检测和鉴别流产布鲁氏菌与猪布鲁氏菌的复合PCR方法的建立

A Multiplex PCR for Detection and Identification of Brucella abortus and Brucella suis

将GenBank上登录的流产布鲁氏菌(Br.abortus)和猪布鲁氏菌(Br.suis)外膜蛋白omp2基因序列进行比对,设计了流产布鲁氏菌特异性引物对A1/A2、猪布鲁氏菌特异性引物对S1/S2,分别用引物A1/A2和S1/S2进行PCR,进而将引物A1、A2、S1、S2置于同一体系内进行复合PCR,建立了一种能区分流产布鲁氏菌和猪布鲁氏菌的复合PCR鉴别诊断方法。应用该方法从流产布鲁氏菌和猪布鲁氏菌染色体基因组中扩增出了大小分别为424bp和535bp的1条特异性片段,从两种染色体基因组混合物中扩增出了大小为424bp和535bp的2条特异性片段。但对大肠杆菌、结核杆菌、坏死杆菌、棒状杆菌染色体基因组进行PCR扩增时均为阴性。该方法可以检测出10个菌/50μL体系。本研究建立的复合PCR可以有效检测布鲁氏菌感染并把流产布鲁氏菌和猪布鲁氏菌区分开。

A multiplex PCR method was developed for detection and identification of Brucella abonus and BruceUa suis. Primers A1/A2 or S1/S2 specific for BruceUa abortus or Brucella suis, respectively, were designed in the PCR. A fragment of 424 bp was am- plified from BruceUa abortus genomic DNA, and a fragment of 535 bp from Brucella suis DNA in the PCR, and two fragments of 424 bp and 535 bp were simultaneously amplified from mixed DNA samples of BruceUa abortus and Brucella suis. No amplification was achieved for Corynebacterium, Pheasianus colchicus linnaeus, Mycobacterium avium, or Mycobacterium bovis BCG. It could detect 10/50 L PCR reaction ofBrucella abortus or Brucella suis. The PCR can be used to effectively detect and identify Brucella abortus and Brucella suis.