Construction of RNA interfering lentivirus vector targeting PIK3CA gene and its identification

Objective To construct the RNA interfering(RNAi) lentivirus vector targeting PIK3CA(P110α).Methods Four specific target sequences and a negative control sequence from PIK3CA-mRNA sequence(NM_006218) were designed to construct pGCL-GFP.293T cells were co-transfected with the successfully constructed RNAi plamid and PIK3CA expression plamid.The transfection efficiency was observed under a fluorescence microscope.Western blot was used to detect the expression of PIK3CA protein.RNAi plasmid with an optimal interfering efficacy was screened and 293T cells were co-transfected with auxiliary packaging vectors to pack lentivirus and detect its titer.Results The RNAi plamid with four targets and a negative control sequence was successfully constructed.Western blot analysis showed that the interfering targets were optimal.The lentivirus with a titer of 2×108TU/ml was successfully packed.Conclusion RNA interfering lentivirus vector targeting PIK3CA gene can be constructed,which provides a foundation for investigating PIK3CA function and gene therapy for ovarian cancer using lentivirus.