| 斑马鱼p53基因原核表达载体的构建及其在大肠杆菌中的表达 |
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| 引用本文： | 赵向忠,刘明,丁丽丽,吴宁,林秀坤.斑马鱼p53基因原核表达载体的构建及其在大肠杆菌中的表达[J].湖北农业科学,2010,49(3). |
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| 作者姓名： | 赵向忠 刘明 丁丽丽 吴宁 林秀坤 |
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| 作者单位： | 1. 中国科学院海洋研究所,山东青岛,266071;中国科学院研究生院,北京,100039
2. 中国科学院海洋研究所,山东青岛,266071 |
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| 基金项目： | 国家自然科学基金，山东省科技攻关项目 |
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| 摘 要： | 利用原核表达系统克隆表达斑马鱼p53基因。RT-PCR法从斑马鱼胚胎中扩增获得p53基因编码区,并将其克隆至原核表达载体pET28a上,构建重组质粒pET28a/z-p53,将重组质粒转化E.coli BL21(DE3)受体菌,IPTG诱导表达,表达产物经镍柱纯化、尿素透析复性,SDS-PAGE电泳分析,结果表明,p53基因在大肠杆菌中成功表达,表达的p53融合蛋白分子量大约为53kD,透析复性后获得了高纯度可溶性的p53蛋白。
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| 关 键 词： | p53基因 斑马鱼 原核表达 |
Prokaryotic Vector Construction and Expression of Zebra Fish p53 Gene |
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| Abstract: | The cloning and expressing of zebra fish p53 gene were studied by using prokaryotic expression system. P53 encoding region was obtained from zebra fish embryos by RT-PCR,and then the p53 fragment was cloned in pET28a vector to construct the recombinant plasmid pET28 a/z-p53. It was transformed into E. Coli BL21, and then induced by lmmol/L IPTG to express p53 protein in E.coli BL21 expression system. The recombinant p53 protein was purified by Ni-NTA affinity chromatography under denaturing conditions and renatured through urea gradient dialysis. Soluble p53 protein with high purity was successful obtained. |
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| Keywords: | p53 gene zebra fish prokaryotic expression |
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