Hello My taxa-bar-plot and taxonomy is not as good as I hoped for. Mostly classification to order level and many sequences only to kingdom level. How can I improve my results? To train the classifier as in the tutorial? I am not sure which artifacts to use besides rep-seqs.qza. Best regards Jasmi…

Hi @Jasmine84 , [image] Jasmine84: Mostly classification to order level and many sequences only to kingdom level. See this post . Depending on the length of your sequences, the primers you are using, etc, deeper than order level might be unreliable. On 250 bp 16S rRNA gene sequences, we can us…

Hello again I have 450 bp long sequences and I am trying to create my own classifier, by using the tutorial train your classifier. However, I receive this error message: (the primers should be correct) [image] What other things should I check to solve this problem? (below the command used) qiim…

[image] Jasmine84: qiime feature-classifier extract-reads –i-sequences 99_otus.qza –p-f-primer TCGTCGGCAGCGTCAGATGTGTATAAGAGACAGCCTACGGGNGGCWGCAG –p-r-primer GTCTCGTGGGCTCGGAGATGTGTATAAGAGACAGGACTACHVGGGTATCTAATCC –p-trunc-len 450 –o-reads 99_450_ref-seqs.qza See this forum thread for a …

It still does not work. qiime feature-classifier extract-reads --i-sequences $dir/rep-seqs_450.qza --p-f-primer CCTACGGGNGGCWGCAG --p-r-primer GACTACHVGGGTATCTAATCC --o-reads $dir/ref-seqs_450.qza Do you have any suggestions to which primer to use? I received this information from my lab technici…

Not it worked! Next error is however this one: [image] Can you help me with this?

Hi @Jasmine84 - I think this error means that your reference reads and your reference taxonomy have no overlapping feature IDs. This can happen if you are accidentally combining files from different reference databases, for example. Have you have a chance to look at this tutorial ? If you need more …

Yes. I wondered over this when I tried to create my classifier. We have downloadet 99_otus_fasta and 99_otu_taxonomy.txt (green genes database) and created 99_ref-taxonomy.qza and classifier.qza through the tutorial you have linked to. However, my rep-seqs.qza are directly linked to NCBI database. S…

[image] Jasmine84: So, how can I further solve this problem. Please see my request from 8 days ago: [image] thermokarst: If you need more assistance, please send us links to download the two QZA files used as inputs in that failing command.

filer.zip (2.8 MB) Here you are :slight_smile:

Thanks @Jasmine84 --- I used the decentralized provenance to look two of the files you provided: rep-seqs_450.qza [01%20PM] ref-seqs_450.qza [12%20PM] It looks like you mixed up your files - your reference sequences should be the sequences provided as part of the taxonomic database - not from…

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