Highly sensitive detection of soybean-derived DNA in soy sauce using an integrated PCR-CRISPR/Cas12a nucleic acid detection platform

Introduction Soy sauce is a traditional fermented condiment with widespread global consumption, and its authenticity depends on the requisite use of soybeans as a primary ingredient. However, for economic gain, soybean has been partially or completely substituted with cheaper materials, including other legumes such as pea or peanut, or non‐soybean nitrogen sources such as acid‐hydrolysed animal protein derived from feathers or hair. Such practices constitute economic fraud and labelling violations and may introduce undeclared allergens, raising legitimate concerns regarding food safety and consumer trust. Conventional DNA‐based detection techniques are often hindered by DNA fragmentation and the presence of PCR inhibitors in the complex, fermented matrix, leading to compromised sensitivity and reliability. Methods To overcome these limitations, we developed a novel nucleic acid detection platform that synergistically combines target pre‐amplification via polymerase chain reaction (PCR) with highly specific sequence recognition and signal amplification utilizing the CRISPR/Cas12a system. Results This integrated approach achieved a detection limit of 10 femtomolar (fM) for a synthetic Le1 target DNA standard, which compares favourably with many existing methods. The practical applicability of this assay was preliminarily evaluated on 22 commercially available soy sauce samples from various markets. Discussion This study establishes a sensitive and reproducible endpoint‐fluorescence tool for the detection of soybean‐derived DNA. The assay holds promise for routine food authentication and for supporting regulatory compliance and the detection of economic fraud within the global food industry.