A Multiplex Pcr And A Taqman Realtime Pcr For The Identification Of Meat Species In Meat Products

Meat adulteration is one of the worldwide challenges for food quality control. In the last decade, methodologies for species identification in meat products are mainly based on the DNA sequences. The precision of analysis is to genus or subgenus level. A multiplex PCR were used to be able to identify4kinds of meat ingredient (chicken, beef, lamb, pork) in food products in this paper. And a Taqman real-time PCR with IAC (Internal amplification control) was developed to identify pork and chicken ingredient. The contents and results were detailed as follows.1. An accurate and reliable universal primers-multiplex PCR (UP-M-PCR) method was developed to be able to identify4kinds of meat ingredient (chicken, beef, lamb, pork) in food products. Two sets of UP-M-PCR primers in different length were designed based on the homologous and specific sites of mitochondrial cytochrome b gene. Each set was comprised of5primers:one general forward primer and four species-specific reverse primers. The UP-M-PCR system, followed by gel electrophoresis assay of spices-specific bands, was established for meat identification. The meat spices were identified according to different length of PCR products. Meanwhile, the accuracy of the method was evaluated by testing of eighty food samples from the market. The optimized UP-M-PCR was established using the set of longer primes, which showed higher specificity than that using the shorter ones. The detection limit was picograms of DNA. The accuracy of the method was verified by examination of80food samples from the market. A sensitive, reliable and convenient method was developed to be able to identify4kinds of meat ingredient in food products.2. A Taqman real-time PCR with IAC (Internal amplification control) was developed to identify pork and chicken ingredient in meat products; Specific primers and probes were designed based on the sequence of beta actin gene of pig DNA and transforming growth factor gene of chicken DNA, respectively. The specificity and sensitivity of the primers and probes were evaluated. An IAC was designed and constructed, and the real-time PCR system with IAC was established by optimizing the concentration of IAC in the system. The DNA of raw and cooked samples was evaluated applying the Taqman real-time PCR system with IAC. Meanwhile,38food samples from market were assayed. The pork and chicken ingredient in food was detected effectively by the Taqman real-time PCR with IAC. No cross-reaction was observed between species specific primer-probe systems in40cycles. The limit of detection was0.5ng of template DNA. No significant difference was observed between the Ct values of raw and cooked samples. The accuracy of the method was verified by examination of38food samples from the market; an accurate and reliable method was developed to identify pork and chicken ingredient in food products.