敲低SET7慢病毒载体的构建及对乳腺癌细胞生长的影响

Construction of a lentiviral vector of SET7 small-interfering RNA and its effect on growth of breast cancer cells

目的为了探讨组蛋白甲基转移酶SET7在乳腺癌发生发展中的功能,构建SET7基因的RNA干扰(RNAi)慢病毒表达载体,研究其对乳腺癌细胞ZR75-1生长的影响。方法根据人SET7的c DNA序列,设计SET7基因短发夹RNA(shRNA)引物,并克隆到p SIH-H1-Puro载体上,包装成慢病毒,感染人乳腺癌细胞ZR75-1,通过实时定量(real-time)PCR以及Western印迹检测干扰效果,并通过生长曲线研究敲低SET7对ZR75-1细胞生长的影响。结果 DNA测序结果表明,SET7 shRNA慢病毒表达载体构建成功。实时定量PCR和Western印迹结果显示,SET7 shRNA能有效抑制SET7基因的表达。生长曲线实验表明,敲低SET7可抑制人乳腺癌细胞ZR75-1的生长。结论成功构建了SET7 shRNA慢病毒表达载体,感染人乳腺癌细胞ZR75-1后,有效抑制了内源SET7基因的表达,敲低SET7能抑制ZR75-1细胞的生长,为进一步研究SET7在乳腺癌中的功能奠定了基础。

Objective To study the biological function of histone methyltransferase SET 7 in the development and pro-gression of breast cancer by constructing the lentiviral vector of SET 7 small-interfering RNA and investigating its effect on the growth of ZR75-1 breast cancer cells.Methods Based on human SET7 cDNA sequence, short hairpin RNA（shRNA） primers targeting SET7 were designed and cloned into the vector pSIH-H1-Puro.Lentivirus was obtained and infected with ZR75-1 breast cancer cells .Real-time PCR and Western blotting were performed to identify the lentivirus-mediated knockdown of SET7.Growth curves were used to study the effect of SET 7 knockdown on the growth of ZR 75-1 cells. Results DNA sequencing indicated that the SET 7 shRNA lentiviral vector was constructed successfully .Real-time PCR and Western blotting showed that SET7 shRNA could suppress SET7 gene expression, which could in turn markedly inhibit the growth of ZR75-1 cells.Conclusion The lentivirus-mediated SET7 shRNA is successfully constructed .Knocking down SET7 could suppress the growth of ZR75-1 cells.