dorado duplex compatibility

[davewabbo]

Aside: I'm running DNAscent on a separate Linux amd64 machine and I've been getting it up and running.

I've been experimenting on dorado duplex outputs.
I use dorado demux to split my barcoded reads
Samtools to split up via dx:1,0,-1 tags (see the commands below).

There are two errors I'm encountering (note there are both pod5 fail and pass reads mixed in the bam):

• For the duplex_stereo reads (dx:1), DNAscent exits immediately almost after starting giving the "Assertion error" also likely because there are two read ids "readid_pair1;readid_pair2" for each read.
• Oddly enough the duplex_simplex reads i.e. simplex reads with duplex offspring (dx:-1) have readIDs that are not present in the index.dnascent file yet the "Assertion error" does not show up and the pipeline completes.
• simplex only reads (dx:0) causes DNAscent to get "killed" at a specific read and does not throw assertion error.

(I have removed the header sections with personally identifiable info. I am happy to share the sequencing files to troubleshoot this error)

Duplex_simplex header
DS:basecall_model=dna_r10.4.1_e8.2_400bps_sup@v5.0.0_dna_r10.4.1_e8.2_5khz_stereo@v1.3 runid=e92471a7a2298044ecc21493c171b83a5a59e86f LB:unknown SM:unknown @PG ID:demux PN:dorado VN:0.9.0+9dc15a85 CL:dorado demux --output-dir /Volumes/SSD_T7_1TB/V26a_DNAscent_workdir/dorado_duplex_reads/demux --kit-name SQK-NBD114-24 duplex.bam @PG ID:samtools PN:samtools PP:demux VN:1.21 CL:samtools view -b -h -q 0 -d dx:-1 duplex_barcode02.bam @PG ID:samtools.1 PN:samtools PP:samtools VN:1.21 CL:samtools view -h duplex_simplex_barcode02.bam

Duplex_stereo header
DS:basecall_model=dna_r10.4.1_e8.2_400bps_sup@v5.0.0_dna_r10.4.1_e8.2_5khz_stereo@v1.3 runid=e92471a7a2298044ecc21493c171b83a5a59e86f LB:unknown SM:unknown @PG ID:demux PN:dorado VN:0.9.0+9dc15a85 CL:dorado demux --output-dir /Volumes/SSD_T7_1TB/V26a_DNAscent_workdir/dorado_duplex_reads/demux --kit-name SQK-NBD114-24 duplex.bam @PG ID:samtools PN:samtools PP:demux VN:1.21 CL:samtools view -b -h -q 0 -d dx:1 duplex_barcode02.bam @PG ID:samtools.1 PN:samtools PP:samtools VN:1.21 CL:samtools view -h duplex_stereo_barcode02.bam

simplex (only) header
DS:basecall_model=dna_r10.4.1_e8.2_400bps_sup@v5.0.0_dna_r10.4.1_e8.2_5khz_stereo@v1.3 runid=e92471a7a2298044ecc21493c171b83a5a59e86f LB:unknown SM:unknown @PG ID:demux PN:dorado VN:0.9.0+9dc15a85 CL:dorado demux --output-dir /Volumes/SSD_T7_1TB/V26a_DNAscent_workdir/dorado_duplex_reads/demux --kit-name SQK-NBD114-24 duplex.bam @PG ID:samtools PN:samtools PP:demux VN:1.21 CL:samtools view -b -h -q 0 -d dx:0 duplex_barcode02.bam @PG ID:samtools.1 PN:samtools PP:samtools VN:1.21 CL:samtools view -h simplex_barcode02.bam