Rapid Detection of Dog-Derived Ingredients by Real-Time Recombinase-Aided Amplification

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.