Fig. 4.

1m-cortex.bam output from Bowtie2 alignment for first mapped read. The meaning of fields in SAM/BAM output reports is described at samtools.github.io/hts-specs/. In 1m-cortex.bam, generated from the Bowtie2 alignment (Subheading 3.2, step 3), the first reported read is assigned a FLAG value of 163. This FLAG value indicates that the read is properly paired, is the second in the pair, and its mate is reverse complemented relative to the reference. The output also indicates that the read aligns to the CDR1as circRNA junction scaffold reference starting at nt 1 (field 4). The mapping quality (MAPQ) field value is 42, and the entire 125-nt (125M) RNA-seq read aligned to the CDR1as junction. An “=“ character is assigned to field 7 to indicate that the mate read aligns to the same reference sequence. Field 8 reports that the mate read aligns at position 29 of the reference sequence. The inferred fragment length of 153 nts from the read1 and read2 alignment is reported in field 9. The sequence and the quality scores for the read are reported in fields 10 and 11, and the alignment score is reported in field 12. A perfect alignment in Bowtie2 end-to-end mode will produce a score of 0, while various alignment penalties (e.g., mismatches, insertions, gaps) will report negative scores