PCR-HRM方法快速鉴定布鲁氏菌的初步应用  

Clinical application of PCR and high resolution melting analysis for rapid identification of Brucellaisolates

作　　者：梁雪妮[1] 崔步云[2] 毛玲玲[3] 任微[4] 于静波[4] 薛文成[4] 孟冬娅[4] 

机构地区：[1]辽宁医学院沈阳军区总医院研究生培养基地,锦州121001 [2]传染病预防控制国家重点实验室,感染性疾病诊治协同创新中心,中国疾病预防控制中心传染病预防控制所,北京102206 [3]辽宁省疾病预防控制中心,沈阳110005 [4]沈阳军区总医院检验科,沈阳110016

出　　处：《中国人兽共患病学报》2015年第3期255-259,共5页Chinese Journal of Zoonoses

基　　金：辽宁省科技攻关计划资助项目(2011225021);国家科技重大专项课题(2009ZX10004-209)联合资助~~

摘　　要：目的本研究旨在建立准确、快速地鉴定布鲁氏菌菌种及生物分型的PCR-HRM(高分辨率熔解曲线)方法。方法根据目的基因序列,参考文献合成6对基因扩增引物(1对布鲁氏菌属特异引物,5对种间特异引物),应用PCR-HRM方法鉴定布鲁氏菌属6个种19个生物型的标准菌株,并初步应用到临床分离的35株布鲁氏菌中。结果采用布鲁氏菌属特异引物(Bspp),6个种19个生物型的标准菌株均扩增出同样形状的溶解曲线,与其它对照菌株的曲线不同;采用5对种特异引物,6个种的标准菌株均有特征性PCR-HRM曲线;临床分离的35株布鲁氏菌的PCR-HRM曲线与羊种布鲁氏菌标准菌株的一致。结论该研究采用的PCR-HRM分析方法,获得了布鲁氏菌属及6个种标准菌株的不同曲线图,可准确鉴定临床分离的羊种布鲁氏菌,可用于临床微生物实验室疑似布鲁氏菌感染的初步鉴定。The aim of this study is to develop a rapid and accurately species typing method for Brucella isolates by using High Resolution Melting （HRM ） analysis .Six pairs of primers were used according to the reference for the sequence of pur‐pose gene .Nineteen biotypes of six species Brucella standard strains were identified by PCR‐HRM analysis and this analysis was used to detect the 35 clinical isolates .Results showed Brucella amplified specific melting curves were different from con‐trasted strains with primer Bspp .The six species Brucella standard strains have own characteristic curve shape from each oth‐ers by PCR‐HRM analysis with five pairs of primers .Thirty‐five clinical isolates of Brucella have entirely consistent with PCR‐HRM curve shape with Brucella melitensis standard strains .So ,PCR‐HRM analysis methods can accurately identify Brucella strains ,especially clinical isolated Brucella melitensis ,and may be used in clinical microbiology laboratories .

关 键 词：布鲁氏菌 分子鉴定 高分辨溶解分析(PCR-HRM) 

分 类 号：R378.5[医药卫生—病原生物学]