Utilizing field collected insects for next generation sequencing: Effects of sampling, storage, and DNA extraction methods.
curated insects
ddRAD
degraded DNA
next generation sequencing
pan traps
propylene glycol
Journal
Ecology and evolution
ISSN: 2045-7758
Titre abrégé: Ecol Evol
Pays: England
ID NLM: 101566408
Informations de publication
Date de publication:
Dec 2019
Dec 2019
Historique:
received:
06
09
2018
revised:
20
08
2019
accepted:
15
09
2019
entrez:
16
1
2020
pubmed:
16
1
2020
medline:
16
1
2020
Statut:
epublish
Résumé
DNA sequencing technologies continue to advance the biological sciences, expanding opportunities for genomic studies of non-model organisms for basic and applied questions. Despite these opportunities, many next generation sequencing protocols have been developed assuming a substantial quantity of high molecular weight DNA (>100 ng), which can be difficult to obtain for many study systems. In particular, the ability to sequence field-collected specimens that exhibit varying levels of DNA degradation remains largely unexplored. In this study we investigate the influence of five traditional insect capture and curation methods on Double-Digest Restriction Enzyme Associated DNA (ddRAD) sequencing success for three wild bee species. We sequenced a total of 105 specimens (between 7-13 specimens per species and treatment). We additionally investigated how different DNA quality metrics (including pre-sequence concentration and contamination) predicted downstream sequencing success, and also compared two DNA extraction methods. We report successful library preparation for all specimens, with all treatments and extraction methods producing enough highly reliable loci for population genetic analyses. Although results varied between species, we found that specimens collected by net sampling directly into 100% EtOH, or by passive trapping followed by 100% EtOH storage before pinning tended to produce higher quality ddRAD assemblies, likely as a result of rapid specimen desiccation. Surprisingly, we found that specimens preserved in propylene glycol during field sampling exhibited lower-quality assemblies. We provide recommendations for each treatment, extraction method, and DNA quality assessment, and further encourage researchers to consider utilizing a wider variety of specimens for genomic analyses.
Identifiants
pubmed: 31938475
doi: 10.1002/ece3.5756
pii: ECE35756
pmc: PMC6953651
doi:
Types de publication
Journal Article
Langues
eng
Pagination
13690-13705Informations de copyright
© 2019 The Authors. Ecology and Evolution published by John Wiley & Sons Ltd.
Déclaration de conflit d'intérêts
None declared.
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