% reads that couldn't be demultiplexed

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1

Hello,

I would like to know if there is a straightforward way to see what percent of my reads could not be demultiplexed. I am looking for this information because I am getting fewer than expected reads per sample, and I want to check whether this is because many of my reads could not be demultiplexed.

Here is my demux command:

qiime demux emp-paired \
    --i-seqs qiime2_output/sequences.qza \
    --m-barcodes-file sample_metadata.txt \
    --m-barcodes-column BarcodeSequence \
    --o-per-sample-sequences qiime2_output/demux.qza \
    --o-error-correction-details qiime2_output/demux-details.qza \
    --p-rev-comp-mapping-barcodes

Attempt #1: I tried adding --verbose, but I don't think this created any new information in my logs.

Attempt #2: I thought this information might be in demux-details.qza, so I tried converting it to a .qzv file with the following command:

qiime metadata tabulate \
    --m-input-file qiime2_output/demux-details.qza \
    --o-visualization qiime2_output/demux-details.qzv

I was only able to make this command work after allocating >16 GB of RAM, and I couldn't open it in the online QIIME2 viewer. The file is 743 Mb, so maybe it's too big? (N.B. I see that you are working on improving this command, and I think that would be great.)

My question is: Is this the right way to find the information I'm looking for? I feel like it would be extremely useful to show this number (% reads that couldn't be demultiplexed) in the output of demux summarize.

Thank you,
Lev

P.S. I am using QIIME version 2021.2.0, which I installed with conda.

2

I believe the simple answer is to divide the total number of sequences that could be demultiplexed (which I get from the "Demultiplexed sequence counts summary" section of demux.qzv) by the number of clusters passing filter (which I get from the sequencer).

This is pretty straightforward, but I do think it could be nice to report this percentage in demux.qzv.

Thanks,
Lev

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