Detection of pork derivatives in meats and suspicious highly processed foods by Real-time PCR

Detection of the residual meat or other components of pork in halal food products particularly in Muslim countries is of prime importance. In this study, real-time polymerase chain reaction based on SyberGreen dye was used for detection of pork DNA in meats and highly processed products such as pork gelatin and gelatin-containing products. SyberGreen based real-time polymerase chain reaction was designed for an assay that can detect pork DNA in meats and highly processed pork derivatives including gelatin and gelatin containing products. The proposed method uses the newly specific primers designed by AlleleID7 software that amplify a 114 bp fragment of porcine mitochondrial D-loop region. After optimization of mixture composition and thermal program for achieving maximum fluorescence and minimum threshold cycle, standard curve of serially diluted DNA extracted from pork meat showed power for detection of 0.001 diluted DNA compared with starting DNA material with the efficiency of 92%. Specificity of this method was evaluated by testing the extracted DNA from sheep, cow, chicken and fish which had no amplification for negative control samples. To analyze the method for detecting highly processed pork derivatives, several pork gelatin powders and sheets were selected and the real-time polymerase chain reaction based on SyberGreen dye proved to successfully detect the traces of the pork residues in the samples tested.