Interval issue with Mouse Variant Calling BaseRecalibrator
I am trying to call mouse variants for grcm39 using gatk and running into the error described below. I've search the snp file and the interval queried does not exist in the file?
Thanks so much for all your assistance.
REQUIRED for all errors and issues:
a) GATK version used:
The Genome Analysis Toolkit (GATK) v4.1.8.1
b) Exact command used:
java -Dsamjdk.use_async_io_read_samtools=false -Dsamjdk.use_async_io_write_samtools=true -Dsamjdk.use_async_io_write_tribble=false -Dsamjdk.compression_level=2 -Xms4G -jar /gatk/gatk-package-4.1.8.1-local.jar BaseRecalibrator -R /cromwell_root/fc-7711ee16-4d3a-44ca-b176-8acc97dff57a/ref/GRCm39/Mus_musculus.GRCm39.dna.primary_assembly.fa -I /cromwell_root/fc-7711ee16-4d3a-44ca-b176-8acc97dff57a/submissions/66a0b5bf-1e94-4e51-bdaf-886055953977/PreProcessingForVariantDiscovery_GATK4/baa27599-1fdf-4500-9238-2f0b8dcfe023/call-SortAndFixTags/cacheCopy/20191210_AN4_BH7589_S4.grcm39.aligned.duplicate_marked.sorted.bam --use-original-qualities -O 20191210_AN4_BH7589_S4.grcm39.recal_data.csv --known-sites /cromwell_root/fc-7711ee16-4d3a-44ca-b176-8acc97dff57a/ref/GRCm39/mgp_REL2021_snps.rsID.vcf.gz --known-sites /cromwell_root/fc-7711ee16-4d3a-44ca-b176-8acc97dff57a/ref/GRCm39/mgp_REL2021_indels.rsID.vcf.gz -L 1:1+
c) Entire program log:
Picked up _JAVA_OPTIONS: -Djava.io.tmpdir=/cromwell_root/tmp.WSPzki 13:31:32.850 INFO NativeLibraryLoader - Loading libgkl_compression.so from jar:file:/gatk/gatk-package-4.1.8.1-local.jar!/com/intel/gkl/native/libgkl_compression.so 13:31:34.721 INFO BaseRecalibrator - ------------------------------------------------------------ 13:31:34.721 INFO BaseRecalibrator - The Genome Analysis Toolkit (GATK) v4.1.8.1 13:31:34.721 INFO BaseRecalibrator - For support and documentation go to https://software.broadinstitute.org/gatk/ 13:31:34.722 INFO BaseRecalibrator - Executing as root@36446dd73f8a on Linux v6.1.75+ amd64 13:31:34.723 INFO BaseRecalibrator - Java runtime: OpenJDK 64-Bit Server VM v1.8.0_242-8u242-b08-0ubuntu3~18.04-b08 13:31:34.723 INFO BaseRecalibrator - Start Date/Time: May 14, 2024 1:31:32 PM GMT 13:31:34.723 INFO BaseRecalibrator - ------------------------------------------------------------ 13:31:34.724 INFO BaseRecalibrator - ------------------------------------------------------------ 13:31:34.726 INFO BaseRecalibrator - HTSJDK Version: 2.23.0 13:31:34.726 INFO BaseRecalibrator - Picard Version: 2.22.8 13:31:34.727 INFO BaseRecalibrator - HTSJDK Defaults.COMPRESSION_LEVEL : 2 13:31:34.727 INFO BaseRecalibrator - HTSJDK Defaults.USE_ASYNC_IO_READ_FOR_SAMTOOLS : false 13:31:34.727 INFO BaseRecalibrator - HTSJDK Defaults.USE_ASYNC_IO_WRITE_FOR_SAMTOOLS : true 13:31:34.728 INFO BaseRecalibrator - HTSJDK Defaults.USE_ASYNC_IO_WRITE_FOR_TRIBBLE : false 13:31:34.728 INFO BaseRecalibrator - Deflater: IntelDeflater 13:31:34.733 INFO BaseRecalibrator - Inflater: IntelInflater 13:31:34.734 INFO BaseRecalibrator - GCS max retries/reopens: 20 13:31:34.734 INFO BaseRecalibrator - Requester pays: disabled 13:31:34.734 INFO BaseRecalibrator - Initializing engine WARNING: BAM index file /cromwell_root/fc-7711ee16-4d3a-44ca-b176-8acc97dff57a/submissions/66a0b5bf-1e94-4e51-bdaf-886055953977/PreProcessingForVariantDiscovery_GATK4/baa27599-1fdf-4500-9238-2f0b8dcfe023/call-SortAndFixTags/cacheCopy/20191210_AN4_BH7589_S4.grcm39.aligned.duplicate_marked.sorted.bai is older than BAM /cromwell_root/fc-7711ee16-4d3a-44ca-b176-8acc97dff57a/submissions/66a0b5bf-1e94-4e51-bdaf-886055953977/PreProcessingForVariantDiscovery_GATK4/baa27599-1fdf-4500-9238-2f0b8dcfe023/call-SortAndFixTags/cacheCopy/20191210_AN4_BH7589_S4.grcm39.aligned.duplicate_marked.sorted.bam 13:31:35.557 INFO FeatureManager - Using codec VCFCodec to read file file:///cromwell_root/fc-7711ee16-4d3a-44ca-b176-8acc97dff57a/ref/GRCm39/mgp_REL2021_snps.rsID.vcf.gz 13:31:35.790 INFO FeatureManager - Using codec VCFCodec to read file file:///cromwell_root/fc-7711ee16-4d3a-44ca-b176-8acc97dff57a/ref/GRCm39/mgp_REL2021_indels.rsID.vcf.gz 13:31:36.062 INFO IntervalArgumentCollection - Processing 195154279 bp from intervals 13:31:36.081 INFO BaseRecalibrator - Done initializing engine 13:31:36.083 WARN BaseRecalibrator - This tool has only been well tested on ILLUMINA-based sequencing data. For other data use at your own risk. 13:31:36.131 INFO BaseRecalibrationEngine - The covariates being used here: 13:31:36.132 INFO BaseRecalibrationEngine - ReadGroupCovariate 13:31:36.132 INFO BaseRecalibrationEngine - QualityScoreCovariate 13:31:36.132 INFO BaseRecalibrationEngine - ContextCovariate 13:31:36.133 INFO BaseRecalibrationEngine - CycleCovariate 13:31:36.145 INFO ProgressMeter - Starting traversal 13:31:36.146 INFO ProgressMeter - Current Locus Elapsed Minutes Reads Processed Reads/Minute 13:31:46.177 INFO ProgressMeter - 1:21583071 0.2 45000 269649.5 13:31:48.143 INFO BaseRecalibrator - Shutting down engine [May 14, 2024 1:31:48 PM GMT] org.broadinstitute.hellbender.tools.walkers.bqsr.BaseRecalibrator done. Elapsed time: 0.26 minutes. Runtime.totalMemory()=4151836672 org.broadinstitute.hellbender.exceptions.GATKException: Error querying file /cromwell_root/fc-7711ee16-4d3a-44ca-b176-8acc97dff57a/ref/GRCm39/mgp_REL2021_snps.rsID.vcf.gz over interval 1:29605331-29605354 at org.broadinstitute.hellbender.engine.FeatureDataSource.refillQueryCache(FeatureDataSource.java:570) at org.broadinstitute.hellbender.engine.FeatureDataSource.queryAndPrefetch(FeatureDataSource.java:536) at org.broadinstitute.hellbender.engine.FeatureManager.getFeatures(FeatureManager.java:352) at org.broadinstitute.hellbender.engine.FeatureContext.getValues(FeatureContext.java:173) at org.broadinstitute.hellbender.engine.FeatureContext.getValues(FeatureContext.java:125) at org.broadinstitute.hellbender.engine.FeatureContext.getValues(FeatureContext.java:263) at org.broadinstitute.hellbender.tools.walkers.bqsr.BaseRecalibrator.apply(BaseRecalibrator.java:185) at org.broadinstitute.hellbender.engine.ReadWalker.lambda$traverse$0(ReadWalker.java:96) at java.util.stream.ForEachOps$ForEachOp$OfRef.accept(ForEachOps.java:183) at java.util.stream.ReferencePipeline$3$1.accept(ReferencePipeline.java:193) at java.util.stream.ReferencePipeline$2$1.accept(ReferencePipeline.java:175) at java.util.stream.ReferencePipeline$3$1.accept(ReferencePipeline.java:193) at java.util.Iterator.forEachRemaining(Iterator.java:116) at java.util.Spliterators$IteratorSpliterator.forEachRemaining(Spliterators.java:1801) at