when to apply trimming parameters

[smzt]

Hi, I've been going through the documentation and the paper but I have not been able to find a clear answer yet to my naive question. When should I apply the values Figaro provides? Should I apply them before trimming the primers or after trimming the primers?

Thanks in advance.

Regards,

Sheila

[alegione]

My understanding if helpful: The tool is written for Dada2, so the output results should be used as truncation lengths in dada2. The tool truncates reads first, then removes the primers if you ask it to also trim the ends.

Example if using dada2 in R (all caps text to be replaced with your data)

fastqPairedFilter(fn = c(FORWARD_IN, REVERSE_IN), fout = c(FORWARD_OUT, REVERSE_OUT), truncLen = c(FIGARO TRIM POSITION 1, FIGARO TRIM POSITION 2), verbose = TRUE, maxN = 0, maxEE = 2, truncQ = 2, trimLeft = c(FORWARD PRIMER LENGTH, REVERSE PRIMER LENGTH), compress = TRUE)

Example if using dada2 in QIIME2

qiime dada2 denoise-paired \
    --i-demultiplexed-seqs demux-paired.qza \
    --p-trunc-len-f FIGARO TRIM POSITION 1 \
    --p-trunc-len-r FIGARO TRIM POSITION 2 \
    --p-trim-left-f FORWARD PRIMER LENGTH \
    --p-trim-left-r REVERSE PRIMER LENGTH \
    --o-representative-sequences representative-sequences.qza \
    --o-table table.qza \
    --o-denoising-stats denoising-stats.qza