【摘要】：為建立腸出血性大腸埃希菌O157:H7的多重PCR檢測方法,針對(duì)O157菌特異性eaeA基因、fliC基因、志賀毒素基因(stx1和stx2)及rfbE基因設(shè)計(jì)5對(duì)引物,構(gòu)建多重PCR反應(yīng)體系,優(yōu)化反應(yīng)的引物濃度和溫度檢測其特異性和敏感性,并對(duì)牛、豬、雞及犬等不同動(dòng)物來源的樣品進(jìn)行了大腸埃希菌O157檢測。結(jié)果顯示,該方法具有良好的特異性和敏感性,敏感性達(dá)到5×10~3CFU。從檢測陽性樣本中均分離到目的菌。應(yīng)用建立的方法在確定樣品中是否含有大腸埃希菌O157的同時(shí),還能對(duì)菌株毒力進(jìn)行初步判定。成功建立了腸出血性大腸埃希菌O157:H7多重PCR檢測方法,為該病原菌感染的預(yù)防和監(jiān)測提供了簡便、快速的技術(shù)手段,具有良好的應(yīng)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.
【作者單位】： 軍事醫(yī)學(xué)科學(xué)院軍事獸醫(yī)研究所吉林省人獸共患病預(yù)防與控制重點(diǎn)實(shí)驗(yàn)室;長春中醫(yī)藥大學(xué)附屬醫(yī)院;
【基金】：吉林省自然科學(xué)基金項(xiàng)目(20150101110JC,20140101032JC) 武漢市科技計(jì)劃項(xiàng)目(2014060101010051)
【分類號(hào)】：S852.612
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本文編號(hào)：2307092