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Evaluation of an automated protocol for efficient and reliable DNA extraction of dietary samples
Molecular techniques have become an important tool to empirically assess feeding interactions. The increased usage of next‐generation sequencing approaches has stressed the need of fast DNA extraction that does not compromise DNA quality. Dietary samples here pose a particular challenge, as these de...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
John Wiley and Sons Inc.
2017
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5574753/ https://www.ncbi.nlm.nih.gov/pubmed/28861241 http://dx.doi.org/10.1002/ece3.3197 |
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author | Wallinger, Corinna Staudacher, Karin Sint, Daniela Thalinger, Bettina Oehm, Johannes Juen, Anita Traugott, Michael |
author_facet | Wallinger, Corinna Staudacher, Karin Sint, Daniela Thalinger, Bettina Oehm, Johannes Juen, Anita Traugott, Michael |
author_sort | Wallinger, Corinna |
collection | PubMed |
description | Molecular techniques have become an important tool to empirically assess feeding interactions. The increased usage of next‐generation sequencing approaches has stressed the need of fast DNA extraction that does not compromise DNA quality. Dietary samples here pose a particular challenge, as these demand high‐quality DNA extraction procedures for obtaining the minute quantities of short‐fragmented food DNA. Automatic high‐throughput procedures significantly decrease time and costs and allow for standardization of extracting total DNA. However, these approaches have not yet been evaluated for dietary samples. We tested the efficiency of an automatic DNA extraction platform and a traditional CTAB protocol, employing a variety of dietary samples including invertebrate whole‐body extracts as well as invertebrate and vertebrate gut content samples and feces. Extraction efficacy was quantified using the proportions of successful PCR amplifications of both total and prey DNA, and cost was estimated in terms of time and material expense. For extraction of total DNA, the automated platform performed better for both invertebrate and vertebrate samples. This was also true for prey detection in vertebrate samples. For the dietary analysis in invertebrates, there is still room for improvement when using the high‐throughput system for optimal DNA yields. Overall, the automated DNA extraction system turned out as a promising alternative to labor‐intensive, low‐throughput manual extraction methods such as CTAB. It is opening up the opportunity for an extensive use of this cost‐efficient and innovative methodology at low contamination risk also in trophic ecology. |
format | Online Article Text |
id | pubmed-5574753 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | John Wiley and Sons Inc. |
record_format | MEDLINE/PubMed |
spelling | pubmed-55747532017-08-31 Evaluation of an automated protocol for efficient and reliable DNA extraction of dietary samples Wallinger, Corinna Staudacher, Karin Sint, Daniela Thalinger, Bettina Oehm, Johannes Juen, Anita Traugott, Michael Ecol Evol Original Research Molecular techniques have become an important tool to empirically assess feeding interactions. The increased usage of next‐generation sequencing approaches has stressed the need of fast DNA extraction that does not compromise DNA quality. Dietary samples here pose a particular challenge, as these demand high‐quality DNA extraction procedures for obtaining the minute quantities of short‐fragmented food DNA. Automatic high‐throughput procedures significantly decrease time and costs and allow for standardization of extracting total DNA. However, these approaches have not yet been evaluated for dietary samples. We tested the efficiency of an automatic DNA extraction platform and a traditional CTAB protocol, employing a variety of dietary samples including invertebrate whole‐body extracts as well as invertebrate and vertebrate gut content samples and feces. Extraction efficacy was quantified using the proportions of successful PCR amplifications of both total and prey DNA, and cost was estimated in terms of time and material expense. For extraction of total DNA, the automated platform performed better for both invertebrate and vertebrate samples. This was also true for prey detection in vertebrate samples. For the dietary analysis in invertebrates, there is still room for improvement when using the high‐throughput system for optimal DNA yields. Overall, the automated DNA extraction system turned out as a promising alternative to labor‐intensive, low‐throughput manual extraction methods such as CTAB. It is opening up the opportunity for an extensive use of this cost‐efficient and innovative methodology at low contamination risk also in trophic ecology. John Wiley and Sons Inc. 2017-07-07 /pmc/articles/PMC5574753/ /pubmed/28861241 http://dx.doi.org/10.1002/ece3.3197 Text en © 2017 The Authors. Ecology and Evolution published by John Wiley & Sons Ltd. This is an open access article under the terms of the Creative Commons Attribution (http://creativecommons.org/licenses/by/4.0/) License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Original Research Wallinger, Corinna Staudacher, Karin Sint, Daniela Thalinger, Bettina Oehm, Johannes Juen, Anita Traugott, Michael Evaluation of an automated protocol for efficient and reliable DNA extraction of dietary samples |
title | Evaluation of an automated protocol for efficient and reliable DNA extraction of dietary samples |
title_full | Evaluation of an automated protocol for efficient and reliable DNA extraction of dietary samples |
title_fullStr | Evaluation of an automated protocol for efficient and reliable DNA extraction of dietary samples |
title_full_unstemmed | Evaluation of an automated protocol for efficient and reliable DNA extraction of dietary samples |
title_short | Evaluation of an automated protocol for efficient and reliable DNA extraction of dietary samples |
title_sort | evaluation of an automated protocol for efficient and reliable dna extraction of dietary samples |
topic | Original Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5574753/ https://www.ncbi.nlm.nih.gov/pubmed/28861241 http://dx.doi.org/10.1002/ece3.3197 |
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