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目的:应用RNA干扰技术抑制NOB1基因的表达,并在卵巢癌HEY细胞上观察NOB1下调对细胞增殖能力和细胞周期的影响。方法:NOB1特异性siRNA构建于慢病毒载体,感染卵巢癌HEY细胞后,Real-time PCR和Western验证NOB1的抑制效率,MTT、Brdu实验观察NOB1基因沉默对HEY细胞增殖的作用,PI染色流式细胞仪检测细胞周期变化。结果:shRNA慢病毒感染HEY细胞能够将NOB1基因mRNA表达抑制82.2%,蛋白表达也显著降低,MTT和Brdu实验显示细胞增殖水平显著变弱,流式细胞仪检测证实细胞出现G1期阻滞。结论:应用RNA干扰技术能在HEY细胞上有效沉默NOB1基因,并降低卵巢癌细胞的增殖能力,使细胞阻滞在G1期。
OBJECTIVE: To study the effect of NOB1 knockdown on cell proliferation and cell cycle in ovarian cancer HEY cells by RNAi technique. Methods: NOB1-specific siRNA was constructed on lentiviral vector and infected with HEY cells. Real-time PCR and Western blotting were used to verify the inhibitory efficiency of NOB1. The effects of NOB1 silencing on proliferation of HEY cells were observed by MTT and Brdu assays. Cytometry to detect cell cycle changes. Results: HEY cells transfected with shRNA lentivirus could inhibit the expression of NOB1 mRNA by 82.2% and the protein expression was significantly decreased. MTT and Brdu experiments showed that cell proliferation was significantly weakened, and G1 arrest was confirmed by flow cytometry. Conclusion: The application of RNAi can effectively silence NOB1 gene in HEY cells and reduce the proliferation of ovarian cancer cells so that cells arrest in G1 phase.