猪种、牛种、羊种、绵羊种布鲁菌多重PCR检测方法的建立及应用

发布时间：2018-06-17 23:29

本文选题：猪种布鲁菌 + 牛种布鲁菌　；参考：《西北农业学报》2017年08期

【摘要】：为建立在临床样本能够同时检测猪种、牛种、羊种、绵羊种布鲁菌的多重PCR方法,根据NCBI已收录的布鲁菌全基因,设计并合成4对特异性引物,通过优化多重PCR的反应条件,建立能够同时检测4种布鲁菌的多重PCR诊断方法。特异性试验结果表明,可以从4种布鲁菌以及4种细菌混合物中扩增出大小分别为276、494、733和976bp的特异性条带,对照组的检测结果为阴性;敏感性试验结果表明,对4种病原菌基因组DNA的检出量为猪种32.2pg、牛种21.3pg、羊种27.5pg、绵羊种43.2pg;人工模拟感染样本检测结果表明,能从混合感染的病料中特异地检测出4种病原菌。应用该方法对200份临床奶山羊乳样进行检测,结果检出4份阳性。建立的多重PCR方法具有特异性强、敏感度高、稳定性好的特点,可以有效地检测4种布鲁菌的感染。
[Abstract]:In order to establish a multiplex PCR method for simultaneous detection of brucellosis in swine, cattle, sheep and sheep in clinical samples, four pairs of specific primers were designed and synthesized according to the whole genes of Brucella collected by NCBI, and the reaction conditions of multiplex PCR were optimized. A multiplex PCR method for simultaneous detection of four Brucella species was established. The specific bands of 276494733 and 976bp could be amplified from four Brucella species and four bacterial mixtures, respectively. The results of the control group were negative, and the sensitivity test showed that the specific bands could be amplified from the four Brucella species and the four bacterial mixtures. The amount of genomic DNA of four pathogens was 32.2pg for pigs, 21.3pg for cattle, 27.5pg for sheep and 43.2pg for sheep. The method was used to detect 200 milk samples of clinical dairy goats and 4 samples were positive. The multiplex PCR method has the characteristics of high specificity, high sensitivity and good stability. It can be used to detect the infection of four Brucella species effectively.
【作者单位】：西北农林科技大学动物医学院;
【基金】：陕西省农业科技创新与攻关(2016NY-092) 陕西省重点产业创新链(2016KTZDNY02-06) 西北农林科技大学试验示范站(基地)科技成果推广(TGZX2015-32)~~
【分类号】：S852.61

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