A simple and effective PCR assay to detect the origin of meat in food using mitochondrial DNA

Adulteration of meat products is a serious global issue. This study aims to establish a multiplex PCR detection system for species authentication in meat products. Six sets of species-specific primers were designed to target mitochondrial DNA variable regions in six animal species, generating 271, 119, 860, 574, 751. and 210 base pairs (bp) long fragments for beef, mutton, pork, chicken, duck, and mouse, respectively. The six pairs of species-specific primers mixture was used to analyze mixed DNA samples. Ingredients of meat products were identified according to the size of the bands which were amplified with specific primers. Based on the performance of serial specific verification tests, the multiplex PCR assay showed high specificity to the corresponding species, and no cross-reactivity with the other five animals. Detection limits for DNA samples from the six animal species varied between 0.01 and 0.001 ng in a 20 μL PCR mixture. Seven processed meat products were successfully identified via the multiplex PCR assay. This study provides a simple, rapid, sensitive, specific and effective molecular technique for meat authentication. It can be directly applied to differentiate between beef, mutton and four more species in animal-derived foods.