大肠杆菌多重耐药调控基因marA的克隆及其原核表达  

Cloning and Prokaryotic Expression of Multi-Drug Resistant Regulator marA Gene of Escherichia coli

作　　者：李春雨[1] 张春艳 丛薇[3] 刘玉堂[3] 马颖[3] 马红霞[3] 

机构地区：[1]新疆农业职业技术学院,昌吉831100 [2]张家产畜牧兽医工作站,文登264407 [3]吉林农业大学动物科学技术学院,长春130118

出　　处：《吉林农业大学学报》2009年第4期452-455,共4页Journal of Jilin Agricultural University

基　　金：国家自然科学基金项目(30400326);吉林省科技发展计划项目(20010538;20030425;20050124);吉林省教育厅项目(2005042)

摘　　要：根据GenBank中大肠杆菌的marA基因序列设计引物,以大肠杆菌ATCC25922的基因组为模板,PCR扩增出约384 bp的marA基因片段,将所得片段与pMD18-T simple vector连接,转化至JM109大肠杆菌中,筛选阳性克隆,其质粒中插入序列测序结果与GenBank中报道一致。从阳性克隆中提取质粒,经BamHⅠ和XhoⅠ酶解,回收384 bp目的片段,定向克隆到pET-28a表达载体中,提取质粒,转化大肠杆菌BL21(DE3)中,获得阳性克隆。经IPTG诱导阳性菌,收集表达产物,通过SDS-PAGE分析证实marA基因得到表达。A construction pMD18-T-marA was generated by inserting the sequence of 384 bp obtained by PCR into pMD18-T simple vector and selecting the sense clones. The result showed that the cloned sequence coincides with the designed sequence by sequencing. This construction was digested with the same enzyme （BamH I and Xho I ） and ligated the pET-28a vector. Then the plasmid pET-28a-marA was transformed to the competent cell of BL21（DE3）. The sense clone was induced with IPTG. The expression of marA was observed on SDS - PAGE.

关 键 词：大肠杆菌 marA基因 克隆 原核表达 

分 类 号：Q785[生物学—分子生物学]