Cutadapt module plot classification of adapters

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Hello,

I’m using MultiQC in my Cutadapt logs. My Cutadapt command looks like this (wildcards {} are due to Snakemake being my workflow management system):

cutadapt \
  {input.fq_f} \
  {input.fq_r} \
  --front Primer_F=^{params.primer_f} \
  -G Primer_R=^{params.primer_r} \
  --adapter Rev_com_primer_R={params.revcom_r} \
  -A Rev_com_primer_F={params.revcom_f} \
  --discard-untrimmed \
  --cores {params.nthreads} \
  --output {output.fq_trim_f} \
  --paired-output {output.fq_trim_r}

Then, in MultiQC I can see the primer of my forward reads (Primer_F) in the plot named " Trimmed Sequence Lengths":

cutadapt_5
cutadapt_5885×627 96.4 KB

And then I see a plot named " Trimmed Sequence Lengths (3’)". So I deduce the previous one is the one for 5’. Anyway, in that 3’ plot I can see a peak corresponding to the primer of the reverse reads (Primer_R). But then I see a long line that corresponds to the aditional adapters I provided Cutadapt with: the reverse complementaries of the forward and the reverse primers:

cutadapt_3
cutadapt_3885×627 31.3 KB

Since this last plot is for 3’ and the previous one is for 5’, shouldn’t the reverese complementary of forward primer be on the 5’ plot, instead of on the 3’?

Many thanks in advance!

2

Hi @salias,

This is more of a Cutadapt question than MultiQC. If something looks odd, I’d recommend going back to the Cutadapt logs themselves and checking the numbers there. If you think that MultiQC is misrepresenting the data there, then please attach some files for us to reproduce and let us know :+1:

Phil

3

Hello @ewels ,

Thank you so much for your response. After thinking about it these last days, I think I understand now how MultiQC represents the data here: First plot (5’) includes primer of forward reads, and 3’ plot includes primer of reverse reads. The reads I specify as adapters (in my case, the reverse complementary sequences of the primers) are included in the 3’ since the are in the 3’ end of the read itself (the forward or the reverse read). My confusion here came from the reverse primer being included in the 3’ plot (since in theory, although it is the 3’ end of the general sequence, if we consider only the reverse reads, the end would be 5’).

Again, thank you so much. Let me know if the question was off-topic enough for me to delete it

Sergio

4

Ah great, thanks for replying with the conclusion!

Not off topic per-se, thanks for posting :slight_smile: