人癌症/睾丸相关蛋白XAGE-1b原核表达载体的构建及融合蛋白的表达  

Construction and fusion protein expression of prokaryotic expression vector of human cancer/testis related protein XAGE-1b

作　　者：郑慧玲[1,2] 刘振杰[3] 陈碧[4] 丁航[2] 刘新光[1,2] 

机构地区：[1]广东省医学分子诊断重点实验室,广东东莞523808 [2]广东医学院生物化学与分子生物学研究所,广东湛江524023 [3]广东省中医院检验科,广东广州510370 [4]广东医学院附属医院,广东湛江524001

出　　处：《广东医学院学报》2014年第4期432-434,438,共4页Journal of Guangdong Medical College

基　　金：国家自然科学基金项目(No.81170327);广东省自然科学基金(No.05011579);东莞市科技计划项目(No.2012108102022);广东医学院科研基金(No.XQ0621)

摘　　要：目的构建人癌症/睾丸相关蛋白XAGE-1b原核表达载体及诱导XAGE-1融合蛋白表达。方法从文库质粒pACT2-XAGE-1b中利用PCR法扩增XAGE-b cDNA编码序列,将其克隆到pET-32a载体中,并将重组质粒pET-32bXAGE-1b转染大肠杆菌DE3中,用异丙基硫代半乳糖苷(IPTG)诱导表达,Western blot检测融合蛋白表达。结果原核表达载体pET-32a-XAGE-1b构建成功;转染菌株经IPTG诱导后,出现约29 kD蛋白过表达,Western-blot检测显示His6融合蛋白表达。结论成功构建原核表达载体pET-32a-XAGE-1b,转染菌株可表达His6融合蛋白。Objective To construct the prokaryotic expression vector of human cancer/testis related protein XAGE-1b and induce XAGE-1 fusion protein expression. Methods The XAGE-1b cDNA was amplified by PCR from pACT2-XAGE-1b and inserted into expression vector pET-32 b.The recombinant plasmid pET-32b-XAGE-1b was transfected into ？DE3 strain of Escheria coli, followed by the induction of isopropyl a-D-1-thiogalactopyranoside（IPTG）. XAGE-1 fusion protein expression was detected by Western blot. Results The prokaryotic expression vector pET-32b-XAGE-1b was successfully constructed. After induced by IPTG, the transfection strain showed overexpression of approximately 29 kD protein, and Western-blot revealed His6 fusion protein expression. Conclusion The prokaryotic expression vector pET-32b-XAGE-1b transfected strain can express His6 fusion protein.

关 键 词：XAGE-1b 原核表达 融合蛋白 

分 类 号：R446.1[医药卫生—诊断学]