草鱼原癌基因c-fos的原核表达载体构建及蛋白表达纯化  

Construction of Prokaryotic Expression Vector and Protein Purification of Proto-oncogene c-fos of Ctenopharyngodon idellus

作　　者：邱倩 QIU Qian(Qibo Medical College,Longdong University,Qingyang 745000,Gansu)

机构地区：[1]陇东学院岐伯医学院,甘肃庆阳745000

出　　处：《陇东学院学报》2022年第2期105-109,共5页Journal of Longdong University

基　　金：甘肃省高等学校创新基金项目(2020A-114);陇东学院青年科技创新项目(XYZK1620)。

摘　　要：为克隆草鱼原癌基因c-fos,构建原核表达载体,诱导表达并纯化c-Fos融合蛋白,为c-Fos蛋白的多克隆抗体制备奠定前期基础。在生物信息学技术方法指导下,利用引物设计软件Primer Premier 5设计c-fos基因的特异性引物,扩增得到883 bp长度的目标核酸片段,构建了原核表达载体pET-28a/c-fos,转化至E.coli Rosetta原核表达系统中通过IPTG诱导表达,检测到连接有His标签的c-Fos蛋白,最终通过His镍柱纯化得到了His-c-Fos融合蛋白。该融合蛋白可作为抗原用于后期c-Fos蛋白的多克隆抗体制备,为其组织和亚细胞定位及生物学功能研究奠定基础。We aim to clone the proto-oncogene c-fos of Ctenopharyngodon idellus and construct its prokaryotic expression vector,induce expressing and purify the c-Fos recombinant protein,and provide a preliminary basis for the preparation of polyclonal antibody of c-Fos protein.Under the guidance of bioinformatics technology methods,we used primer design software Primer Premier 5 to design specific primers for c-fos gene,and 883 bp target nucleic acid fragment was amplified,prokaryotic expression vector pET-28a/c-fos was constructed and transformed into E.coli Rosetta.The expression of recombinant protein was induced by IPTG,and c-Fos protein with his tag detected.Finally,His-c-Fos recombinant protein was purified by Ni-NTA His Bind.It can be used as an antigen for the preparation of polyclonal antibodies against c-Fos protein,which lays a foundation for the study of tissue and subcellular localization and biological function of the c-Fos protein.

关 键 词：草鱼 C-FOS 基因克隆 原核表达 蛋白纯化 

分 类 号：Q[生物学]