The employment of q-PCR using specific primer targeting on mitochondrial cytochrome-b gene for identification of wild boar meat in meatball samples
Document Type
Article
Publication Date
1-1-2019
Abstract
Objective: The objective of this study was to employ real-time or quantitative polymerase chain reaction (q-PCR) using novel species specific primer (SSP) targeting on mitochondrial cytochrome-b of wild boar species (CYTBWB2-wb) gene for the identification of non-halal meat of wild boar meat (WBM) in meatball products. Materials and Methods: The novel SSP of CYTBWB2-wb was designed by our group using PRIMERQUEST and NCBI software. DNA was extracted using propanol-chloroform-isoamyl alcohol method. The designed SSP was further subjected for validation protocols using DNA isolated from fresh meat and from meatball, which include specificity test, determination of efficiency, limit of detection and repeatability, and application of developed method for analysis of commercially meatball samples Results: The results showed that CYTBWB2-wb was specific to wild boar species against other animal species with optimized annealing temperature of 59°C. The efficiency of q-PCR obtained was 91.9% which is acceptable according to the Codex Allimentarius Commission (2010). DNA, with as low as 5 pg/μl, could be detected using q-PCR with primer of CYTBWB2-wb. The developed method was also used for DNA analysis extracted from meatball samples commercially available. Conclusion: q-PCR using CYTBWB2-wb primers targeting on mitochondrial cytochrome-b gene (forward: CGG TTC CCT CTT AGG CAT TT; Reverse: GGA TGA ACA GGC AGA TGA AGA) can be fruitfully used for the analysis of WBM in commercial meatball samples. © 2019 Network for the Veterinarians of Bangladesh.
Keywords
CYTBWB2-wb primer, Halal authentication, Meatball, Q-PCR, Wild boar
Divisions
nanotechnology
Funders
Kementerian Riset dan Pendidikan Tinggi (Kemenristek Dikti) through research scheme of Penelitian Unggulan Perguruan Tinggi year 2017-2018 with contract number of 185/UN1/DITLIT/DIT-LIT/LT/2018
Publication Title
Journal of Advanced Veterinary and Animal Research
Volume
6
Issue
3
Publisher
Network for the Veterinarians of Bangladesh