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目的验证HBeAgTP基因在细胞内的表达及表达蛋白的亚细胞定位。方法构建HBeAgTP基因的酵母表达诱饵质粒pGBKT7-HBeAgTP及绿色荧光蛋白表达质粒pEGFP-C1-HBeAgTP,pEGFP-C1-HBeAgTP转染HepG2细胞,24h后荧光显微镜下观察蛋白表达的亚细胞定位。pGBKT7-HBeAgTP转化AH109酵母细胞,提取转化了质粒的酵母蛋白质,进行Western免疫印迹分析。结果成功构建出HBeAgTP基因。HBeAgTP基因的酵母表达诱饵质粒pGBKT7-HBeAgTP及绿色荧光蛋白表达质粒pEGFP-C1-HBeAgTP,Western免疫印迹法印证HBeAgTP可表达蛋白,其表达的蛋白亚细胞定位于细胞质。结论HBeAgTP基因可表达蛋白,其表达蛋白亚细胞定位于细胞质。
Objective To verify the expression of HBeAgTP gene in cells and the subcellular location of the expressed protein. Methods The bait plasmid pGBKT7-HBeAgTP and green fluorescent protein plasmid pEGFP-C1-HBeAgTP were constructed and transfected into HepG2 cells with pEGFP-C1-HBeAgTP. The subcellular localization of the protein was observed under a fluorescence microscope 24 hours later. pGBKT7-HBeAgTP was transformed into AH109 yeast cells and the plasmid-transformed yeast protein was extracted for Western immunoblot analysis. Results HBeAgTP gene was successfully constructed. HBeAgTP yeast expression plasmid bait plasmid pGBKT7-HBeAgTP and green fluorescent protein plasmid pEGFP-C1-HBeAgTP, Western immunoblotting confirmed HBeAgTP expression of protein, the expression of protein subcellular localization in the cytoplasm. Conclusion The HBeAgTP gene can express the protein, and the subunits of the expressed protein are located in the cytoplasm.