Can I use sepp fragment insertion with gg 13_8, then classify my reads with the updated gg 2024.09 ?
If I should not be doing that?
How should I be building the tree using the updated greengenes database?
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Hi @RanaAbdelaal,
If placement is necessary, a SEPP reference for 2022.10 is available here. The backbone phylogeny is identical to what's in 2024.09.
Best,
Daniel
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Will you kindly suggest the optimal approach?
I tried the Greengenes non-V4-16S plugin, but I found the tree incompatible with the rest of the downstreem analysis. (The core metrics phylogenetics command failed because of the tree)
qiime greengenes2 non-v4-16s
--i-table dada2_table.qza
--i-sequences dada2_rep_set.qza
--i-backbone 2024.09.backbone.full-length.fna.qza
--o-mapped-table mapped-table.qza
--o-representatives mapped-sequences.qza
qiime phylogeny align-to-tree-mafft-fasttree
--i-sequences mapped-sequences.qza
--o-alignment aligned-rep-seqs.qza
--o-masked-alignment masked-aligned-rep-seqs.qza
--o-tree unrooted-tree.qza
--o-rooted-tree tree.qza
qiime diversity core-metrics-phylogenetic
--i-table ./dada2_table.qza
--i-phylogeny ./tree.qza
--m-metadata-file ./metadata.tsv
--p-sampling-depth 81498
--output-dir ./core-metrics-results
I would recommend just using the table and Greengenes2 phylogeny following the non-v4-16s step. No need to re-estimate a phylogeny, and doing so from short sequences can yield a poor quality phylogeny anyway
Best,
Daniel
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Thank you, but I just feel more comfortable using sepp and fragment insertion.
If I do so using the sepp reference for 2022.10.
It's still a valid approach correct?
One more question.
I already did my analysis using sepp fragment insertion with 13_8 greengenes.
And classified using gg 2024.09 full length.
Do I have to redo the whole analysis, or are my reaults valid and I should go with it?
Thank you 
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They aren't invalid, but using Greengenes 13_8 could draw reviewer scrutiny as that database is over 10 years old.
Best,
Daniel
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