Development and Application of a Polymerase Spiral Reaction (PSR)-Based Isothermal Assay for Rapid Detection of Yak...

The growing demand for robust food authentication methods has driven the establishment of fast, sensitive, and field-based detection systems for identifying meat species. This study presents a colorimetric-based PSR approach for identifying yak (Bos grunniens) meat within fresh, thermally processed, and blended meat samples. Targeting the mitochondrial D-loop locus, the assay incorporates a simple alkaline lysis (AL) procedure for efficient DNA extraction, eliminating the requirement for specialized instrumentation. The PSR assay demonstrated high specificity, showing no evidence of cross-reactivity with closely associated food animals such as buffalo, cattle, goat, sheep, mithun, and pig. Sensitivity assessment revealed the assay's capability to detect 1 pg of yak DNA, with reliable performance in samples exposed to thermal conditions up to 121 °C. Additionally, the technique detected yak meat down to a concentration of 0.1% in binary beef mixtures. This method provides a significant improvement in sensitivity over end-point PCR and is particularly well-suited for field applications due to its practical simplicity, affordability, as well as no reliance on sophisticated instrument. This is, to the best of our understanding, the first reported PSR-based approach developed for the identification of yak meat, offering a robust tool for food origin verification, regulatory enforcement, and product integrity monitoring.