鸡胚胎性病原菌多重PCR检测方法的建立  

Establishment of multiplex PCR detection for pathogens of chicken embryos

作　　者：谭燕玲[1] 朱瑞良[1] 王慧[1] 王新建[1] 魏凯[1] 孙振红[1] 盛鹏程[1] 

机构地区：[1]山东农业大学动物科技学院山东省动物生物技术与疫病防治重点实验室,山东泰安271018

出　　处：《中国预防兽医学报》2011年第5期374-377,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家自然科学基金(30972183;30740077;30670114);山东省自然科学基金项目(Y2006D09)

摘　　要：为建立同时检测禽波氏杆菌(Bordetella avium)、沙门氏菌(Salmonella)、大肠杆菌(Escherichia coli)和绿脓杆菌(Pseudomonas aeruginosa)4种导致鸡胚死亡病原菌的多重PCR方法,本研究根据B.avium的ompA基因、Salmonella的invA基因、E.coli的phoA基因和P.aeruginosa的toxR基因序列,各设计一对特异性引物进行多重PCR反应,并对反应体系和条件进行优化。结果显示,4对引物分别扩增出597bp、724bp、372bp和278bp的目的条带;并且特异性强不与其他非目的菌发生反应。经优化B.avium、P.aeruginosa和E.coli多重PCR检测灵敏度达到104cfu/mL,而Salmonella为103cfu/mL。本研究建立的多重PCR方法为相关病原菌的快速检测提供方法。To establish a multiplex PCR assay for simultaneous detecation of Bordetella avium, Escherichia coli, Salmonella and Pseudomonas aeruginosa in infected embryos, four pairs of specific primers were designed and synthesized based on the genes of ompA in B. avium, invA in Salmonella, phoA in E. coli and toxR in P. aeruginosa for detection of these pathogenic bacteria by multiplex PCR. Test results indicated that the PCR products were 597 bp for B. avium, 724 bp for Salmonella, 372 bp for E. coli, and 278 bp for P. aeruginosa, and the limit detections were approximate 104 cfu/mL for B. avium, E. coli and P. aeruginosa and 103 cfu/mL for Salmonella, respectively, without any amplification from other related bacteria. The detection results from clinical embryos samples showed that the multiplex PCR were consistent with that of bacteria culture isolation. The multiplex PCR assay could be used for rapid detection of pathogenic bactera.

关 键 词：禽波氏杆菌 沙门氏菌 大肠杆菌 绿脓杆菌 多重PCR 

分 类 号：S852.61[农业科学—基础兽医学]