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文章摘要
恒温实时荧光法快速检测不同样品中的单增李斯特菌
Rapid Detection of Listeria monocytogenes in Different Samples by Real Time Fluorescence Isothermal Amplification
  
DOI:10.3969/j.issn.1673-1689.2019.05.007
中文关键词: 恒温实时荧光法  快速检测  单增李斯特菌
英文关键词: real time fluorescence isothermal amplification assay,rapid detection,Lis. Monocytegenes
基金项目:
作者单位
张文敏 上海理工大学 医疗器械与食品学院上海 200093 
石育娇 上海理工大学 医疗器械与食品学院上海 200093 
戚成 上海理工大学 医疗器械与食品学院上海 200093 
田明胜 上海食品药品监督管理局上海 200021 
朱智 上海捷易生物科技有限公司上海 201210 
殷荣永 上海捷易生物科技有限公司上海 201210 
钟少水 上海捷易生物科技有限公司上海 201210 
王大鹏 上海交通大学 农业与生物学院上海 200240 
何更生 复旦大学 公共卫生学院上海 200433 
冯元琦 广州迪澳生物科技有限公司广州 510663 
董庆利 上海理工大学 医疗器械与食品学院上海 200093 
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中文摘要:
      为实现乳制品中、临床病人腹泻物中单增李斯特菌的快速检测,选用外引物F3和B3、内引物FIP和BIP作为环介导等温扩增(loop-mediated isothermal amplification,LAMP)的特异性引物,采用便携式荧光恒温扩增仪作为检测平台,选取单增李斯特菌标准菌株进行基因组DNA灵敏度和最低检测限测定;选用人工污染的15份乳制品样品和20份腹泻样品进行适用性实验;以上样品同时利用国标法GB4789.30-2010进行菌落计数。结果表明:恒温实时荧光法对纯培养的单增李斯特菌基因组DNA、乳制品和腹泻样品中的单增李斯特菌基因组DNA灵敏度均达到102 CFU/mL、检测限均达到103 CFU/mL;阴性样本出现假阳性的概率分别为0、0.03%和0;单增李斯特菌污染浓度在检出限以上的阳性样本检出率分别为100%、99%和92%。研究表明恒温实时荧光法的检测结果与传统国标培养结果基本一致,恒温实时荧光法适用于乳制品中和临床腹泻样本中单增李斯特菌的快速检测。
英文摘要:
      In order to detect the Listeria monocytogenes in dairy products and clinical diarrhea rapidly,the outer primer F3 and B3,inter primer FIP and BIP were chosen as the loop-mediated isothermal amplification (LMAP) specific primers. Then a portable thermostat fluorescence detector was used as the detection platform,while the sensitivity and detection limit on genomic DNA were also determined with a standard strain of Lis. monocytegenes;Artificially contaminated 15 dairy products and 20 clinical diarrhea were served to measuring the applicability of this assay,and a Chinese national standard method (GB4789.30-2010) was used at the same time. The results showed that the sensitivity and detection limit respectively reached to 102 CFU/mL and 103 CFU/mL,while the probabilities of false positive were 0,0.03 and 0,the positive detection rate were 100%,99% and 92%. The results were essentially in agreement with the culture method of GB4789.30-2010,indicating that this real time fluorescence isothermal amplification assay is suitable for the rapid detection of Lis. monocytegenes in dairy products and clinical diarrhea.
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