来源于放线多孢菌的CRISPR/Cas系统的分析及功能鉴定

项目名称： 来源于放线多孢菌的CRISPR/Cas系统的分析及功能鉴定

项目编号： No.31500009

项目类型： 青年科学基金项目

立项/批准年度： 2016

项目学科： 生物科学

项目作者： 罗晓霞

作者单位： 塔里木大学

项目金额： 20万元

中文摘要： CRISPR/Cas系统对DNA分子具有靶向切割活性，可以被用于定向的基因修饰，它在细菌、古菌中广泛存在，但在放线菌中还未见报道。对来自罗布泊的新物种新疆放线多孢菌TRM40136进行全基因组测序分析表明，其中蕴藏着13个CRISPR/Cas 位点，因此研究放线多孢菌中的CRISPR/Cas具有首创性。本项目以TRM40136菌株为研究材料，对放线菌CRISPR/Cas进行多样性及系统进化分析，揭示放线多孢菌中CRISPRs的特异性。通过分子生物学手段在大肠杆菌中构建来自于放线多孢菌的CRISPR/Cas系统，以放线菌中广泛存在的抗生素合成基因的负调控基因TetR为靶标，筛选能够切割TetR基因的CRISPR/Cas系统，这一工作的实施首次建立来自于放线菌的CRISPR/ Cas基因修饰系统，为构建放线菌基因簇的激活模型奠定理论基础。

中文关键词： 放线多孢菌；；基因结构和功能；CRISPR/Cas系统；分析和鉴定

英文摘要： Due to CRISPR/Cas system possess the characters of targeted cutting DNA molecules, it is used for targeted genetic modification. The system exists in Bacteria and Archaea widely, but it has not been reported in Actinomycetes. Actinopolyspora xinjiangensis TRM4013 was isolated from lop nur, it is a novel species. After analysis of the whole genome sequence, the results showed that it has 13 CRISPR/Cas sites, it has innovation to research the CRISPR/Cas system from Actinopolyspora xinjiangensis. In this project we will use the TRM40136 strains as the research material, research the diversity and phylogenetic analysis of the Actinomycetes CRISPR/Cas, reveals the specificity of CRISPRs in Actinomycetes. By means of molecular biology, we will construct the CRISPR/Cas system from Actinomycetes in E. coli, then apply the TetR genes to identify the function of CRISPR/Cas system, TetR gene is the negative regulation factor of Actinomycetes. Then screen the CRISPR/Cas system, which can cut TetR nucleic acid. It is the first report of CRISPR/Cas system in Actinomycetes to implement this project. This research provides the basic data of the evolution and function mechanism about the Actinomycetes CRISPRs. It established the theoretical base for building activation model of the gene cluster in Actinomycets.

英文关键词： Actinopolyspora;Gene structure and function;CRISPR/Cas system ;Analysis and identification