Silencing Of PCNA Gene Expression In Colon Cancer Cell Line Using Designed SiRNA

Objective:To design and screen out functional small interfering RNA (siRNA) for specifically silencing PCNA gene expressing and effectively inhibiting cell proliferation in human digestive cancer cell lines, evaluate the significance of mRNA local structures in siRNA design based on RNAstructure 3.7 and Mfold software analysis, and to study the anti-proliferation and apoptosis inducing effects on human colorectal carcinoma Caco2 cells following PCNA silencing induced by PCNA siRNA.MethodsPCNA siRNA were designed based on previous studies about design guidelines and transcribed through T7 polymerase methods. Four in vitro synthesized double-stand siRNA (dsRNA) were respectively transfected into three types of digestive carcinoma cell lines by lipofectamine?2000 reagent . The subsequent alterlations in PCNA mRNA and protein levels were determined by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) and immunocytochemistry, respectively. WST-8 assays and colony forming inhibition assay were employed to examine the changes of cell proliferation. Flow cytomy and fluorescence microscopy were used to check the changes in cell cycle and cell apoptosis. The correlation between siRNA efficacy and values about mRNA local secondary structure...