PRL-3基因的克隆及其原核表达载体的构建表达  

Cloning of human PRL-3 gene and construction of its prokaryotic expression vector

作　　者：周军[1] 李建明[1] 柳玉红[1] 丁彦青[1] 

机构地区：[1]南方医科大学南方医院病理科/南方医科大学基础医学院病理学教研室/教育部广东省共建人类重大疾病转录组学和蛋白组学重点实验室/广东省分子肿瘤病理重点实验室,广东广州510515

出　　处：《南方医科大学学报》2007年第5期641-643,共3页Journal of Southern Medical University

基　　金：国家自然科学基金(30500241;30670968)~~

摘　　要：目的克隆人PRL-3基因并构建其原核表达载体。方法设计PRL-3特异性引物,提取人大肠癌细胞系SW480总RNA,用RT-PCR方法获取人PRL-3cDNA。经T-A克隆,通过双酶切鉴定及测序确认后,构建人PRL-3基因的原核表达载体pGEX-4T-1-PRL-3。结果成功扩增出人PRL-3全长cDNA,并构建pGEX-4T-1-PRL-3原核表达载体,测序结果表明PRL-3全长cDNA与GenBank中PRL-3序列完全一致,SDS-PAGE胶分析表明,在大肠杆菌BL21(DE3)中成功表达了重组蛋白。结论获得人PRL-3基因全长cDNA并构建其原核表达载体,使其在大肠杆菌中获得了高效表达,为深入研究PRL-3基因在大肠癌发生发展中的作用奠定了基础。Objective To obtain the entire coding sequence of human PRL-3 gene and construct its prokaryotic expression vector. Methods With total RNA extracted from the human colorectal carcinoma cell line SW480 as the template, PRL-3 gene was amplified by RT-PCR with primers designed according to the published sequence of GenBank, and the product was inserted into pGEM-T Easy vector. The recombinant plasmid pGEM-T-PRL-3 was identified by restriction endonuclease analysis and DNA sequencing. After digestion with restriction endonuclease, PRL-3 gene was cloned into the multicloning sites of the prokaryotic expression vector pGEX-4T-1. Results and conclusion The entire coding region of human PRL-3 gene was cloned, and the recombinant pGEX-4T-1-PRL-3 vector was successfully constructed and expressed, which may provide the basis for further study of the relationship between human colorectal carcinoma and PRL-3 gene.

关 键 词：大肠癌 PRL-3 基因克隆 原核表达 

分 类 号：R346[医药卫生—基础医学]