检测汞的全细胞生物传感器构建  

The Construction of Whole-Cell Biosensors for Monitoring Mercury

作　　者：杨锋 齐显尼[2] 王钦宏[2] 张雪洪[3] 

机构地区：[1]北京维德丰贸易有限公司,北京100026 [2]中国科学院天津工业生物技术研究所,天津300308 [3]上海交通大学生命科学技术学院,上海200240

出　　处：《科技创新导报》2014年第24期205-209,共5页Science and Technology Innovation Herald

摘　　要：录(Hg)是环境中主要的重金属污染物,暴露于环境中的汞对人体健康有严重危害。汞的精确检测对控制环境污染和减少对健康的危害有着重要意义。该实验是基于MerR蛋白调节的启动子对应答汞毒性基因的调控机理,首先通过试验确定红色荧光蛋白mCherry为报告基因。将MerR基因片段链接到质粒pUC57得到质粒pUC57-MerR,以其为模版得到质粒pUCRR,转入到E.coli DH5α中,得到工程菌株E.coli UCRR。为提高菌株的性能和灵敏度,通过易错PCR技术获得MerR基因突变体库,筛选出菌株V109E在培养4 h时,荧光信号强度远远高于其他菌株荧光信号,对Hg^(2+)有高度专一性,检测Hg^(2+)的最低浓度为0.5nM。Mercury has been well known as an environmental pollutant for several decades. Emissions of mercury to the environment could have serious effects on human health. To control mercury pollution and reduce mercury damage to human health, sensitive determination of mercury is important. Our biosensor design for mercury detection was based on MerR whice is an activator that controls genes involved in the response to mercury poisoning. we selected mCheery out for fluorescent detection signal ,Then cloned MerR into pUC57 to obtain pUC57-MerR, and utilized pUC57-MerR as a template to obtain the recombinant plasmid pUCRR. This plasmid was transformed into E. coli DH5α in order to obtain engineered strain E. coli UCRR. We established a library of mutated MerR gene by Error-prone PCR in order to improve MerR protein derived biosensors to detect and monitor mercury more efficiently and with higher sensitivity. The mutant MerR genes V109E which have improved functionalities been selected. V109E had highest sensitivity increasing than other mutated mercury biosensors after 4-hour incubation time.The whole-cell biosensors based on V109E had high sensitivity and Specificity, the lowest concentration of mercury detection had reached to 0.5nM.

关 键 词：汞检测 全细胞生物传感器 MerR蛋白 易错PCR 

分 类 号：R3[医药卫生—基础医学]