LAMP实时浊度法快速检测肠出血性大肠杆菌O157的研究  

Rapid Detection of Enterohemorrhagic Escherichia coli O157 by LAMP Real-time Turbidity Method

作　　者：李碧霞[1,2] 游淑珠[2] 邝筱珊[2] 冯家望[2] 王小玉[2] 唐食明[2] 成晓维[2] 许喜林[1] 

机构地区：[1]华南理工大学轻工与食品学院,广东广州510640 [2]珠海出入境检验检疫局技术中心,广东珠海519015

出　　处：《现代食品科技》2014年第7期268-272,305,共6页Modern Food Science and Technology

基　　金：国家质检总局科技计划项目(2010IK174)

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。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 2.8×10^3 CFU/mL, and 100 times more sensitive than the PCR, and the minimal adding quantity of 2.8×10^0 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.

关 键 词：环介导等温扩增技术 实时浊度仪 大肠杆菌O157 检测 

分 类 号：TS207.4[轻工技术与工程—食品科学]