Making an OTU Table with Barcodes

Hi,

I'm having issues getting an OTU table that I can use to BLAST against different databases. I don't want to use the I have demultiplexed single-end sequences that I denoised in DADA2:

qiime dada2 denoise-single
--i-demultiplexed-seqs single-end-demux.qza
--p-trim-left 13
--p-trunc-len 120
--o-representative-sequences rep-seqs.qza
--o-table table1.qza
--o-denoising-stats stats1.qza

Then I filtered sequences with less than 10 reads and that fell in less than 2 samples:
qiime feature-table filter-features
--i-table table1.qza
--p-min-frequency 10
--o-filtered-table feature-freq-filter-table.qza

and qiime feature-table filter-features
--i-table feature-freq-filter-table.qza
--p-min-samples 2
--o-filtered-table filtered-samples-table.qza

and filtered sequences by the newest feature-table:

qiime feature-table filter-seqs
--i-data rep-seqs1.qza
--i-table filtered-samples-table.qza
--o-filtered-data clean-rep-seqs.qza

Now I have clustered sequences at 99% similarity:
qiime vsearch cluster-features-de-novo
--i-table filtered-samples-table.qza
--i-sequences clean-rep-seqs.qza
--p-perc-identity 0.99
--o-clustered-table table-dn-99.qza
--o-clustered-sequences rep-seqs-dn-99.qza

I want to get a table that tells me sample ID, frequency of OTU per sample, and gives me the barcodes of OTUs so I can BLAST them. I also want to add my metadata table, but not sure during which step. Is something like that possible?

Thank you,
Logan

Hi @lnovak4 ,

The q2-feature-classifier plugin has a taxonomic classification method that is based on blastn, so you can just use your sequences as input to that method to achieve the first part of what you want (i.e., instead of using BLAST manually). The benefit of this method is that it automatically finds a consensus taxonomy (much better than taking the top blast hit or manually finding the consensus).

use qiime metadata tabulate with the feature table, sequences, and taxonomy as inputs — see here for an example:
https://docs.qiime2.org/2021.11/tutorials/metadata/#exploring-feature-metadata

technically QIIME 2 can be used for this as well (also with metadata tabulate), but this is beginning to sound like a customized table that you should probably prepare outside of QIIME 2, e.g., to export these files and merge/modify per your specifications. QIIME 2 cannot add metadata to both the sample and feature axes simultaneously, so creating such a custom report will require some custom work.

Good luck!

2 Likes

Hi,

The issue I have is that I don't want to use BLAST for taxonomy, I have another database I want to use. Is there a way to get the clustered sequence barcodes in a table?

QIIME 2 can accept any database as input, provided that it is supplied in the correct format (see the online tutorials for formatting and importing details)

Sure, see the exporting tutorial:
https://docs.qiime2.org/2021.11/tutorials/exporting/

good luck!

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