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GATK version 4.beta.3 (i.e. the third beta release) is out. See the GATK4 beta page for download and details.

ERROR in Running GATK Unified Genotyper module?

Dear GATK Users,

I am using GATK for first time. I am doing Targeted reseq Analysis. while i am running Unified Genotyper in Galaxy interface. it gives error message as mentioned below..

"ERROR MESSAGE: The fasta file you specified (/tmp/tmp-gatk-WBrGTf) does not exist."
But i places the Fasta file path in .loc file.
Also could anyone tell what are the preprocessing steps before using Bam file for Unified Genotyper??

Your answers are very much Appericiated.



  • Geraldine_VdAuweraGeraldine_VdAuwera Cambridge, MAMember, Administrator, Broadie

    Hi Sridhar,

    To your first question, I think this is a problem specific to the Galaxy interface. You can probably get more helpful advice to deal with this from the Galaxy support mailing list.

    To your second question, have a look at our Best Practices workflow. Basically, once you have mapped your data, you need to mark duplicates, perform local realignment around indels and base recalibration. Optionally, you can also compress your data with ReduceReads.

  • Thanks Geraldine..
    i am facing new ERROR in running the UnifiedGenotyper
    java -jar galaxy/apps/GenomeAnalysisTK-2.5-2-gf57256b/GenomeAnalysisTK.jar -R ens_BRCA1.fa -T UnifiedGenotyper -I picard.sort.bam
    INFO 10:10:41,862 HelpFormatter - --------------------------------------------------------------------------------
    INFO 10:10:41,864 HelpFormatter - The Genome Analysis Toolkit (GATK) v2.5-2-gf57256b, Compiled 2013/05/01 09:27:02
    INFO 10:10:41,864 HelpFormatter - Copyright (c) 2010 The Broad Institute
    INFO 10:10:41,864 HelpFormatter - For support and documentation go to
    INFO 10:10:41,869 HelpFormatter - Program Args: -R ens_BRCA1.fa -T UnifiedGenotyper -I picard.sort.bam
    INFO 10:10:41,869 HelpFormatter - Date/Time: 2013/06/19 10:10:41
    INFO 10:10:41,869 HelpFormatter - --------------------------------------------------------------------------------
    INFO 10:10:41,870 HelpFormatter - --------------------------------------------------------------------------------
    INFO 10:10:41,959 GenomeAnalysisEngine - Strictness is SILENT
    INFO 10:10:42,044 GenomeAnalysisEngine - Downsampling Settings: Method: BY_SAMPLE, Target Coverage: 250
    INFO 10:10:42,051 SAMDataSource$SAMReaders - Initializing SAMRecords in serial
    INFO 10:10:42,066 SAMDataSource$SAMReaders - Done initializing BAM readers: total time 0.01
    INFO 10:10:44,401 GATKRunReport - Uploaded run statistics report to AWS S3

    ERROR ------------------------------------------------------------------------------------------
    ERROR A USER ERROR has occurred (version 2.5-2-gf57256b):
    ERROR The invalid arguments or inputs must be corrected before the GATK can proceed
    ERROR Please do not post this error to the GATK forum
    ERROR See the documentation (rerun with -h) for this tool to view allowable command-line arguments.
    ERROR Visit our website and forum for extensive documentation and answers to
    ERROR commonly asked questions
    ERROR MESSAGE: Input files reads and reference have incompatible contigs: Found contigs with the same name but different lengths:
    ERROR contig reads = chr17 / 81195210
    ERROR contig reference = chr17 / 125979.
    ERROR reads contigs = [chrM, chr1, chr2, chr3, chr4, chr5, chr6, chr7, chr8, chr9, chr10, chr11, chr12, chr13, chr14, chr15, chr16, chr17, chr18, chr19, chr20, chr21, chr22, chrX, chrY]
    ERROR reference contigs = [chr17]
    ERROR ------------------------------------------------------------------------------------------

    Could you please suggest the same


  • I foud the solution for this error..

  • Geraldine_VdAuweraGeraldine_VdAuwera Cambridge, MAMember, Administrator, Broadie

    Feel free to share your solution so that others who have the same problem can benefit from your findings :)

  • Hello Geraldine,
    I am using Particular target in the genome for the Analysis, but while mapping i used whole genome and converted to sam format and later i used same file and run it on GATK so i got this error..

    Now i used the Target region Fasta file for mapping and pcoceeded and founf no error.


  • Geraldine_VdAuweraGeraldine_VdAuwera Cambridge, MAMember, Administrator, Broadie

    Thanks for sharing!

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