志贺菌成簇的规律间隔短回文重复序列的检测及同源性分析

doi:10.13481/j.1671-587x.20150211

吉林大学学报(医学版) ›› 2015, Vol. 41 ›› Issue (02): 261-268.doi: 10.13481/j.1671-587x.20150211

志贺菌成簇的规律间隔短回文重复序列的检测及同源性分析

王鹏飞¹, 王颖芳², 段广才^1,3, 王琳琳¹, 郭向娇¹, 薛泽润¹, 郗园林¹, 杨海燕¹

1. 郑州大学公共卫生学院流行病学教研室, 河南郑州 450001;
2. 河南科技大学医学院公共卫生教研室, 河南洛阳 471003;
3. 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, 河南新乡 453003

收稿日期:2014-07-22 出版日期:2015-03-28 发布日期:2015-04-04
通讯作者: 段广才, 教授, 博士研究生导师(Tel:0371-67781964, E-mail:gcduan@zzu.edu.cn) E-mail:gcduan@zzu.edu.cn
作者简介:王鹏飞(1989-), 男, 河南省洛阳市人, 在读医学硕士, 主要从事分子流行病学方面的研究。
基金资助:
国家科技重大专项基金资助课题(2013ZX10004607)

Detection and homology analysis of clustered regularly interspaced short palindromic repeats in Shigella

WANG Pengfei¹, WANG Yingfang², DUAN Guangcai^1,3, WANG Linlin¹, GUO Xiangjiao¹, XUE Zerun¹, XI Yuanlin¹, YANG Haiyan¹

1. Department of Epidemiology, College of Public Health, Zhengzhou University, Zhengzhou 450001, China;
2. Department of Public Health, College of Medical Sciences, Henan University of Science and Technology, Luoyang 471003, China;
3. Henan Innovation Center of Molecular Diagnosis and Laboratory Medicine, Xinxiang Medical University, Xinxiang 453003, China

Received:2014-07-22 Online:2015-03-28 Published:2015-04-04

摘要/Abstract

摘要：

目的:检测志贺菌成簇的规律间隔短回文重复序列(CRISPR),并与GenBank全基因组中志贺菌重复序列和间隔序列对比,分析其结构特征及同源性。方法:利用PCR法扩增获得志贺菌CRISPR,采用生物信息学方法对重复序列及间隔序列进行同源性分析;运用多序列比对分析间隔序列特点及其与侧翼序列间的联系;BLAST分析间隔序列与质粒和噬菌体的同源性;weblogo分析重复序列碱基频率及RNAfold预测其RNA二级结构;重复序列的同源聚类分析并用BLAST分析重复序列与其他细菌的同源性。结果:被测3株临床志贺菌及9株全基因组志贺菌中均含有不同数量CRISPR;同一个CRISPR中,间隔序列有的相似、有的完全不同,某些CRISPR间隔序列的部分序列与CRISPR侧翼序列存在同源性,某些间隔序列可能是信息基因和操纵基因的拼接重组;重复序列保守性不一致,可能与细菌进化存在一定的联系,重复序列碱基的差异影响茎的长度,从而影响二级结构的稳定性及CRISPR的功能。重复序列与个别远缘菌种具有较高的同源性。结论:志贺菌存在多样的CRISPR结构,不同CRISPR的重复序列保守性不同。某些间隔序列部分与CRISPR侧翼序列同源,某些间隔序列是多基因拼接而成。

关键词: 志贺菌, 成簇的规律间隔短回文重复序列, 重复序列, 间隔序列, 同源性

Abstract:

Objective To detect the clustered regularly interspaced short palindromic repeats (CRISPR) of Shigella,and to analyze the characteristics and homology of their repeats and spacers with complete genome Shigella in GenBank.Methods The CRISPR were obtained by PCR amplification method,and the homology of repeats and spacers was analyzed by bioinformatics;multiple sequence alignment was used to analyze the features of spacers and the relationship between the spacers and flanking sequences;BLAST was used to analyze the homology between the spacers and plasmids and phages;weblogo was used to analyze the frequency of the bases of the repeats and RNAfold was used to predict the secondary structures of the repeats RNA;homology clustering analysis of repeats was performed and BLAST was used to analyze the homology of the repeats with other bacteria.Results All 3 studied clinical strains and 9 complete genome Shigella contained different number of CRISPR.In the same CRISPR,the spacers were similar or different;the partial sequences of the spacers in some CRISPR and flanking sequences of CRISPR had the same sequences.Some spacers possible came from the splicing and gene recombination between the informational genes and operational genes.There may be relationship between the repeats which were not completely conserved and bacterial evolution.The differences of the bases of the repeats affected the length of stem,thus affected the stability of the secondary structure and the function of CRISPR.The repeats had high similarity with the several individual species of distant relationships.Conclusion The different CRISPR structures exist in Shigella,and the conservation of the repeats in different CRISPR is different.The partial sequences of some spacers have homology with the flanking sequences of CRISPR,and some spacers come from the gene splicing.

Key words: Shigella, clustered regularly interspaced short palindromic repeats, repeats, spacers, homology

中图分类号:

王鹏飞, 王颖芳, 段广才, 王琳琳, 郭向娇, 薛泽润, 郗园林, 杨海燕. 志贺菌成簇的规律间隔短回文重复序列的检测及同源性分析[J]. 吉林大学学报(医学版), 2015, 41(02): 261-268.

WANG Pengfei, WANG Yingfang, DUAN Guangcai, WANG Linlin, GUO Xiangjiao, XUE Zerun, XI Yuanlin, YANG Haiyan. Detection and homology analysis of clustered regularly interspaced short palindromic repeats in Shigella[J]. Journal of Jilin University Medicine Edition, 2015, 41(02): 261-268.

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志贺菌成簇的规律间隔短回文重复序列的检测及同源性分析

Detection and homology analysis of clustered regularly interspaced short palindromic repeats in Shigella

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