Identification of Meat Species by PCR-RFLP Method using Single Set of Degenerative Primers

Abstract

Electrophoresis, chromatography and enzyme-linked immunoassay are common analytical techniques (Hsieh, 2000;Floren et al., 2015). Different muscle proteins are separated by electrophoresis, but the requirement of baseline data and uniformity of protein patterns make their results elusive (Ilhak and Arslan, 2007). Chromatographic techniques differentiate among origin of meat by their fatty acid and protein composition (Chou et al., 2007), enzyme-linked immunoassay by reactivity of specific antibodies with Abstract | Due to religious and social concerns, identification of meat and meat products is a matter of great interest for general public. DNA-based identification methods are ideal and reliable. DNA was extracted from muscle tissues of cow, buffalo, goat, donkey and a dog. A set of degenerative primers was designed for Polymerase Chain Reaction (PCR) of the cytochrome b gene fragment. All PCR was accomplished at the same reaction conditions. Amplified products had a length of 400 bp, which was our estimated length. After the sequencing of these amplicons, In silico restriction analysis was performed for each specie. Restriction endonuclease TfiI was selected for further analysis. Restriction fragment length polymorphism (RFLP) was performed at the same reaction conditions for all species. RFLP analysis produced species specific restriction profiles. Comparing these profiles, we have identified the species origin of meat unambiguously.

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

“…Furthermore, the developed direct PCR method is coupled with RFLP assay which is much more desirable for molecular based meat identification, especially, in cases where large number of samples have to be processed as it does not require DNA sequencing and/or specialized equipment and reduces the cost and time for post-PCR processing of samples (Guan et al, 2019;Gargouri et al, 2021;Vaithiyanathan et al, 2021;Taha et al, 2021). It has been mostly applied for species discrimination in processed and unprocessed meat products because of its simplicity, quicker detection and reduced cost (Al et al, 2020;Asghar et al, 2022;Farag et al, 2022).…”

A systematic review of DNA‐based methods in authentication of game and less common meat species

“…In Islamic communities, eating donkey meat is forbidden, and mislabeling donkey meat as halal (permissible) is considered a heinous offense (Al- . Despite this, there are still cases of adulterated donkey in beef products in countries such as Pakistan (Asghar et al, 2021) and Tunisia (Gargouri et al, 2021). Zhao et al (2019b) tested the applicability of their developed authentication method on 46 commercial samples labeled as donkey burgers and found that four products contained horse meat and no donkey meat.…”

“…In Islamic communities, eating donkey meat is forbidden, and mislabeling donkey meat as halal (permissible) is considered a heinous offense (Al‐Ali & A, 2019). Despite this, there are still cases of adulterated donkey in beef products in countries such as Pakistan (Asghar et al., 2021) and Tunisia (Gargouri et al., 2021). Zhao et al.…”

Optimizing PCR using mtDNA Universal Primers for Detecting Indigenous Domestic Animal Species from Pakistan

“…Pakistan is facing drastic peak in meat adulteration cases in past few years. The scenario clearly projects non-compliance of halal food laws [4]. This concrns the public health as sometimes edible and inedible meat can elicit the allergies, metabolic disorders and infectious diseases specially in sensitive and immunocompromised individuals [5].…”