Long-fragment targeted capture for long read sequencing of plastomes

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1 UMR DIADE, Institut de Recherche pour le Développement, Montpellier, France
2 UMR AGAP, Institut National de la Recherche Agronomique, Montpellier, France
3 Centre de biologie et gestion des populations (CBGP), Institut National de la Recherche Agronomique, Montferrier-sur-Lez, France
4 Facultad de Ciencias Exactas y Naturales, Pontificia Universidad Católica del Ecuador, Quito, Ecuador
DOI
10.7287/peerj.preprints.27411v1
Published
Accepted
Subject Areas
Biodiversity, Evolutionary Studies, Genomics
Keywords
MinION, DNA probes, long-range PCR, whole chloroplast sequencing, De Novo assembly
Copyright
© 2018 Bethune et al.
Licence
This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ Preprints) and either DOI or URL of the article must be cited.
Cite this article
Bethune K, Mariac C, Couderc M, Scarcelli N, Santoni S, Ardisson M, Martin J, Montufar R, Klein V, Sabot F, Vigouroux Y, Couvreur TL. 2018. Long-fragment targeted capture for long read sequencing of plastomes. PeerJ Preprints 6:e27411v1

Abstract

Third generation sequencing methods generate significantly longer reads than those produced using alternative sequencing methods. This provides increased possibilities to better study biodiversity, phylogeography and population genetics. We developed a protocol for in-solution enrichment hybridization capture of long DNA fragments applicable to complete chloroplast genomes. The protocol uses cost effective in-house probes developed via long-range PCR and was used in six non-model monocot species (Poaceae: African rice, pearl millet, fonio; and three palm species). DNA was extracted from fresh and silicagel dried leaves. Our protocol successfully captured long read chloroplast fragments (up to 4264 bp median) with an enrichment rate ranging from 15% to 98%. DNA extracted from silicagel dried leaves led to low quality plastome assemblies when compared to freshly extracted DNA. Our protocol could also be generalized to capture long sequences from specific nuclear fragments.

Author Comment

This paper documents a protocol for long DNA fragment (> 2 kbp) capturing and sequencing of chloroplast DNA. This paper has been submitted to Applications in Plant Sciences

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