-1-SRY was successfully obtained. The result of SDS-PAGE assay indicated that SRY was successfully expressed in E.coli Rosetta DE3 and expressed a 49 KD SRY protein.Conclus ion The successful construction of SRY gene recombinant prokaryotic expression vector and expression of SRY protein will be helpful for further studies of SRY gene function. |
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