Failed to execute /usr/bin/perl /opt/Augustus/scripts/join_aug_pred.pl

[M-Asim-Javed]

Hello,

I am running BRAKER3 using Singularity container but encountered an error during Augustus gene prediction twice. Below are the details: Please assist me with resolving this issue.

Thank you.

Error

ERROR in file /opt/BRAKER/scripts/braker.pl at line 7895 Failed to execute /usr/bin/perl /opt/Augustus/scripts/join_aug_pred.pl < /home/mujav/braker3/braker/augustus.tmp1.gff > /home/mujav/braker3/braker/augustus.hints.gff

My command:

singularity exec braker3.sif braker.pl --useexisting --genome=pbe3_genomic.fa --species=pb --prot_seq=proteins.fa --rnaseq_sets_ids=S001A93_ID1,S001E7A_ID2,21dpi_ID3,7dpi_ID4 --rnaseq_sets_dirs=/home/mujav/braker3/rnaseq --threads 20 --busco_lineage eukaryota_odb10 >& pbe3_3.log

braker.log file

# BRAKER CALL: /opt/BRAKER/scripts/braker.pl --useexisting --genome=pbe3_genomic.fa --species=pb --prot_seq=proteins.fa --rnaseq_sets_ids=S001A93_ID1,S001E7A_ID2,21dpi_ID3,7dpi_ID4 --rnaseq_sets_dirs=/home/mujav/braker3/rnaseq --threads 20 --busco_lineage eukaryota_odb10
# Sat Apr 20 10:47:51 2024: braker.pl version 3.0.8
# Sat Apr 20 10:47:51 2024:Both protein and RNA-Seq data in input detected. BRAKER will be executed in ETP mode (BRAKER3).
#*********
# Sat Apr 20 10:47:51 2024: Configuring of BRAKER for using external tools...
# Sat Apr 20 10:47:51 2024: Searching for local files of RNA-Seq sets in /home/mujav/braker3/rnaseq ...
# Sat Apr 20 10:47:51 2024 Found paired RNA-Seq reads /home/mujav/braker3/rnaseq/S001A93_ID1_1.fastq, /home/mujav/braker3/rnaseq/S001A93_ID1_2.fastq for S001A93_ID1.
# Sat Apr 20 10:47:51 2024 Found paired RNA-Seq reads /home/mujav/braker3/rnaseq/S001E7A_ID2_1.fastq, /home/mujav/braker3/rnaseq/S001E7A_ID2_2.fastq for S001E7A_ID2.
# Sat Apr 20 10:47:51 2024 Found unpaired RNA-Seq reads /home/mujav/braker3/rnaseq/21dpi_ID3.fastq for 21dpi_ID3.
# Sat Apr 20 10:47:51 2024 Found unpaired RNA-Seq reads /home/mujav/braker3/rnaseq/7dpi_ID4.fastq for 7dpi_ID4.
# Sat Apr 20 10:47:51 2024: Trying to set $AUGUSTUS_CONFIG_PATH...
# Sat Apr 20 10:47:51 2024: Found environment variable $AUGUSTUS_CONFIG_PATH.
# Sat Apr 20 10:47:51 2024: Checking /opt/Augustus/config/ as potential path for $AUGUSTUS_CONFIG_PATH.
# Sat Apr 20 10:47:51 2024: Success! Setting $AUGUSTUS_CONFIG_PATH to /opt/Augustus/config/!
# Sat Apr 20 10:47:51 2024: WARNING: in file /opt/BRAKER/scripts/braker.pl at line 1931
AUGUSTUS_CONFIG_PATH/species (in this case /opt/Augustus/config//species) is not writeable. BRAKER will try to copy the AUGUSTUS config directory to a writeable location.
# Sat Apr 20 10:47:51 2024: Trying to set $AUGUSTUS_BIN_PATH...
# Sat Apr 20 10:47:51 2024: Found environment variable $AUGUSTUS_BIN_PATH.
# Sat Apr 20 10:47:51 2024: Checking /opt/Augustus/bin/ as potential path for $AUGUSTUS_BIN_PATH.
# Sat Apr 20 10:47:51 2024: Success! Setting $AUGUSTUS_BIN_PATH to /opt/Augustus/bin/!
# Sat Apr 20 10:47:51 2024: Trying to set $AUGUSTUS_SCRIPTS_PATH...
# Sat Apr 20 10:47:51 2024: Found environment variable $AUGUSTUS_SCRIPTS_PATH.
# Sat Apr 20 10:47:51 2024: Checking /opt/Augustus/scripts/ as potential path for $AUGUSTUS_SCRIPTS_PATH.
# Sat Apr 20 10:47:51 2024: Success! Setting $AUGUSTUS_SCRIPTS_PATH to /opt/Augustus/scripts/!
# Sat Apr 20 10:47:51 2024: WARNING: BRAKER will copy the
 AUGUSTUS_CONFIG folder into your home directory!
