摘要
目的利用小片段干扰RNA(small interfering RNA,siRNA)抑制S100A6蛋白的表达,为研究CacyBP/SIP核转位机制提供有利的工具。方法设计S100A6的siRNA序列,用脂质体LipofectamineTM 2000将siRNA转染至人结肠癌SW480细胞中,在荧光显微镜下观察转染效率,用Real-time PCR及Western blot方法检测S100A6在人结肠癌SW480细胞内的mRNA和蛋白表达。结果构建了3对S100A6-siRNA,荧光显微镜下检测转染人结肠癌SW480细胞成功;RT-PCR证明了在SW480细胞内S100A6的mRNA表达显著降低(P<0.05);Western blot证明了SW480细胞内S100A6蛋白表达显著降低(P<0.05);与S100A6si-1、S100A6si-3组相比,S100A6si-2组对S100A6的mRNA和蛋白抑制效果最明显(P<0.05)。结论成功构建了S100A6-siRNA,为CacyBP/SIP核转位机制研究提供了有利的工具。
Objective To inhibit the expression of S100A6 protein with short interfering RNA（siRNA）so as to provide a useful tool for studying calcyclin（S100A6）-binding-protein（CacyBP）/SIP nuclear translocation mechanism.Methods The siRNAs of the S100A6 coding sequence were designed and transiently transfected to human colon cancer SW480 cells by Lipo-fectamine 2000.Western blot and semi-quantitative reverse transcription polymerase chain reaction（RT-PCR）analyses were used to examine the expression of S100A6 in these transfectants.Results S100A6-siRNA was constructed and transiently transfected successfully to human colon cancer SW480 cells under the fluorescence microscope.Silent expression of S100A6 at the mRNA and protein levels was confirmed by RT-PCR and Western blot,respectively（both P〈0.05）.The inhibitory effect was the most obvious in S100A6si-2group（P〈0.05）.Conclusion The available tool of the CacyBP/SIP nuclear translocation mechanism has been provided by constructing the S100A6-siRNA.
出处
《西安交通大学学报（医学版）》
CAS
CSCD
北大核心
2016年第5期754-757,761,共5页
Journal of Xi’an Jiaotong University（Medical Sciences）
基金
国家自然科学基金资助项目(No.81072040)
宁夏自然科学基金资助项目(No.NZ0897))
宁夏医科大学重点资助项目(No.XZ201413)~~
