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Variant Discovery in High-Throughput Sequencing Data

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Haploytpe caller shows me that 0 read(s) were filtered by: MappingQualityAvailableReadFilter etc.

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    Anthony Dias-Ciarla

    Hi pb,

    Thank you for writing to the GATK forum! We hope that we can help you clarify this question.

    High MappingQualityReadFilter results don't necessarily indicate a problem with GATK. Sometimes a lot of poorly mapped reads are related to the reference/organism you are using.

    Could you please first run CountReads on your BAM file and compare the result to the filtering result you included above? From there we'll be better equipped to determine the source of your initial result.

    Best,

    Anthony

     

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    pb

    Hi Anthony,

     

    Thank you very much for the swift response from your side. I ran the CountReads on my BAM file and this was the output (see below).

     From this documentations, WellformedReadFilter – GATK (broadinstitute.org), does this then indicate that none of my reads were removed by the filter set? 

    09:51:25.276 INFO  CountReads - ------------------------------------------------------------
    09:51:25.277 INFO  CountReads - HTSJDK Version: 2.24.1
    09:51:25.277 INFO  CountReads - Picard Version: 2.27.1
    09:51:25.277 INFO  CountReads - Built for Spark Version: 2.4.5
    09:51:25.277 INFO  CountReads - HTSJDK Defaults.COMPRESSION_LEVEL : 2
    09:51:25.277 INFO  CountReads - HTSJDK Defaults.USE_ASYNC_IO_READ_FOR_SAMTOOLS : false
    09:51:25.277 INFO  CountReads - HTSJDK Defaults.USE_ASYNC_IO_WRITE_FOR_SAMTOOLS : true
    09:51:25.277 INFO  CountReads - HTSJDK Defaults.USE_ASYNC_IO_WRITE_FOR_TRIBBLE : false
    09:51:25.277 INFO  CountReads - Deflater: IntelDeflater
    09:51:25.277 INFO  CountReads - Inflater: IntelInflater
    09:51:25.278 INFO  CountReads - GCS max retries/reopens: 20
    09:51:25.278 INFO  CountReads - Requester pays: disabled
    09:51:25.278 INFO  CountReads - Initializing engine
    09:51:25.679 INFO  CountReads - Done initializing engine
    09:51:25.679 INFO  ProgressMeter - Starting traversal
    09:51:25.679 INFO  ProgressMeter -        Current Locus  Elapsed Minutes       Reads Processed     Reads/Minute
    09:51:35.681 INFO  ProgressMeter -   Pf3D7_08_v3:699293              0.2               4902000       29409059.1
    09:51:45.681 INFO  ProgressMeter -  Pf3D7_13_v3:1479591              0.3              10406000       31216439.2
    09:51:50.862 WARN  IntelInflater - Zero Bytes Written : 0
    09:51:50.865 INFO  CountReads - 0 read(s) filtered by: WellformedReadFilter 

    09:51:50.865 INFO  ProgressMeter -    Pf3D7_MIT_v3:5934              0.4              13303080       31691606.4
    09:51:50.866 INFO  ProgressMeter - Traversal complete. Processed 13303080 total reads in 0.4 minutes.
    09:51:50.866 INFO  CountReads - CountReads counted 13303080 total reads
    09:51:50.866 INFO  CountReads - Shutting down engine
    [July 15, 2022 9:51:50 AM CEST] org.broadinstitute.hellbender.tools.CountReads done. Elapsed time: 0.43 minutes.
    Runtime.totalMemory()=2076049408



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    Anthony Dias-Ciarla

    Hi pb,

    Thank you for getting back with your CountReads output! All looks perfectly normal to me. A filter output of 600,000 poorly mapped reads out of 13,000,000 is not out of the ordinary in this context.

    We generally expect a WellformedReadFilter output of zero, as it only checks for totally illegal/invalid reads that break the spec/fail quality checks.

    I hope this helps clear up any doubts. Thank you again for submitting this question to our GATK forum. Please feel free to reach out with any further questions in the future!

    Best,

    Anthony

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    pb

    Hi Anthony Dias-Ciarla,

     

    Oh I see, thanks a lot for looking into it! All is clear from my side

     

    Best,

    Patricia

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