人BNIP3基因在Hela细胞中的表达及定位  

Human BNIP3 Gene Expression and Its Intracellular Localization in Hela Cells

作　　者：陈登宇[1] 刘芸[1] 钟田雨[1] 王蔚[1] 赵明哲[1] 刘靖华[1] 姜勇[1] 

机构地区：[1]南方医科大学病理生理学教研室广东省功能蛋白质组学重点实验室,广东广州510515

出　　处：《贵阳医学院学报》2009年第1期5-8,共4页Journal of Guiyang Medical College

基　　金：长江学者和创新团队发展计划(No:IRT0731);国家自然科学基金委员会-广东省人民政府自然科学联合基金重点项目(No:U0632004);国家自然科学基金项目(No:30670829)

摘　　要：目的:构建BNIP3-HA融合蛋白的真核表达载体,观察BNIP3在Hela细胞中表达及其定位。方法:采用两步克隆法将HA和BNIP3的编码序列以融合表达的形式克隆到载体pcDNA3上,随后转染Hela细胞。BNIP3经AlexaFluor 488免疫荧光标记,线粒体用MitoFluor Red 589染色后在荧光显微镜下观察BNIP3的表达和定位。结果:重组质粒经酶切、聚合酶链反应(PCR)和测序鉴定构建正确,并在Hela细胞中能够表达,在荧光显微镜下观察,BNIP3-HA融合蛋白分布于线粒体。结论:成功构建BNIP3-HA融合蛋白表达载体并在Hela细胞线粒体中表达。Objective： To construct human BNIP3 fuse protein vector, express it in Hela cells, and determine its location in the cells. Methods： The expression vector was constructed by cloning the coding sequences of fusion protein of human BNIP3 and HA onto vector pcDNA3 by two-step method; The constructed vector was then transfected into Hela cells and observed with fluorescence microscope. The recombinant plasmid was verified by enzyme digestion, polymerase chain reaction （PCR） and sequence analysis. Results： The fusion protein was highly expressed in Hela cells. Being observed with fluorescence microscopy, the fusion protein of human BNIP3 and HA was found to distribute in the mitoehondria. Conclusion： The expression vector of BNIP3 fusing to HA is successfully constructed and the fusion protein has expressed in the mitoehondria of Hela cells, which would facilitate the further study of BNIP3.

关 键 词：基因 BCL-2 HELA细胞 线粒体 荧光免疫测定 细胞内定位 

分 类 号：R73-75[医药卫生—肿瘤] Q753[医药卫生—临床医学]