Combining nanopore and illumina sequencing permits detailed analysis of insertion mutations and structural variations produced by PEG-Mediated transformation in ostreococcus tauri

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• dc.contributor.author Thomy, Julie
• dc.contributor.author Sanchez, Frederic
• dc.contributor.author Gut, Marta
• dc.contributor.author Cruz, Fernando
• dc.contributor.author Alioto, Tyler
• dc.contributor.author Piganeau, Gwenael
• dc.contributor.author Grimsley, Nigel
• dc.contributor.author Yau, Sheree
• dc.date.accessioned 2021-05-14T07:25:21Z
• dc.date.available 2021-05-14T07:25:21Z
• dc.date.issued 2021
• dc.description.abstract Ostreococcus tauri is a simple unicellular green alga representing an ecologically important group of phytoplankton in oceans worldwide. Modern molecular techniques must be developed in order to understand the mechanisms that permit adaptation of microalgae to their environment. We present for the first time in O. tauri a detailed characterization of individual genomic integration events of foreign DNA of plasmid origin after PEG-mediated transformation. Vector integration occurred randomly at a single locus in the genome and mainly as a single copy. Thus, we confirmed the utility of this technique for insertional mutagenesis. While the mechanism of double-stranded DNA repair in the O. tauri model remains to be elucidated, we clearly demonstrate by genome resequencing that the integration of the vector leads to frequent structural variations (deletions/insertions and duplications) and some chromosomal rearrangements in the genome at the insertion loci. Furthermore, we often observed variations in the vector sequence itself. From these observations, we speculate that a nonhomologous end-joining-like mechanism is employed during random insertion events, as described in plants and other freshwater algal models. PEG-mediated transformation is therefore a promising molecular biology tool, not only for functional genomic studies, but also for biotechnological research in this ecologically important marine alga.en
• dc.description.sponsorship This research received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No. 824110—EASI-Genomics and the French National Research Agency project grant (ANR Algalvirus ANR-17-CE02-0012). We acknowledge the support of the Spanish Ministry of Science and Innovation through the Instituto de Salud Carlos III and the 2014–2020 Smart Growth Operating Program, to the EMBL partnership and cofinancing with the European Regional Development Fund (MINECO/FEDER, BIO2015-71792-P). We also acknowledge the support of the Centro de Excelencia Severo Ochoa, and the Generalitat de Catalunya through the Departament de Salut, Departament d’Empresa i Coneixement and the CERCA Programmeen
• dc.format.mimetype application/pdf
• dc.identifier.citation Thomy J, Sanchez F, Gut M, Cruz F, Alioto T, Piganeau G et al. Combining nanopore and illumina sequencing permits detailed analysis of insertion mutations and structural variations produced by PEG-Mediated transformation in ostreococcus tauri. Cells. 2021 Mar 17;10(3):664. DOI: 10.3390/cells10030664
• dc.identifier.doi http://dx.doi.org/10.3390/cells10030664
• dc.identifier.issn 2073-4409
• dc.identifier.uri http://hdl.handle.net/10230/47564
• dc.language.iso eng
• dc.publisher MDPI
• dc.relation.projectID info:eu-repo/grantAgreement/ES/1PE/BIO2015-71792-P
• dc.relation.projectID info:eu-repo/grantAgreement/EC/H2020/824110
• dc.rights © 2021 Julie Thomy et al. Licensee MDPI, Basel, Switzerland. This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited
• dc.rights.accessRights info:eu-repo/semantics/openAccess
• dc.rights.uri http://creativecommons.org/licenses/by/4.0/
• dc.subject.other Genèticaca
• dc.subject.other Genòmicaca
• dc.subject.other Algues verdesca
• dc.subject.other Fitoplàncton maríca
• dc.subject.other Mutagènesica
• dc.title Combining nanopore and illumina sequencing permits detailed analysis of insertion mutations and structural variations produced by PEG-Mediated transformation in ostreococcus taurien
• dc.type info:eu-repo/semantics/article
• dc.type.version info:eu-repo/semantics/publishedVersion