RNA干扰抑制结肠癌细胞血管内皮生长因子表达  

Inhibition of Vascular Endothelial Growth Factor by RNA Interference in HT29 Cells

作　　者：李铁军[1] 康楷[2] 宋建宁[3] 胡瓒斓[3] Tong-Chuan He 张秉强[1] 张才全[1] 

机构地区：[1]重庆医科大学附属第一医院,重庆400016 [2]重庆医科大学附属第二医院,重庆400010 [3]重庆市第九人民医院,重庆400700 [4]Molecular Oncology Laboratory The University of Chicago Medical Center IL60637, USA

出　　处：《中国生物工程杂志》2007年第8期14-18,共5页China Biotechnology

基　　金：国家青年科学基金资助项目(30300298)

摘　　要：目的:应用RNA干扰技术抑制结肠癌血管内皮生长因子(VEGF)表达。方法:将VEGF基因作为RNA干扰的靶区,通过E-RNAi网上提供的服务,设计两个特异的RNA干扰序列,将其装入含U6启动子的载体上,构建成抗VEGF基因的小发夹样RNA(shRNA)表达载体,再转染人结肠癌细胞HT29,通过RT-PCR、Northern blotting、免疫荧光和Western blotting,观察VEGF表达受抑的程度。结果:成功构建了两种抗VEGF基因的shRNA表达载体,RT-PCR、Northern blotting、免疫荧光和Western blotting,均发现其能明显抑制HT29细胞VEGF基因的表达,抑制率分别达42%、88%、73%和82%。结论:针对VEGF基因的shRNA表达载体能够明显抑制结肠癌细胞VEGF基因的表达。Objective：To inhibit the expression of Vascular Endothelial Growth Factor （VEGF） by RNA interference in HT29 cells. Methods：Using the service of E-RNAi, we designed and constructed two kinds of shRNA expression vectors which were aimed at the VEGF gene, then transfected them into colon cancer HT29 ceils by lipofectamineTM 2000. The level of VEGF mRNA was determined by RT-PCR and Northern blotting. The protein expression of VEGF was examined by Immunolluoresence staining and Western blotting. Results ： The two kinds of VEGF specific shRNA expression were found to efficiently inhibit the expression of VEGF in HT29 ceils, the inhibition rate showed by RT-PCR, Northern blotting, Immunofluoresence staining and Western vectors blotting were 42%, 88% ,73% and 82%, respectively. Conclusion ：The expression of VEGF could be inhibited by RNA interference in HT29 ceils.

关 键 词：RNA干扰 血管内皮生长因子(VEGF) 结肠癌细胞 NORTHERN BLOTTING 

分 类 号：Q78[生物学—分子生物学]