Development of Duplex Real‐time Polymerase Chain Reaction for Simultaneous Detection of Oilfish and Escolar derived Components

Background Oilfish (Ruvettus pretiosus) and escolar (Lepidocybium flavobrunneum) are often used to adulterate high-value aquatic products, causing serious economic losses to the consumers and even keriorrhea in severe cases. It was difficult to identify them by morphological features for these two fish were processed into steak or fillet. The purpose of this study is therefore to develop an accurate and efficient method for detecting the oilfish and escolar derived components. Results By comparing the mitochondrial 16S rRNA gene sequences of oilfish, escolar and 16 varieties susceptible to adulteration, specific primers/probes were designed, and a duplex real-time PCR method was established to detect oilfish and escolar-derived components. The specificity and sensitivity of the method were analyzed, and the method was used to analyze 70 commercial samples to evalutate its applicability to actual samples in the market. The method developed was highly specific, without any cross-reaction on other 16 species, with 0.0002 ng μL-1 (R. pretiosus) and 0.002 ng μL-1 (L. flavobrunneum) of limit of detection (LOD) for DNA. In addition, the LOD for mixed muscle tissues was 0.1 g kg-1 . Oilfish and escolar-derived components were detected in 12 of the 70 commercial samples, this result was consistent with the classic DNA barcoding test results. Conclusion The developed duplex real-time PCR method can be used to detect oilfish and escolar specifically, sensitively and accurately; it provides a good technical support for the identification of authentic aquatic products. This article is protected by copyright. All rights reserved.