Explanation needed for POD5 files- channel ID's, PromethION run

Question

[carla.finn]

Hello! I am new to processing ONT data. I am running a split-read-by-channel command (below), so that I can run dorado duplex basecalling on the reads more efficiently.

pod5 view /path/to/pod5s  --include "read_id, channel" --output summary.tsv

I have, say, 2000 pod5 files. However when I check the amount of channels I have (cut -f 2 summary.tsv | sort | uniq | wc -l) I get 2500 unique channels.

My point is, how do I have significantly more channels- am I fundamentally misunderstanding something about how channels/etc work? Samples were sequenced with PromethION / P2 Kit V14 (SQK-LSK114) Flow cell: R10.4.1 (FLO-PRO114M) Pointers towards an explanation or tutorial would be really apprecited. Thank you!

Answer 1

[GenoMax]

PromethION flowcells are supposed to have 2675 channels so 2500 unique channels seems normal. Looks like only 2000 of those may be producing usable data.

[carla.finn]

Pretend there's a 'facepalm' emoji here. That's an easy explanation. Thanks for clearing it up!