# Sat Apr 20 10:47:51 2024: WARNING: $AUGUSTUS_CONFIG_PATH/species (in this case /opt/Augustus/config//species ) is not writeable.
*** IMPORTANT: Resetting $AUGUSTUS_CONFIG_PATH=/home/mujav/.augustus because BRAKER requires a writable location!
# Sat Apr 20 10:47:51 2024: Trying to set $PYTHON3_PATH...
# Sat Apr 20 10:47:51 2024: Did not find environment variable $PYTHON3_PATH.
# Sat Apr 20 10:47:51 2024: Trying to guess PYTHON3_PATH from location of python3 executable that is available in your $PATH
# Sat Apr 20 10:47:51 2024: Checking /opt/conda/bin as potential path for $PYTHON3_PATH.
# Sat Apr 20 10:47:51 2024: Success! Setting $PYTHON3_PATH to /opt/conda/bin!
# Sat Apr 20 10:47:51 2024: Trying to set $COMPLEASM_PATH...
# Sat Apr 20 10:47:51 2024: Did not find environment variable $COMPLEASM_PATH.
# Sat Apr 20 10:47:51 2024: Trying to guess COMPLEASM_PATH from location of compleasm.py executable that is available in your $PATH
# Sat Apr 20 10:47:51 2024: Checking /opt/compleasm_kit as potential path for $COMPLEASM_PATH.
# Sat Apr 20 10:47:51 2024: Success! Setting $COMPLEASM_PATH to /opt/compleasm_kit!
# Sat Apr 20 10:47:51 2024: Trying to set $GENEMARK_PATH...
# Sat Apr 20 10:47:51 2024: Found environment variable $GENEMARK_PATH.
# Sat Apr 20 10:47:51 2024: Checking /opt/ETP/bin as potential path for $GENEMARK_PATH.
# Sat Apr 20 10:47:51 2024: Success! Setting $GENEMARK_PATH to /opt/ETP/bin!
# Sat Apr 20 10:47:51 2024: Trying to set $HISAT2_PATH...
# Sat Apr 20 10:47:51 2024: Did not find environment variable $HISAT2_PATH.
# Sat Apr 20 10:47:51 2024: Trying to guess HISAT2_PATH from location of hisat2 executable that is available in your $PATH
# Sat Apr 20 10:47:51 2024: Checking /opt/ETP/tools as potential path for $HISAT2_PATH.
# Sat Apr 20 10:47:51 2024: Success! Setting $HISAT2_PATH to /opt/ETP/tools!
# Sat Apr 20 10:47:51 2024: Trying to set $BAMTOOLS_PATH...
# Sat Apr 20 10:47:51 2024: Did not find environment variable $BAMTOOLS_PATH.
# Sat Apr 20 10:47:51 2024: Trying to guess BAMTOOLS_PATH from location of bamtools executable that is available in your $PATH
# Sat Apr 20 10:47:51 2024: Checking /usr/bin as potential path for $BAMTOOLS_PATH.
# Sat Apr 20 10:47:51 2024: Success! Setting $BAMTOOLS_PATH to /usr/bin!
# Sat Apr 20 10:47:51 2024: Trying to set $SAMTOOLS_PATH...
# Sat Apr 20 10:47:51 2024: Did not find environment variable $SAMTOOLS_PATH.
# Sat Apr 20 10:47:51 2024: Trying to guess SAMTOOLS_PATH from location of samtools executable that is available in your $PATH
# Sat Apr 20 10:47:51 2024: Checking /usr/bin as potential path for $SAMTOOLS_PATH.
# Sat Apr 20 10:47:51 2024: Success! Setting $SAMTOOLS_PATH to /usr/bin!
# Sat Apr 20 10:47:51 2024: Trying to set $DIAMOND_PATH...
# Sat Apr 20 10:47:51 2024: Did not find environment variable $DIAMOND_PATH.
