all features are unclassified

I have a problem. I am trying to get taxonomy classification for my dataset which used primers in V5 and V6 regions. I have tried using the general classifier, but no classification was done by the classfier, the species were all unassigned.Please suggest ways to go about it.

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Which classifier did you use from https://docs.qiime2.org/2020.2/data-resources/ ? You will need to use one of the full-length sequence classifiers or train your own on your amplified region I believe.

Additionally, your dataset may not have the proper resolution for species level classification. Did you get genera level classification?

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Hi @oast,

Welcome the :qiime2: forum!

Did you, by chance, use the silva 138 classifier? What’s your enviroment? How is your genus/family level coverage?

Additionally, can I ask why not having species is an issue when you have an ASV or centroid? That gives you species/sub species resolution without having to worry about pesky things like uncultured organisms,
polyphyletic clades (okay, fewer polyphyletic clades as you go up), and names that change.

Best,
Justine

I used the Greengenes 13_8 99% OTUs full-length sequences classifier.

I did not get the genera level classification

Hi @oast,

What is the lowest taxonomic level where you did get classification? Could you please share the taxonomy artifact?

Best,
Justine

The lowest classification I got was level 1.From level 2, it did not show any classification.
Please find attached pictures. Thank you.

Hi @oast,

And, can i assume you’re not working in a rare or weird enviroment?
What’s the orientation of your reads?

Best,
Justine

I am not working in a weird environment.
My reads are forward single-end reads

Hi @oast,

Can you please share your taxonomy barchart artifact or your taxonomy artifact?
You can either post it here or DM it to me if you’re uncomfortable with sharing it publically.

Best,
Justine

Please find attached the two artifacts.

taxa-bar-plots.qzv (1.2 MB) taxonomy.qzv (1.5 MB)

So I analyzed two datasets, one of the datasets was classified successfully by the classifier but the other dataset which has primer in regions V5 and V6 was not successfully classified by the classfier

Hi @oast,

I just checked the provinance on the taxonomy file you sent me. It looks like you used a classifier trained for V4 hypervariable region (515-805) which would explain why you’re not getting any results. Could you please download the full length classifier again and try classification. You can double check the reads with the provenience, which Im having trouble explaining, but sometimes a picture is worth a thousand words. (Drop your artifact into view.qiime2.org, click on provenience, and then here you go.)

Best,
Justine

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I finally resolved the issue. Thank you for the assistance.

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