应用TaqMan荧光定量PCR检测牛分枝杆菌

摘要

为建立快速检测牛分枝杆菌(M.bovis)的TaqMan荧光定量PCR方法,本研究以GenBank登录的M.bovis特有229 bp基因为研究对象,设计并合成引物及探针.该方法具有较好的特异性,与标准质控菌株呈阳性反应,与其他微生物样品呈阴性反应;灵敏性最低检测值可达1 pg/mL;对20阳性临床样品进行荧光定量PCR检测,均为阳性;而对培养为阴性的20份临床样品进行检测,6份为阳性.该研究结果表明,建立的方法特异性强,敏感性高,稳定性好,能够用于M.bovis的鉴别检测,对牛分枝杆菌病的快速检测和早期诊断具有重要意义.%A TaqMan real-time PCR assay was developed for detection of Mycobacterium bovis infection in cattle based on primers and TaqMan probe derived from M. bovis sequence in GenBank. The specific test showed that the assay had positive results for detection of M. bovis strains and negative for other bacteria. This assay could detect single bacteria. Comparing with other methed on 20 PPD positive clinical samples which were negative by bacteria isolation, six samples were positive detected by the real-time PCR. Indicating the real-time PCR is a rapid and specific assay for detection of M. bovis infection and could be used in the early diagnosis of bovine tuberculosis.