custom DB causes ParseError thrown: Unexpected character

[we32i225]

Hi, very new to Lotus2 so this might be trivial. I have been trying to use the AF_full_region database produced by the Anaerobic Fungi Network (https://anaerobicfungi.org/databases/) but i got the error below

[M::main] CMD: /root/lotus2//bin//minimap2-2.17_x64-linux/minimap2 -x sr --sr -u both --secondary=no -N 30 -c -t 1 -o Will_AF/output_AFN/tmpFiles//otu_seeds.fna.phiX.0.cont_hit.paf /root/lotus2//DB//phiX.fasta Will_AF/output_AFN/tmpFiles//otu_seeds.fna [M::main] Real time: 0.005 sec; CPU: 0.004 sec; Peak RSS: 0.004 GB Loading Subject Sequences and Ids... ParseError thrown: Unexpected character '-' found. Make sure that the file is standards compliant. If you get an unexpected character warning make sure you have set the right program parameter (-p), i.e. Lambda expected nucleic acid alphabet, maybe the file was protein?

in response I substituted the '-' for '' thinking Lotus2 couldn't understand '-'. this still didn't work the same error appeared but with ParseError thrown: Unexpected character '' found.

is the problem the characters in my DB files? if so what character can i substitute with that Lotus2 can read. Many thanks :) (i cant submit the AF_full_region FASTA file to github apologies)

[hildebra]

Hey, no this wasn't the "-" char in the command args, probably relates to a char in the .fas file.. can you try to run lotus2 with the same parameters, but the std ITS database (UNITE)? If the error is not occurring, it's a problem with the new .fas file. Also note that you need a specific format for the .tax file you are submitting, please see here: https://github.com/hildebra/lotus2#coustom-reference-database best, Falk

[we32i225]

Hi Falk thanks for getting back so quick! The sequencing is actually on the LSU region (unusual for fungi i know), i used the SLV LSU DB and it ran just fine, so i think it is my custom .fas and taxonomy file then. Just to clarify, can Lotus2 read '-' characters or will i have to substitute them, and does the FASTA file need to be formatted in a certain way as well, or just the taxonomy file?

Many thanks

[hildebra]

Hey Wei, I suspect that the error occurred inside the aligner used.. can you check in the log file with the commands called by Lotus2 (LotuSLogs/LotuS_commands.log (or similar, currently not on the cluster), and just try to rerun the last command there? As far as I remember, LotuS2 doesn't have specific formatting requirements for the fas&tax files, just matching headers of course (everything beyond a space character in the fas header will be ignored). best, Falk

[hildebra]

Hey Wei, I think the error occurred in the aligner, can you check the LotuSLogs/Lotus_cmd.log (or similar) file, and rerun the last logged command? There shouldn't be any specfici requirements for the formatting of the fasta and tax file, beyond matching labels. best, Falk

[we32i225]

Hi Falk thanks for the help. I tried re-running the last command line, but that didn't fix it, i also tried using UNITE (which worked fine). I think it is likely my DB files are not formatted correctly, and not actually an issue with Lotus2, apologies. best Wei

[hildebra]

Sorry to hear Wei, it might be an unsupported character (eg "-") is being used in the DB fasta string. I hope you still find LotuS2 useful though. best, Falk