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Silencing efficiency of Hsp90α gene expression mediated by RNA interference in HEK293 cell line detected by real-time fluorescence quantitative PCR

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Author:
No author available
Journal Title:
JOURNAL OF THE FOURTH MILITARY MEDICAL UNIVERSITY
Issue:
24
DOI:
10.3321/j.issn:1000-2790.2008.24.005
Key Word:
热休克蛋白90α;RNA干扰;逆转录聚合酶链反应

Abstract: 目的:采用实时荧光定量PCR方法观察Hsp90αRNAi的沉默效率.方法:通过T-A克隆构建扩增Hsp90α,Hsp90β,GAPDH的标准品,建立实时荧光定量PCR方法;采用脂质体转染法将Hsp90αRNAi瞬时转染至HEK293细胞24 h后,检测Hsp90α,Hsp90β-mRNA水平的变化.结果:建立的实时荧光定量PCR标准曲线相关系数大于0.99,PCR扩增效率在95%以上.与对照组相比较.Hsp90αRNAi可使HEK293细胞中的Hsp90α mRNA表达量降低61%,而Hsp90β表达未受影响.结论:成功建立检测Hsp90α,Hsp90β实时荧光定量PCR方法,且Hsp90αRNAi具有高效特异的沉默效率,为下一步研究Hsp90α在创伤应激中的作用奠定基础.

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