Construction of SGK3 Gene Lentiviral RNA Interference Vector and Effects on Cell proliferation and Apoptosis of Breast Cancer Cell Line MB-474

doi:10.13560/j.cnki.biotech.bull.1985.2017-0557

Abstract

Abstract: This study is aimed to construct the lentiviral RNA interference（RNAi）vector of SGK3 gene,and to observe its effect on the proliferation and apoptosis of human breast cancer cell line MB-474. The lentiviral vector of shRNA sequence targeting SGK3 was constructed and transfected into human breast cancer MB-474 cells to silence the SGK3 gene. The expression of SGK3 mRNA and protein in transfected MB-474 cells was detected by real-time PCR and Western bloting,and the effect of SGK3 gene silencing on the proliferation,cell cycle and apoptosis of MB-474 cells was detected by MTT assay and flow cytometry. Gene sequencing showed that 4 RNAi lentiviral vectors targeting the SGK3 gene were successfully constructed. The lentiviral vectors were packed successfully by 293T cells and the virus titer was（3-8）×10⁸ TU/mL. After the recombinant lentiviral vector was transfected into MB-474 cells,real-time PCR and Western bloting demonstrated that the expression of SGK3 in interference group was lower than that of control group. Among the 4 lentiviral vectors,the one with best interference effect is the PGC-LV3-SGK3-1 sequence. SGK3 silencing inhibited MB-474 cell proliferation and promoted apoptosis,but had no significant effect on cell cycle progression. In short,RNAi lentivirus vector targeting SGK3 gene was successfully constructed,and SGK3 gene silencing affected cell proliferation and apoptosis of breast cancer.

Key words: RNAi lentivirus, SGK3, cell proliferation, apoptosis

GUO Hong-yan, GAO Han, WU Qi, SUN Xiao-jie, LIU Xiu-cai, ZHAO Li-qun. Construction of SGK3 Gene Lentiviral RNA Interference Vector and Effects on Cell proliferation and Apoptosis of Breast Cancer Cell Line MB-474[J]. Biotechnology Bulletin, 2018, 34(1): 247-252.

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