【摘要】 采用环介导等温扩增(Loop Mediated Isothermal Amplification,LAMP)技术,利用实时浊度仪实时检测LAMP反应过程中所产生的焦磷酸镁白色沉淀,实现对扩增反应全过程的监控,建立食品中肠出血性大肠杆菌(Enterohemorrhagic Escherichia coli,EHEC)O157的LAMP实时浊度法快速检测方法。针对EHEC O157抗原基因rfbE的序列设计4条特异性LAMP引物。通过实时浊度仪在恒温63℃下1小时完成检测,对方法的特异性、灵敏度、稳定性进行了评价,并进行了食品样品添加试验以评价其应用于实际样品的效果。结果显示,经优化后,该方法的最低检出限为2.8×103 CFU/mL,与PCR法灵敏度比较高100倍。样品添加试验能检出的最低添加菌量为2.8×102 CFU/25 g(或mL)。LAMP实时浊度法具有快速、灵敏、特异性高、操作简便的优势,为食源致病菌快速筛查提供了一种良好的检测模式。更多还原

【Abstract】 Real-time monitoring of magnesium pyrophosphate produced from the loop-mediated isothermal amplification(LAMP) reaction was achieved by the turbidimeter. Four specific primers were designed according to the sequences of antigen gene rfbE of EHEC O157. Detection was completed at 63 ℃ on the real-time turbidimeter in 1 h. The specificity, sensitivity, stability of the method was evaluated. Results showed that after optimized, the lowest detection limit was 2.8×103 CFU/mL, and 100 times more sensitive than the PCR, and the minimal adding quantity of 2.8×100 CFU/25 g(or mL) could be detected. LAMP real-time turbidity method is rapid, sensitive, high specificity and easy to operate, which is a good pattern for fast screening of foodborne pathogens.更多还原