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  

Expression of RegⅣ mRNA in colorectal cancer cells and the transfection of constructed recombinant plasmid with pcDNA-RegⅣ

作　　者：高峰[1] 郑声琴[1] 张宇伟[1] 金月玲[1] 田小强[1] 黄培林[1] 

机构地区：[1]东南大学基础医学院病理学与病理生理学系,江苏南京210009

出　　处：《东南大学学报（医学版）》2006年第3期147-150,共4页Journal of Southeast University(Medical Science Edition)

基　　金：国家自然科学基金资助项目(30400534);江苏省青年科技创新人才项目(BK2004407)

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 pcDNA-Regiv 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 pcDNA-Reg Ⅳ 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 coiorectal cancer cells selected by G418 were identified by RT-PCR. Results There was a high expression of RegⅣ in HT-29 and Caco-2, 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.

关 键 词：RegⅣ基因 结直肠癌 重组质粒 基因表达 基因转染 

分 类 号：Q786[生物学—分子生物学] Q785