V3-v4 classifier

Hi all

I am trying to train a classifier and assign taxonomy to my V3V4 sequences. the commands work but they only classify to the kingdom level in my rep_seq.qza file…

qiime tools import
–type ‘FeatureData[Sequence]’
–input-path 99_otus.fasta
–output-path 99_otus.qza

qiime tools import
–type ‘FeatureData[Taxonomy]’
–source-format HeaderlessTSVTaxonomyFormat
–input-path 99_otu_taxonomy.txt
–output-path ref-taxonomy.qza

qiime feature-classifier extract-reads
–i-sequences 99_otus.qza
–p-trunc-len 285
–o-reads ref-seqs.qza

Hi Gal,
See here. Kingdom-level classification usually indicates:

  1. the wrong database was used
  2. the wrong primers were used for extraction or the wrong type of data was input to the classifier (e.g., training a V4 classifier but attempting to classify V3-V4)
  3. inappropriate trim lengths are being used in extract-reads. This should either match the length of your sequences, or just don’t use it at all (e.g., if you have overlapping paired-end reads)

Mixed classification (e.g., lots of kingdom-level but also deeper classifications) is a more complicated problem that can indicate user error or data issues, but kingdom-only is almost always user errors (and it happens to all of us :smile:)

Please double-check your inputs and parameters to make sure that the primers, trim length, and database are all appropriate for the input data.

Please let us know if you are still having issues!

1 Like

Thank you so much! It was a human error :frowning: unfortunately the error was at the start of my processing so it took me forever to find it! I found that I used the wrong importing format…

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