特异性siRNA质粒的构建及其抑制癌细胞中mcl-1表达的研究  

The Specific siRNA Plasmids Construction and their Inhibition Effect on Expression of mcl-1 Protein in Liver Cancer Cell

作　　者：徐爱群[1] 曹以诚[1] 区镜深[1] 张珍武[1] 

机构地区：[1]华南理工大学生物科学与工程学院,广东广州510006

出　　处：《现代食品科技》2010年第8期789-792,783,共5页Modern Food Science and Technology

基　　金：国家自然科学基金重大研究项目(90412015);教育部中国网格计划生物信息网格平台子项目(B12137040130);国家科技基础条件平台项目(2005DKA64001);科技部基础学科平台建设生物计算网络平台项目(2005DKA64001);广州市科技攻关计划(2005Z12E4023)资助项目

摘　　要：本文构建了靶向mcl-l基因构建的特异性siRNA重组质粒,在细胞水平检测的其抑制效果,并筛选出能高效抑制mcl-l基因表达的siRNA重组质粒,针对mcl-1的特异性siRNA重组质粒,将其转染到人肝癌细胞HepG2,用Rt-PCR(实时荧光定量PCR)方法和WesternBlot方法分别检测转染前后mcl-1mRNA水平和Mcl-1蛋白表达水平,比较对应不同位点的两个siRNA重组质粒对mcl-1的抑制效果。结果表明,Rt-PCR结果显示对应不同位点的两个siRNA重组质粒对mcl-1均可降低mcl-1mRNA水平,最大抑制效率达70.0%,远高于作为对照非相关组;Western-Blot结果显示转染特异性siRNA重组质粒后Mcl-1蛋白表达受到抑制,最大抑制率为44.2%。合理设计的特异性siRNA重组质粒可大幅度下调mcl-1mRNA水平,能有效抑制Mcl-1蛋白表达。The mcl-l gene-specific siRNA plasmids were constructed and transfected into HepG2.The levels of mcl-1 mRNA and Mcl-1 protein expression were detected with the Rt-PCR（real time PCR） and WesternBlot methods,respectively.And the inhibitory effects of siRNA plasmid corresponding to two different sites on mcl-1 expression were compared.Rt-PCR results showed that both the two siRNA plasmids correspomding to two sites could reduce the levels of mcl-1 mRNA and the maximum inhibition efficiency was of 70.0%,which was much higher than that of a control group.Western-Blot results were showed that transfection of mcl-l gene-specific siRNA plasmids into HepG2.The expression of Mcl-1 protein obviously inhibit the expression of mcl-l protein with the maximum inhibition rate was 44.2%.It was concluded that the rational design of specific siRNA plasmid can significantly lower levels of mcl-1mRNA and inhibit the expression of Mcl-1 protein.

关 键 词：肝癌 MCL-1 基因沉默 实时荧光定量PCR 

分 类 号：R730.2[医药卫生—肿瘤]