Development and Evaluation of Multiplex PCR Detection Kit for Detection of Three Types of Diarrheagenic Escherichia coli
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Abstract:
In order to develop a multiplex PCR detection kit for detecting three types of Diarrheagenic Escherichia coli (DEC), which includes Enteropathogenic E. coli (EPEC), Shigatoxin-producing E. coli (STEC) and Enterohemorrhagic E. coli (EHEC), multiplex PCR reaction system was optimized and their components of the kit were confirmed. The main detection reagents were produced into powder pattern by freeze drying technology, and the specificity, sensitivity, repeatability and shelf life of the kit were evaluated. The results showed that EP and ST reaction systems were successfully established for detection of three types of DEC. All standard strains and isolated strains belonging to the three types of DEC were detected positive and presented obvious specific amplification bands, while other types of DEC were all detected negative. The limit of detection of detection tube EP and ST in the kit were calculated to 1.5×103 cfu/mL and 2.1×103 cfu/mL, respectively, and the testing accordance rates were all 100%. The kit can be stored under 4°C condition for 12 months and transported under 42 ℃ condition for 120 h without efficacy loss. In conclusion, the kit had a good performance with stable and reliable detection results, and it could be applicable for rapid detection of three types of DEC in food samples. Key words:
