RegⅣ基因在结直肠癌细胞系中的mRNA表达及pcDNA-RegⅣ的构建与转染-东南大学学报医学版

RegⅣ基因在结直肠癌细胞系中的mRNA表达及pcDNA-RegⅣ的构建与转染

单位：东南大学基础医学院,病理学与病理生理学系,江苏,南京,210009

分类号：Q786, Q785

出版年·卷·期（页码）：2006·25·第三期（147-150）

摘要：

目的:探讨RegⅣ基因在4种结直肠癌细胞系中的表达水平,构建并转染pcDNA-RegⅣ重组质粒,为阐明RegⅣ基因在结直肠癌发生发展中的作用机制奠定基础.方法:采用RT-PCR方法检测HT-29、Caco-2、Sw480、LoVo细胞系的RegⅣ基因表达水平,将获得目的基因RegⅣ克隆于pcDNA3.1/(-)质粒上,用PCR、酶切进行鉴定后,脂质体转染至低表达甚至不表达RegⅣ的结直癌细胞系,用G418筛选后进行RT-PCR鉴定.结果:HT-29、Caco-2细胞系RegⅣ基因高表达,Sw480细胞系表达较前两者弱,LoVo细胞系RegⅣ阴性表达;构建的重组质粒中含有RegⅣ基因,经pcDNA-RegⅣ转染的LoVo细胞RegⅣ基因呈阳性表达.结论:RegⅣ基因在不同的结直肠癌细胞系中表达水平不同.成功构建和转染了pcDNA-RegⅣ,可使得RegⅣ(-)的LoVo细胞系RegⅣ基因呈阳性表达.

Objective To investigate the expression of RegⅣ in four colorectal cancer cells and construct recombinant plasmid pcDNARegⅣ and transfect it for further study of RegⅣ in colorectal cancer cells.Methods RegⅣ obtained from colorectal cancer cells by RT-PCR was cloned into pcDNA3.1/(-) to form pcDNARegⅣ which was transfected into colorectal cancer cell with less or no expression of RegⅣ via liposome after identification of PCR,restriction enzyme digesting and DNA sequencing.The colorectal cancer cells selected by G418 were identified by RT-PCR.Results There was a high expression of RegⅣ in HT-29 and Caco2,less in Sw480,no expression in LoVo.The recombinant plasmid had RegⅣ gene which was expressed in RegⅣ-regative LoVo after transfection.Conclusions The different expression levels of RegⅣ in four colorectal cancer cells are confirmed and the pcDNA-RegⅣ has been constructed and transfected successfully,which makes RegⅣ highly expressed in RegⅣ-regenerating LoVo cell.

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