{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,23]],"date-time":"2026-10-23T20:07:35Z","timestamp":1792786055422,"version":"4.5.0"},"reference-count":79,"publisher":"Oxford University Press (OUP)","issue":"11","license":[{"start":{"date-parts":[[2026,6,17]],"date-time":"2026-06-17T00:00:00Z","timestamp":1781654400000},"content-version":"vor","delay-in-days":9,"URL":"https:\/\/creativecommons.org\/licenses\/by-nc\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100000780","name":"European Union","doi-asserted-by":"publisher","award":["956810"],"award-info":[{"award-number":["956810"]}],"id":[{"id":"10.13039\/501100000780","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100010663","name":"European Research Council","doi-asserted-by":"publisher","award":["101042103"],"award-info":[{"award-number":["101042103"]}],"id":[{"id":"10.13039\/100010663","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100010663","name":"European Research Council","doi-asserted-by":"publisher","award":["101187456"],"award-info":[{"award-number":["101187456"]}],"id":[{"id":"10.13039\/100010663","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100015439","name":"CERCA","doi-asserted-by":"publisher","id":[{"id":"10.13039\/100015439","id-type":"DOI","asserted-by":"publisher"}]},{"name":"CRG Core Technologies Programme"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2026,6,8]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>Nanopore direct RNA sequencing holds promise for advancing our understanding of the epitranscriptome. Recently, Oxford Nanopore Technologies released basecalling models capable of detecting N6-methyladenosine (m6A), inosine (I), pseudouridine (\u03a8), and 5-methylcytosine (m5C). However, their performance and cross-reactivity with other modifications remain largely unexplored. Here, we systematically benchmark four available modification-aware basecalling models by evaluating their per-read and per-site predictions across synthetic molecules and biological samples from diverse species. Models performed well on highly modified, balanced synthetic constructs (AUC = 0.93\u20130.97, PR-AUC = 0.84\u20130.91), but their performance dropped sharply on unbalanced datasets that reflect modification abundances in biological samples (PR-AUC: 0.04\u20130.09). Analysis of\u00a0in vivo rRNA samples confirmed this limitation, with false-discovery rate ranging from 50% to 100%, even after filtering with modification-free controls. We identify two major sources of false positives: cross-reactivities with other modifications and current alterations at sites neighbouring a modified residue. Finally, we demonstrate that basecalling error- and current-based methods can accurately detect modifications, offering effective alternatives for modifications lacking dedicated models. Our results highlight the utility and limitations of modification-aware basecalling models for RNA modification detection, and underscore the importance of including control samples to mitigate false-positive predictions.<\/jats:p>","DOI":"10.1093\/nar\/gkag349","type":"journal-article","created":{"date-parts":[[2026,4,7]],"date-time":"2026-04-07T11:32:16Z","timestamp":1775561536000},"source":"Crossref","is-referenced-by-count":7,"title":["Systematic benchmarking of\n                    <i>dorado<\/i>\n                    basecalling models for RNA modification detection with highly multiplexed nanopore sequencing"],"prefix":"10.1093","volume":"54","author":[{"given":"Gregor","family":"Diensthuber","sequence":"first","affiliation":[{"name":"Center for Genomic Regulation (CRG), The Barcelona Institute of Science and Technology , Dr Aiguader 88, Barcelona 08003 ,","place":["Spain"]},{"name":"Universitat Pompeu Fabra (UPF) , Barcelona 08003 ,","place":["Spain"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ivan","family":"Milenkovic","sequence":"additional","affiliation":[{"name":"Center for Genomic Regulation (CRG), The Barcelona Institute of Science and Technology , Dr Aiguader 88, Barcelona 08003 ,","place":["Spain"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Laia","family":"Llovera","sequence":"additional","affiliation":[{"name":"Center for Genomic Regulation (CRG), The Barcelona Institute of Science and Technology , Dr Aiguader 88, Barcelona 08003 ,","place":["Spain"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ana","family":"Milovanovic","sequence":"additional","affiliation":[{"name":"Center for Genomic Regulation (CRG), The Barcelona Institute of Science and Technology , Dr Aiguader 88, Barcelona 08003 ,","place":["Spain"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Francesco","family":"Pelizzari","sequence":"additional","affiliation":[{"name":"Center for Genomic Regulation (CRG), The Barcelona Institute of Science and Technology , Dr Aiguader 88, Barcelona 08003 ,","place":["Spain"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9367-6311","authenticated-orcid":false,"given":"Eva Maria","family":"Novoa","sequence":"additional","affiliation":[{"name":"Center for Genomic Regulation (CRG), The Barcelona Institute of Science and Technology , Dr Aiguader 88, Barcelona 08003 ,","place":["Spain"]},{"name":"Universitat Pompeu Fabra (UPF) , Barcelona 08003 ,","place":["Spain"]},{"name":"ICREA, Passeig Llu\u00eds Companys 23 , Barcelona 08010 ,","place":["Spain"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"286","published-online":{"date-parts":[[2026,6,17]]},"reference":[{"key":"2026061705270276000_B1","doi-asserted-by":"publisher","first-page":"365","DOI":"10.1038\/nrg.2016.47","article-title":"RNA modifications: what have we learned and where are we headed?","volume":"17","author":"Frye","year":"2016","journal-title":"Nat Rev Genet"},{"key":"2026061705270276000_B2","doi-asserted-by":"publisher","first-page":"375","DOI":"10.1038\/s41580-021-00342-0","article-title":"The expanding world of tRNA modifications and their disease relevance","volume":"22","author":"Suzuki","year":"2021","journal-title":"Nat Rev Mol Cell Biol"},{"key":"2026061705270276000_B3","doi-asserted-by":"publisher","first-page":"201","DOI":"10.1038\/nmeth.4577","article-title":"Highly parallel direct RNA sequencing on an array of nanopores","volume":"15","author":"Garalde","year":"2018","journal-title":"Nat 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