# Sat Apr 20 10:47:51 2024: Trying to guess DIAMOND_PATH from location of diamond executable that is available in your $PATH
# Sat Apr 20 10:47:51 2024: Checking /opt/ETP/tools as potential path for $DIAMOND_PATH.
# Sat Apr 20 10:47:51 2024: Success! Setting $DIAMOND_PATH to /opt/ETP/tools!
# Sat Apr 20 10:47:51 2024: Trying to set $PROTHINT_PATH...
# Sat Apr 20 10:47:51 2024: Did not find environment variable $PROTHINT_PATH.
# Sat Apr 20 10:47:51 2024: Trying to guess PROTHINT_PATH from location of prothint.py executable that is available in your $PATH
# Sat Apr 20 10:47:51 2024: Checking /opt/ETP/bin/gmes/ProtHint/bin as potential path for $PROTHINT_PATH.
# Sat Apr 20 10:47:51 2024: Success! Setting $PROTHINT_PATH to /opt/ETP/bin/gmes/ProtHint/bin!
# Sat Apr 20 10:47:51 2024: Trying to set $TSEBRA_PATH...
# Sat Apr 20 10:47:51 2024: Did not find environment variable $TSEBRA_PATH.
# Sat Apr 20 10:47:51 2024: Trying to guess TSEBRA_PATH from location of tsebra.py executable that is available in your $PATH
# Sat Apr 20 10:47:51 2024: Checking /opt/TSEBRA/bin as potential path for $TSEBRA_PATH.
# Sat Apr 20 10:47:51 2024: Success! Setting $TSEBRA_PATH to /opt/TSEBRA/bin!
# Sat Apr 20 10:47:51 2024: Trying to set $CDBTOOLS_PATH...
# Sat Apr 20 10:47:51 2024: Did not find environment variable $CDBTOOLS_PATH.
# Sat Apr 20 10:47:51 2024: Trying to guess CDBTOOLS_PATH from location of cdbfasta executable that is available in your $PATH
# Sat Apr 20 10:47:51 2024: Checking /opt/cdbfasta as potential path for $CDBTOOLS_PATH.
# Sat Apr 20 10:47:51 2024: Success! Setting $CDBTOOLS_PATH to /opt/cdbfasta!
#**********************************************************************************
#                               CREATING DIRECTORY STRUCTURE                       
#**********************************************************************************
# Sat Apr 20 10:47:51 2024: create working directory /home/mujav/braker3/braker.
mkdir /home/mujav/braker3/braker
# Sat Apr 20 10:47:51 2024: create working directory /home/mujav/braker3/braker/rnaseq for RNA-Seq libraries.
mkdir /home/mujav/braker3/braker/rnaseq
# Sat Apr 20 10:47:51 2024: creating file that contains citations for this BRAKER run at /home/mujav/braker3/braker/what-to-cite.txt...
# Sat Apr 20 10:47:51 2024: create working directory /home/mujav/braker3/braker/GeneMark-ETP.
mkdir /home/mujav/braker3/braker/GeneMark-ETP
# Sat Apr 20 10:47:51 2024: create working directory /home/mujav/braker3/braker/species
mkdir /home/mujav/braker3/braker/species
# Sat Apr 20 10:47:51 2024: create working directory /home/mujav/braker3/braker/errors
mkdir /home/mujav/braker3/braker/errors
# Sat Apr 20 10:47:52 2024: changing into working directory /home/mujav/braker3/braker
cd /home/mujav/braker3/braker
# Sat Apr 20 10:47:52 2024: getting GC content of the genome
/opt/BRAKER/scripts/get_gc_content.py --sequences /home/mujav/braker3/pbe3_genomic.fa --print_sequence_length 1> /home/mujav/braker3/braker/gc_content.out 2> /home/mujav/braker3/braker/errors/gc_content.stderr
