Enhanced detection of RNA modifications and read mapping with high-accuracy nanopore RNA basecalling models

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  • dc.contributor.author Diensthuber, Gregor
  • dc.contributor.author Pryszcz, Leszek Piotr, 1985-
  • dc.contributor.author Llovera Nadal, Laia
  • dc.contributor.author Lucas, Morghan C.
  • dc.contributor.author Delgado-Tejedor, Anna
  • dc.contributor.author Cruciani, Sonia
  • dc.contributor.author Roignant, Jean-Yves
  • dc.contributor.author Begik, Oguzhan
  • dc.contributor.author Novoa, Eva Maria
  • dc.date.accessioned 2024-10-01T06:30:49Z
  • dc.date.embargoEnd info:eu-repo/date/embargoEnd/2025-03-13
  • dc.date.issued 2024
  • dc.description Data de publicació electrònica: 13-09-2024
  • dc.description.abstract In recent years, nanopore direct RNA sequencing (DRS) became a valuable tool for studying the epitranscriptome, due to its ability to detect multiple modifications within the same full-length native RNA molecules. While RNA modifications can be identified in the form of systematic basecalling 'errors' in DRS datasets, N6-methyladenosine (m6A) modifications produce relatively low 'errors' compared to other RNA modifications, limiting the applicability of this approach to m6A sites that are modified at high stoichiometries. Here, we demonstrate that the use of alternative RNA basecalling models, trained with fully unmodified sequences, increases the 'error'signal of m6A, leading to enhanced detection and improved sensitivity even at low stoichiometries. Moreover, we find that high-accuracy alternative RNA basecalling models can show up to 97% median basecalling accuracy, outperforming currently available RNA basecalling models, which show 91% median basecalling accuracy. Notably, the use of high-accuracy basecalling models is accompanied by a significant increase in the number of mapped reads -especially in shorter RNA fractions- and increased basecalling error signatures at pseudouridine (Ψ) and N1-methylpseudouridine (m1Ψ) modified sites. Overall, our work demonstrates that alternative RNA basecalling models can be used to improve the detection of RNA modifications, read mappability, and basecalling accuracy in nanopore DRS datasets.
  • dc.description.sponsorship GD is part of the ROPES ITN which received funding from the European Union’s Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement no. 956810. OB is supported by funds from Merck Innovation 2020. LPP was supported by funding from the European Union’s H2020 research and innovation programme under Marie Sklodowska-Curie grant agreement No. 754422, and is currently supported by ERC funds (ERC-StG-2021 No 101042103 to EMN). JYR is supported by funds from the Swiss National Science Foundation (310030_197906) and the Deutsche Forschungsgemeinschaft (RO 4681/4-2, RO 4681/6-1, and RO 4681/12-1, TRR319 RMaP). This work was supported by funds from the Spanish Ministry of Economy, Industry and Competitiveness (MEIC) (PID2021-128193NB-100 to EMN) and the European Research Council (ERC-StG-2021 No 101042103 to EMN). We acknowledge the support of the MEIC to the EMBL partnership, Centro de Excelencia Severo Ochoa and CERCA Programme / Generalitat de Catalunya.
  • dc.embargo.liftdate 2025-03-13
  • dc.format.mimetype application/pdf
  • dc.identifier.citation Diensthuber G, Pryszcz LP, Llovera L, Lucas MC, Delgado-Tejedor A, Cruciani S, et al. Enhanced detection of RNA modifications and read mapping with high-accuracy nanopore RNA basecalling models. Genome Res. 2024 Sep 13:gr.278849.123. DOI: 10.1101/gr.278849.123
  • dc.identifier.doi http://dx.doi.org/10.1101/gr.278849.123
  • dc.identifier.issn 1088-9051
  • dc.identifier.uri http://hdl.handle.net/10230/61274
  • dc.language.iso eng
  • dc.publisher Cold Spring Harbor Laboratory Press (CSHL Press)
  • dc.relation.ispartof Genome Res. 2024 Sep 13:gr.278849.123
  • dc.relation.projectID info:eu-repo/grantAgreement/EC/H2020/956810
  • dc.relation.projectID info:eu-repo/grantAgreement/EC/H2020/754422
  • dc.relation.projectID info:eu-repo/grantAgreement/ES/3PE/PID2021-128193NB-100
  • dc.relation.projectID info:eu-repo/grantAgreement/EC/HE/101042103
  • dc.rights This article is distributed exclusively by Cold Spring Harbor Laboratory Press for the first six months after the full-issue publication date (see https://genome-cshlp-org.sare.upf.edu/site/misc/terms.xhtml). After six months, it is available under a Creative Commons License (Attribution-NonCommercial 4.0 International), as described at http://creativecommons.org/licenses/by-nc/4.0/.
  • dc.rights.accessRights info:eu-repo/semantics/embargoedAccess
  • dc.rights.uri http://creativecommons.org/licenses/by-nc/4.0/
  • dc.subject.other RNA
  • dc.subject.other Nanoporus
  • dc.title Enhanced detection of RNA modifications and read mapping with high-accuracy nanopore RNA basecalling models
  • dc.type info:eu-repo/semantics/article
  • dc.type.version info:eu-repo/semantics/publishedVersion

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