马流产沙门菌双重PCR检测方法的建立  

Establishment of a duplex PCR method for detection of Salmonella abortus equi

作　　者：刘香 梁佳平 张灿[1] 邹玲[1] 任慧英[1] 刘文华[1] LIU Xiang;LIANG Jia-ping;ZHANG Can;ZOU Ling;REN Hui-ying;LIU Wen-hua(College of Veterinary Medicine,Qingdao A gricultural University,Qingdao 266109,China)

机构地区：[1]青岛农业大学动物医学院,山东青岛266109

出　　处：《中国兽医科学》2023年第3期292-297,共6页Chinese Veterinary Science

基　　金：山东省产业技术体系驴产业创新团队项目(SDAIT-27);2022年青岛农业大学大学生创新创业训练项目(2022-580)。

摘　　要：为建立一种检测马流产沙门菌的双重PCR方法,以实现对驴场中马流产沙门菌和其他血清型沙门菌感染的快速检测。首先针对马流产沙门菌Ⅰ相鞭毛基因FliC和沙门菌保守毒力基因invA分别设计引物,建立单特异性PCR方法,在此基础上建立双重PCR,并对该双重PCR方法进行特异性、敏感性检测。同时利用该双重PCR方法对临床样品进行检测,通过符合率比较验证该方法的实用性。结果显示,马流产沙门菌能分别扩增出针对特异基因的747 bp的目的条带和针对保守基因的175 bp的目的条带,其他血清型沙门菌只能扩增出针对保守基因的175 bp的目的条带,非沙门菌均无扩增条带;对模板DNA的最低检出限为34.8 pg;利用建立的双重PCR方法对186份临床样品进行检测,结果显示,与细菌分离培养方法和单重PCR方法的符合率均为100%。说明本试验建立的双重PCR方法特异性强、灵敏性高。该方法的建立为马流产沙门菌的快速检测提供了有力的技术支撑。A duplex PCR method was established for the rapid detection of the infection of Salmonella abortus equi and other Salmonella serotypes in donkey farms.Firstly,primers were designed by theflagellargene FliC of Salmonella abortus equi and the conservative virulence gene invA of Salmonella to establish a monospecific PCR method respectively,then a duplex PCR was established according to the monospecific PCR method.Specificity and sensitivity of the duplex PCR method were tested.At the same time,the duplex PCR method was used to detect clinical samples,and the practicability of the method was identified by comparison of coincidence rates.Results showed that Salmonella abortus equi could specifically amplify a 747 bp target band for specific genes and a 175 bp target band for conserved genes,respectively.Other serotypes of Salmonella could only amplify the 175 bp target band,and non-Salmonella strains had no amplification band.The minimum detection quantity of template DNA was 34.8 pg;the established duplex PCR method was used to detect 186 clinical samples,and results showed that the coincidence rate was 100% with methods of bacterial isolation and identification and the single PCR detection.The above experiments indicated thatthe established duplex PCR method has good specificity and high sensitivity.which provides effective technical support for the rapid detection of Salmonella abortus equi.

关 键 词：马流产沙门菌 驴 FLIC invA 双重PCR 

分 类 号：S852.612[农业科学—基础兽医学]