# Sat Apr 20 10:47:55 2024: Creating parameter template files for AUGUSTUS with new_species.pl
# Sat Apr 20 10:47:55 2024: check_fasta_headers(): Checking fasta headers of file /home/mujav/braker3/pbe3_genomic.fa
# Sat Apr 20 10:47:57 2024: check_fasta_headers(): Checking fasta headers of file /home/mujav/braker3/proteins.fa
# Sat Apr 20 10:47:57 2024: Assuming that this is not a DNA fasta file because other characters than A, T, G, C, N, a, t, g, c, n were contained. If this is supposed to be a DNA fasta file, check the content of your file! If this is supposed to be a protein fasta file, please ignore this message!
# Sat Apr 20 10:47:57 2024: Assuming that this is not a protein fasta file because other characters than AaRrNnDdCcEeQqGgHhIiLlKkMmFfPpSsTtWwYyVvBbZzJjOoUuXx were contained. If this is supposed to be DNA fasta file, please ignore this message.
#*********
 WARNING: empty line was removed! This warning will be supressed from now on!
#*********
#*********
# WARNING: Detected | in fasta header of file /home/mujav/braker3/proteins.fa. This may later on cause problems! The pipeline will create a new file without spaces or "|" characters and a genome_header.map file to look up the old and new headers. This message will be suppressed from now on!
#*********
# Sat Apr 20 10:51:40 2024: create working directory /home/mujav/braker3/braker/rnaseq/hisat2/ for downloading raw RNA-Seq sets.
mkdir /home/mujav/braker3/braker/rnaseq/hisat2/
# Sat Apr 20 10:51:40 2024: Building genome index for HISAT2.
hisat2-build -p 20 /home/mujav/braker3/braker/genome.fa genome 1> /home/mujav/braker3/braker/errors/hisat2-build.stdout 2> /home/mujav/braker3/braker/errors/hisat2-build.stderr
# Sat Apr 20 10:51:58 2024: Mapping RNA-Seq reads to the genome...
# Sat Apr 20 10:51:58 2024: Mapping 21dpi_ID3 ...
hisat2 -x genome -U /home/mujav/braker3/rnaseq/21dpi_ID3.fastq --dta -p 20 -S/home/mujav/braker3/braker/rnaseq/hisat2//21dpi_ID3.sam  1> /home/mujav/braker3/braker/errors/hisat2.21dpi_ID3.stdout 2> /home/mujav/braker3/braker/errors/hisat2.21dpi_ID3.stderr
# Sat Apr 20 11:05:08 2024: Converting SAM file into a sorted BAM file for 21dpi_ID3.
/usr/bin//samtools sort -o /home/mujav/braker3/braker/rnaseq/hisat2//21dpi_ID3.bam -@ 20 /home/mujav/braker3/braker/rnaseq/hisat2//21dpi_ID3.sam 1> /home/mujav/braker3/braker/errors/samtools.21dpi_ID3.stdout 2> /home/mujav/braker3/braker/errors/samtools.21dpi_ID3.stderr
# Sat Apr 20 11:05:47 2024: Mapping 7dpi_ID4 ...
hisat2 -x genome -U /home/mujav/braker3/rnaseq/7dpi_ID4.fastq --dta -p 20 -S/home/mujav/braker3/braker/rnaseq/hisat2//7dpi_ID4.sam  1> /home/mujav/braker3/braker/errors/hisat2.7dpi_ID4.stdout 2> /home/mujav/braker3/braker/errors/hisat2.7dpi_ID4.stderr
# Sat Apr 20 11:21:18 2024: Converting SAM file into a sorted BAM file for 7dpi_ID4.
/usr/bin//samtools sort -o /home/mujav/braker3/braker/rnaseq/hisat2//7dpi_ID4.bam -@ 20 /home/mujav/braker3/braker/rnaseq/hisat2//7dpi_ID4.sam 1> /home/mujav/braker3/braker/errors/samtools.7dpi_ID4.stdout 2> /home/mujav/braker3/braker/errors/samtools.7dpi_ID4.stderr
