Hi, I run all the steps below to run GATK
gatk HaplotypeCaller -R chrX.fa -I ERR188044_chrX.bam -O ERR188044_chrX.vcf -ERC GVCF
But I got the following error
Using GATK jar /home/sahar/apps/gatk-4.1.9.0/gatk-package-4.1.9.0-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 -jar /home/sahar/apps/gatk-4.1.9.0/gatk-package-4.1.9.0-local.jar HaplotypeCaller -R chrX.fa -I ERR188044_chrX.bam -O ERR188044_chrX.vcf -ERC GVCF
23:14:55.161 INFO NativeLibraryLoader - Loading libgkl_compression.so from jar:file:/home/sahar/apps/gatk-4.1.9.0/gatk-package-4.1.9.0-local.jar!/com/intel/gkl/native/libgkl_compression.so
Feb 18, 2021 11:15:03 PM shaded.cloud_nio.com.google.auth.oauth2.ComputeEngineCredentials runningOnComputeEngine
INFO: Failed to detect whether we are running on Google Compute Engine.
23:15:03.691 INFO HaplotypeCaller - ------------------------------------------------------------
23:15:03.700 INFO HaplotypeCaller - The Genome Analysis Toolkit (GATK) v4.1.9.0
23:15:03.703 INFO HaplotypeCaller - For support and documentation go to https://software.broadinstitute.org/gatk/
23:15:03.704 INFO HaplotypeCaller - Executing as sahar@sahar-VirtualBox on Linux v5.8.0-43-generic amd64
23:15:03.711 INFO HaplotypeCaller - Java runtime: Java HotSpot(TM) 64-Bit Server VM v15.0.2+7-27
23:15:03.712 INFO HaplotypeCaller - Start Date/Time: February 18, 2021 at 11:14:54 PM IRST
23:15:03.712 INFO HaplotypeCaller - ------------------------------------------------------------
23:15:03.712 INFO HaplotypeCaller - ------------------------------------------------------------
23:15:03.733 INFO HaplotypeCaller - HTSJDK Version: 2.23.0
23:15:03.733 INFO HaplotypeCaller - Picard Version: 2.23.3
23:15:03.734 INFO HaplotypeCaller - HTSJDK Defaults.COMPRESSION_LEVEL : 2
23:15:03.734 INFO HaplotypeCaller - HTSJDK Defaults.USE_ASYNC_IO_READ_FOR_SAMTOOLS : false
23:15:03.739 INFO HaplotypeCaller - HTSJDK Defaults.USE_ASYNC_IO_WRITE_FOR_SAMTOOLS : true
23:15:03.739 INFO HaplotypeCaller - HTSJDK Defaults.USE_ASYNC_IO_WRITE_FOR_TRIBBLE : false
23:15:03.739 INFO HaplotypeCaller - Deflater: IntelDeflater
23:15:03.748 INFO HaplotypeCaller - Inflater: IntelInflater
23:15:03.748 INFO HaplotypeCaller - GCS max retries/reopens: 20
23:15:03.749 INFO HaplotypeCaller - Requester pays: disabled
23:15:03.749 INFO HaplotypeCaller - Initializing engine
23:15:03.775 INFO HaplotypeCaller - Shutting down engine
[February 18, 2021 at 11:15:03 PM IRST] org.broadinstitute.hellbender.tools.walkers.haplotypecaller.HaplotypeCaller done. Elapsed time: 0.15 minutes.
Runtime.totalMemory()=62980096
***********************************************************************
A USER ERROR has occurred: Fasta dict file file:///home/sahar/project/chrX/chrX_data/gatk/chrX.dict for reference file:///home/sahar/project/chrX/chrX_data/gatk/chrX.fa does not exist. Please see http://gatkforums.broadinstitute.org/discussion/1601/how-can-i-prepare-a-fasta-file-to-use-as-reference for help creating it.
***********************************************************************
Set the system property GATK_STACKTRACE_ON_USER_EXCEPTION (--java-options '-DGATK_STACKTRACE_ON_USER_EXCEPTION=true') to print the stack trace.
What should I do to fix this error?
The GATK version 4.1.9.0, The python version 3.8.5, The java version java-8-openjdk-amd64 and java-15-oracle which is set to java-8-openjdk-amd64.
Thank you for your help.
the answer to your problem is quite clear to me:
A USER ERROR has occurred: Fasta dict file file:///home/sahar/project/chrX/chrX_data/gatk/chrX.dict for reference file:///home/sahar/project/chrX/chrX_data/gatk/chrX.fa does not exist. Please see http://gatkforums.broadinstitute.org/discussion/1601/how-can-i-prepare-a-fasta-file-to-use-as-reference for help creating it.
I have both orders
samtools dict chrX.fa
@HD VN:1.0 SO:unsorted
@SQ SN:chrX LN:156040895 M5:49527016a48497d9d1cbd8e4a9049bd3 UR:file:///home/sahar/project/chrX/chrX_data/gatk/chrX.fa
and
samtools faidx chrX.fa
I run the same error again.
You need to create a sequence dictionary: https://gatk.broadinstitute.org/hc/en-us/articles/360036729911-CreateSequenceDictionary-Picard-
samtools dict output must be written in a file chrX.dict
I executed the command like this,
gatk HaplotypeCaller -R chrX.dict -I ERR188044_chrX.bam -O ERR188044_chrX.vcf -ERC GVCF
but it still has the same error.
gatk HaplotypeCaller -R chrX.fa -I ERR188044_chrX.bam -O ERR188044_chrX.vcf -ERC GVCF
with an existing chrX.dict in the same directory of chrX.fa
By executing this command,
gatk HaplotypeCaller -R chrX.fa -I ERR188044_chrX.bam -O ERR188044_chrX.vcf -ERC GVCF
I encountered this error.
A USER ERROR has occurred: Argument emit-ref-confidence has a bad value: Can only be used in single sample mode currently. Use the --sample-name argument to run on a single sample out of a multi-sample BAM file.
there are several samples in the lines @RG SM:xxx in the header of the bam file.
what it the output of
samtools view -H ERR188044_chrX.bam | grep '^RG'
how many SM: do you see ?
Only one sample is allowed in GVCF mode...
There is only one item inside the ERR188044_chrX.bam file.
Data is related to the ftp://ftp.ccb.jhu.edu/pub/RNAseq_protocol link.
can I see the output of the samtools view command above please
Sorry, answered late, becuase I can only send 5 posts in 6 hours. I did not see the output, but I used the
samtools view -Sb ERR188044_chrX.sam > ERR188044_chrX.bam
command to convert the sam file to bam. And I answered the previous question according to the bam file created.
Log in to answer this question.
