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Unified Genotyper gets stuck

eliasozioloreliasoziolor Baylor UniversityMember

Hello,

I used bwa to map my samples to a mitochondrial genome of a non-model organism. Afterwards I careated a merged .bam file from multiple (288) sample .bams (used samtools merge and re-assigned RG tags), but when I run UnifiedGenotyper on that file it gets stuck at 32.1% and never moves forward from there. I also wanted to run RealignerTargetCreator, but I always get a truncated realigned.bam file. Any suggestions for how to troubleshoot this?

Thanks you.

Answers

  • SheilaSheila Broad InstituteMember, Broadie, Moderator

    @eliasoziolor
    Hi,

    How many contigs are in your reference genome? What do you mean you get a truncated bam file from RealignerTargetCreator? Does it give an error message? Have you tried running the tools on a single sample bam file?

    -Sheila

  • eliasozioloreliasoziolor Baylor UniversityMember

    1)There is 1 contig in my reference (it's a small mitochondrial genome).
    2)When I try to run UG on the realigned file it tells me that there is a missing EOF marker and is likely a truncated file, and it does not give me that error if I just run on the merged sorted bam file, so I assumed the error is coming from the realignment around indels since that's the only step between.
    3)yes I have done that before, but for ease of analysis I decided to merge the files before variant calling, since I wasn't able to conveniently export the data from non-merged vcf files.

  • SheilaSheila Broad InstituteMember, Broadie, Moderator

    @eliasoziolor
    Hi,

    Thanks. Can you tell me the version of GATK you are using and the exact command you ran for Realigner Target Creator and Indel Realigner?

    Thanks,
    Sheila

  • eliasozioloreliasoziolor Baylor UniversityMember
    edited July 2015

    Hello,

    I am using the latest version of GATK 3.4-46. Commands as follows:

    java -jar pathtoGATK -T RealignerTargetCreator -R pathtoreference -I pathtofile -o pathtooutput

    java -jar pathtoGATK -T IndelRealigner -R pathtoreference -I pathtofile -targetIntervals pathto.listfile -o pathtorealignedfile

    java -Xmx16g -jar pathtoGATK\
    -T UnifiedGenotyper\
    -R pathtoreference\
    -I pathtorealignedfile\
    -ploidy 1\
    -glm BOTH\
    -stand_call_conf 30.0\
    -stand_emit_conf 10.0\
    -o pathtovcf

    Thank you!

  • SheilaSheila Broad InstituteMember, Broadie, Moderator

    @eliasoziolor
    Hi,

    So, neither Realigner Target Creator nor Indel Realigner are giving an error message? They both run to completion? Can you try running Picard's Validate Sam File on the bam file output by Indel Realigner?

    Thanks,
    Sheila

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