Single-cell mode

Starting with version 0.6.1 oarfish incorporates the first single-cell quantification capabilities. Given a bam file, collated by cell barcode and with already (UMI) deduplicated reads, this mode, enabled with the --single-cell flag, will allow oarfish to produce a single-cell quantification matrix.

Single-cell mode requires a transcriptome-aligned BAM, supplied with --alignments

Unlike bulk quantification — which can take a transcriptome BAM (--alignments), raw reads (--reads), a genome FASTA + reads (--genome), or a genome BAM (--genome-alignments) — single-cell mode supports only the transcriptome-BAM path. It does not support raw-read mapping or either of the genome (projection) modes. The CLI enforces this: --single-cell requires --alignments and conflicts with --reads, --genome, and --genome-alignments. The input BAM must therefore be reads aligned to the transcriptome (e.g. with minimap2/pbmm2), not to the genome.

Formatting requirements of BAM input in single-cell mode: All alignment records for the same cell barcode should be adjacent in the bam file, and a count will be obtained for each read record, so UMI de-duplication should have been performed if those are the counts you want. In the future, counting UMIs directly may be supported, and some of these other restrictions may be lifted.