It is possible to detect modified bases directly from the signal data when the appropriate PCR-free protocols have been used, providing additional insight into biological data even when not directly the goal of an experiment. Tools to detect modified bases for several different methylated states currently exist. To detect a modified base typically requires a model for the detection tool and the specific modified base.

Where a detection model exists, modified bases may be detectable through basecalling of the raw signal data. Both Dorado and MinKNOW include models and options for detecting certain types of methylated states from the signal data generated during sequencing. ONT tools have been developed for this purpose and have been shown to be accurate and highly repeatable. When detecting methylation through Guppy, Dorado or MinKNOW please ensure to output a BAM formatted file as methylation is stored in the BAM format. For more information, please see both the applications page and the accuracy page.

(For third-party tools, please see their GitHub pages for installation and usage. To address any issues with these and other third-party tools, please post to the Issues tab on the individual GitHub repositories as we cannot offer support for third-party software.)