RNA干扰GPC3基因对肺癌A549细胞生物学行为的影响

发布时间：2018-04-03 09:11

本文选题：肺腺癌　切入点：GPC3　出处：《重庆医科大学》2016年硕士论文

【摘要】：目的:研究用靶向GPC3基因的si RNA干扰人肺腺癌A549细胞中GPC3基因的表达后,对A549细胞生物学行为产生的影响,说明GPC3基因在A549细胞株中的作用,进而为人类肺腺癌的早期诊断、预测预后及治疗手段提供理论基础。方法:将细胞接种于不含抗生素的培养基中,24h后用Opti-MEM稀释Lipofectamine 2000和siRNA的混合液,再加入到需要转染的细胞中。37℃培养24h后,通过western blot和荧光定PCR检测空白组,NC组及干扰组的细胞GPC3蛋白/基因表达量。通过CCK-8实验、划痕实验以及流式细胞术检测干扰了GPC3基因之后的A549细胞的生物学行为的变化。结果:干扰组GPC 3基因的蛋白及核算表达水平明显低于空白组和NC组,差异具有显著性(P0.05);而NC组和空白组的表达量没有显著差异。与NC组和空白组A549细胞相比,干扰组的A549细胞迁移和增殖能力明显降低,而细胞凋亡率明显增加(P0.01)。结论:利用Lipofectamine 2000介导的基因转染技术成功的干扰了A549细胞中GPC3基因的表达。当GPC3的表达降低时,A549细胞的增殖和迁移能力都减低;相反,A549细胞的凋亡率增加。
[Abstract]:Objective: To study the expression of GPC3 targeting GPC3 gene by Si interference of RNA human lung adenocarcinoma cell A549 gene, the influence on the biological behavior of A549 cells, indicating that the effect of GPC3 gene in A549 cell line, and then for the early diagnosis of human lung adenocarcinoma, and provide a theoretical basis for prognosis and treatment. Methods: the cells were seeded in culture medium without antibiotics, 24h with a mixture of Opti-MEM diluted Lipofectamine 2000 and siRNA, and then added to the 24h culture.37 C need transfected cells after western and blot by fluorescence quantitative PCR detection of the blank group, the expression of NC group and interference group of cells GPC3 protein / gene the amount of CCK-8. Through the experiment, changes of scratch test and flow cytometry interfere with the biological behavior of GPC3 gene in A549 cells. Results: the protein and accounting interference group GPC 3 gene expression levels were significantly lower than the blank group and In NC group, the difference was significant (P0.05); and the expression of NC group and blank group was no significant difference. Compared with the NC and A549 cells in the blank group, A549 cell migration and proliferation ability of interference group was significantly decreased, and the apoptosis rate increased significantly (P0.01). Conclusion: the use of Lipofectamine 2000 mediated gene transfection technique successfully interfered with the expression of GPC3 gene in A549 cells. The decreased expression of GPC3, A549 cell proliferation and migration are reduced; on the contrary, the apoptosis rate of A549 cells increased.

【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R734.2

【参考文献】