Error with Dada2 denoising process

Technical Support
1

Hello, i wanted to ask if i can run the Dada2 algorithm in moving pictures using fastq files that i produced from fasta files. (My data were in fasta format and they were 16s single-end trimmed seqs. I converted them in fastq using a script and i made every base with quality score "I"). My purpose was to run the moving pictures pipeline. The files were imported just fine producing the "demux-single-end.qza" file, but when i run the dada2 algorithm there was an error "Plugin error from dada2: An error was encountered while running DADA2 in R (return code 1), please inspect stdout and stderr to learn more. See above for debug info."
I run the same command with the --verbose command and got this result:
R version 4.1.3 (2022-03-10)
Loading required package: Rcpp
DADA2: 1.22.0 / Rcpp: 1.0.8.3 / RcppParallel: 5.1.5

  1. Filtering ...................................................................
  2. Learning Error Rates
    430569243 total bases in 1000484 reads from 44 samples will be used for learning the error rates.
    Error rates could not be estimated (this is usually because of very few reads).
    Error in getErrors(err, enforce = TRUE) : Error matrix is NULL.
    Execution halted

The original command was this:
qiime dada2 denoise-single --i-demultiplexed-seqs demux-single-end.qza --p-trunc-len 0 --o-representative-sequences rep-seqs-dada2.qza --o-table table-dada2.qza --o-denoising-stats stats-dada2.qza --verbose

Does this have to do with the fact that all my bases in the fastq files got the same quality score? Or is it something else?
Thanks a lot

2

Hi @t.gousdovas,

Welcome to the :qiime2: forum!

Dada2 requires variation in the quality score since the model is based on learning about your sequencing quality. Because every basepair has the same error rate, there's no variability to model.

You can't run any of the denoising techniques on data without quality information; regardless of how you try to spoof semantic types. You should be able to cluster the data into OTUs, though, since those don't require quality information.

Best,
Justine

1 Like
3

Thanks a lot.
Is there any tutorial on how to do this with OTUs?

4

Hi @t.gousdovas,

Check out the OTU clustering tutorial.

Best,
Justine

1 Like
5

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