构建一种基于双启动子模型的特异性检测镉离子的大肠杆菌传感器  

Detection of cadmium by a double-promoters based Escherichia coli biosensor

作　　者：吕攀攀[1] 肖芳兰[1] 严锡娟[1] 卢彬彬[1] 林炜炜[1] 许清清[1] 张珍珍[1] 王伍[1] 吕建新[1] 

机构地区：[1]温州医科大学检验医学院生命科学学院浙江省模式生物技术与应用重点实验室,浙江温州325000

出　　处：《生物工程学报》2015年第11期1601-1611,共11页Chinese Journal of Biotechnology

基　　金：国家高技术研究发展计划(863计划)(No.2014AA06A514);浙江省自然科学基金(No.LY13B070012)资助~~

摘　　要：为构建一种能够特异性检测镉离子的大肠杆菌荧光报告菌株,本研究通过对检测元件模式和荧光蛋白的筛选后,以蓝色荧光蛋白mTagBFP2基因作为报告基因,使用双启动子模式,构建融合报告载体Pmer::merR-m-Pmer::mTagBFP2-pMD19-T。然后将报告载体转化至E.coli MC4100菌中,构建特异性检测镉离子的微生物传感器,并对此微生物传感器进行了最佳培养基、起始诱导浓度和时间、最适检测范围等相关参数的确定,以及对特异性进行了测定。研究表明,此微生物传感器检测镉离子背景信号低,选定起始菌液浓度为OD600=0.1,于IHMM培养基中诱导2 h为最佳检测条件,检测镉离子线性浓度范围为0.1-75μmol/L,并且只对Cd2+响应,特异性较高。因此,本研究成功构建了能够特异性检测镉离子的生物传感器,为重金属微生物传感器的优化研究工作提供了有用方案。To detect cadmium ions, we constructed a specific microbial sensor and screened detecting cassettes and different fluorescence proteins. Blue fluorescence protein mTagBFP2 was selected as a reporter and a double-promoters model was used in the construction of the fusion reporter vector Pmer：：merR-m-Pmer：：mTagBFP2-pMD19-T. The reporter vector was then transformed into Escherichia coli MC4100 wild type strain. The medium, incubation time, initial density for induction, and the optimal detection range were determined. The specificity of the biosensor was also checked. The biosensor responded specifically to cadmium irons with low background, and the linear concentration range detection ranged from 0.1 to 75 μmol/L at the initial OD600=0.1 with 2 h incubation in IHMM medium. Thus we successfully constructed a specific biosensor to detect cadmium irons and provided useful strategies for development and optimization of microbial sensors to detect heavy metals.

关 键 词：微生物传感器 镉离子检测 双启动子 蓝色荧光蛋白 

分 类 号：TP212[自动化与计算机技术—检测技术与自动化装置] Q81[自动化与计算机技术—控制科学与工程]