嗜铬颗粒蛋白B-GST融合蛋白表达载体的构建及其在大肠杆菌中的表达  

Recombination and Expression of Chromogranin B Fusion Protein in E.coli

作　　者：油红捷[1] 毋晓涛[1] 潘颖[1] 王泽生[1] 

机构地区：[1]首都医科大学基础医学院生物化学与分子生物学系

出　　处：《首都医科大学学报》2006年第2期206-209,共4页Journal of Capital Medical University

基　　金：北京市教委(KM200410025006);教育部重点项目(204005)资助项目

摘　　要：目的构建小鼠嗜铬颗粒蛋白B(chromogranin B,CgB)重组原核表达载体。方法先用RT-PCR方法从小鼠脑基因组中扩增出CgB基因cDNA序列,应用T-A克隆技术,克隆到质粒pMD-T vector,经DNA测序证实基因碱基无误后,亚克隆到原核表达质粒pGEX-4T-1中进行表达。结果表达产物经Western Blot分析,在140处有一明显特异带,与预期结果相符。结论构建重组融合表达载体pGEX-CgB,在原核细胞中成功表达出小鼠CgB-GST融合蛋白。Objective To construct the prokaryotic expression vector for mouse chromogranin B gene. Methods Reverse transcriptase-polymerase chain reaction （RT-PCR） was performed on the total RNA extracted from the mouse brain to obtain the cDNA of chromogranin B. The cDNA fragment of CgB was inserted into pMD-T vector. DNA sequencing was performed before the amplified products were cloned into the prokaryotic expression vector pGEX identified by endonuclease digestion. The amplified products were confirmed as the cDNA of chromogranin B by DNA sequencing. The chromogranin B fusion protein expressed in E. coli was identified by Western Blot. Results There was only one obvious band at the position of relative molecular weight 140, and it was equivalent to the expected value. Conclusion The recombinant fusion expression vector pGEX is constructed, and chromogranin B fusion protein gene is expressed in E. coli successfully.

关 键 词：RT-PCR 嗜铬颗粒蛋白B 基因表达 

分 类 号：Q51[生物学—生物化学]