日本七鳃鳗Lj-RGD3缺失突变体Lj-113人工合成序列的基因克隆与蛋白表达  

Cloning and expression of the synthesized sequence of deletion mutant Lj-113 of Lj-RGD3 from Lampetra japonica

作　　者：张亚前[1] 李庆伟[1] 吕莉[2] 刘欣[1] 王继红[1] 

机构地区：[1]辽宁师范大学海洋生物功能基因与蛋白质组学研究所,辽宁大连116029 [2]大连医科大学药学院药理教研室,辽宁大连116044

出　　处：《吉林医药学院学报》2011年第2期73-76,共4页Journal of Jilin Medical University

基　　金：国家高技术研究发展863计划资助项目(2007AA09Z428);国家自然科学基金(30770297);辽宁省博士启动基金(20061050);辽宁省博士启动基金(20081080);辽宁省教育厅创新团队项目(No.2008T102);大连市重大科技攻关项目(2007E11SF051);大连市科学技术局留学回国人员科研基金(2008J22JH010)

摘　　要：目的获得突变体Lj-113的基因重组蛋白,为其与野生型活性比较打下物质基础。方法对日本七鳃鳗RGD缺失突变体Lj-113进行基因全序列人工合成,并以NdeI与HindIII为酶切位点构建于pET23b载体上,获得阳性基因重组质粒pET23b-Lj113。CaC l2法将重组质粒转化入大肠杆菌BL21表达菌中,对其进行终浓度为1 mmol/L的IPTG诱导表达,并对表达蛋白进行组氨酸亲和层析纯化蛋白。结果成功获得七鳃鳗RGD缺失突变体的基因重组蛋白的可溶性表达。结论人工合成基因序列可通过分子克隆方法获得其蛋白的表达。Objective To get the recombinant protein of deletion mutant Lj-113 and provide material basis for making comparison in activity to the wild types.Methods Gene sequence of deletion mutant Lj-113 was synthesized and built onto pET23b carrier,with Nde I and Hind III being Enzyme cutting site,to get positive gene recombination plasmid pET23b-Lj113 which was transformed into E.coli BL21 by CaCl2 method and expressed with IPTG whose concentration was 1 mmol/L and purified with histidine affinity chromatography.Results Recombinant rLj-113 protein was obtained successfully.Conclusion Synthesized gene sequence could be expressed by molecular cloning.

关 键 词：七鳃鳗 人工合成序列 缺失突变体 基因克隆 蛋白表达 

分 类 号：Q78[生物学—分子生物学]