GeneMark-ETP didn't receive enough evidence from the input data

[RezwanCAAS]

Hi braker team, I am running this code using iso-seq file, having 100% mapping with the genome file can I add all RNA-Seq and iso-seq files in bam format in --bam ?

module load singularity/3.9.7 braker/3.0.3/singularity

singularity exec -B $PWD,$HOME,/ibex,/sw /ibex/sw/rl9c/braker/3.0.3/singularity/braker3_latest.sif braker.pl \
 --workingdir=~/braker_isoseq4/ \
 --genome=~/genome.fasta.masked \
 --prot_seq=~/annotation/protein_sequences/ghb_db_cg_os_le.fa \
 --bam=~/annotation/isoseq/isoseq_cleaning/isoseq_aligned.bam \
 --crf --threads=32 --species=isoseq2 --useexisting --gff3 --Augustus_ab_initio --busco_lineage=eudicots_odb10

I have this error

ERROR in file /opt/BRAKER/scripts/braker.pl at line 5575
The most common problem is that GeneMark-ETP didn't receive enough evidence from the input data, in this case, see errors/GeneMark-ETP.stderr!

[tariqr@login509-02-r errors]$ cat GeneMark-ETP.stderr
FASTA index file /genome.softmasked.fasta.fai created.
Use of uninitialized value $ph1 in addition (+) at /opt/ETP/bin/gmes/parse_set.pl line 205.
Use of uninitialized value $ph0 in addition (+) at /opt/ETP/bin/gmes/parse_set.pl line 205.
Use of uninitialized value $ph2 in addition (+) at /opt/ETP/bin/gmes/parse_set.pl line 205.
Use of uninitialized value $ph0 in division (/) at /opt/ETP/bin/gmes/parse_set.pl line 208.
Illegal division by zero at /opt/ETP/bin/gmes/parse_set.pl line 208.
Illegal division by zero at /opt/ETP/bin/train_super.pl line 184.
error, file/folder not found: /ibex/project/c2141/dragon-fruit/hifireads_condor/path/proteins.fa/model/output.mod

Please suggest how to fix this

[yyliang12]

Hi,

You can just simply add multiple .bam files after arg --bam with comma separated (eg --bam aln1.bam,aln2.bam). Or you can use samtools merge to combine all .bam files together as a single input for --bam option. Btw, be aware of the incorrect args which placed after singularity -B. The right one might be /path/on/the/host:/path/to/bind/in/the/container.

Best, Yanyang.

[KatharinaHoff]

Please be aware that there's a separate container for isoseq processing. I am not sure what image you are using (if it is latest, it is the wrong one). BRAKER currently does not perform co-assembly of short and long read, together. So pick one type of data source, use the correct container for it. You could run BRAKER twice, once for each expression data type, and then merge with TSEBRA.

[zilov]

Hello! I am also getting same error message on short-reads RNA + proteins mode.

Build with: singularity build braker3.sif docker://teambraker/braker3:latest

Run with: singularity exec -B ${PWD}:${PWD} braker3.sif braker.pl --genome=assembly_FINAL_softmasked_denovo_and_rodent.fasta --prot_seq=Vertebrata.fa --bam=./bams/merged.bam -threads=48 --AUGUSTUS_CONFIG_PATH=${PWD}/augustus_config --species=sr2 --verbosity=4 --gff3

Bam files were generated with hisat2 and merged with samtools, here is an example of command: hisat2 -p 50 -x assembly_FINAL_softmasked_denovo_and_rodent -U AE1.fq.gz --no-spliced-alignment | samtools view --threads 50 -Sbh -o bams/AE1.fq.bam

I've tried running with both a single merged BAM file and a list of BAM files, but the error persists.

Could you please advise on how to resolve this issue?

[zilov]

The issue turned out to be on my side. I was using the wrong type of data for genome annotation. Previous massage is not relevant.