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MinION-Based DNA Barcoding of Preserved and Non-Invasively Collected Wildlife Samples

The ability to sequence a variety of wildlife samples with portable, field-friendly equipment will have significant impacts on wildlife conservation and health applications. However, the only currently available field-friendly DNA sequencer, the MinION by Oxford Nanopore Technologies, has a high err...

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Autores principales: Seah, Adeline, Lim, Marisa C.W., McAloose, Denise, Prost, Stefan, Seimon, Tracie A.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7230362/
https://www.ncbi.nlm.nih.gov/pubmed/32325704
http://dx.doi.org/10.3390/genes11040445
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author Seah, Adeline
Lim, Marisa C.W.
McAloose, Denise
Prost, Stefan
Seimon, Tracie A.
author_facet Seah, Adeline
Lim, Marisa C.W.
McAloose, Denise
Prost, Stefan
Seimon, Tracie A.
author_sort Seah, Adeline
collection PubMed
description The ability to sequence a variety of wildlife samples with portable, field-friendly equipment will have significant impacts on wildlife conservation and health applications. However, the only currently available field-friendly DNA sequencer, the MinION by Oxford Nanopore Technologies, has a high error rate compared to standard laboratory-based sequencing platforms and has not been systematically validated for DNA barcoding accuracy for preserved and non-invasively collected tissue samples. We tested whether various wildlife sample types, field-friendly methods, and our clustering-based bioinformatics pipeline, SAIGA, can be used to generate consistent and accurate consensus sequences for species identification. Here, we systematically evaluate variation in cytochrome b sequences amplified from scat, hair, feather, fresh frozen liver, and formalin-fixed paraffin-embedded (FFPE) liver. Each sample was processed by three DNA extraction protocols. For all sample types tested, the MinION consensus sequences matched the Sanger references with 99.29%–100% sequence similarity, even for samples that were difficult to amplify, such as scat and FFPE tissue extracted with Chelex resin. Sequencing errors occurred primarily in homopolymer regions, as identified in previous MinION studies. We demonstrate that it is possible to generate accurate DNA barcode sequences from preserved and non-invasively collected wildlife samples using portable MinION sequencing, creating more opportunities to apply portable sequencing technology for species identification.
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spelling pubmed-72303622020-05-22 MinION-Based DNA Barcoding of Preserved and Non-Invasively Collected Wildlife Samples Seah, Adeline Lim, Marisa C.W. McAloose, Denise Prost, Stefan Seimon, Tracie A. Genes (Basel) Article The ability to sequence a variety of wildlife samples with portable, field-friendly equipment will have significant impacts on wildlife conservation and health applications. However, the only currently available field-friendly DNA sequencer, the MinION by Oxford Nanopore Technologies, has a high error rate compared to standard laboratory-based sequencing platforms and has not been systematically validated for DNA barcoding accuracy for preserved and non-invasively collected tissue samples. We tested whether various wildlife sample types, field-friendly methods, and our clustering-based bioinformatics pipeline, SAIGA, can be used to generate consistent and accurate consensus sequences for species identification. Here, we systematically evaluate variation in cytochrome b sequences amplified from scat, hair, feather, fresh frozen liver, and formalin-fixed paraffin-embedded (FFPE) liver. Each sample was processed by three DNA extraction protocols. For all sample types tested, the MinION consensus sequences matched the Sanger references with 99.29%–100% sequence similarity, even for samples that were difficult to amplify, such as scat and FFPE tissue extracted with Chelex resin. Sequencing errors occurred primarily in homopolymer regions, as identified in previous MinION studies. We demonstrate that it is possible to generate accurate DNA barcode sequences from preserved and non-invasively collected wildlife samples using portable MinION sequencing, creating more opportunities to apply portable sequencing technology for species identification. MDPI 2020-04-18 /pmc/articles/PMC7230362/ /pubmed/32325704 http://dx.doi.org/10.3390/genes11040445 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Seah, Adeline
Lim, Marisa C.W.
McAloose, Denise
Prost, Stefan
Seimon, Tracie A.
MinION-Based DNA Barcoding of Preserved and Non-Invasively Collected Wildlife Samples
title MinION-Based DNA Barcoding of Preserved and Non-Invasively Collected Wildlife Samples
title_full MinION-Based DNA Barcoding of Preserved and Non-Invasively Collected Wildlife Samples
title_fullStr MinION-Based DNA Barcoding of Preserved and Non-Invasively Collected Wildlife Samples
title_full_unstemmed MinION-Based DNA Barcoding of Preserved and Non-Invasively Collected Wildlife Samples
title_short MinION-Based DNA Barcoding of Preserved and Non-Invasively Collected Wildlife Samples
title_sort minion-based dna barcoding of preserved and non-invasively collected wildlife samples
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7230362/
https://www.ncbi.nlm.nih.gov/pubmed/32325704
http://dx.doi.org/10.3390/genes11040445
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