基于土壤细菌宏基因组克隆新型群体感应信号降解酶基因

项目名称： 基于土壤细菌宏基因组克隆新型群体感应信号降解酶基因

项目编号： No.31272082

项目类型： 面上项目

立项/批准年度： 2013

项目学科： 农业科学

项目作者： 张力群

作者单位： 中国农业大学

项目金额： 80万元

中文摘要： 群体感应（QS）是细菌的一种调控机制，即通过分泌和感应特定的信号分子来感知细菌自身群体密度，调控特定基因表达以适应环境变化。许多植物病原细菌利用以N-乙酰高丝氨酸内酯（AHL）为信号的QS系统调控致病性的表达。破坏AHL信号分子，即可造成细菌致病性丧失或减弱。已知的AHL信号降解酶基因多克隆自有降解AHL信号能力的可培养细菌，其中包括本课题组前期研究中克隆的源自苍白杆菌的新型AHL内酯酶基因aidH。在此基础上，本项目以土壤中不可培养细菌为主要DNA资源，构建细菌宏基因组文库，以染色体整合pcoI基因的大肠杆菌为宿主，建立高通量的AHL降解酶基因筛选体系，克隆获得新型AHL降解酶基因。对纯化的降解酶蛋白进行AHL降解机制的生化分析，并检测其对病原细菌致病力和生防细菌防病能力的影响。基于土壤宏基因组大量分离AHL降解酶基因资源，有望发现新的AHL降解机制及阐明AHL降解酶基因的系统发育关系。

中文关键词： 群体感应淬灭；宏基因组；高通量；AidL；AidE

英文摘要： Quorum sensing (QS) is a regulatory system employed by bacteria to monitor their own population density by means of small signals and to coordinate the expression of specialized genes in respond to the environmental changes. Many phytopathogenic bacteria use the N-acyl homoserine lacton(AHL)-dependent QS system to regulate their virulence expression. Inactivation of the AHL molecules leads to a reduction of the bacterial virulence. Most of the characterized AHL degradase genes so far, are cloned from culturable bacterial strains, including a novel type of AHL lactonase gene aidH cloned from Ochrobacterum sp. by our research team. The present project, based on our previous quorum-quenching study, focuses on cloning and characterization of novel AHL degradase genes from soil metagenomic library, which is constructed by using genomic DNA from unculturable bacteria. A high throughput system for metagenomic library screening will be established based on a Escherichia coli host strain containing chromosome-integrated AHL biosynthetic gene pcoI. The cloned AHL degradase genes will be oeverexpressed in E. coli, and the action model of the recombiant proteins will be further characterized. Genes encoding novel metagenome-derived AHL degradases will be selected to test their quorum quenching functions in phytopathogenic

英文关键词： quoruming quenching；metagenome；high throughput；AidL；AidE