# Sat Apr 20 11:22:03 2024: Mapping S001A93_ID1 ...
hisat2 -x genome -1 /home/mujav/braker3/rnaseq/S001A93_ID1_1.fastq -2 /home/mujav/braker3/rnaseq/S001A93_ID1_2.fastq --dta -p 20 -S/home/mujav/braker3/braker/rnaseq/hisat2//S001A93_ID1.sam 1> /home/mujav/braker3/braker/errors/hisat2.S001A93_ID1.stdout 2> /home/mujav/braker3/braker/errors/hisat2.S001A93_ID1.stderr
# Sat Apr 20 11:28:51 2024: Converting SAM file into a sorted BAM file for S001A93_ID1.
/usr/bin//samtools sort -o /home/mujav/braker3/braker/rnaseq/hisat2//S001A93_ID1.bam -@ 20 /home/mujav/braker3/braker/rnaseq/hisat2//S001A93_ID1.sam 1> /home/mujav/braker3/braker/errors/samtools.S001A93_ID1.stdout 2> /home/mujav/braker3/braker/errors/samtools.S001A93_ID1.stderr
# Sat Apr 20 11:29:37 2024: Mapping S001E7A_ID2 ...
hisat2 -x genome -1 /home/mujav/braker3/rnaseq/S001E7A_ID2_1.fastq -2 /home/mujav/braker3/rnaseq/S001E7A_ID2_2.fastq --dta -p 20 -S/home/mujav/braker3/braker/rnaseq/hisat2//S001E7A_ID2.sam 1> /home/mujav/braker3/braker/errors/hisat2.S001E7A_ID2.stdout 2> /home/mujav/braker3/braker/errors/hisat2.S001E7A_ID2.stderr
# Sat Apr 20 11:37:24 2024: Converting SAM file into a sorted BAM file for S001E7A_ID2.
/usr/bin//samtools sort -o /home/mujav/braker3/braker/rnaseq/hisat2//S001E7A_ID2.bam -@ 20 /home/mujav/braker3/braker/rnaseq/hisat2//S001E7A_ID2.sam 1> /home/mujav/braker3/braker/errors/samtools.S001E7A_ID2.stdout 2> /home/mujav/braker3/braker/errors/samtools.S001E7A_ID2.stderr
# Sat Apr 20 11:38:19 2024: Running compleasm and converting the output to hints
/opt/conda/bin/python3 /opt/Augustus/scripts/compleasm_to_hints.py -p /opt/compleasm_kit/compleasm.py -g /home/mujav/braker3/braker/genome.fa -d eukaryota_odb10 -t 20 -o /home/mujav/braker3/braker/compleasm_hints.gff 1> /home/mujav/braker3/braker/errors/compleasm_to_hints.stderr 2>&1
# Sat Apr 20 11:55:33 2024: Generating hints from compleasm (genome level) finished.
#**********************************************************************************
#                              RUNNING GENEMARK-EX                                 
#**********************************************************************************
# Sat Apr 20 11:55:33 2024: Preparing genemark_evidence file hints from manual hints...
# Sat Apr 20 11:55:33 2024: Running GeneMark-ETP
# Sat Apr 20 11:55:33 2024: changing into GeneMark-ETP directory /home/mujav/braker3/braker/GeneMark-ETP
cd /home/mujav/braker3/braker/GeneMark-ETP
# Sat Apr 20 11:55:33 2024: sorting RNA-Seq BAM files
/usr/bin/samtools sort /home/mujav/braker3/braker/rnaseq/hisat2//21dpi_ID3.bam -@ 19 -o /home/mujav/braker3/braker/GeneMark-ETP/etp_data/21dpi_ID3.bam 1> /home/mujav/braker3/braker/errors/samtools.sort.21dpi_ID3.stdout 2> /home/mujav/braker3/braker/errors/samtools.sort.21dpi_ID3.stderr
/usr/bin/samtools sort /home/mujav/braker3/braker/rnaseq/hisat2//7dpi_ID4.bam -@ 19 -o /home/mujav/braker3/braker/GeneMark-ETP/etp_data/7dpi_ID4.bam 1> /home/mujav/braker3/braker/errors/samtools.sort.7dpi_ID4.stdout 2> /home/mujav/braker3/braker/errors/samtools.sort.7dpi_ID4.stderr
