Establishment and Application of Recombinase-Mediated Strand Displacement Isothermal Amplification Assay for Rapid Detection of Horse-Derived Components in Meat Products

Abstract

Objective: A recombinase-mediated strand displacement isothermal amplification assay for rapid detection of horse-derived components in meat and meat products was established. Methods: A series of specific primers and Exo probes were designed using the horse-derived ATpase 6 gene as the target gene. The primers were screened and the reaction parameters were optimized. The specificity, sensitivity and stability of the assay were evaluated, and the detection limit, applicability and accuracy were analyzed by detecting simulated samples with different mixing ratios and different processing technologies, and commercial samples. Results: The assay was characterized by rapid response, high sensitivity and specificity. The reaction was completed within 16 minutes under a constant temperature of 39 ℃. The system had good specificity to 23 non-target sources. The detection sensitivity of the target DNA was 1.8 copies/μL. The detection limit was 0.01% for raw meat (mass fraction), and 0.1% (mass fraction) for processed meat products. For 90 commercial samples, the results of this method were consistent with those of the standard method. Conclusion: The recombinase-mediated isothermal amplification assay can be used for the detection of horse-derived components in meat and meat products.

Keywords

• recombinase aided chain displacement isothermal amplification; adulteration identification; horse-derived component; meat and meat products