人BNIP3基因在Hela细胞中的表达及定位

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Alexa Fluor488免疫荧光标记，线粒体用MitoFluor Red589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。

Human BNIP3 Gene Expression and Its Intracellular Localization in Hela Cells

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.