转染CD147干扰质粒肝癌细胞7721的鉴定

目的构建CD147干扰质粒,对肝癌细胞7721内源性CD147表达的抑制效果进行检测。方法设计及构建3对pGenesil-1-CD147/shRNA及1对阴性对照干扰质粒,通过测序鉴定。将干扰质粒用Lipofec-tamineTM2000转染肝癌细胞,通过瞬时转染获得细胞系,实时PCR和蛋白印迹法检测3对干扰质粒、阴性对照质粒的mRNA及蛋白表达水平。结果实时PCR和蛋白印迹法结果显示干扰质粒pGenesil-1-CD147/shRNA3对肝癌细胞7721内CD147mRNA及蛋白的抑制效果最显著。结论成功构建了CD147干扰真核表达载体,筛选出有效干扰质粒,为进一步研究低表达CD147的肝癌细胞7721在肝癌发生发展中的机制提供基础。

Identification of hepatocellular carcinoma cell 7721 transfected with CD147 recombinant plasmid

Objective To construct the CD147 recombinant plasmid and to detect the inhibitory effects of CD147 endogenous expression in hepatocellular carcinoma cells 7721. Methods Three pairs of pGenesil-l-CD147/ shRNA and negative control recombinant plasmids were designed and constructed for further identification via gene sequencing. The hepatocellular carcinoma cells 2000 were transfected with LipofectamineTM, the plasmid for gene in- terference, to obtain cell lines by transient transfection. This was followed by detection of mRNA and protein expres- sion of the three pairs of plasmids and negative control plasmid v/a real-time polymerase chain reaction and Western blotting. Results The pGenesil-l-CD147/shRNA3 plasmid conferred marked inhibition of CD147 mRNA and protein expression in hepatocellular carcinoma cells 7721, as revealed by real-time polymerase chain reaction and Western blotting. Conclusion Successful construction of CD147 eukaryotic expression interference vector and screening of effective interference plasmids may allow for further studies regarding the mechanism of low-expression CD147 cells 7721 in the development of hepatocellular carcinoma.