靶向FBXL20基因的siRNA片段的筛选及其验证  

Screening and Verification of the siRNA Targeted for the FBXL20 Gene

作　　者：朱剑军[1] 蒋琴[1] 袁粒星[2] 陈尧[1] 

机构地区：[1]四川大学基础医学与法医学院人体解剖学教研室,成都610041 [2]四川大学华西附属第二医院公共实验室,成都610041

出　　处：《中国生物化学与分子生物学报》2011年第3期237-243,共7页Chinese Journal of Biochemistry and Molecular Biology

基　　金：四川大学口腔疾病国家重点实验室自主研究课题资助(No.SKLODSCU20090021)~~

摘　　要：在人结肠腺癌SW480细胞株及口腔鳞癌HSC3细胞株中,筛选有效干扰fbxl 20的siRNA片段.设计合成3对靶向fbxl 20 mRNA的小干扰RNA片段,通过阳离子脂质体介导转染SW480细胞和HSC3细胞.结果发现siRNA-1,siRNA-2,siRNA-3均能抑制SW480细胞和HSC3细胞fbxl20 mRNA的表达,siRNA-2抑制作用最明显,抑制率分别为86.8%和100%.MTT试验显示:siRNA-2转染SW480细胞和HSC3细胞后,细胞增殖能力明显降低,细胞增殖抑制率分别为29.5%和43.4%.Transwell小室迁移实验表明,HSC3细胞在转染24 h后,细胞迁移能力明显减弱,迁移抑制率高达72.4%.流式细胞术检测细胞凋亡结果显示:转染36 h后,SW480细胞实验组细胞凋亡率为21.9%;转染24 h后,HSC3细胞实验组细胞凋亡率为44.7%.本实验成功的筛选出了特异性的抑制fbxl 20基因的siRNA片段,并初步鉴定了fbxl 20基因的功能.To obtain effective siRNA fragement of fbxl 20 in the SW480 and HSC3 cell lines.Three pairs of small interfering RNA targeting to the fbxl 20 mRNA were designed and introduced into SW480 human colorectal adenocarcinoma and HSC3 oral squamous carcinoma cells in cationic liposomes.The mRNA expression of the fbxl 20 were detected by semi-quantitative RT-PCR,the cell proliferation and migration were assayed by MTT and transwell chamber,and the apoptosis was measured by flow-cytometry.The mRNA expression of the fbxl 20 was inhibited by the three siRNA in both cell lines,among which siRNA2 gave the strongest inhibiting of fbxl 20 by 86.8% and 100% in SW480 and HSC3,respectively.The inhibition of cell proliferation were 29.5% in SW480 and 43.4% in HSC3.The migration was inhibited by 72.4% in HSC3.The apoptosis rates were 21.9% in SW480 and 44.7% in HSC3.We successfully validated a siRNA pair to specifically degrade fbxl 20 mRNA that could be used to study the function of fbxl 20 gene.

关 键 词：fbxl20 RNA干扰 SW480细胞 HSC3细胞 

分 类 号：Q51[生物学—生物化学] Q52