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Optimizing the lysis step in CTAB DNA extractions of silica‐dried and herbarium leaf tissues
PREMISE: The use of cetyltrimethylammonium bromide (CTAB) is an effective and inexpensive method of extracting DNA from plants. The CTAB protocol is frequently modified to optimize DNA extractions, but experimental approaches rarely perturb a single variable at a time to systematically infer their e...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
John Wiley and Sons Inc.
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10278933/ https://www.ncbi.nlm.nih.gov/pubmed/37342163 http://dx.doi.org/10.1002/aps3.11522 |
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author | Carey, S. James Becklund, L. Ellie Fabre, Paige P. Schenk, John J. |
author_facet | Carey, S. James Becklund, L. Ellie Fabre, Paige P. Schenk, John J. |
author_sort | Carey, S. James |
collection | PubMed |
description | PREMISE: The use of cetyltrimethylammonium bromide (CTAB) is an effective and inexpensive method of extracting DNA from plants. The CTAB protocol is frequently modified to optimize DNA extractions, but experimental approaches rarely perturb a single variable at a time to systematically infer their effect on DNA quantity and quality. METHODS AND RESULTS: We investigated how chemical additives, incubation temperature, and lysis duration affected DNA quantity and quality. Altering those parameters influenced DNA concentrations and fragment lengths, but only extractant purity was significantly affected. CTAB and CTAB plus polyvinylpyrrolidone buffers produced the highest DNA quality and quantity. Extractions from silica gel–preserved tissues had significantly higher DNA yield, longer DNA fragments, and purer extractants compared to herbarium‐preserved tissues. CONCLUSIONS: We recommend DNA extractions of silica gel–preserved tissues that include a shorter and cooler lysis step, which results in purer extractions compared to a longer and hotter lysis step, while preventing fragmentation and reducing time. |
format | Online Article Text |
id | pubmed-10278933 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | John Wiley and Sons Inc. |
record_format | MEDLINE/PubMed |
spelling | pubmed-102789332023-06-20 Optimizing the lysis step in CTAB DNA extractions of silica‐dried and herbarium leaf tissues Carey, S. James Becklund, L. Ellie Fabre, Paige P. Schenk, John J. Appl Plant Sci Protocol Note PREMISE: The use of cetyltrimethylammonium bromide (CTAB) is an effective and inexpensive method of extracting DNA from plants. The CTAB protocol is frequently modified to optimize DNA extractions, but experimental approaches rarely perturb a single variable at a time to systematically infer their effect on DNA quantity and quality. METHODS AND RESULTS: We investigated how chemical additives, incubation temperature, and lysis duration affected DNA quantity and quality. Altering those parameters influenced DNA concentrations and fragment lengths, but only extractant purity was significantly affected. CTAB and CTAB plus polyvinylpyrrolidone buffers produced the highest DNA quality and quantity. Extractions from silica gel–preserved tissues had significantly higher DNA yield, longer DNA fragments, and purer extractants compared to herbarium‐preserved tissues. CONCLUSIONS: We recommend DNA extractions of silica gel–preserved tissues that include a shorter and cooler lysis step, which results in purer extractions compared to a longer and hotter lysis step, while preventing fragmentation and reducing time. John Wiley and Sons Inc. 2023-05-27 /pmc/articles/PMC10278933/ /pubmed/37342163 http://dx.doi.org/10.1002/aps3.11522 Text en © 2023 The Authors. Applications in Plant Sciences published by Wiley Periodicals LLC on behalf of Botanical Society of America. https://creativecommons.org/licenses/by/4.0/This is an open access article under the terms of the http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Protocol Note Carey, S. James Becklund, L. Ellie Fabre, Paige P. Schenk, John J. Optimizing the lysis step in CTAB DNA extractions of silica‐dried and herbarium leaf tissues |
title | Optimizing the lysis step in CTAB DNA extractions of silica‐dried and herbarium leaf tissues |
title_full | Optimizing the lysis step in CTAB DNA extractions of silica‐dried and herbarium leaf tissues |
title_fullStr | Optimizing the lysis step in CTAB DNA extractions of silica‐dried and herbarium leaf tissues |
title_full_unstemmed | Optimizing the lysis step in CTAB DNA extractions of silica‐dried and herbarium leaf tissues |
title_short | Optimizing the lysis step in CTAB DNA extractions of silica‐dried and herbarium leaf tissues |
title_sort | optimizing the lysis step in ctab dna extractions of silica‐dried and herbarium leaf tissues |
topic | Protocol Note |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10278933/ https://www.ncbi.nlm.nih.gov/pubmed/37342163 http://dx.doi.org/10.1002/aps3.11522 |
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