De novo basecalling of RNA modifications at single molecule and nucleotide resolution

Abstract RNA modifications influence RNA function and fate, but detecting them in individual molecules remains challenging for most modifications. Here we present a novel methodology to generate training sets and build modification-aware basecalling models. Using this approach, we develop the m 6 AB...

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Main Authors: Sonia Cruciani, Anna Delgado-Tejedor, Leszek P. Pryszcz, Rebeca Medina, Laia Llovera, Eva Maria Novoa
Format: Article
Language:English
Published: BMC 2025-02-01
Series:Genome Biology
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Online Access:https://doi.org/10.1186/s13059-025-03498-6
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Summary:Abstract RNA modifications influence RNA function and fate, but detecting them in individual molecules remains challenging for most modifications. Here we present a novel methodology to generate training sets and build modification-aware basecalling models. Using this approach, we develop the m 6 ABasecaller, a basecalling model that predicts m6A modifications from raw nanopore signals. We validate its accuracy in vitro and in vivo, revealing stable m6A modification stoichiometry across isoforms, m6A co-occurrence within RNA molecules, and m6A-dependent effects on poly(A) tails. Finally, we demonstrate that our method generalizes to other RNA and DNA modifications, paving the path towards future efforts detecting other modifications.
ISSN:1474-760X