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Variant Discovery in High-Throughput Sequencing Data

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Developed in the Data Sciences Platform at the Broad Institute, the toolkit offers a wide variety of tools with a primary focus on variant discovery and genotyping. Its powerful processing engine and high-performance computing features make it capable of taking on projects of any size. Learn more

Call SNPs from mapped reads with MAPQ zero (bwa-mem, hg38, GATK4 Mutect2)



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    Genevieve Brandt (she/her)

    Hi Juliab, please post the error message so that we can get Mutect2 to work for you. There have been many changes to Mutect2 since version, I would recommend updating to our current version,

    Unfortunately we don't have any recommendations for your first request, since we do not support this usage. However, if any members of the community have experience with this, please chime in here! Here is our support policy for context.

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    This is my code for Mutect2 command:

    gatk Mutect2 -R ${ref_fasta_hs} -I ${sample}_recal_sorted.bam -tumor ${sample} -pon ${pon_hg38} -L ${intervals_idt} --interval-padding 100 -O ${sample}_mutect2_disfilt.vcf --max-population-af 0.10 --base-quality-score-threshold 20 --disable-tool-default-read-filters true --min-pruning 3 --read-validation-stringency LENIENT

    This is what I obtain after INFO lines:

    [September 29, 2020 6:32:04 PM EDT] done. Elapsed time: 0.04 minutes.



    at org.broadinstitute.hellbender.utils.locusiterator.ReadStateManager.readStartsAtCurrentPosition(

    at org.broadinstitute.hellbender.utils.locusiterator.ReadStateManager.collectPendingReads(

    at org.broadinstitute.hellbender.utils.locusiterator.LocusIteratorByState.lazyLoadNextAlignmentContext(

    at org.broadinstitute.hellbender.utils.locusiterator.LocusIteratorByState.hasNext(

    at org.broadinstitute.hellbender.utils.locusiterator.IntervalAlignmentContextIterator.advanceAlignmentContext(

    at org.broadinstitute.hellbender.utils.locusiterator.IntervalAlignmentContextIterator.advanceAlignmentContextToCurrentInterval(



    at org.broadinstitute.hellbender.engine.AssemblyRegionIterator.loadNextAssemblyRegion(

    at org.broadinstitute.hellbender.engine.AssemblyRegionIterator.<init>(

    at org.broadinstitute.hellbender.engine.AssemblyRegionWalker.processReadShard(

    at org.broadinstitute.hellbender.engine.AssemblyRegionWalker.traverse(

    at org.broadinstitute.hellbender.engine.GATKTool.doWork(

    at org.broadinstitute.hellbender.cmdline.CommandLineProgram.runTool(

    at org.broadinstitute.hellbender.cmdline.CommandLineProgram.instanceMainPostParseArgs(

    at org.broadinstitute.hellbender.cmdline.CommandLineProgram.instanceMain(

    at org.broadinstitute.hellbender.Main.runCommandLineProgram(

    at org.broadinstitute.hellbender.Main.mainEntry(

    at org.broadinstitute.hellbender.Main.main(

    It is not repoted as "USER ERROR" and it generates an output file but the output file doesn't contain variant calls.

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    Genevieve Brandt (she/her)

    Juliab I'll get more information about this and let you know what the error might be.

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    Genevieve Brandt (she/her)

    Juliab, the read triggering the java.lang.NullPointerException has no contig filled in which is consistent with trying to run on MQ0 data. One solution would be to make sure that all the variants have positions, even if MQ = 0. 

    You can also run bwa mem with the -q flag to preserve qualities for supplementary/secondary alignments, that should help.

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