Modern on-site tool for monitoring contamination of halal meat with products from five non-halal animals using multiplex polymerase chain reaction coupled with DNA strip

Meat adulteration is a persistent problem in the food industry in many countries. This unfair practice can potentially not only affect the economy of a country but also the confidence and health of its citizens. Additionally, it can aggravate allergies or damage religious beliefs (e.g., Muslims). The aim of this study was to develop a simple and rapid technique for the on-site inspection of several targets using multiplex polymerase chain reaction (PCR) coupled with DNA strip for detecting contamination with products from five non-halal animals (monkeys, dogs, rats, pigs, and cats) in foods certified to be halal. Species-specific primers were retrieved from the mitochondrial genes of each non-halal animal and modified with a tag sequence and biotin. The selected primers showed high specificity for each non-halal animal. The PCR products obtained after amplification were detected using DNA strip. The limit of detection ranged from 0.01 ppm to 1 ppm (dogs, 1 ppm; cats, 0.1 ppm; pigs, 0.01 ppm; monkeys, 0.01 ppm; and rats, 0.01 ppm). Beef meatballs spiked with non-halal meat in different ratios were prepared to validate the developed technique, which presented 100% accuracy in detection. In addition, the developed technique was applied to verify 115 samples of meat products commercially available in Thailand. Sixteen samples were contaminated with pig DNA, but contamination with the DNA of other non-halal animals was not found. Therefore, this technique has high potential for on-site monitoring of non-halal animal contaminants in halal products in Thailand.