人雌激素相关受体α配体结合域的原核表达载体构建·表达及鉴定  

Construction,Expression and Identification of Prokaryotic Expression Vector of Human ERRα-LBD

作　　者：王燕飞[1] 黄志伟[2] 

机构地区：[1]中国科学院华南植物园,广东广州510650 [2]中国科学院广州生物医药与健康研究院,广东广州510663

出　　处：《安徽农业科学》2008年第23期9912-9913,9920,共3页Journal of Anhui Agricultural Sciences

摘　　要：[目的]构建含人雌激素相关受体α配体结合域区段的原核表达载体,并在大肠杆菌表达系统中表达。[方法]用Trizol试剂提取人肺腺癌A549细胞总RNA,逆转录为cDNA;应用PCR扩增人EERα-LBD基因片段,插入表达载体pET-30a-c(+)中;转化大肠杆菌BL2 l(DE3)PLysS,经IPTG诱导后表达并鉴定。[结果]应用PCR方法扩增出约687 bp的基因片段,酶切、纯化、连接至载体后,经测序与GenBank中的一致,表达蛋白相对分子质量约32 kD,纯化后用生物大分子相互作用系统鉴定为目的蛋白。[结论]已成功获得人EERα-LBD融合蛋白,为进一步的研究和应用奠定了基础。[ Objective ] The research aimed to clone human ERRα-LBD and purify its fusion protein. [ Method ] The gene encoding ERRα- LBD was amplified by PCR,and first inserted into cloning vector pUC-19. After sequenced, ERRα-LBD was then cloned into expression vector pET-30a-c （ ＋ ）and transformed to E. coli BL2 1 （DE31 ） for expression. After induction with IPTG, the strain expressed the fused ERRα-LBD protein. The target protein was purified under denaturing condition and confirmed. [ Result] A gene fragment at a length of 687 bp was amplified. And its sequence was identical to that reported in GenBank. The protein with a relative molecular weight of 32 kD was expressed and was identified after purification. [ Conclusion] ERRα-LBD fusion protein was successfully expressed which laid a foundation for further study and the application of ERRα-LBD.

关 键 词：雌激素相关受体α配体 基因克隆 原核表达 

分 类 号：Q78[生物学—分子生物学]