Real-Time PCR-based detection of bovine DNA by specific targeting on cytochrome-B
The design of specific primers is an interesting research topic such that it offers selective, specific, and effective DNA analysis using real-time PCR. This research was intended to detect bovine DNA using real-time PCR and specific primers to ensure the halal authenticity of food products. Primers...
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Universitas Ahmad Dahlan
2019-11-01
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doaj-85601498ab3b43d6974d930c0eee9f7c2021-09-20T06:40:22ZengUniversitas Ahmad DahlanPharmaciana2088-45592477-02562019-11-019220121010.12928/pharmaciana.v9i2.14070Real-Time PCR-based detection of bovine DNA by specific targeting on cytochrome-BNina Salamah0Yuny Erwanto1Sudibyo Martono2Abdul Rohman3Department of Analytical Chemistry, Faculty of Pharmacy, Universitas Ahmad DahlanDivision of Animal Products Technology, Faculty of Animal Science, Universitas Gadjah MadaDepartment of Pharmaceutical Chemistry, Faculty of Pharmacy, Universitas Gadjah MadaDepartment of Pharmaceutical Chemistry, Faculty of Pharmacy, Universitas Gadjah MadaThe design of specific primers is an interesting research topic such that it offers selective, specific, and effective DNA analysis using real-time PCR. This research was intended to detect bovine DNA using real-time PCR and specific primers to ensure the halal authenticity of food products. Primers of bovine DNA sequences were designed in the NCBI and Primer-BLAST programs. The outcome validation was assessed using several parameters, namely specificity, repeatability, and linearity by real-time PCR. Primer specificity test was performed on fresh tissue (pork and negative control), while the repeatability test used six replications and was based on the calculated coefficient of variation (CV). In the linearity test, six different DNA concentrations (50000, 10000, 5000, 500, 100, and 50 pg/μL) were examined to obtain the efficiency value. Using the specific primer from Cytochrome-B, the real-time PCR could specifically identify the presence of bovine DNA at the optimum annealing temperature of 58.70C. The repeatability analysis yielded a coefficient of variation (CV) of 0.57 %, while the linearity test produced an efficiency value of 206 %. These figures confirm that the method employed in this study is not only specific but also sensitive and reliable for detecting bovine DNA. Real-time PCR using specific primer targeting on the cytochrome-B region of bovine DNA (forward: CTACTGACACTCACATGAATTGG; reverse CACTAGGATGAGGAGAAAGTATAGG) can be used to identify bovine DNA and distinguish it from porcine DNA.http://journal.uad.ac.id/index.php/PHARMACIANA/article/view/14070/pdf_120primercytochrome-bbovine dnaspecificreal-time pcr |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Nina Salamah Yuny Erwanto Sudibyo Martono Abdul Rohman |
spellingShingle |
Nina Salamah Yuny Erwanto Sudibyo Martono Abdul Rohman Real-Time PCR-based detection of bovine DNA by specific targeting on cytochrome-B Pharmaciana primer cytochrome-b bovine dna specific real-time pcr |
author_facet |
Nina Salamah Yuny Erwanto Sudibyo Martono Abdul Rohman |
author_sort |
Nina Salamah |
title |
Real-Time PCR-based detection of bovine DNA by specific targeting on cytochrome-B |
title_short |
Real-Time PCR-based detection of bovine DNA by specific targeting on cytochrome-B |
title_full |
Real-Time PCR-based detection of bovine DNA by specific targeting on cytochrome-B |
title_fullStr |
Real-Time PCR-based detection of bovine DNA by specific targeting on cytochrome-B |
title_full_unstemmed |
Real-Time PCR-based detection of bovine DNA by specific targeting on cytochrome-B |
title_sort |
real-time pcr-based detection of bovine dna by specific targeting on cytochrome-b |
publisher |
Universitas Ahmad Dahlan |
series |
Pharmaciana |
issn |
2088-4559 2477-0256 |
publishDate |
2019-11-01 |
description |
The design of specific primers is an interesting research topic such that it offers selective, specific, and effective DNA analysis using real-time PCR. This research was intended to detect bovine DNA using real-time PCR and specific primers to ensure the halal authenticity of food products. Primers of bovine DNA sequences were designed in the NCBI and Primer-BLAST programs. The outcome validation was assessed using several parameters, namely specificity, repeatability, and linearity by real-time PCR. Primer specificity test was performed on fresh tissue (pork and negative control), while the repeatability test used six replications and was based on the calculated coefficient of variation (CV). In the linearity test, six different DNA concentrations (50000, 10000, 5000, 500, 100, and 50 pg/μL) were examined to obtain the efficiency value. Using the specific primer from Cytochrome-B, the real-time PCR could specifically identify the presence of bovine DNA at the optimum annealing temperature of 58.70C. The repeatability analysis yielded a coefficient of variation (CV) of 0.57 %, while the linearity test produced an efficiency value of 206 %. These figures confirm that the method employed in this study is not only specific but also sensitive and reliable for detecting bovine DNA. Real-time PCR using specific primer targeting on the cytochrome-B region of bovine DNA (forward: CTACTGACACTCACATGAATTGG; reverse CACTAGGATGAGGAGAAAGTATAGG) can be used to identify bovine DNA and distinguish it from porcine DNA. |
topic |
primer cytochrome-b bovine dna specific real-time pcr |
url |
http://journal.uad.ac.id/index.php/PHARMACIANA/article/view/14070/pdf_120 |
work_keys_str_mv |
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