<abstract xmlns="http://www.w3.org/1999/xhtml"><bold>Introduction:</bold> 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 <italic>Bacillus anthracis</italic> genes in contaminated liver and blood samples. The goals for detection were <italic>rpoB</italic> gene as a chromosomal marker, <italic>pag</italic> gene located on plasmid pXO1, and <italic>capC</italic> gene located on plasmid pXO2.<bold>Material and Methods:</bold> Five <italic>B. anthracis</italic> strains were used for the experiments. Additionally, single strains of other species of the genus <italic>Bacillus</italic>, <italic>i.e</italic>. <italic>B. cereus</italic>, <italic>B. brevis</italic>, <italic>B. subtilis</italic>, and <italic>B. megaterium</italic>, 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 <italic>B. anthracis</italic> in the biological samples.<bold>Results:</bold> The observation of amplification curves in real-time PCRs enabled the detection of the chromosomally encoded <italic>rpoB</italic> gene, <italic>pag</italic> gene, and <italic>capC</italic> gene of <italic>B. anthracis</italic>. 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.<bold>Conclusion:</bold> All real-time PCRs for the detection of <italic>B. anthracis</italic> in biological samples demonstrated a significant sensitivity and high specificity.</abstract>

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.

Evaluation of real-time PCR based on SYBR Green I fluorescent dye for detection of <italic>Bacillus anthracis</italic> strains in biological samples

Department of Microbiology, National Veterinary Research Institute

Journal of Veterinary Research

This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License.

Evaluation of real-time PCR based on SYBR Green I fluorescent dye for detection of  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...

Evaluation of real-time PCR based on SYBR Green I fluorescent dye for detection of Bacillus anthracis strains in biological samples

Publicado en línea: 31 dic 2018

Páginas: 549 - 554

Recibido: 09 oct 2018

Aceptado: 07 dic 2018

DOI: https://doi.org/10.2478/jvetres-2018-0075

Palabras clave
real-time PCR, SYBR Green I

© 2018 Agnieszka Kędrak-Jabłońska et al., published by Sciendo

This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License.

Evaluation of real-time PCR based on SYBR Green I fluorescent dye for detection of Bacillus anthracis strains in biological samples

Publicado en línea: 31 dic 2018

Páginas: 549 - 554

Recibido: 09 oct 2018

Aceptado: 07 dic 2018

DOI: https://doi.org/10.2478/jvetres-2018-0075

Palabras clavereal-time PCR, SYBR Green I

© 2018 Agnieszka Kędrak-Jabłońska et al., published by Sciendo

This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License.

Palabras clave
real-time PCR, SYBR Green I