Effective double‐digest RAD sequencing and genotyping despite large genome size.
Resource type
Journal article
Date published
December 22, 2020
Abstract
Obtaining informative data is the ambition of any genomic project, but in nonmodel species with very large genomes, pursuing such a goal requires surmounting a series of analytical challenges. Double‐digest RAD sequencing is routinely used in nonmodel organisms and offers some control over the volume of data obtained. However, the volume of data recovered is not always an indication of the reliability of data sets, and quality checks are necessary to ensure that true and artefactual information is set apart. In the present study, we aim to fill the gap existing between the known applicability of RAD sequencing methods in plants with large genomes and the use of the retrieved loci for population genetic inference. By analysing two populations of , a nonmodel orchid species with a large genome size (1C ~ 31.6 Gbp), we provide a complete workflow from library preparation to bioinformatic filtering and inference of genetic diversity and differentiation. We show how filtering strategies to dismiss potentially misleading data need to be explored and adapted to data set‐specific features. Moreover, we suggest that the occurrence of organellar sequences in libraries should not be neglected when planning the experiment and analysing the results. Finally, we explain how, in the absence of prior information about the genome of the species, seeking high standards of quality during library preparation and sequencing can provide an insurance against unpredicted technical or biological constraints.
Funder
| Funder name | Awards |
British Ecological Society, United Kingdom | SR18/1418 |
Royal Botanic Gardens, Kew, United Kingdom | Kew Pilot Study Fund 2018 |
Ministry of Education and Research, Estonia | Institutional research funding (IUT 21-1) |
Herbarium, Eesti Maaülikool, Estonia | |
Journal title
Molecular Ecology Resources
Volume
21
Issue
4
Publisher
John Wiley & Sons Ltd.
Place of publication
UK
ISSN
1755-098X
eISSN
1755-0998
Date accepted
December 14, 2020
Official URL
Rights statement
In Copyright