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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...

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Autores principales: Wallinger, Corinna, Staudacher, Karin, Sint, Daniela, Thalinger, Bettina, Oehm, Johannes, Juen, Anita, Traugott, Michael
Formato: Online Artículo Texto
Lenguaje:English
Publicado: John Wiley and Sons Inc. 2017
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.
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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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