恒温实时荧光法快速检测奶粉中阪崎肠杆菌方法的建立

发布时间：2018-05-30 18:58

本文选题：恒温实时荧光法 + 阪崎肠杆菌　；参考：《现代食品科技》2016年09期

【摘要】：为实现奶粉中快速检测阪崎肠杆菌,本文建立了检测阪崎肠杆菌的恒温实时荧光法。针对阪崎肠杆菌16S r RNA设计三组LAMP引物,采用Deaou-308C恒温实时荧光检测平台,选取常见病原菌标准株进行引物特异性检测;选取阪崎肠杆菌标准菌株进行基因组DNA灵敏度和最低检测限测定,同时利用人工污染方式检测此方法在脱脂和全脂奶粉中的灵敏度和最低检测限,利用Real Amp法和国标法对20份市售奶粉进行对比实验。结果显示,引物组16S-11扩增效率最优,与常见病原菌无交叉反应,对阪崎肠杆菌基因组DNA、阪崎肠杆菌污染的脱脂和全脂奶粉的灵敏度分别达到102 CFU/m L、102 CFU/m L和103 CFU/m L;对阪崎肠杆菌基因组DNA和阪崎肠杆菌污染的脱脂和全脂奶粉的最低检测限分别达到103 CFU/m L、103 CFU/m L和104 CFU/m L;在20份市售奶粉样品中Real Amp检测结果与传统国标培养结果一致,表明本文建立的阪崎肠杆菌Real Amp检测方法适用于阪崎肠杆菌的快速检测。
[Abstract]:In order to rapidly detect Enterobacter sakazakii in milk powder, a real-time isothermal fluorescence method was developed for the detection of Enterobacter sakazakii. Three sets of LAMP primers were designed for Enterobacter sakazakii 16s r RNA. The standard strains of common pathogenic bacteria were selected for specific detection using Deaou-308C real-time fluorescence detection platform. Enterobacter sakazakii standard strains were selected to detect the sensitivity and minimum detection limit of genomic DNA, and the sensitivity and minimum detection limit of this method in defatted and full-fat milk powder were detected by artificial contamination. Real Amp method and national standard method were used to compare 20 samples of milk powder sold in the market. The results showed that the primer group had the best amplification efficiency and had no cross reaction with common pathogens. The sensitivity to Enterobacter sakazakii genome DNA, Enterobacter sakazakii contaminated defatted milk powder and whole milk powder were 102 CFU/m L ~ (10 ~ 2) CFU/m L and 10 ~ 3 CFU/m / L, respectively, while the defatted and whole milk powder contaminated by Enterobacter sakazakii genome DNA and Enterobacter sakazakii were the most sensitive. The low detection limit was 103 CFU/m / L 103 CFU/m / L and 104 CFU/m / L, respectively, and the results of Real Amp detection in 20 samples of marketable milk powder were consistent with those of the traditional national standard. This method is suitable for rapid detection of Enterobacter sakazakii.
【作者单位】：华南理工大学轻工与食品学院;广州迪澳生物科技有限公司;
【基金】：中央高校基本科研业务费专项资金资助(2015ZM063) 中国博士后科学基金(2015M580723)
【分类号】：R155.5

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