绿色荧光蛋白融合人核糖核酸酶抑制因子病毒感染NIH3T3细胞的研究

目的：鉴定NIH3T3细胞中绿色荧光蛋白融合人核糖核酸酶抑制因子基因（egfp-hRI）的表达情况。方法：采用上清感染法,将产生绿色荧光蛋白融合人核糖核酸酶抑制因子反转录病毒的PA317细胞上清感染NIH3T3细胞,使反转录病毒介导的绿色荧光蛋白融合人核糖核酸酶抑制因子载体（pLNCX-EGFP-C1-hRI）稳定整合于NIH3T3细胞。用荧光显微镜检测目的基因的表达,采用蛋白质印迹（Western blot）法检测目的基因在NIH3T3细胞的表达。结果：在荧光显微镜下可见绿色荧光在细胞浆内表达,Western blot法检测到目的基因表达。结论：建立表达绿色荧光蛋白融合人核糖核酸酶抑制因子的NIH3T3细胞株。

Study on NIH3T3 cells infected by retrovirus encoding EGFP fusion of human ribonuclease inhibitor protein

Objective： To identify the expression of egfp-hri fusion on NIH3T3 cells which is carried by recombinant retrovirus. Methods： The egfp-hri fusion gene was integrated intoNIH3T3 cells by the infection of recombinant viral supernatant from the G418 resistent PAS17 cells transfected stably with the plasmid of pLNCX-EGFP-CI-hri. The expression effect of egfp-hri fusion was visualized under the fluorescent microscope, while the expression levels of egfp-hri fusion protein by Western blot. Results： The observation from fluorescent microscope showed the green fluorescent was obviously observed in the cytoplasm, western-blotting showed the expression level of egfp-hri fusion gene increased by 83% and 81%, as compared with those uninfected cells and in the cells infected with empty vector, respectively. Conclusion： The infected NIH3T3 cells can express egfp-hri fusion protein successfully.