Hi,

I am working with Nanopore sequencing data that was base called and demultiplexed with guppy (version unknown, but likely rather recent). Alignment was done using bwa tool.

Now I want to sub-sample an existing bam so that I end up with a new bam file which only contains reads that were sequenced up hour 5 into the sequencing process. I did so by first extracting all read_id values that full fill this criteria.

awk '$10 <= 1800 {print $5}' 20230503_1351_MC-112161_AIB461_29a045a6.sequencing_summary.txt  > f31583_5h_parent_ID_5h.txt

Then I use the resulting file as input for samtools view.

samtools view -b -N f31583_5h_parent_ID_5h.txt REF_aln_trim.bam > sub_sampled.bam

However, after a closer investigation I discovered that not all qname values in REF_aln_trim.bam have a matching value in 20230503_1351_MC-112161_AIB461_29a045a6.sequencing_summary.txt. This is counter intuitive for me as I assumed that xxx.sequencing_summary.txt contains information about every read that was produced during sequencing. Or is the read_id column in xxx.sequencing_summary.txt not equivalent to the qname tag in a bam file. I would appreciate any input or suggestions how to do the aimed sub-sampling better.

Cheers!