# Supermatrix alignment has varying length -\> error in tree build

**URL:** https://forum.biobakery.org/t/supermatrix-alignment-has-varying-length-error-in-tree-build/1488
**Category:** PhyloPhlAn
**Created:** [January 7, 2021, 4:29am UTC](https://forum.biobakery.org/t/supermatrix-alignment-has-varying-length-error-in-tree-build/1488 "2021-01-07T04:29:16Z")
**Posts on this page:** 3
**Page:** 1

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### Author: ![tanaes](https://yyz1.discourse-cdn.com/flex027/user_avatar/forum.biobakery.org/tanaes/32/551_2.png) [@tanaes](https://forum.biobakery.org/u/tanaes)
#### Post date: [January 7, 2021, 4:29am UTC](https://forum.biobakery.org/t/supermatrix-alignment-has-varying-length-error-in-tree-build/1488/1 "2021-01-07T04:29:16Z")

</div>

I’m attempting to use PhyloPhlan to build a tree from some MAGs, but obtaining an error in tree building (with IQTREE):  
ERROR: Sequence Ppolionotus\_5F\_Bin\_77\_1 contains not enough characters (4869)  
ERROR: Sequence Ppolionotus\_3F\_Bin\_38\_2 contains not enough characters (4520)  
ERROR: Sequence Ppolionotus\_3F\_Bin\_53\_4 contains not enough characters (5539)

The alignments indeed all seem to be of varied length.

I’m using version 3.0.58, and running the following configuration:

```
[db_aa]
program_name = /home/jgs286/miniconda3/envs/phylophlan/bin/diamond
params = makedb
threads = --threads
input = --in
output = --db
version = version
command_line = #program_name# #params# #threads# #input# #output#

[map_dna]
program_name = /home/jgs286/miniconda3/envs/phylophlan/bin/diamond
params = blastx --quiet --threads 1 --outfmt 6 --more-sensitive --id 50 --max-hsps 35 -k 0
input = --query
database = --db
output = --out
version = version
command_line = #program_name# #params# #input# #database# #output#

[map_aa]
program_name = /home/jgs286/miniconda3/envs/phylophlan/bin/diamond
params = blastp --quiet --threads 1 --outfmt 6 --more-sensitive --id 50 --max-hsps 35 -k 0
input = --query
database = --db
output = --out
version = version
command_line = #program_name# #params# #input# #database# #output#

[msa]
program_name = /home/jgs286/miniconda3/envs/phylophlan/bin/mafft
params = --quiet --anysymbol --thread 1 --auto
version = --version
command_line = #program_name# #params# #input# > #output#
environment = TMPDIR=/tmp

[trim]
program_name = /home/jgs286/miniconda3/envs/phylophlan/bin/trimal
params = -gappyout
input = -in
output = -out
version = --version
command_line = #program_name# #params# #input# #output#

[tree1]
program_name = /home/jgs286/miniconda3/envs/phylophlan/bin/iqtree
params = -quiet -nt AUTO -m LG
input = -s
output = -pre
version = -version
command_line = #program_name# #params# #input# #output#

```

Any clues? I have also uploaded the complete output to Dropbox and could send a link if helpful.

Thanks!!!

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<div class="post-metadata">

### Author: ![tanaes](https://yyz1.discourse-cdn.com/flex027/user_avatar/forum.biobakery.org/tanaes/32/551_2.png) [@tanaes](https://forum.biobakery.org/u/tanaes)
#### Post date: [January 7, 2021, 4:56am UTC](https://forum.biobakery.org/t/supermatrix-alignment-has-varying-length-error-in-tree-build/1488/2 "2021-01-07T04:56:53Z")

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…updating, I’ve retried with the Amphora marker set and it seems to be working. Problem seems constrained to Phylophlan markers…

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<div class="post-metadata">

### Author: ![f.asnicar](https://yyz1.discourse-cdn.com/flex027/user_avatar/forum.biobakery.org/f.asnicar/32/196_2.png) [@f.asnicar](https://forum.biobakery.org/u/f.asnicar)
#### Post date: [January 8, 2021, 1:54pm UTC](https://forum.biobakery.org/t/supermatrix-alignment-has-varying-length-error-in-tree-build/1488/3 "2021-01-08T13:54:18Z")

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Hi @tanaes thank you for reporting this.  
Can you provide me with the Dropbox link and the command line so that I can test it?

Many thanks,  
Francesco