Mutect w/ panel of normal throwing errors, even though mutect w/o panel of normal works fine.
I am getting an error running mutect2 with panel of normals on some samples, but not others. Mutect2 w/o a panel of normals works well on all the samples, and when I run validatesamfiles on the bam files that fail, I get that there are no errors. Not sure what could be causing this error.
I run the following command on all the samples:
On the samples that have an error I get the following log:
[eam255@r206u22n03.mccleary scripts]$ nano KNPH99-1.mutect.out
[eam255@r206u22n03.mccleary scripts]$ cat KNPH99-1.mutect.out
19:11:00.737 INFO NativeLibraryLoader - Loading libgkl_compression.so from jar:file:/vast/palmer/apps/avx2/software/GATK/4.4.0.0-GCCcore-10.2.0-Java-17/gatk-package-4.4.0.0-local.jar!/com/intel/gkl/native/libgkl_compre
ssion.so
19:11:00.832 INFO Mutect2 - ------------------------------------------------------------
19:11:00.838 INFO Mutect2 - The Genome Analysis Toolkit (GATK) v4.4.0.0
19:11:00.838 INFO Mutect2 - For support and documentation go to https://software.broadinstitute.org/gatk/
19:11:00.838 INFO Mutect2 - Executing as eam255@r209u11n01.mccleary.ycrc.yale.edu on Linux v4.18.0-372.52.1.el8_6.x86_64 amd64
19:11:00.838 INFO Mutect2 - Java runtime: OpenJDK 64-Bit Server VM v17.0.4+8
19:11:00.838 INFO Mutect2 - Start Date/Time: September 8, 2023 at 7:11:00 PM EDT
19:11:00.838 INFO Mutect2 - ------------------------------------------------------------
19:11:00.838 INFO Mutect2 - ------------------------------------------------------------
19:11:00.839 INFO Mutect2 - HTSJDK Version: 3.0.5
19:11:00.839 INFO Mutect2 - Picard Version: 3.0.0
19:11:00.839 INFO Mutect2 - Built for Spark Version: 3.3.1
19:11:00.839 INFO Mutect2 - HTSJDK Defaults.COMPRESSION_LEVEL : 2
19:11:00.839 INFO Mutect2 - HTSJDK Defaults.USE_ASYNC_IO_READ_FOR_SAMTOOLS : false
19:11:00.840 INFO Mutect2 - HTSJDK Defaults.USE_ASYNC_IO_WRITE_FOR_SAMTOOLS : true
19:11:00.840 INFO Mutect2 - HTSJDK Defaults.USE_ASYNC_IO_WRITE_FOR_TRIBBLE : false
19:11:00.840 INFO Mutect2 - Deflater: IntelDeflater
19:11:00.840 INFO Mutect2 - Inflater: IntelInflater
19:11:00.840 INFO Mutect2 - GCS max retries/reopens: 20
19:11:00.840 INFO Mutect2 - Requester pays: disabled
19:11:00.841 INFO Mutect2 - Initializing engine
19:11:00.913 INFO Mutect2 - Shutting down engine
[September 8, 2023 at 7:11:00 PM EDT] org.broadinstitute.hellbender.tools.walkers.mutect.Mutect2 done. Elapsed time: 0.00 minutes.
Runtime.totalMemory()=19327352832
java.lang.RuntimeException: Cannot use index file with textual SAM file
at htsjdk.samtools.SamReaderFactory$SamReaderFactoryImpl.open(SamReaderFactory.java:442)
at htsjdk.samtools.SamReaderFactory.open(SamReaderFactory.java:106)
at org.broadinstitute.hellbender.engine.ReadsPathDataSource.<init>(ReadsPathDataSource.java:245)
at org.broadinstitute.hellbender.engine.ReadsPathDataSource.<init>(ReadsPathDataSource.java:181)
at org.broadinstitute.hellbender.engine.GATKTool.initializeReads(GATKTool.java:453)
at org.broadinstitute.hellbender.engine.GATKTool.onStartup(GATKTool.java:724)
at org.broadinstitute.hellbender.engine.AssemblyRegionWalker.onStartup(AssemblyRegionWalker.java:79)
at org.broadinstitute.hellbender.cmdline.CommandLineProgram.runTool(CommandLineProgram.java:147)
at org.broadinstitute.hellbender.cmdline.CommandLineProgram.instanceMainPostParseArgs(CommandLineProgram.java:198)
at org.broadinstitute.hellbender.cmdline.CommandLineProgram.instanceMain(CommandLineProgram.java:217)
at org.broadinstitute.hellbender.Main.runCommandLineProgram(Main.java:160)
at org.broadinstitute.hellbender.Main.mainEntry(Main.java:203)
at org.broadinstitute.hellbender.Main.main(Main.java:289)
Using GATK jar /vast/palmer/apps/avx2/software/GATK/4.4.0.0-GCCcore-10.2.0-Java-17/gatk-package-4.4.0.0-local.jar
-
Based on the error message you posted It looks like some of your input files are in sam format rather than bam format. You may be able to check their bgzip compression status by just typing
cat filename | head -n 10
If you get a meaningful text header then the file is not bam but sam format. You may be able to convert them to bam using gatk PrintReads or samtools view.
I hope this helps.
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