PGEFP-C1/N载体的构建及其在转染细胞中的初步定位

Construction of PGEFP-C1/N and its localization in transfected cell

目的观察SARS冠状病毒(SARS-CoV)N蛋白的亚细胞定位。方法将SARS-CoV N基因片段克隆入融合绿色荧光蛋白载体(PGEFP-C1)中。采用瞬时转染技术,在A549、VeroE6细胞株中利用荧光显微镜观察GPT-N蛋白的亚细胞定位,Westernblot鉴定N蛋白的表达。结果成功构建了PGEFP-C1/N表达载体,在A549、VeroE6细胞中观察到N蛋白定位于细胞胞浆中,Western blot证实了N蛋白表达。结论SARS-CoV N蛋白在真核细胞中定位于细胞胞质中。

Objective To identify the subcellular location of SARS-CoV N protein in mammalian cells. Methods The gene fraction of SARS-CoV was cloned into the PGEFP-C1 plasmid to construct expression vector PGEFP-C1/N. The subcellular location of N in A549 and VeroE6 cells was observed under fluorescence microscope with the aid of transient transfection technique. The expression of the fusion protein （GFP N） was detected by Western blot. Results The PGEFP-C1/N was constructed. N protein was localized in the cytoplasm of transfected cells and detected by Western blot. Conclusion N protein was localized in the cytoplasm of mammalian cells.