Rad51基因沉默对人肺腺癌A549细胞体内外增殖能力的影响.pdfVIP

Abstract ABSTRACT Objective: To silence gene Rad51 in A549 cells by Rad51 siRNA, and to observe variation of the colony-forming ability, the cell viability, the cell cycle progression and the early growth status of transplanted tumor in nude mouse after transfected by Rad51 siRNA. Then investigate the role of Rad51 in the survival of lung cancer with the aim to provide experimental data and theory data for targeted therapy of NSCLC. Methods: A549 cells were cultured as normal, Rad51 siRNA was designed and constructed to Lentivirus. The experiment was divided into three groups ：①Normal group ：the human A549 cells were cultured with DMEM medium suppl- emented with 10% Fetal Bovine Serum ； ②Lenti-NC group ：the A549 cells were infected with lentivirus of empty vector ；③Lenti-Rad51si group ：infected with lentivirus of Rad51siRNA； Real-time quantitative PCR analysis and Western blot were used to check the mRNA and protein level of Rad51 in A549 cells; cell proliferation ability was decided by Colony-forming assay; cell cycle were analyzed by fluorescence-activated cell sorter; the early growth status of transplanted tumor in nude mouse were observed. Result: 1. Lenti-Rad51si and Lenti-NC virus were manufactured by Shanghai Genechem Company. The virus Lenti-Rad51 titer was 2×108 TU/ml and Lenti-NC titer was 8×108 TU/ml packaging; 2. A high transfection efficiency could be obtained after infection of Lenti-Rad51 siRNA and Lenti-NC in A549 cells; 3. After 48h of the Lenti-Rad51si infection, the mRNA and protein expression of Rad51 were downregulated detected by Quantitative RT-PCR and Western Blot; 4. Silencing gene Rad51 resulted in a re