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Comparative evaluation of different molecular methods for DNA extraction from individual Teladorsagia circumcincta nematodes

BACKGROUND: The purpose of this study was to develop a reliable DNA extraction protocol to use on individual Teladorsagia circumcincta nematode specimens to produce high quality DNA for genome sequencing and phylogenetic analysis. Pooled samples have been critical in providing the groundwork for T....

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Autores principales: Sloan, S., Jenvey, C. J., Piedrafita, D., Preston, S., Stear, M. J.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8130168/
https://www.ncbi.nlm.nih.gov/pubmed/34001079
http://dx.doi.org/10.1186/s12896-021-00695-6
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author Sloan, S.
Jenvey, C. J.
Piedrafita, D.
Preston, S.
Stear, M. J.
author_facet Sloan, S.
Jenvey, C. J.
Piedrafita, D.
Preston, S.
Stear, M. J.
author_sort Sloan, S.
collection PubMed
description BACKGROUND: The purpose of this study was to develop a reliable DNA extraction protocol to use on individual Teladorsagia circumcincta nematode specimens to produce high quality DNA for genome sequencing and phylogenetic analysis. Pooled samples have been critical in providing the groundwork for T. circumcincta genome construction, but there is currently no standard method for extracting high-quality DNA from individual nematodes. 11 extraction kits were compared based on DNA quality, yield, and processing time. RESULTS: 11 extraction protocols were compared, and the concentration and purity of the extracted DNA was quantified. Median DNA concentration among all methods measured on NanoDrop 2000™ ranged between 0.45–11.5 ng/μL, and on Qubit™ ranged between undetectable – 0.962 ng/μL. Median A260/280 ranged between 0.505–3.925, and median A260/230 ranged − 0.005 – 1.545. Larval exsheathment to remove the nematode cuticle negatively impacted DNA concentration and purity. CONCLUSIONS: A Schistosoma sp. DNA extraction method was determined as most suitable for individual T. circumcincta nematode specimens due to its resulting DNA concentration, purity, and relatively fast processing time. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12896-021-00695-6.
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spelling pubmed-81301682021-05-18 Comparative evaluation of different molecular methods for DNA extraction from individual Teladorsagia circumcincta nematodes Sloan, S. Jenvey, C. J. Piedrafita, D. Preston, S. Stear, M. J. BMC Biotechnol Research Article BACKGROUND: The purpose of this study was to develop a reliable DNA extraction protocol to use on individual Teladorsagia circumcincta nematode specimens to produce high quality DNA for genome sequencing and phylogenetic analysis. Pooled samples have been critical in providing the groundwork for T. circumcincta genome construction, but there is currently no standard method for extracting high-quality DNA from individual nematodes. 11 extraction kits were compared based on DNA quality, yield, and processing time. RESULTS: 11 extraction protocols were compared, and the concentration and purity of the extracted DNA was quantified. Median DNA concentration among all methods measured on NanoDrop 2000™ ranged between 0.45–11.5 ng/μL, and on Qubit™ ranged between undetectable – 0.962 ng/μL. Median A260/280 ranged between 0.505–3.925, and median A260/230 ranged − 0.005 – 1.545. Larval exsheathment to remove the nematode cuticle negatively impacted DNA concentration and purity. CONCLUSIONS: A Schistosoma sp. DNA extraction method was determined as most suitable for individual T. circumcincta nematode specimens due to its resulting DNA concentration, purity, and relatively fast processing time. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12896-021-00695-6. BioMed Central 2021-05-17 /pmc/articles/PMC8130168/ /pubmed/34001079 http://dx.doi.org/10.1186/s12896-021-00695-6 Text en © The Author(s) 2021 https://creativecommons.org/licenses/by/4.0/Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) . The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/ (https://creativecommons.org/publicdomain/zero/1.0/) ) applies to the data made available in this article, unless otherwise stated in a credit line to the data.
spellingShingle Research Article
Sloan, S.
Jenvey, C. J.
Piedrafita, D.
Preston, S.
Stear, M. J.
Comparative evaluation of different molecular methods for DNA extraction from individual Teladorsagia circumcincta nematodes
title Comparative evaluation of different molecular methods for DNA extraction from individual Teladorsagia circumcincta nematodes
title_full Comparative evaluation of different molecular methods for DNA extraction from individual Teladorsagia circumcincta nematodes
title_fullStr Comparative evaluation of different molecular methods for DNA extraction from individual Teladorsagia circumcincta nematodes
title_full_unstemmed Comparative evaluation of different molecular methods for DNA extraction from individual Teladorsagia circumcincta nematodes
title_short Comparative evaluation of different molecular methods for DNA extraction from individual Teladorsagia circumcincta nematodes
title_sort comparative evaluation of different molecular methods for dna extraction from individual teladorsagia circumcincta nematodes
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8130168/
https://www.ncbi.nlm.nih.gov/pubmed/34001079
http://dx.doi.org/10.1186/s12896-021-00695-6
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