orthofinder3 "assign" option failed

[uguyet]

I'm very interested in the new --core and --assign options added to orthofinder3.

I launched orthofinder3 on 1000 genomes with the following command:
orthofinder -t 40 -M 'msa' -f input/core/ -o orthofinder_Core

Then I tried to use the "--assign" command to add among 1500 new genomes with the following command:
orthofinder --core orthofinder_Core/Results_Nov10/ --assign input/assign -t 40 -a 40 -os -M 'msa'

I got the following error:

40 thread(s) for highly parallel tasks (BLAST searches etc.)
40 thread(s) for OrthoFinder algorithm

INFO: For --assign defaulting to 'mafft --memsave' to reduce RAM usage

INFO: For --assign defaulting to 'FastTree -fastest' to reduce RAM usage

Results directory: /env/export/05_orthofinder_v2/orthofinder_Core/Results_Nov10/../Results_Nov11/

Checking required programs are installed
----------------------------------------
Test can run "astral-pro -i /env/export/05_orthofinder_v2/orthofinder_Core/Results_Nov10/../Results_Nov11/WorkingDirectory/dependencies/astral.input.nwk -o /env/export/05_orthofinder_v2/orthofinder_Core/Results_Nov10/../Results_Nov11/WorkingDirectory/dependencies/astral.output.nwk -t 2" - ok

Creating orthogroup profiles
----------------------------
2024-11-11 12:29:18 : Done 0 of 4905
Sequences must all be the same length

What should I do to correct this problem ?
Thanks in advance

[lauriebelch]

orthofinder --core orthofinder_Core/Results_Nov10/ --assign input/assign -t 40 -a 40 -os -M 'msa'

It might be that the path to the --core folder is slightly wrong, try

orthofinder --core orthofinder_Core/OrthoFinder/Results_Nov10/ --assign input/assign -t 40 -a 40 -os -M 'msa'

[uguyet]

Hello,
the path to the "core" folder seems good. When I do ls orthofinder_Core/Results_Nov10/, I got the following output:

Citation.txt  Log.txt 
Orthogroups
Orthogroup_Sequences
Orthologues
WorkingDirectory

[lauriebelch]

It looks like core run is maybe still going? It hasn't produced any sequence alignments or gene trees

One thing that might speed it up is using famsa for multiple sequence alignment (instead of mafft). This will become the default in the final release of orthofinder3.

There are some instructions on how to do this in https://github.com/davidemms/OrthoFinder?tab=readme-ov-file#configjson--adding-addtional-programs-for-tree-inference-local-alignment-or-msa

[uguyet]

You were right, there was a problem with the core run, thank you for pointing it out to me.

However, I have now an error during the creation of the Unassigned diamond database during the --assign run:

2024-11-25 17:36:58 : Creating diamond database 1 of 50
2024-11-25 17:36:58 : Creating diamond database 2 of 50
2024-11-25 17:36:58 : Creating diamond database 3 of 50
2024-11-25 17:36:58 : Creating diamond database 4 of 50
2024-11-25 17:36:58 : Creating diamond database 5 of 50
2024-11-25 17:36:58 : Creating diamond database 6 of 50
2024-11-25 17:36:58 : Creating diamond database 7 of 50
2024-11-25 17:36:58 : Creating diamond database 8 of 50
2024-11-25 17:36:58 : Creating diamond database 9 of 50
2024-11-25 17:36:58 : Creating diamond database 10 of 50
2024-11-25 17:36:58 : Creating diamond database 11 of 50
2024-11-25 17:36:58 : Creating diamond database 12 of 50
2024-11-25 17:36:58 : Creating diamond database 13 of 50
2024-11-25 17:36:58 : Creating diamond database 14 of 50
2024-11-25 17:36:58 : Creating diamond database 15 of 50
2024-11-25 17:36:58 : Creating diamond database 16 of 50
2024-11-25 17:36:58 : Creating diamond database 17 of 50
2024-11-25 17:36:58 : Creating diamond database 18 of 50
2024-11-25 17:36:58 : Creating diamond database 19 of 50
2024-11-25 17:36:58 : Creating diamond database 20 of 50
2024-11-25 17:36:58 : Creating diamond database 21 of 50
2024-11-25 17:36:58 : Creating diamond database 22 of 50
2024-11-25 17:36:58 : Creating diamond database 23 of 50
2024-11-25 17:36:58 : Creating diamond database 24 of 50
2024-11-25 17:36:58 : Creating diamond database 25 of 50
2024-11-25 17:36:58 : Creating diamond database 26 of 50
2024-11-25 17:36:58 : Creating diamond database 27 of 50
2024-11-25 17:36:58 : Creating diamond database 28 of 50
2024-11-25 17:36:58 : Creating diamond database 29 of 50
2024-11-25 17:36:58 : Creating diamond database 30 of 50
2024-11-25 17:36:58 : Creating diamond database 31 of 50
2024-11-25 17:36:58 : Creating diamond database 32 of 50
2024-11-25 17:36:58 : Creating diamond database 33 of 50
2024-11-25 17:36:58 : Creating diamond database 34 of 50
2024-11-25 17:36:58 : Creating diamond database 35 of 50

ERROR: external program called by OrthoFinder returned an error code: 1

Command: diamond makedb --in OrthoFinder/Results_Nov25/../Results_Nov25_1/WorkingDirectory/Unassigned.Species34.fa -d OrthoFinder/Results_Nov25/../Results_Nov25_1/WorkingDirectory/diamondDBSpecies34

stdout:
b''
stderr:
b'diamond v2.1.10.164 (C) Max Planck Society for the Advancement of Science, Benjamin Buchfink, University of Tuebingen\nDocumentation, support and updates available at http://www.diamondsearch.org\nPlease cite: http://dx.doi.org/10.1038/s41592-021-01101-x Nature Methods (2021)\n\n#CPU threads: 36\nScoring parameters: (Matrix=BLOSUM62 Lambda=0.267 K=0.041 Penalties=11/1)\nDatabase input file: OrthoFinder/Results_Nov25/../Results_Nov25_1/WorkingDirectory/Unassigned.Species34.fa\nOpening the database file... Error: Error detecting input file format. First line seems to be blank.\n'
ERROR: diamond makedb failed
ERROR: An error occurred, ***please review the error messages*** they may contain useful information about the problem.

I have the suspicion that Species34 does not have any unassigned genes, which causes the diamond makedb to crash as it tries to create a db on an empty fasta file...