Potential authentication of various meat‐based products using simple and efficient DNA extraction method

Abstract

BACKGROUND: The growth of halal food consumption worldwide has resulted in an increase in the request for halal authentication. DNA-based detection using powerful real-time polymerase chain reaction (PCR) technique has been shown to be highly specific and sensitive authentication tool. The efficient DNA extraction method in terms of quality and quantity is a backbone step to obtain successful real-time PCR assays. In this study, different DNA extraction methods using three lysis buffers were evaluated and developed to recommend a much more efficient method as well as achieve a successful detection using real-time PCR.RESULTS: The lysis buffer 2 (LB2) has been shown to be the best lysis buffer for DNA extraction from both raw and processed meat samples comparing to other lysis buffers tested. Hence, the LB2 has been found to be ideal to detect meat and porcine DNAs by real-time PCR using pairs of porcine specific primers and universal primers which amplified at 119 bp fragment and 93 bp fragment, respectively. This assay allows detection as low as 0.0001 ng of DNA. Higher efficiency and sensitivity of real-time PCR via a simplified DNA extraction method using LB2 have been observed, as well as a reproducible and high correlation coefficient (R 2 = 0.9979) based on the regression analysis of the standard curve have been obtained.CONCLUSION: This study has established a fast, simple, inexpensive and efficient DNA extraction method that is feasible for raw and processed meat products. This extraction technique allows an accurate DNA detection by real-time PCR and can also be implemented to assist the halal authentication of various meat-based products available in the market.

Comparison of different DNA preparatory methods for development of a universal direct PCR-RFLP workflow for identification of meat origin in food products

“…The efficiency of species identification assays mostly relies on two main components; first, the extracted genomic DNA and second, the selected genomic region for identification. With time several methods have been developed either by following different PCR approaches (Song et al, 2018;Skouridou et al, 2019;Mokhtar et al, 2020;Batule et al, 2020;Yan et al, 2022) or by varying the genomic regions (Marchetti et al, 2020;Suryawan et al, 2020;Li et al, 2021;Tao et al, 2022) or both. Among such attempts, the direct PCR approach allows PCR amplification without prior DNA extraction.…”

“…It is well-acknowledged that extraction of genomic DNA is an imperative step to ensure amplifiable DNA template (Mokhtar et al, 2020). However, conventional DNA extraction protocols for molecular meat testing are complicated, labor-intensive, time-consuming and expensive (Besbes et al, 2022;Sajali et al, 2018;Yue & Orban, 2001).…”

A Simple and Reliable Single Tube Septuple PCR Assay for Simultaneous Identification of Seven Meat Species

“…Meat authentication is an important concern to protect consumers from illegal and unwanted ingredients [ 1 , 2 , 3 , 4 ]. However, meat adulteration such as unlisted, mislabeled or fraudulent ingredients has frequently been reported around the world and has become a severe global issue [ 4 , 5 ]. Although some laws have been enacted for ensuring the quality and safety of meat products, adulteration is still widespread due to the purpose of economic pursuit [ 1 , 6 ].…”

“…Many analytical methods of biological, immunological, physical, chemical, anatomical and histological analyses have been developed [ 2 , 3 , 4 , 7 , 12 ]. Among them, protein-based methods are widely used to identify meat species in composite mixtures [ 5 , 13 ]. Proteins are likely to be degraded, denatured or damaged in processed meat products, limiting the accuracy of the identification of meat species in thermally treated foods [ 13 , 14 ].…”

“…Proteins are likely to be degraded, denatured or damaged in processed meat products, limiting the accuracy of the identification of meat species in thermally treated foods [ 13 , 14 ]. In comparison, DNA-based analytical methods coupled with polymerase chain reaction (PCR) present a reliable alternative to protein-based methods in the discrimination and mislabeling detection of meat species, as DNA molecules possess high stability and are present in every type of cell [ 4 , 5 ]. Both multiplex and real-time PCR techniques are highly specific and efficient in the identification of meat adulteration [ 15 ].…”