java.util.stream.AbstractPipeline.copyInto(AbstractPipeline.java:482) at java.util.stream.AbstractPipeline.wrapAndCopyInto(AbstractPipeline.java:472) at java.util.stream.ForEachOps$ForEachOp.evaluateSequential(ForEachOps.java:150) at java.util.stream.ForEachOps$ForEachOp$OfRef.evaluateSequential(ForEachOps.java:173) at java.util.stream.AbstractPipeline.evaluate(AbstractPipeline.java:234) at java.util.stream.ReferencePipeline.forEach(ReferencePipeline.java:485) at org.broadinstitute.hellbender.engine.ReadWalker.traverse(ReadWalker.java:94) at org.broadinstitute.hellbender.engine.GATKTool.doWork(GATKTool.java:1049) at org.broadinstitute.hellbender.cmdline.CommandLineProgram.runTool(CommandLineProgram.java:140) at org.broadinstitute.hellbender.cmdline.CommandLineProgram.instanceMainPostParseArgs(CommandLineProgram.java:192) at org.broadinstitute.hellbender.cmdline.CommandLineProgram.instanceMain(CommandLineProgram.java:211) at org.broadinstitute.hellbender.Main.runCommandLineProgram(Main.java:160) at org.broadinstitute.hellbender.Main.mainEntry(Main.java:203) at org.broadinstitute.hellbender.Main.main(Main.java:289) Caused by: java.io.IOException: Invalid argument at java.io.RandomAccessFile.seek0(Native Method) at java.io.RandomAccessFile.seek(RandomAccessFile.java:557) at htsjdk.samtools.seekablestream.SeekableFileStream.seek(SeekableFileStream.java:63) at htsjdk.tribble.TribbleIndexedFeatureReader$BlockStreamWrapper.<init>(TribbleIndexedFeatureReader.java:579) at htsjdk.tribble.TribbleIndexedFeatureReader$QueryIterator.advanceBlock(TribbleIndexedFeatureReader.java:470) at htsjdk.tribble.TribbleIndexedFeatureReader$QueryIterator.<init>(TribbleIndexedFeatureReader.java:440) at htsjdk.tribble.TribbleIndexedFeatureReader.query(TribbleIndexedFeatureReader.java:297) at org.broadinstitute.hellbender.engine.FeatureDataSource.refillQueryCache(FeatureDataSource.java:567) ... 27 more Using GATK jar /gatk/gatk-package-4.1.8.1-local.jar Running: java -Dsamjdk.use_async_io_read_samtools=false -Dsamjdk.use_async_io_write_samtools=true -Dsamjdk.use_async_io_write_tribble=false -Dsamjdk.compression_level=2 -Xms4G -jar /gatk/gatk-package-4.1.8.1-local.jar BaseRecalibrator -R /cromwell_root/fc-7711ee16-4d3a-44ca-b176-8acc97dff57a/ref/GRCm39/Mus_musculus.GRCm39.dna.primary_assembly.fa -I /cromwell_root/fc-7711ee16-4d3a-44ca-b176-8acc97dff57a/submissions/66a0b5bf-1e94-4e51-bdaf-886055953977/PreProcessingForVariantDiscovery_GATK4/baa27599-1fdf-4500-9238-2f0b8dcfe023/call-SortAndFixTags/cacheCopy/20191210_AN4_BH7589_S4.grcm39.aligned.duplicate_marked.sorted.bam --use-original-qualities -O 20191210_AN4_BH7589_S4.grcm39.recal_data.csv --known-sites /cromwell_root/fc-7711ee16-4d3a-44ca-b176-8acc97dff57a/ref/GRCm39/mgp_REL2021_snps.rsID.vcf.gz --known-sites /cromwell_root/fc-7711ee16-4d3a-44ca-b176-8acc97dff57a/ref/GRCm39/mgp_REL2021_indels.rsID.vcf.gz -L 1:1+
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Can you try running this command with at least version 4.3.0.0 to get a better error message? Additionally if you can add the following parameter to command that may bring more detailed error message.
--java-options '-DGATK_STACKTRACE_ON_USER_EXCEPTION=true'
Regards.
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