Place the ASVs against the PICRUSt2 reference multiple-sequence alignment and phylogeny.

[EugeniaSH (Eugenia Silva Herzog)]

Hello,
I am trying to follow the tutorial to use qiime2 output to analyze in picrust2 plugin.
qiime2-session/hands-on-pricust2.md at master · mbcarbonetto/qiime2-session · GitHub
I am stuck in the first step of placing te ASV against the Picrust2 references
I am using Qiime2-2021.11,
This is what I add
qiime fragment-insertion sepp
--i-representative-sequences representative_sequences.qza
--p-threads 1
--i-reference-database reference.fna.qza
--o-tree phylogeny
--output-dir picrust2/results

and this is the error
There were some problems with the command:
(1/2) Invalid value for '--i-reference-database': Expected an artifact of at
least type SeppReferenceDatabase. An artifact of type
FeatureData[AlignedSequence] was provided.
(2/2) Missing option '--o-placements'. ("--output-dir" may also be used)

Thanks a lot

[cherman2 (Chloe Herman)]

Hello @EugeniaSH,

The second error is the easier one to address. If you put --output-dir picrust2/results you can not put any parameter with a --o at the beginning.

I think that tutorial you are using is a bit outdated. Its last commit on github was 2019 and q2-fragment-insertion has been updated since then. The tutorial you linked uses this command:

qiime fragment-insertion sepp \
--i-representative-sequences DADA2/representative_sequences.qza \
--p-threads 1 \
--i-reference-alignment picrust2/reference.fna.qza \
--i-reference-phylogeny picrust2/reference.tre.qza \
--output-dir picrust2/results

However,

--i-reference-alignment picrust2/reference.fna.qza \
--i-reference-phylogeny picrust2/reference.tre.qza \

are not valid inputs to q2-fragment-insertion at this moment. I think you realized this and changed your input to:

--i-reference-database reference.fna.qza 

but now instead of a reference sequences and an alignment, q2-fragment-insertion takes a SeppReferenceDatabase type.

I would recommend using a more up to date tutorial on the QIIME2 website.
Here are a couple:

https://library.qiime2.org/plugins/q2-fragment-insertion/16/

https://docs.qiime2.org/2022.2/tutorials/pd-mice/

Hope that helps!

[EugeniaSH (Eugenia Silva Herzog)]

Thank you I will try them.