Multiplex PCR and 12S rRNA gene sequencing for detection of meat adulteration: A case study in the Egyptian markets

•Meat adulteration has become a critical case, worldwide.•Frozen liver samples were correctly identified through 12S rRNA gene sequencing.•Multiplex PCR revealed that processed beef samples contained poultry contaminants.•Sequencing and mPCR are sensitive methodologies for controlling meat industry. Recently, DNA-based methods have proved to be accurate, fast and sensitive for meat authentication. According to the European Union, the food safety standards require accurate and detailed composition information of the meat products. Therefore, an accurate, fast and cost-effective identification methodology is needed. In this study, multiplex PCR coupled with 12S rDNA sequencing was employed for the detection of meat adulteration in two red meat products (frozen beef liver and cold cut samples, respectively) in Egypt. Multiplex PCR allowed the identification of ruminant, poultry, pork, and donkey residuals in processed red meat products (cold cuts) in a single step PCR reaction. Preliminary uniplex PCR was performed to evaluate primers specificity using DNA extracted from the positive control samples. The primers produced specific fragments for ruminant, poultry, pork, and donkey as follows: 271, 183, 531 and 145 bp, respectively. Multiplex PCR revealed that none of the samples was contaminated by porcine or donkey residuals, but 62.5% of all tested processed beef samples contained poultry contaminants. The sensitivity of this method was 0.01 ng/μL for beef, poultry and donkey and 0.1 ng/μL for pig. Another promising finding is the identification of all frozen beef liver samples as a cattle species (Bos taurus) through PCR-sequencing of a short fragment of 12S rRNA gene. Finally, we recommend the employment of multiplex PCR and PCR-sequencing of 12S rDNA for quality control in routine analysis of processed and frozen meat products.