Validating duplex-PCR targeting ND2 for bovine and porcine detection in meat products

Food authentication is a mandatory effort to assure the fair-trade. This study developed a duplex polymerase chain reaction (PCR) from the NADH dehydrogenase subunit 2 (ND2) gene to amplify specific segments of a cattle and porcine DNA. A universal forward primer composed of nineteen base pairs (bp) (3 & PRIME;-CCAAACA-CAACTCCGAAAA-5 & PRIME;) and species-specific reverse primers composed of twenty (3 & PRIME;-CCAAACACAACTCCGAAAA-5 & PRIME;) and twenty-one (3 & PRIME;-TGGCAAGAATTAGGACGGTTA-5 & PRIME;) bp were used to limit the amplified DNA segment for porcine and cattle. The PCR reaction would generate a product with a profile of 168 and 227 bp, respectively. To investigate the accuracy and limit of detection, an in vitro experiment was conducted using simplex and duplex PCR on commercial meatballs randomly purchased from a commercial market in Surakarta, Indonesia. The findings of this study indicated that ND2 could be used as an alternative genetic marker for the identification of porcine and beef species in meat-derived products.