Extended Data Fig. 3: Signal-to-read and signal-to-reference alignment.

(a) Per-k-mer current means from signal-to-read and signal-to-reference Uncalled4 alignments and uncorrected basecaller moves. (b) Alignment dotplot of a D. melanogaster r10.4.1 read to a reference containing a spiked-in G- > T substitution at the location indicated by the red lines, causing increased read-model current MAD. (c) Alignment dotplot of the same read to a reference with a 10 nucleotide deletion with boundaries indicated by red lines. Uncalled4 masks signal around insertions or deletions 10 nucleotides or larger based on the ref-moves coordinates, meaning the signal corresponding to the deleted sequence is not included. (d) Reference- and read-aligned signal of a read which features a likely sequencing error causing a two nucleotide insertion in the basecalled sequence. A slight jump in signal is observed within the signal mapping to ‘A’ in the signal-to-reference alignment, indicated by the green arrow. This nucleotide is broken into ‘GAG’ in the basecalled sequence, making the signal-to-read alignment erroneously more similar to the pore model.