Development of Bovine-Derived β-Actin Genomic DNA Reference Materials for Meat Adulteration

Meat quality and authenticity are critical food safety issues, especially for processed meat products prone to species mislabeling and adulteration. Accurate molecular identification of beef relies on species-specific DNA markers, and nucleic acid reference materials (RMs) serve as the "yardstick" in molecular metrology. However, the lack of single-copy gene DNA RMs has limited detection accuracy and reliability. To address this, we developed and validated certified reference materials (CRMs) for identifying beef in food products using bovine genomic DNA. The β-actin gene in beef was detected by digital PCR to establish these CRMs. The certified values of β-actin in the CRMs were determined to be (6.85 ± 0.63) × 10³ copies/μL and (8.00 ± 0.77) × 10² copies/μL, with extended uncertainty (coverage factor k = 2), based on β-actin quantification by 9 independent collaborating laboratories. Furthermore, these CRMs were used to evaluate the standard suitability, analyze quantitative deviations between single-copy gene and multi-gene approaches, and test with real samples. These efforts provide robust support for expanding the national catalog of animal food ingredient CRMs and enhancing detection accuracy.