腺病毒载体介导的GFP基因在鸭胚成纤维细胞中的表达及RNA干扰  

Expression of GFP Gene Mediated by Adenovirus Vector and RNA Interference in Duck Embryo Fibroblast

作　　者：孟宇航[1] 汤承[1] 杨发龙[1] 邓书[1] 岳华[1] 于学辉[1] 

机构地区：[1]西南民族大学生命科学与技术学院,四川成都610041

出　　处：《生物技术通讯》2008年第3期404-406,共3页Letters in Biotechnology

基　　金：四川省应用基础项目(05JY029-007-5)

摘　　要：目的:建立在鸭胚成纤维细胞(DEF)中进行RNA干扰(RNAi)的技术平台,为鸭基因组功能的研究提供新的技术手段。方法:以绿色荧光蛋白(GFP)基因为报告基因,脂质体转染化学合成的GFP特异小干扰RNA(GFP-siRNA),用流式细胞仪测定GFP-siRNA对重组腺病毒(Adv-GFP)介导的GFP基因在DEF中表达的干扰效果。结果:200MOI(感染复数)Adv-GFP介导的GFP基因在DEF中表达效率最高,为31.20%±3.11%,对DEF的活力无明显影响;GFP-siRNA能有效干扰GFP基因在DEF中的表达,相对抑制率为98.56%。结论:在DEF中进行RNAi是可行的,Adv-GFP是介导外源基因在DEF中表达较为理想的载体;首次建立了在DEF中进行RNAi的技术平台,为鸭基因组的功能等研究提供了新的技术手段。Objective： To establish a methodological platform for RNA interference（RNAi） studies in duck embryo fibroblast（DEF）, so as to provide a novel way for the studies of genome functions of ducks. Methods： Green fluorescent protein （GFP） gene was used as report gene and DEF was transfected by synthesized anti-GFP small interfering RNA（siRNA） （GFP-siRNA） using the method of cationic liposome reagent. Interference effect of GFP-siRNA to GFP gene expression that was mediated by adenovirus（Adv-GFP） in DEF was evaluated by flow cytometer method. Results 200 multiplicity of infection（MOI） Adv-GFP could get highest expression rates of GFP gene（31.20%±3.11%）, and wouldn＇t influence cell activity of DEF. GFP-siRNA could significantly interfere with the expression of GFP gene in the DEF, and the relative inhibitory rate was 98.56%. Conclusion： The results showed that it is feasible to carry out RNAi in DEF and Adv-GFP is an ideal vector for exogenous gene expression in DEF. The present study established a methodological platform for RNAi studies in DEF for the first time, and provided a novel way for the studies of genome functions of ducks.

关 键 词：鸭胚成纤维细胞 RNA干扰 腺病毒载体 绿色荧光蛋白 鸭 

分 类 号：Q78[生物学—分子生物学] R394[医药卫生—医学遗传学]