QIIME 2 de novo tree creation

These commands are based on QIIME 2 tutorials and are on this page simply for convenience.

Making multiple-sequence alignment

We'll need to make a multiple-sequence alignment of the ASVs before running FastTree. First, we'll make a folder for the output files.

mkdir tree_out

We'll use MAFFT to make a de novo multiple-sequence alignment of the ASVs.

qiime alignment mafft --i-sequences deblur_output/rep_seqs_final.qza \
                      --p-n-threads 1 \
                      --o-alignment tree_out/rep_seqs_final_aligned.qza

Filtering multiple-sequence alignment

Variable positions in the alignment need to be masked before FastTree is run, which can be done with this command:

qiime alignment mask --i-alignment tree_out/rep_seqs_final_aligned.qza \
  --o-masked-alignment tree_out/rep_seqs_final_aligned_masked.qza

Running FastTree

Finally FastTree can be run on this masked multiple-sequence alignment:

qiime phylogeny fasttree --i-alignment tree_out/rep_seqs_final_aligned_masked.qza \
                         --p-n-threads 1 \
                         --o-tree tree_out/rep_seqs_final_aligned_masked_tree

Add root to tree

FastTree returns an unrooted tree. One basic way to add a root to a tree is to add it add it at the midpoint of the largest tip-to-tip distance in the tree, which is done with this command:

qiime phylogeny midpoint-root --i-tree tree_out/rep_seqs_final_aligned_masked_tree.qza \
                              --o-rooted-tree tree_out/rep_seqs_final_aligned_masked_tree_rooted.qza

Re-name file

To keep this output filename consistent with the SOP you can simply make a copy of this output tree.

cp tree_out/rep_seqs_final_aligned_masked_tree_rooted.qza asvs-tree.qza