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HaplotypeCaller does not call any variant in my targeted sequencing data

LauraDomenechLauraDomenech BarcelonaMember


I am working with targeted sequencing data and, after align it, I started running HaplotypeCaller to generate gvcf files, using the same line that I used before for exome sequencing analysis (changing the bedfile), which worked correctly. However, with the targeted sequencing data, although there are variant positions when looking at the bamfiles with igv, haplotype caller calls all positions 0/0. Those positions that are clearly variant with igv have PL values of 0,0,0 and calls 0/0, despite being 100% the same alternate allele. Alternate allele in the gvcf file is called .

This is the HaplotypeCaller line:

java -jar /GATK/GenomeAnalysisTK-3.3-0-g37228af/GenomeAnalysisTK.jar -T HaplotypeCaller -R hg19/hg19.fasta -I /$IN --dbsnp dbsnp_138.hg19.vcf -o /$INdir/$SAMPLE.haplotype.vcf.gz -L targetedbedfile.bed -A AlleleBalance -A BaseCounts -A BaseQualityRankSumTest -A ChromosomeCounts -A ClippingRankSumTest -A Coverage -A FisherStrand -A GCContent -A GenotypeSummaries -A HaplotypeScore -A HardyWeinberg -A HomopolymerRun -A InbreedingCoeff -A LikelihoodRankSumTest -A LowMQ -A MVLikelihoodRatio -A MappingQualityRankSumTest -A MappingQualityZero -A NBaseCount -A PossibleDeNovo -A QualByDepth -A RMSMappingQuality -A ReadPosRankSumTest -A SampleList -A SnpEff -A SpanningDeletions -A StrandOddsRatio -A TandemRepeatAnnotator -A TransmissionDisequilibriumTest -A VariantType -A AlleleBalanceBySample -A AlleleCountBySample -A DepthPerAlleleBySample -A DepthPerSampleHC -A MappingQualityZeroBySample -A StrandBiasBySample --emitRefConfidence GVCF -variant_index_type LINEAR -variant_index_parameter 128000 >> /users/project/PanCanRisk_005555_no_backup/PanCanRisk_Targeted/Haplotype_annotation.log_file 2>&1

Could you help me with this issue?

Let me know if you need more information.

Thank you very much,



  • SheilaSheila Broad InstituteMember, Broadie, Moderator admin

    Hi Laura,

    Can you post some IGV screenshots of sites where you think a variant should be called, but is called 0/0 with PLs all 0? Please post ~300 bases before and after the sites.


    P.S. Can you check the base qualities at those sites?

  • LauraDomenechLauraDomenech BarcelonaMember

    Sorry, I solved this issue! It was a problem of our queue nodes that where not playing well with GATK (3.3 at least).

    Thanks a lot!

    Laura Domènech

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