Helping decision making for reliable and cost-effective 2b-RAD sequencing and genotyping analyses in non-model species.
Caretta caretta
Diplodus puntazzo
conservation genomics
genotyping-by-sequencing
high-throughput sequencing
sequencing simulations
Journal
Molecular ecology resources
ISSN: 1755-0998
Titre abrégé: Mol Ecol Resour
Pays: England
ID NLM: 101465604
Informations de publication
Date de publication:
May 2020
May 2020
Historique:
received:
30
10
2019
revised:
04
02
2020
accepted:
10
02
2020
pubmed:
16
2
2020
medline:
20
5
2021
entrez:
16
2
2020
Statut:
ppublish
Résumé
High-throughput sequencing has revolutionized population and conservation genetics. RAD sequencing methods, such as 2b-RAD, can be used on species lacking a reference genome. However, transferring protocols across taxa can potentially lead to poor results. We tested two different IIB enzymes (AlfI and CspCI) on two species with different genome sizes (the loggerhead turtle Caretta caretta and the sharpsnout seabream Diplodus puntazzo) to build a set of guidelines to improve 2b-RAD protocols on non-model organisms while optimising costs. Good results were obtained even with degraded samples, showing the value of 2b-RAD in studies with poor DNA quality. However, library quality was found to be a critical parameter on the number of reads and loci obtained for genotyping. Resampling analyses with different number of reads per individual showed a trade-off between number of loci and number of reads per sample. The resulting accumulation curves can be used as a tool to calculate the number of sequences per individual needed to reach a mean depth ≥20 reads to acquire good genotyping results. Finally, we demonstrated that selective-base ligation does not affect genomic differentiation between individuals, indicating that this technique can be used in species with large genome sizes to adjust the number of loci to the study scope, to reduce sequencing costs and to maintain suitable sequencing depth for a reliable genotyping without compromising the results. Here, we provide a set of guidelines to improve 2b-RAD protocols on non-model organisms with different genome sizes, helping decision-making for a reliable and cost-effective genotyping.
Identifiants
pubmed: 32061018
doi: 10.1111/1755-0998.13144
doi:
Substances chimiques
DNA
9007-49-2
DNA Restriction Enzymes
EC 3.1.21.-
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Subventions
Organisme : Ministerio de Ciencia, Innovación y Universidades, Agencia Estatal de Investigación (AEI) and Fondo Europeo de Desarrollo Regional (FEDER)
ID : CTM2017-88080
Organisme : Generalitat de Catalunya
ID : SGR2017-1120
Organisme : Generalitat de Catalunya
ID : SGR2017-378
Organisme : Agència de Gestió d'Ajuts Universitaris i de Recerca
ID : FI_B 00997
Organisme : Ministerio de Ciencia, Innovación y Universidades
ID : FPU15/02390
Informations de copyright
© 2020 John Wiley & Sons Ltd.
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