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用于检测即食水果和蔬菜中沙门氏菌的可视化环介导等温扩增试剂盒的研制。

Development of a test kit for visual loop-mediated isothermal amplification of Salmonella in spiked ready-to-eat fruits and vegetables.

机构信息

College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095, People's Republic of China.

College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095, People's Republic of China.

出版信息

J Microbiol Methods. 2020 Feb;169:105830. doi: 10.1016/j.mimet.2019.105830. Epub 2019 Dec 28.

Abstract

The purpose of this study was to assemble two types of loop-mediated isothermal amplification (LAMP) kit that have the ability to visually detect Salmonella in ready-to-eat fruits and vegetables. The reaction results were obtained within 20-40 min after addition of DNA and can be discerned by the naked eye or an amplification plot. The stability of the LAMP wet kit was evident after multiple freezing and thawing cycles, and the one-step LAMP lyophilized kit was further evolved to allow ambient temperature transport for deployment in resource-limited settings. The cost-effective wet kit had the ability to detect minimum amounts of 1.8 CFU/ml Salmonella DNA without enrichment, while the sensitivity of the one-step LAMP lyophilized kit was only 9.8 × 10 CFU/ml. They both have good anti-interference, as they were both able to detect 2.1 × 10 CFU/ml Salmonella mixed with 10 CFU/ml four non-Salmonella strain mixture. Moreover, cucumber and lettuce that were contaminated with an initial inoculation of 1.7 CFU of Salmonella/10 g showed detection within a reaction time of 30 min after 10 h enrichment. The present research setup is a convenient and practical kit for Salmonella rapid detection that has good application prospects in food safety monitoring.

摘要

本研究的目的是组装两种具有可视化检测即食型水果和蔬菜中沙门氏菌能力的环介导等温扩增(LAMP)试剂盒。加入 DNA 后 20-40 分钟即可得到反应结果,可通过肉眼或扩增图谱进行辨别。LAMP 湿试剂盒经过多次冻融循环后稳定性明显,一步法 LAMP 冻干试剂盒进一步改进,可在常温下运输,以便在资源有限的环境中部署。具有成本效益的湿试剂盒无需富集即可检测到低至 1.8 CFU/ml 的沙门氏菌 DNA,而一步法 LAMP 冻干试剂盒的灵敏度仅为 9.8×10 CFU/ml。它们都具有良好的抗干扰能力,因为它们都能够检测到 2.1×10 CFU/ml 的沙门氏菌与 10 CFU/ml 的四种非沙门氏菌菌株混合物。此外,用初始接种量为 1.7 CFU/10g 的黄瓜和生菜进行污染,在富集 10 小时后 30 分钟的反应时间内即可检测到沙门氏菌。本研究建立了一种方便实用的沙门氏菌快速检测试剂盒,在食品安全监测方面具有良好的应用前景。

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