shRNA抑制SGC7901胃癌细胞绿色荧光蛋白基因的表达

目的：观察针对绿色荧光蛋白(GFP)基因的发夹结构RNA(small hairpin RNA，shRNA)表达载体(SHi—pU6-GFP)对SGC7901胃癌细胞中GFP的抑制作用。方法：设计针对GFP基因的shRNA序列．构建SHi—pU6-GFP质粒。采用质粒pCX—GFP(5510bp)和含有鼠U6启动子pU6（3．3kb)质粒，应用Lipofectamine^TM2000将pCX-GFP质粒和SHi—pU6-GFP质粒转染SGC7901细胞。荧光显微镜观察转染效果和不同时间的转染效率。结果：质粒SHi-pU6-GFP经酶切电泳后，出现3．3kh和约350bp条带，与实验设计的针对GFP的shRNA长度相符。说明本实验已成功构建了针对绿色荧光蛋白GFP基因的SHi—pU6-GFP质粒。SGC7901细胞中观察到转染pCX—EGFP质粒和SHi—pU6-GFP质粒前后发绿色荧光的细胞数目明显减少：结论：shRNA可以有效地抑制SGC7901细胞中GFP基因的表达。

Inhibition of GFP expression by small hairpin RNA in SGC 7901 gastric cancer cells

Objective To study the inhibition of GFP gene expression in SGC 7901 cells by using an expression vector carrying small hairpin RNA (SHi,pU6,GFP). Methods Plasmids pCX,GFP(5510 bp), pU6 (3.3 kb)and SHi,pU6,GFP carried hairpin RNA for GFP gene were transfected to SGC7901 cells by using LipofectamineTM2000. Results The constructed SHi,pU6,GFP plasmids carried hairpin RNA for GFP gene as designed. The expression of GFP gene was inhibited by SHi,pU6,GFP in SGC 7901 cells. Conclusions Short hairpin RNA (shRNA) could induce sequence,specific GFP gene silencing in SGC 7901 cells. shRNA,induced silencing may shed light on the application of RNA interference in the treatment of gastric cancer.