Dynamont: A comprehensive cross-species comparison of ONT segmentation tools.

Oxford Nanopore Technologies (Oxford Nanopore Technologies (ONT)) sequencing enables direct, long-read sequencing of DNA and RNA, preserving nucleotide modifications. During basecalling, deep neural networks translate raw nanopore signals into nucleotide sequences, internally segmenting the signal to align it with the corresponding bases. This is a challenging task due to uneven motor protein rotation, signal variability, low-quality reads, and the presence of nucleotide modifications. However, the signal to nucleotide assignment is critical for novel downstream signal analysis. Existing tools, such as Tombo Resquiggle, f5c Eventalign, f5c Resquiggle, and Uncalled4, operate after basecalling and rely on event-based segmentation and mapping approaches, that often fail to align low-quality or modified reads and lack confidence estimates for segmentation accuracy.