摘要
目的探讨基于成簇规律间隔的短回文重复序列(CRJSPR)1上游侧翼序列对大肠埃希菌和志贺菌鉴定和评价效果。方法通过BLAST重复序列识别并获得全基因组测序大肠埃希菌和志贺菌的CRISPRs和CRISPR相关基因(CRISPR-associated,cas),并分析其种系;选取CRISPRs上、下游各500bp侧翼序列,使用ClustalX进行序列比对;采用PCR方法扩增CRISPR1上游侧翼序列,以确定其对大肠埃希菌和志贺菌鉴定和评价效果。结果73.4%(149/203)的大肠埃希菌存在I-E型CRISPR/Cas系统,包含了A、B1、D种系;8.4%(17/203)的大肠埃希菌存在I-F型CRISPR/Cas、17.2%(35/203)的大肠埃希菌不存在CRISPR/Cas,这2种大肠埃希菌均属于B2种系;9株志贺菌均存在I-E型CRISPR/Cas。在大肠埃希菌(B2种系外)、志贺菌CRISPR1上游和大肠埃希菌(B2种系)各存在61bp侧翼序列,序列一致性为99%,且有种属特异性,PCR扩增此区域鉴定大肠埃希菌和志贺菌的灵敏度和特异度均>91%。结论基于CRISPR1上游序列可用来鉴定大肠埃希菌和志贺菌,且具有良好的效果。
Objective To analyze the effect of the identification and evaluation of Escherichia (E.)coli and Shigella,based on the upstream flanking sequences of CRISPR1.Methods Both CRISPR and cas sequences were obtained through the BLAST with repeating sequences against the publicly complete genome in GenBank that related to E.coli and Shigella.Clustal X was used to perform multi-sequences alignment of the flanking sequences.PCR.method was used to amplify the upstream flanking sequences of CRISPR1in order to appraise the effect of identification and evaluation of upstream flanking sequences on E.coli and Shigella,which were based on the upstream flanking sequences of CRISPR1.Results The results showed that 73.4%of the strains containing the I-E CRISPR/Cas that belonged to the phylogroups A,B1,D while 8.4%strains carried the I-F CRISPR/ Cas.Another 17.2% of the strains owned CRISPR3-4(non-CRISPR/Cas)only belonged to the phylogroups B2.All the Shigella strains carried I-E CRISPR/Cas.More than 99% of similarity the CRISPR1upstream-flanking sequences was seen in E.coli (except B2)and Shigella and E.coli (B2). Both sensitivity and specificity were greater than 91%after PCR amplification in the region to identify the E.coli and Shigella.Conclusion The upstream of CRISPR1could achieve a preliminary identification effect on E.coli and Shigella.
作者
梁文娟
崔聪聪
段广才
刘慧莹
徐亚珂
郗园林
杨海燕
陈帅印
Liang Wenjuan;Cui Congcong;Duan Guangcai;Liu Huiying;Xu Yake;Xi Yuanlin;Yang Haiyan;Chen Shuaiyin(Department of Epidemiology and Statistics,School of Public Health,Xinxiang Medical University,Xinxiang 453003,China;Department of Epidemiology,College of Public Health,Zhengzhou University,Zhengzhou 450001,China;Henan Innovation Center of Molecular Diagnosis and Laboratory Medicine,Xinxiang Medical University,Xinxiang 453003,China)
出处
《中华流行病学杂志》
CAS
CSCD
北大核心
2018年第12期1607-1610,共4页
Chinese Journal of Epidemiology
基金
国家重大科技专项(2013ZX10004607)
新乡医学院分子诊断与医学检验技术河南省协同创新中心(XTCX一2015一PY4)
新乡医学院博士启动课题(XYBSKYZZ201805).
关键词
大肠埃希菌
志贺菌
成簇规律间隔的短回文重复序列
侧翼序列
鉴定
Escherichia coli
Shigella
Clustered regularly interspaced short palindromic repeats
Flanking sequence
Identification
