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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

Mutect2 variant calling output with tumor and matched normal: 0/1 in all variants called

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    David Benjamin

    @RicPin You need to specify which sample is normal, in this case with `-normal ESO08N`.  Mutect2 treats any sample not specified as normal as a tumor.  That is, you were unwittingly running Mutect2 in multiple-tumor mode.

    By the way, `--callable-depth` does something different from what you would expect.  The Javadoc describes it as: "Minimum depth to be considered callable for Mutect stats. Does not affect genotyping."

    Also, we strongly recommend using our best practices resources for the `--germline-resource` and `--pon` (panel of normals) arguments.  You can obtain them at gs://gatk-best-practices/somatic-b37/Mutect2-WGS-panel-b37.vcf / gs://gatk-best-practices/somatic-b37/Mutect2-exome-panel.vcf and gs://gatk-best-practices/somatic-b37/af-only-gnomad.raw.sites.vcf.

    Finally, don't forget to run FilterMutectCalls.  Mutect2 outputs unfiltered calls, the great majority of which are false positives.

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    RicPin

    Thanks for your reply @David Benjamin. It has helped a lot.

    However, the links you've posted before are suitable for the b37 reference genome, and I am working with the h19. Can I use them anyway in my analyses?

    Best regards

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    David Benjamin

    The resources will work with hg19.  Occasionally one encounters versions with a "chr1" vs "1" naming convention.  In such cases, you can manually delete all "chrs" -- for example, in vim the command :%s/chr//g does the job -- and then reindex.

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    RicPin

    Sorry, but I can't find the file gs://gatk-best-practices/somatic-b37/Mutect2-exome-panel.vcf in GATK FTP Server. Is that the correct filename?

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    David Benjamin

    Anything starting with gs:// is in a google cloud bucket.  GATK tools can read from them directly, or you can download them with gsutil cp.

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