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Evaluation of Real-time PCR Based on SYBR Green I Fluorescent Dye for Detection of Bacillus Anthracis Strains in Biological Samples
INTRODUCTION: The aim of the study was the application and evaluation of real-time PCRs based on the fluorescence of SYBR Green I intercalating dye for the detection of three Bacillus anthracis genes in contaminated liver and blood samples. The goals for detection were rpoB gene as a chromosomal mar...
Autores principales: | , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Sciendo
2018
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6364171/ https://www.ncbi.nlm.nih.gov/pubmed/30729215 http://dx.doi.org/10.2478/jvetres-2018-0075 |
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author | Kędrak-Jabłońska, Agnieszka Budniak, Sylwia Szczawińska, Anna Reksa, Monika Krupa, Marek Szulowski, Krzysztof |
author_facet | Kędrak-Jabłońska, Agnieszka Budniak, Sylwia Szczawińska, Anna Reksa, Monika Krupa, Marek Szulowski, Krzysztof |
author_sort | Kędrak-Jabłońska, Agnieszka |
collection | PubMed |
description | INTRODUCTION: The aim of the study was the application and evaluation of real-time PCRs based on the fluorescence of SYBR Green I intercalating dye for the detection of three Bacillus anthracis genes in contaminated liver and blood samples. The goals for detection were rpoB gene as a chromosomal marker, pag gene located on plasmid pXO1, and capC gene located on plasmid pXO2. MATERIAL AND METHODS: Five B. anthracis strains were used for the experiments. Additionally, single strains of other species of the genus Bacillus, i.e. B. cereus, B. brevis, B. subtilis, and B. megaterium, and strains of six other species were used for evaluation of the specificity of the tests. Three SYBR Green I real-time PCRs were conducted allowing confirmation of B. anthracis in the biological samples. RESULTS: The observation of amplification curves in real-time PCRs enabled the detection of the chromosomally encoded rpoB gene, pag gene, and capC gene of B. anthracis. The specificity of the tests was confirmed by estimation of the melting temperature of the PCR products. The sensitivity and linearity of the reactions were determined using regression coefficients. Strains of other microbial species did not reveal real-time PCR products. CONCLUSION: All real-time PCRs for the detection of B. anthracis in biological samples demonstrated a significant sensitivity and high specificity. |
format | Online Article Text |
id | pubmed-6364171 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | Sciendo |
record_format | MEDLINE/PubMed |
spelling | pubmed-63641712019-02-06 Evaluation of Real-time PCR Based on SYBR Green I Fluorescent Dye for Detection of Bacillus Anthracis Strains in Biological Samples Kędrak-Jabłońska, Agnieszka Budniak, Sylwia Szczawińska, Anna Reksa, Monika Krupa, Marek Szulowski, Krzysztof J Vet Res Research Article INTRODUCTION: The aim of the study was the application and evaluation of real-time PCRs based on the fluorescence of SYBR Green I intercalating dye for the detection of three Bacillus anthracis genes in contaminated liver and blood samples. The goals for detection were rpoB gene as a chromosomal marker, pag gene located on plasmid pXO1, and capC gene located on plasmid pXO2. MATERIAL AND METHODS: Five B. anthracis strains were used for the experiments. Additionally, single strains of other species of the genus Bacillus, i.e. B. cereus, B. brevis, B. subtilis, and B. megaterium, and strains of six other species were used for evaluation of the specificity of the tests. Three SYBR Green I real-time PCRs were conducted allowing confirmation of B. anthracis in the biological samples. RESULTS: The observation of amplification curves in real-time PCRs enabled the detection of the chromosomally encoded rpoB gene, pag gene, and capC gene of B. anthracis. The specificity of the tests was confirmed by estimation of the melting temperature of the PCR products. The sensitivity and linearity of the reactions were determined using regression coefficients. Strains of other microbial species did not reveal real-time PCR products. CONCLUSION: All real-time PCRs for the detection of B. anthracis in biological samples demonstrated a significant sensitivity and high specificity. Sciendo 2018-12-31 /pmc/articles/PMC6364171/ /pubmed/30729215 http://dx.doi.org/10.2478/jvetres-2018-0075 Text en © 2018 A. Kędrak-Jabłońska et al. published by Sciendo http://creativecommons.org/licenses/by-nc-nd/3.0 This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License. |
spellingShingle | Research Article Kędrak-Jabłońska, Agnieszka Budniak, Sylwia Szczawińska, Anna Reksa, Monika Krupa, Marek Szulowski, Krzysztof Evaluation of Real-time PCR Based on SYBR Green I Fluorescent Dye for Detection of Bacillus Anthracis Strains in Biological Samples |
title | Evaluation of Real-time PCR Based on SYBR Green I Fluorescent Dye for Detection of Bacillus Anthracis Strains in Biological Samples |
title_full | Evaluation of Real-time PCR Based on SYBR Green I Fluorescent Dye for Detection of Bacillus Anthracis Strains in Biological Samples |
title_fullStr | Evaluation of Real-time PCR Based on SYBR Green I Fluorescent Dye for Detection of Bacillus Anthracis Strains in Biological Samples |
title_full_unstemmed | Evaluation of Real-time PCR Based on SYBR Green I Fluorescent Dye for Detection of Bacillus Anthracis Strains in Biological Samples |
title_short | Evaluation of Real-time PCR Based on SYBR Green I Fluorescent Dye for Detection of Bacillus Anthracis Strains in Biological Samples |
title_sort | evaluation of real-time pcr based on sybr green i fluorescent dye for detection of bacillus anthracis strains in biological samples |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6364171/ https://www.ncbi.nlm.nih.gov/pubmed/30729215 http://dx.doi.org/10.2478/jvetres-2018-0075 |
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