[Establishment of rapid detection method for multiple real-time fluorescent PCR of toxin-producing fungi].

OBJECTIVE: To establish a multiplex real-time fluorescent PCR rapid detection method for simultaneous detection of aflatoxin-producing fungi and three latent toxin-producing fungi in a single system. METHODS: Primers and probes were designed based on the protein activation genes AflR, ITS sequence, ß-tubulin coding region and LS rRNA of aflatoxin-producing fungi, Aspergillusochraceus, Penicillium and Fusarium, respectively. A real-time fluorescent PCR method for simultaneous detection of commontoxin-producing fungi was performed, and the sensitivity and specificity of the method were analyzed by optimizing the reaction components and reaction conditions. RESULTS: The detection limits of the optimized reaction system for aflatoxin-producing fungi, Aspergillusochraceus, Penicillium and Fusarium were 3. 37×10~4, 1. 91×10~4, 1. 53×10~4, 3. 95×10~4 copies/reaction, respectively. CONCLUSION: The multiplex real-time fluorescent PCR assay established in this study is highly specific and sensitive. It can detect toxin-producing fungi within 2 hours and provide technical support for fungal monitoring in food.