摘要
目的探讨短发夹状RNA对糖调节蛋白78(GRP78)在结直肠癌RKO细胞中表达抑制作用。方法以脂质体Lipofectamine2000介导GRP78短发卡状RNA(shRNA-GRP78)质粒表达载体转染RKO细胞,G418筛选得到稳定表达株,半定量RT-PCR和Westernblooting方法检测GRP78mRNA和蛋白水平变化。结果转染成功并筛选出稳定表达RKO细胞株。与对照组相比,shRNA-GRP78转染RKO细胞GRP78的mRNA及蛋白水平均显著下降(t=7.079、7.510,P<0.05)。结论 shRNA-GRP78能明显抑制RKO细胞GRP78mRNA及蛋白的表达,为进一步在体内外研究GRP78在结直肠癌中的作用奠定了基础。
Objective To explore the supression of glucose-regulated protein 78（GRP78）expression in human colorectal carcinoma cells RKO by short hairpin RNA（shRNA）interference.Methods The shRNA plasmid expression vector（shRNA-GRP78）was transfected into cells with Lipofectamine 2000.After selection with G418,the stable cell clones were attained.GRP78 expression was assayed with real-time quantitative PCR and Western blotting at both mRNA and protein levels.ResultsThe transfection was successful and stable RKO cell clones with shRNA-GRP78 plasmid were obtained.Compared with the control,the shRNA-GRP78 plasmid caused significant reduction of GRP78 expression at mRNA and protein levels（t=7.079,7.510;P〈0.01）.Conclusion The shRNA-GRP78 plasmid can significantly inhibit the expressions of GRP78 mRNA and protein le-vels,which establishes a foundation for further study,in vivo and in vitro,of the role of GRP78 in colorectal carcinoma.
出处
《青岛大学医学院学报》
CAS
2010年第5期387-389,共3页
Acta Academiae Medicinae Qingdao Universitatis
基金
山东省优秀中青年科学家奖励基金资助项目(2006BSB14001)
山东省自然科学基金资助项目(2009ZRA02112)