/usr/bin/samtools sort /home/mujav/braker3/braker/rnaseq/hisat2//S001A93_ID1.bam -@ 19 -o /home/mujav/braker3/braker/GeneMark-ETP/etp_data/S001A93_ID1.bam 1> /home/mujav/braker3/braker/errors/samtools.sort.S001A93_ID1.stdout 2> /home/mujav/braker3/braker/errors/samtools.sort.S001A93_ID1.stderr
/usr/bin/samtools sort /home/mujav/braker3/braker/rnaseq/hisat2//S001E7A_ID2.bam -@ 19 -o /home/mujav/braker3/braker/GeneMark-ETP/etp_data/S001E7A_ID2.bam 1> /home/mujav/braker3/braker/errors/samtools.sort.S001E7A_ID2.stdout 2> /home/mujav/braker3/braker/errors/samtools.sort.S001E7A_ID2.stderr
# Sat Apr 20 11:59:45 2024: Running gmetp.pl
/usr/bin/perl /opt/ETP/bin/gmetp.pl --cfg /home/mujav/braker3/braker/GeneMark-ETP/etp_config.yaml --workdir /home/mujav/braker3/braker/GeneMark-ETP --bam /home/mujav/braker3/braker/GeneMark-ETP/etp_data/ --cores 20 --softmask  1>/home/mujav/braker3/braker/errors/GeneMark-ETP.stdout 2>/home/mujav/braker3/braker/errors/GeneMark-ETP.stderr
# Sat Apr 20 17:51:47 2024: change to working directory /home/mujav/braker3/braker
cd /home/mujav/braker3/braker
# Sat Apr 20 17:51:47 2024: Getting hints for AUGUSTUS from GeneMark-ETP
/opt/conda/bin/python3 /opt/BRAKER/scripts/get_etp_hints.py --genemark_scripts /opt/ETP/bin --out /home/mujav/braker3/braker/hintsfile.gff --etp_wdir /home/mujav/braker3/braker/GeneMark-ETP 1> /home/mujav/braker3/braker/errors/get_etp_hints.stdout 2> /home/mujav/braker3/braker/errors/get_etp_hints.stderr
# Sat Apr 20 17:51:48 2024: Checking for hints of src=C and with grp tags that should not be joined according to multiplicity
# Sat Apr 20 17:51:48 2024: Joining hints that are identical (& from the same source) into multiplicity hints (input file /home/mujav/braker3/braker/tmp_merge_hints.gff)
# Sat Apr 20 17:51:48 2024: sort hints of type all
cat /home/mujav/braker3/braker/tmp_merge_hints.gff | sort -n -k 4,4 | sort -s -n -k 5,5 | sort -s -n -k 3,3 | sort -s -k 1,1 >/home/mujav/braker3/braker/hints.all.temp.sort.gff
# Sat Apr 20 17:51:50 2024: join multiple hints
/usr/bin/perl /opt/Augustus/scripts/join_mult_hints.pl </home/mujav/braker3/braker/hints.all.temp.sort.gff >/home/mujav/braker3/braker/tmp_merge_hints.gff 2>/home/mujav/braker3/braker/errors/join_mult_hints.all.stderr
cat /home/mujav/braker3/braker/tmp_merge_hints.gff /home/mujav/braker3/braker/tmp_no_merge_hints.gff > /home/mujav/braker3/braker/hintsfile.gff
# Sat Apr 20 17:51:53 2024: GeneMark-ETP run finished.
# Sat Apr 20 17:51:53 2024: link GeneMark-ETP training genes to /home/mujav/braker3/braker/GeneMark-ETP/training.gtf
ln -s /home/mujav/braker3/braker/GeneMark-ETP/proteins.fa/model/training.gtf /home/mujav/braker3/braker/GeneMark-ETP/training.gtf
# Sat Apr 20 17:51:53 2024: sorting GeneMark predictions in /home/mujav/braker3/braker/GeneMark-ETP/training.gtf
/opt/conda/bin/python3 /opt/BRAKER/scripts/sortGeneMark.py /home/mujav/braker3/braker/GeneMark-ETP/training.gtf > /home/mujav/braker3/braker/GeneMark-ETP/training.gtf.sorted 2> /home/mujav/braker3/braker/errors/sortGeneMark.err
mv /home/mujav/braker3/braker/GeneMark-ETP/training.gtf.sorted /home/mujav/braker3/braker/GeneMark-ETP/training.gtf
#**********************************************************************************
