Sam Kovaka, a PhD candidate from Johns Hopkins University, spoke at the Nanopore Community Meeting 2021 about “Visualization and analysis of nanopore RNA and DNA signal alignments with UNCALLED4.” They explained adaptive sampling (AKA “ReadUntil“): “software-based targeted sequencing via selective ejection of reads” Kovaka explained that this requires rapid identification based on the beginning of each read which can be done with basecalling or directly from the raw signal. Basecalling methods are ReadFish and MinKNOW’s built in adaptive sampling system. However, the basecalling is computationally intensive. Kovaka explained that UNCALLED which is based on “real-time mapping of raw electrical signal.” Kovaka noted that raw signals can be used to detect modifications. Uncalled4 is a toolkit for “nanopore signal alignment, analysis, and visualization.” The toolkit is in active development and has several features to plot, compare, and align. Kovaka went over several examples that highlighted the features of the toolkit and the richness of the raw signals from Nanopore sequencing. They explained that future work will include more types of alignment and trying to integrate and improve adaptive sampling.
