Effects of RNAi on HER2/neu gene and proliferation of SK-OV-3 ovarian cancer cells in vitro

Objective: To investigate the effects of HER2/neu gene silencing on SK-OV-3 ovarian cancer cells by RNA interference.Methods: Design and synthesize two siRNAs based on the sequence of HER2/neu mRNA.They were separately subcloned into the plasmid of pGenesil-1 containing U6 promoter.The pGenesil-1 vectors of the RNA interference eukaryotic expression specific to HER2/neu gene were constructed.The pGenesil-1 vectors were transfected into SK-OV-3 ovarian cancer cells with METAFECTENE in vitro.The positive cell clones were obtained after being screened with(250?μg/ml) G418.Then the effects of RNAi on the expression of HER2/neu gene were detected by RT-PCR and Western Blot.The cellular growth activities were assayed by CCK-8 colorimetry and flow cytometry in vitro.Results: DNA sequencing of the plasmids verified the successful construction of the HER2/neu siRNA vectors. Compared with the blank and the control group,the expressions of HER2/neu mRNA and protein 185 were remarkably down-regulated in the RNAi groups;and the cell proliferations decreased in the RNAi groups.Conclusion: We successfully construct the recombinant plasmids of siRNAs specific to HER2/neu gene and transfect them into the SK-OV-3 ovarian cancer cells in vitro.The RNAi inhibits the expression of HER2/neu mRNA and protein 185,and subsequently suppresses the proliferation of SK-OV-3 cells.