#                               TRAIN AUGUSTUS                                     
#**********************************************************************************
# Sat Apr 20 17:51:54 2024: training AUGUSTUS
# Sat Apr 20 17:51:54 2024: Copying optimized parameters to working directory /home/mujav/braker3/braker/species
cp -r /home/mujav/.augustus/species/pb /home/mujav/braker3/braker/species
#**********************************************************************************
#                               PREDICTING GENES WITH AUGUSTUS (NO UTRS)           
#**********************************************************************************
# Sat Apr 20 17:51:54 2024: RUNNING AUGUSTUS
# Sat Apr 20 17:51:54 2024: Preparing genome for running AUGUSTUS in parallel
# Sat Apr 20 17:51:54 2024: Creating directory for storing AUGUSTUS files (hints, temporarily) /home/mujav/braker3/braker/genome_split.
# Sat Apr 20 17:51:54 2024: splitting genome file in smaller parts for parallel execution of AUGUSTUS prediction
/usr/bin/perl /opt/Augustus/scripts/splitMfasta.pl /home/mujav/braker3/braker/genome.fa --outputpath=/home/mujav/braker3/braker/genome_split 2>/home/mujav/braker3/braker/errors/splitMfasta.stderr
cd /home/mujav/braker3/braker/genome_split; for f in genome.split.*; do NAME=`grep ">" $f`; mv $f ${NAME#>}.fa; done; cd ..
# Sat Apr 20 17:51:55 2024: Split genome file in 20 parts, finished.
# Sat Apr 20 17:51:55 2024: copy extrinsic file /opt/BRAKER/scripts/cfg/etp.cfg to working directory
cp /opt/BRAKER/scripts/cfg/etp.cfg /home/mujav/braker3/braker/species/pb/ex1.cfg
# Sat Apr 20 17:51:55 2024: Making AUGUSTUS jobs with hintsfile /home/mujav/braker3/braker/hintsfile.gff, cfgFile /opt/BRAKER/scripts/cfg/etp.cfg, UTR status off, and hintId hints
# Sat Apr 20 17:51:55 2024: Creating directory for storing AUGUSTUS files (hints, temporarily) /home/mujav/braker3/braker/augustus_tmp.
# Sat Apr 20 17:51:55 2024: creating /home/mujav/braker3/braker/aug_hints.lst for AUGUSTUS jobs
# Sat Apr 20 17:51:55 2024: creating AUGUSTUS jobs (with hints)
cd /home/mujav/braker3/braker
/usr/bin/perl /opt/Augustus/scripts/createAugustusJoblist.pl --sequences=/home/mujav/braker3/braker/aug_hints.lst --wrap="#!/bin/bash" --overlap=500000 --chunksize=2500000 --outputdir=/home/mujav/braker3/braker/augustus_tmp --joblist=/home/mujav/braker3/braker/hints.job.lst --jobprefix=aug_hints_ --partitionHints --command "/opt/Augustus/bin//augustus --species=pb --AUGUSTUS_CONFIG_PATH=/home/mujav/.augustus --extrinsicCfgFile=/opt/BRAKER/scripts/cfg/etp.cfg --alternatives-from-evidence=true --UTR=off --exonnames=on --codingseq=on --allow_hinted_splicesites=gcag,atac  --softmasking=1" 2>/home/mujav/braker3/braker/errors/createAugustusJoblist_hints.stderr
cd ..
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS jobs from /home/mujav/braker3/braker/hints.job.lst
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 1
bash /home/mujav/braker3/braker/aug_hints_1
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 2
bash /home/mujav/braker3/braker/aug_hints_2
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 3
