Can anyone help me with data from sequencing center

[timanix (Timur Yergaliyev)]

@sdpapet

I can provide the primers sequences, but I don't know barcodes. So, there is no way that I can provide the barcode sequences. You mentioned, "I would suggest to use cutadapt to remove forward and reverse primers that were used in library preparation and discard any reads without those sequences. "

Is it enough to remove both barcodes and primer sequences, if I only provide primer sequences.

Yes, as I already wrote and as it is written in the q2-cutadapt description it will remove the primers (provided) and any preceding sequenses (including barcodes in that case).

@sdpapet

I check the q2-cutadpt manual here (cutadapt — QIIME 2 2022.8.3 documentation). I suppose I should use trim-paired script but not demux-paired.

That's right - this time you should use trim-paired script since the purpose is to delete primers from already demultiplexed reads.

@sdpapet

Also, is there any "real example" of cutadpt scripts? Based on the manual, it keeps saying 5' - 3'. I am quite confused about the orientation of primer sequence. I think everyone write it from 5' to 3'? Can you provide a cutadpt scripts that you usually run to remove barcodes/primers etc.?

That's an example from my pipeline

qiime cutadapt trim-paired \
    --i-demultiplexed-sequences demux.qza \
    --o-trimmed-sequences trimmed.qza \
    --p-cores 6 \
    --p-front-f CAAGRGTTHGATYMTGGCTCAG \
    --p-front-r TGCTGCCTCCCGTAGGAGT \
    --p-match-adapter-wildcards \
    --p-discard-untrimmed \
    --p-match-read-wildcards

All primers are provided in the 5'-3' orientation.

PS. Since this thread is becoming too long and the point of discussion already diverging from the original question, please create new topic for new questions (after performing quick search on the forum to make sure that this question is not already answered). This approach may help other users who'll encounter the same issues in future).

Best,