应用载体介导的RNAi技术抑制Bcl-2的表达  

RNAi-mediated gene silencing of Bcl-2 expression

作　　者：蹇锐[1] 程小星[1] 安静[1] 陈炜[1] 王嘉丽[1] 张俊磊[1] 万颖杰[1] 陈宗涛[1] 

机构地区：[1]第三军医大学基础医学部微生物学教研室

出　　处：《第三军医大学学报》2004年第4期294-297,共4页Journal of Third Military Medical University

基　　金：国家自然科学基金资助项目 ( 30 370 6 0 3)~~

摘　　要：目的　应用载体介导的RNAi技术特异地干扰抗凋亡基因bcl 2在HeLa细胞中的表达。方法　构建利用H1启动子转录功能性siRNA的载体 ;在H1启动子下游分别插入含不同bcl 2基因特异性序列的 64nt寡核苷酸片段 ;将构建的质粒转染HeLa细胞 ,以间接免疫荧光和免疫印迹检测转染后细胞中Bcl 2的表达水平。结果　 3种含不同bcl 2特异性序列的质粒转染后均能干扰该基因在细胞中的表达 ,但干扰的效果存在差异 ,从 45 %～ 83 5 %。结论　本研究建立的载体介导的RNAi技术可成功地用于干扰BclObjective To silence the expression of antiapoptotic gene bcl 2 in HeLa cells using vector based RNA interference (RNAi) technique. Methods A vector used to transcribe functional short interfering RNA (siRNA) was constructed, and 64 nt oligonucleotide fragment was inserted into the downstream of the H1 promoter. Plasmids containing different bcl 2 target sequences were transfected into HeLa cells, and then the expression level of Bcl 2 was detected by indirect immunofluorescence assay (IFA) and Western blotting. Results Vector based RNAi had advantages over antisense RNA and synthesized siRNA because it could be delivered to the target cell more efficiently, and its effect could last longer. bcl 2 expression could be inhibited by plasmid expressed siRNA. Three different targeting sequences were selected from bcl 2 gene, and the inhibitory effect ranged from 45% to 83 5%. Conclusion Bcl 2 expression in HeLa cells can be inhibited significantly using plasmid based RNAi.

关 键 词：载体介导 RNAI BCL-2 表达 

分 类 号：Q782[生物学—分子生物学] Q786