TEM-1型β-内酰胺酶编码基因的克隆表达  

Cloning and expression of TEM-1 gene

作　　者：张翔[1] 刘刚[1] 赵廷坤[1] 袁斌[1] 凌保东[1] 雷军[1] 

机构地区：[1]川北医学院药物研究所,南充637007

出　　处：《四川生理科学杂志》2005年第2期52-53,共2页Sichuan Journal of Physiological Sciences

基　　金：四川省重点科技攻关项目(编号02SG022-030)

摘　　要：目的:构建TEM-l型β-内酰胺酶基因的表达载体。方法:通过PCR扩增TEM-l全编码基因,将其连接入pBK-CMV载体中,并在大肠杆菌JM109中表达。采用琼脂二倍稀释法对TEM-l克隆菌株进行MIC检测,双纸片法筛选及确证实验检测其表型,等电聚焦(IEF)电泳测定其等电点(pIs)。结果:经酶切测序鉴定TEM-l基因的表达载体构建正确。重组菌产pI为5.4的TEM-l广谱酶,仅对青霉素类耐药。结论:TEM-l基因原核表达载体正确构建,为进一步研究打下了基础。Objective： To clone TEM-1 gene with full length open reading frame for a recombinant prokaryotic expression vector. Methods： TEM-1 gene was amplified by Polymerase chain reaction（PCR）, The target amplification fragment was cloned into pBK-CMV vector. The recombinant vector was sequenced by Sanger＇ s dideoxy chain termination composition method. Antimicrobial susceptibility of the clone strain was detected by the disc diffusion method and by the agar dilution method. Crude β-1actamases were extracted and then isoelectric point of TEM-1 was determinded by IEF. Results： Sequence and restriction anallysis revealed TEM-1 gene was cloned in frame into pBK-CMV. The clone strain producing TEM-1 with pl value of 5.4 was resistant to ampicillin and piperacillin, only. Conclusions： TEM-1 gene has been cloned and expressed, to prepare for the site-directed mutagenesis study.

关 键 词：Β内酰胺酶 TEM-1 基因 克隆 表达 TEM-1型 Β-内酰胺酶 克隆表达 编码基因 表达载体构建 

分 类 号：R346[医药卫生—基础医学]