The Fiber-seq BAM format

The Fiber-seq BAM format adds to m6A BAM data from a PacBio (ft predict-m6a) or ONT (dorado) sequencing run with m6A.

The following tags are added to the BAM file:

• ns:B:I,: Nucleosome start sites (0-based) on the forward strand of the sequencing read (u32).
• nl:B:I,: Equal length array to ns of nucleosome lengths (u32).
• as:B:I,: MSP start sites (0-based) on the forward strand of the sequencing read (u32).
• al:B:I,: Equal length array to as of MSP lengths (u32).
• aq:B:C,: Quality scores for the MSP [0, 255] (u8). This tag is optional and added by ft fire.

The ns/as/nl/al tags are with respect to the unaligned sequencing read (forward strand) and are not affected by alignment or the orientation of the read after alignment. This also means that the starts and lengths encoded in these tags may change when ft lifts these molecular coordinates to reference coordinates.

The ns or as tag do not need to begin or end at the start or end of the read; however, once begun, they must be contiguous. i.e. the ns and as tags must combine to form a contiguous set of alternating nucleosome and MSP sites. The ns, nl, as, and al tags are added to the BAM file automatically using ft predict-m6a or later using the ft add-nucleosomes command.

The aq tag is added using the ft fire command and represents the estimated precision of the MSP being a FIRE. Specifically, the estimated precision of a FIRE is the value of the aq tag divided by 255.