RNA干扰技术建立人PARP-1缺陷细胞株  

Construction of hPARP1-deficient cell strain by RNA interference

作　　者：沙焱[1] 曾玉云 庄志雄[3] 何云[1] 胡大林[1] 胡恭华[1] 杨建平[1] 涂晓志[1] 

机构地区：[1]中山大学公共卫生学院,广州510089 [2]深圳沙井卫生监督所 [3]深圳市疾病预防控制中心

出　　处：《卫生研究》2005年第6期650-652,共3页Journal of Hygiene Research

基　　金："973"国家重点基础研究发展计划基金资助项目(No.2002CB512904;2002CB512903);国家自然科学基金项目(No.30571592)

摘　　要：目的建立并鉴定聚ADP核糖聚合酶1(hPARP1)缺陷细胞株,用于研究hPARP1基因的作用机制及其缺陷与DNA损伤发生的关系。方法将用已经构建的pEGFPC1shRNA(包含两个PARP1基因及一个阴性对照)转染支气管上皮细胞(16HBE),使之在16HBE中表达,转染后命名为16HBEP1、16HBEP2及16HBEN。用蛋白免疫印迹法鉴定转染细胞中hPARP1基因的表达水平。结果pEGFPC1shRNA在真核细胞成功表达;16HBEP1和16HBEP2的蛋白表达水平分别下降了84.3%及63.7%。结论hPARP1缺陷细胞株的成功建立和鉴定为hPARP1基因功能研究提供了一种有效手段。Objective To construct hPARPl-deficient cell strain which can be used in studying the functions and action mechanisms of hPARP1 gene, Methods 16-HBEs were transfected with two kinds of eukaryotic expression plasmids of hPARP1 gene short hairpin RNA to construct hPARPl-deficient cells （named as＂16-HBEP1 and 16-HBEP2＂） and a negative control plasmids （named as 16-HBEN）. The protein expression levels of hPARP1 gene in 16-HBE, 16- HBEP1, 16-HBEP2 and 16-HBEN were detected by the western blotting to estimate the effects of RNA interference. Results The construction of hPARPl-deficient cells train was successful and the protein expression level of hPARP1 gene in the 16-HBEP1 and 16-HBEP2 were decreased 84,3% and 63.7% respectively as compared with that in 16-HBE. Conclusion The successes in constructing hPARPl-deficient strain offered an effective cell strain for studying the function of hPARP1 gene.

关 键 词：RNA干扰 短发夹-RNA hPARP1 绿色荧光蛋白栽体 真核细胞转染 

分 类 号：Q522[生物学—生物化学] Q343