同时检测鸭疫里默氏菌和大肠杆菌的环介导间接PCR方法的建立

发布时间：2018-01-25 06:55

本文关键词： 鸭疫里默氏菌大肠杆菌环介导间接PCR　出处：《中国预防兽医学报》2017年08期 　论文类型：期刊论文

【摘要】：为建立同时检测鸭疫里默氏菌和大肠杆菌的环介导间接PCR方法,本研究根据鸭疫里默氏菌外膜蛋白A基因和大肠杆菌的gapA基因的保守序列设计两对特异性探针,分别标记于猪线粒体基因片段的两端形成两种不同的捕获探针,将捕获探针与待检测菌的基因组杂交、补平缺口、环化后,采用1对引物反向扩增捕获探针,建立检测鸭疫里默氏菌和大肠杆菌的环介导间接PCR方法,扩增片段分别为540 bp和328 bp,本研究建立的方法对金黄色葡萄球菌、禽多杀性巴氏杆菌、沙门氏菌的检测结果为阴性;能够检测出10 pg的鸭疫里默氏菌和大肠杆菌的基因组DNA,与常规PCR的符合率为100%。本研究建立的环介导间接PCR方法是一种特异性好、灵敏度高的快速病原学诊断方法,为同时检测鸭疫里默氏菌和大肠杆菌提供了一种可行的方法。
[Abstract]:To establish an indirect cycle-mediated PCR method for simultaneous detection of Riemer's disease and Escherichia coli in ducks. In this study, two pairs of specific probes were designed according to the conserved sequences of the outer membrane protein A gene of Riemer's disease and the gapA gene of Escherichia coli. Two different kinds of capture probes were formed at the two ends of pig mitochondrial gene fragments respectively. The catch probe was hybridized with the genome of the bacteria to be detected. After cyclization, a pair of primers were used to reverse amplify the capture probe. An indirect PCR method was established for detection of Riemer's disease and Escherichia coli. The amplified fragments were 540bp and 328bprespectively. The method was developed for the detection of Staphylococcus aureus (Staphylococcus aureus). The detection results of Pasteurella multocida and Salmonella were negative. The genomic DNAs of 10 PG of Riemer's disease and Escherichia coli could be detected, and the coincidence rate with conventional PCR was 100. The indirect PCR method established in this study was a good specificity. The rapid etiological diagnosis method with high sensitivity provides a feasible method for simultaneous detection of Riemer's disease and Escherichia coli.
【作者单位】：福建省农业科学院畜牧兽医研究所福建省畜禽疫病防治工程技术研究中心/福建省禽病防治重点实验室;
【基金】：现代农业产业技术体系建设专项资金(CARS-42) 福建省属公益类项目(2015R1023-2) 福建省畜禽疫病防控技术重大研发平台(2014N2003)
【分类号】：S852.61
【正文快照】： 鸭疫里默氏菌病和大肠杆菌病是鸭、鹅临床上出入境检验检疫局郑腾研究员惠赠;金黄色葡萄球最常见的疾病,两种病原的耐药特点不同,治疗时菌261111株购自中国药品生物制品所;多杀性巴氏选择的用药方案也不相同,但临床上仅凭病变特点杆菌CVCC44801株购自中国兽医药品监察所;肠炎

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