肠出血性大肠埃希菌O157:H7多重PCR检测方法的建立
发布时间：2018-11-03 07:18
【摘要】：为建立肠出血性大肠埃希菌O157:H7的多重PCR检测方法,针对O157菌特异性eaeA基因、fliC基因、志贺毒素基因(stx1和stx2)及rfbE基因设计5对引物,构建多重PCR反应体系,优化反应的引物浓度和温度检测其特异性和敏感性,并对牛、猪、鸡及犬等不同动物来源的样品进行了大肠埃希菌O157检测。结果显示,该方法具有良好的特异性和敏感性,敏感性达到5×10~3CFU。从检测阳性样本中均分离到目的菌。应用建立的方法在确定样品中是否含有大肠埃希菌O157的同时,还能对菌株毒力进行初步判定。成功建立了肠出血性大肠埃希菌O157:H7多重PCR检测方法,为该病原菌感染的预防和监测提供了简便、快速的技术手段,具有良好的应用前景。
[Abstract]:In order to establish a multiplex PCR detection method for O157:H7 of Escherichia coli, five pairs of primers were designed for O157 specific eaeA gene, fliC gene, Shigella toxin gene (stx1 and stx2) and rfbE gene to construct multiple PCR reaction system. The sensitivity and specificity of primer concentration and temperature were optimized, and Escherichia coli O157 was detected from different animal samples, such as cattle, pig, chicken and dog. The results showed that the method had good specificity and sensitivity, and the sensitivity was 5 脳 10 ~ (-3) CFU. The target bacteria were isolated from positive samples. The established method can also determine the virulence of Escherichia coli O157 at the same time. A multiplex PCR method for the detection of enterohemorrhagic Escherichia coli (O157:H7) was successfully established, which provides a simple and rapid technique for the prevention and monitoring of the pathogen infection, and has a good prospect of application.
【作者单位】： 军事医学科学院军事兽医研究所吉林省人兽共患病预防与控制重点实验室;长春中医药大学附属医院;
【基金】：吉林省自然科学基金项目(20150101110JC,20140101032JC) 武汉市科技计划项目(2014060101010051)
【分类号】：S852.612
，
本文编号：2307092
[Abstract]:In order to establish a multiplex PCR detection method for O157:H7 of Escherichia coli, five pairs of primers were designed for O157 specific eaeA gene, fliC gene, Shigella toxin gene (stx1 and stx2) and rfbE gene to construct multiple PCR reaction system. The sensitivity and specificity of primer concentration and temperature were optimized, and Escherichia coli O157 was detected from different animal samples, such as cattle, pig, chicken and dog. The results showed that the method had good specificity and sensitivity, and the sensitivity was 5 脳 10 ~ (-3) CFU. The target bacteria were isolated from positive samples. The established method can also determine the virulence of Escherichia coli O157 at the same time. A multiplex PCR method for the detection of enterohemorrhagic Escherichia coli (O157:H7) was successfully established, which provides a simple and rapid technique for the prevention and monitoring of the pathogen infection, and has a good prospect of application.
【作者单位】： 军事医学科学院军事兽医研究所吉林省人兽共患病预防与控制重点实验室;长春中医药大学附属医院;
【基金】：吉林省自然科学基金项目(20150101110JC,20140101032JC) 武汉市科技计划项目(2014060101010051)
【分类号】：S852.612
，
本文编号：2307092
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