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黄偌颖, 方梅, 罗影殊等. EMA-PCR技术检测肠出血性大肠杆菌O157∶H7[J]. koko体育app 学报（医学版）, 2014, 45(1): 152-155.
引用本文: 黄偌颖, 方梅, 罗影殊等. EMA-PCR科技判断肠大出血性结肠杆菌O157∶H7[J]. 上海高中学报（临床版）, 2014, 45(1): 152-155.
HUANG Ruo-ying, FANG Mei, LUO Ying-shu. et al. EMA-PCR Detection of Enterohemorrhagic Eschrichia ColiO157∶H7[J]. Journal of Sichuan University (Medical Sciences), 2014, 45(1): 152-155.
Citation: HUANG Ruo-ying, FANG Mei, LUO Ying-shu. et al. EMA-PCR Detection of Enterohemorrhagic Eschrichia ColiO157∶H7[J]. Journal of Sichuan University (Medical Sciences), 2014, 45(1): 152-155.

EMA-PCR技术检测肠出血性大肠杆菌O157∶H7

EMA-PCR Detection of Enterohemorrhagic Eschrichia ColiO157∶H7

  • 摘要: 目的 将叠氮溴化乙锭 （ethidium monoazide bromide，EMA）与PCR技术相结合，建立一种有效、快速检测活肠出血性大肠杆菌O157∶H7的方法。方法 以rfbE为PCR检测靶基因，O157∶H7培养物经EMA处理后制备模板进行PCR检测，并对EMA使用浓度、作用时间等进行优化。结果 不抑制O157∶H7活菌PCR扩增的最大EMA浓度为10 μg/mL；抑制2×107 CFU/mL死菌PCR反应的最小EMA浓度为0.5 μg/mL；该法对O157∶H7检测的灵敏度为2×104 CFU/mL，结果显示能检出混合体系中含有的1%的活菌。结论 EMA-PCR技术能有效检测活的O157∶H7，在突发公共卫生事件的快速检测中具有良好的应用前景。  
    Abstract: Objective?A rapid and effective method with ethidium monoazide bromide (EMA) in combination with PCR (EMA-PCR) was established to detect live Enterohemorrhagic Eschrichia Coli O157∶H7. Methods?The rfbE gene was used as the target gene for PCR detection of Eschrichia Coli O157∶H7 by utilizing its pure isolates after the treatment of EMA as the template. The EMA concentration and reaction time was optimized. Results?The use of 10 μg/mL or less EMA did not inhibit the PCR amplification of DNA derived from viable bacteria. The PCR amplification of DNA derived from 2×107 CFU/mL dead cells can be inhibited by 0.5 μg/mL EMA. The sensitivity of the method was 2×104CFU/mL. The results demonstrated that it could detect 1% live bacteria from a mixed bacterial population. Conclusion?EMA-PCR can effectively detect live bacteria of O157∶H7, it could be a potential rapid detection method applied in public health emergent events.  
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