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The employment of q-PCR using specific primer targeting on mitochondrial cytochrome-b gene for identification of wild boar meat in meatball samples

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) i...

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Autores principales: Aina, Ganea Qorry, Erwanto, Yuny, Hossain, Motalib, Johan, Mohd Rafie, Ali, Md. Eaqub, Rohman, Abdul
Formato: Online Artículo Texto
Lenguaje:English
Publicado: A periodical of the Network for the Veterinarians of Bangladesh (BDvetNET) 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6760509/
https://www.ncbi.nlm.nih.gov/pubmed/31583226
http://dx.doi.org/10.5455/javar.2019.f348
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author Aina, Ganea Qorry
Erwanto, Yuny
Hossain, Motalib
Johan, Mohd Rafie
Ali, Md. Eaqub
Rohman, Abdul
author_facet Aina, Ganea Qorry
Erwanto, Yuny
Hossain, Motalib
Johan, Mohd Rafie
Ali, Md. Eaqub
Rohman, Abdul
author_sort Aina, Ganea Qorry
collection PubMed
description 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.
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spelling pubmed-67605092019-10-03 The employment of q-PCR using specific primer targeting on mitochondrial cytochrome-b gene for identification of wild boar meat in meatball samples Aina, Ganea Qorry Erwanto, Yuny Hossain, Motalib Johan, Mohd Rafie Ali, Md. Eaqub Rohman, Abdul J Adv Vet Anim Res Short Communication 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. A periodical of the Network for the Veterinarians of Bangladesh (BDvetNET) 2019-07-06 /pmc/articles/PMC6760509/ /pubmed/31583226 http://dx.doi.org/10.5455/javar.2019.f348 Text en Copyright: © Journal of Advanced Veterinary and Animal Research http://creativecommons.org/licenses/by-nc/4.0 This is an open-access article distributed under the terms of the Creative Commons Attribution-Noncommercial-Share Alike 4.0 Unported, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Short Communication
Aina, Ganea Qorry
Erwanto, Yuny
Hossain, Motalib
Johan, Mohd Rafie
Ali, Md. Eaqub
Rohman, Abdul
The employment of q-PCR using specific primer targeting on mitochondrial cytochrome-b gene for identification of wild boar meat in meatball samples
title The employment of q-PCR using specific primer targeting on mitochondrial cytochrome-b gene for identification of wild boar meat in meatball samples
title_full The employment of q-PCR using specific primer targeting on mitochondrial cytochrome-b gene for identification of wild boar meat in meatball samples
title_fullStr The employment of q-PCR using specific primer targeting on mitochondrial cytochrome-b gene for identification of wild boar meat in meatball samples
title_full_unstemmed The employment of q-PCR using specific primer targeting on mitochondrial cytochrome-b gene for identification of wild boar meat in meatball samples
title_short The employment of q-PCR using specific primer targeting on mitochondrial cytochrome-b gene for identification of wild boar meat in meatball samples
title_sort employment of q-pcr using specific primer targeting on mitochondrial cytochrome-b gene for identification of wild boar meat in meatball samples
topic Short Communication
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6760509/
https://www.ncbi.nlm.nih.gov/pubmed/31583226
http://dx.doi.org/10.5455/javar.2019.f348
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