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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 investigate the function of SET7 in the development and progression of breast cancer and to construct RNA interference (RNAi) lentivirus vector of SET7 gene to study its effect on the growth of breast cancer cell line ZR75-1. Methods The short hairpin RNA (shRNA) of SET7 gene was designed based on the human DNA sequence of human SET7 and cloned into p SIH-H1-Puro vector and packaged into lentivirus to infect human breast cancer cell line ZR75-1. Real-time PCR and Western blot were used to detect the interference effect. The effect of SET7 knockdown on the growth of ZR75-1 cells was studied by growth curve. Results DNA sequencing results showed that SET7 shRNA lentivirus expression vector was successfully constructed. Real-time quantitative PCR and Western blotting results showed that SET7 shRNA can effectively inhibit the expression of SET7 gene. Growth curve experiments showed that SET7 knockdown inhibited the growth of human breast cancer cell line ZR75-1. Conclusions SET7 shRNA lentiviral vector was successfully constructed. After infection with human breast cancer cell line ZR75-1, the expression of endogenous SET7 gene was effectively inhibited. Knockdown of SET7 inhibited the growth of ZR75-1 cells. To further investigate the role of SET7 in breast cancer In the function laid the foundation.