In order to establish a real-time recombinase aided amplification (Rt-RAA) method to rapidly detect dog-derived ingredients, three pairs of primers and one exonuclease (exo) probe were designed targeting the conserved region of the cytochrome B (cytb) gene. The designed primer combinations and probe concentration were optimized. The specificity and stability of the method were evaluated by cross-reactivity and repeatability experiments. The sensitivity of the Rt-RAA method and its results for simulated mixed samples and commercial samples were compared with those of the national standard method. The results showed that the Rt-RAA method had strong specificity and good stability and could detect the target sequence in less than 30 min. The detection limit for dog-derived ingredients was 0.2%, which was significantly better than that of real-time fluorescence polymerase chain reaction (PCR). The results of the Rt-RAA method for simulated mixed samples and commercial samples were completely consistent with those of the national standard method. Therefore, Rt-RAA is suitable for the detection of dog-derived ingredients, providing a fast and accurate new method for the identification of meat-derived ingredients.

Digital polymerase chain reaction (dPCR) is a novel nucleic acid amplification technology that enables absolute quantitative analysis of nucleic acid at the single-molecule level through extreme dilution and Poisson distribution statistics without any need for internal reference genes or standard curves. It has the advantages of high sensitivity, accuracy and tolerance, and has great development potential in the field of food safety detection. This paper introduces the technical principle, advantages and disadvantages, and commercial platforms of dPCR, and summarizes recent progress in the application of dPCR in the fields of foodborne pathogen detection, animal- and plant-derived component detection, genetically modified component detection, and foodborne virus detection and the current status of the standardization of dPCR. The problems existing in this technology and possible future development directions are discussed. We hope that this review can provide ideas for promoting the application of dPCR in food safety detection and formulating a standard for dPCR.