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目的:利用多重PCR技术,建立可以同时检测多种食源性致病菌的多重PCR方法。方法:分别选择沙门氏菌invA基因,志贺氏菌的ipaH基因,单核细胞增生李斯特氏菌的hlyA基因,大肠杆菌O157:H7的eaeA基因,副溶血弧菌的toxR基因,设计多重PCR引物,建立多重PCR检测体系,并对该体系进行特异性和灵敏度实验。结果:通过对19株菌株进行实验,所有的目标菌株均为阳性,而其余菌株为阴性。对多重PCR体系的灵敏度进行考察,沙门菌的灵敏度为5000 CFU/mL;志贺氏菌的灵敏度为5500CFU/mL;单核细胞增生李斯特氏菌的灵敏度为5200 CFU/mL;O157:H7的灵敏度为5000CFU/mL;副溶血弧菌的灵敏度为6300CFU/mL。结论:建立的多重PCR体系能实现多种致病菌同时检测。
Objective: To establish a multiplex PCR method that can detect a variety of food-borne pathogens simultaneously using multiplex PCR. Methods: The invA gene of Salmonella, the ipaH gene of Shigella, the hlyA gene of Listeria monocytogenes, the eaeA gene of Escherichia coli O157: H7 and the toxR gene of Vibrio parahaemolyticus were selected respectively. Multiplex PCR primers were designed, To establish a multiplex PCR detection system, and the system specificity and sensitivity experiments. Results: By testing 19 strains, all of the target strains were positive and the rest of the strains were negative. The sensitivity of multiplex PCR system was investigated. The sensitivity of Salmonella was 5000 CFU / mL. The sensitivity of Shigella was 5500 CFU / mL. The sensitivity of Listeria monocytogenes was 5200 CFU / mL. The sensitivity of O157: H7 Sensitivity of 5000CFU / mL; Vibrio parahaemolyticus sensitivity of 6300CFU / mL. Conclusion: The established multiplex PCR system can detect multiple pathogens simultaneously.