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[Establishment of rapid detection method for multiple real-time fluorescent PCR of toxin-producing fungi]
Qing Zhang1, Xiaomei Zhao1, Ying Chen1
1Agro-Product Safety Research Center, Chinese Academy of Inspection and Quarantine, Beijing 100176, China.
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, β-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.