bash /home/mujav/braker3/braker/aug_hints_3
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 4
bash /home/mujav/braker3/braker/aug_hints_4
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 5
bash /home/mujav/braker3/braker/aug_hints_5
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 6
bash /home/mujav/braker3/braker/aug_hints_6
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 7
bash /home/mujav/braker3/braker/aug_hints_7
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 8
bash /home/mujav/braker3/braker/aug_hints_8
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 9
bash /home/mujav/braker3/braker/aug_hints_9
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 10
bash /home/mujav/braker3/braker/aug_hints_10
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 11
bash /home/mujav/braker3/braker/aug_hints_11
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 12
bash /home/mujav/braker3/braker/aug_hints_12
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 13
bash /home/mujav/braker3/braker/aug_hints_13
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 14
bash /home/mujav/braker3/braker/aug_hints_14
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 15
bash /home/mujav/braker3/braker/aug_hints_15
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 16
bash /home/mujav/braker3/braker/aug_hints_16
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 17
bash /home/mujav/braker3/braker/aug_hints_17
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 18
bash /home/mujav/braker3/braker/aug_hints_18
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 19
bash /home/mujav/braker3/braker/aug_hints_19
# Sat Apr 20 17:51:56 2024: Running AUGUSTUS job 20
bash /home/mujav/braker3/braker/aug_hints_20
# Sat Apr 20 17:51:57 2024: Joining AUGUSTUS predictions in directory /home/mujav/braker3/braker/augustus_tmp
# Sat Apr 20 17:51:57 2024: Concatenating AUGUSTUS output files in /home/mujav/braker3/braker/augustus_tmp
/usr/bin/perl /opt/Augustus/scripts/join_aug_pred.pl < /home/mujav/braker3/braker/augustus.tmp1.gff > /home/mujav/braker3/braker/augustus.hints.gff

[KatharinaHoff]

This is an AUGUSTUS issue. We would need the output of /home/mujav/braker3/braker/augustus.tmp1.gff, /home/mujav/braker3/braker/augustus.hints.gff, and we may require the input files to that step in order to investigate the cause. (This may not be resolved very fast.)

[M-Asim-Javed]

Hello,

Thanks for the immediate response.

The directory /home/mujav/braker3/braker/augustus.tmp is empty, and neither the file augustus.tmp1.gff nor augustus.hints.gff was generated. I am sharing screenshots of the output files and attaching two documents: tmp_merge_hints.gff and hintsfile.gff, which were generated up to this step. Please let me know if there are any other files I should share.

braker3.zip

[M-Asim-Javed]

Hello,

There was an installation issue where Augustus couldn't create the species directory. So, I removed everything and reinstalled it with the Singularity container, and now it works perfectly. Thanks for the immediate response and assistance.