I'm using Qiime2: qiime2-2021.11
I already used the dada2 script with no problems but now I have one with a new dataset.
I have a problem with the qiime dada2 denoise-paired step.
When I do this step:
qiime dada2 denoise-paired --i-demultiplexed-seqs aguas.qza --p-trim-left-f 0 --p-trim-left-r 0 --p-trunc-len-f 235 --p-trunc-len-r 235 --p-n-threads 31 --o-table aguas_table.qza \ --o-representative-sequences aguas_rep-seqs.qza \ --o-denoising-stats aguas_stats.qza \ --verbose
I have the following error message appearing.
Command: run_dada_paired.R /tmp/tmp1ubwn1ot/forward /tmp/tmp1ubwn1ot/reverse /tmp/tmp1ubwn1ot/output.tsv.biom /tmp/tmp1ubwn1ot/track.tsv /tmp/tmp1ubwn1ot/filt_f /tmp/tmp1ubwn1ot/filt_r 235 235 0 0 2.0 2.0 2 12 independent consensus 1.0 31 1000000 R version 4.0.5 (2021-03-31) Loading required package: Rcpp DADA2: 1.18.0 / Rcpp: 1.0.7 / RcppParallel: 5.1.4 1) Filtering The filter removed all reads: /tmp/tmp1ubwn1ot/filt_f/Up01_10_L001_R1_001.fastq.gz and /tmp/tmp1ubwn1ot/filt_r/Up01_10_L001_R2_001.fastq.gz not written. The filter removed all reads: /tmp/tmp1ubwn1ot/filt_f/Up45_10_L001_R1_001.fastq.gz and /tmp/tmp1ubwn1ot/filt_r/Up45_10_L001_R2_001.fastq.gz not written. The filter removed all reads: /tmp/tmp1ubwn1ot/filt_f/Down45_10_L001_R1_001.fastq.gz and /tmp/tmp1ubwn1ot/filt_r/Down45_10_L001_R2_001.fastq.gz not written. The filter removed all reads: /tmp/tmp1ubwn1ot/filt_f/Down01_10_L001_R1_001.fastq.gz and /tmp/tmp1ubwn1ot/filt_r/Down01_10_L001_R2_001.fastq.gz not written. Some input samples had no reads pass the filter. x.xx.x 2) Learning Error Rates 456320650 total bases in 1941790 reads from 2 samples will be used for learning the error rates. 456320650 total bases in 1941790 reads from 2 samples will be used for learning the error rates. 3) Denoise samples .. .. 4) Remove chimeras (method = consensus) 6) Write output Saved FeatureTable[Frequency] to: aguas_table.qza Saved FeatureData[Sequence] to: aguas_rep-seqs.qza Saved SampleData[DADA2Stats] to: aguas_stats.qza
Notice that my 6 samples appear in the filter step, but in the result I only have 2 samples.
I tried do do the following: changing
--p-trunc-len-r for possible merging problems, but nothing.
Why is it not working ? How can I know where is the problem in my data?
When I looked at my demux.qzv file, all the counts were good in "Per-sample sequence counts".
Thank you in advance for your help.