Extract reads - SILVA 128 issue

[Azra_Music (Azra Musić)]

Hi,

I am having an issue with extracting appropriate reads from SILVA 97 OTU 16S. As a result of the feature-classifier, I have obtained taxonomy consisting of level 1 only (classifies sequences as Bacteria).

qiime tools import
–type ‘FeatureData[Sequence]’
–input-path 97_otus_16S.fasta
–output-path 97_otus_silva.qza

qiime tools import
–type ‘FeatureData[Taxonomy]’
–source-format HeaderlessTSVTaxonomyFormat
–input-path taxonomy_7_levels.txt
–output-path ref-taxonomy.qza

qiime feature-classifier extract-reads
–i-sequences 97_otus_silva.qza
–p-f-primer GTGCCAGCMGCCGCGGTAA
–p-r-primer GGACTACHVGGGTWTCTAAT
–p-trunc-len 100
–o-reads ref-seqs.qza

qiime feature-classifier fit-classifier-naive-bayes
–i-reference-reads ref-seqs.qza
–i-reference-taxonomy ref-taxonomy.qza
–o-classifier classifier_silva_97_otu.qza

I guess the issue could be wrong primers(?). I have found the topic including 18S classifier, but it did not help solve my problem.

[Nicholas_Bokulich]

Hi @Azra_Music,
It looks like you are using the 515f/806r primer pair for 16S rRNA gene sequences. Why not just use the pretrained classifiers for SILVA or Greengenes that are available on the QIIME 2 website?

@Azra_Music

I have obtained taxonomy consisting of level 1 only (classifies sequences as Bacteria).

Does this post or this post describe your issue? Those sound like what you describe — the tips in those posts may help resolve your issue.

If those posts do not help resolve this issue, please give us some more details:

1. What read length do you have?
2. single-end or paired-end reads?
3. What sequencing protocol are you using?

Thanks!