Problem with vcf format to run gatk FastaAlternateReferenceMaker

Question

[pablo]

Hello,

I'm trying to run gatk FastaAlternateReferenceMaker to get the FASTA files from my vcf file.

I run : gatk FastaAlternateReferenceMaker -R my_reference.fa -O my_output.fasta -V my_file.vcf

I get that error message :

Using GATK jar /opt/apps/gcc-8.1.0/gatk-4.1.0.0/gatk-package-4.1.0.0-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 /opt/apps/gcc-8.1.0/gatk-4.1.0.0/gatk-package-4.1.0.0-local.jar FastaAlternateReferenceMaker -R my_reference.fa -O my_output.fasta -V my_file.vcf    14:15:23.712 INFO  NativeLibraryLoader - Loading libgkl_compression.so from jar:file:/opt/apps/gcc-8.1.0/gatk-4.1.0.0/gatk-package-4.1.0.0-local.jar!/com/intel/gkl/native/libgkl_compression.so
14:15:30.426 INFO  FastaAlternateReferenceMaker - ------------------------------------------------------------
14:15:30.427 INFO  FastaAlternateReferenceMaker - The Genome Analysis Toolkit (GATK) v4.1.0.0
14:15:30.427 INFO  FastaAlternateReferenceMaker - For support and documentation go to https://software.broadinstitute.org/gatk/
14:15:30.430 INFO  FastaAlternateReferenceMaker - Executing as *** on Linux v3.10.0-1127.18.2.el7.x86_64 amd64
14:15:30.430 INFO  FastaAlternateReferenceMaker - Java runtime: Java HotSpot(TM) 64-Bit Server VM v1.8.0_45-b14
14:15:30.430 INFO  FastaAlternateReferenceMaker - Start Date/Time: November 3, 2020 2:15:23 PM CET
14:15:30.430 INFO  FastaAlternateReferenceMaker - ------------------------------------------------------------
14:15:30.430 INFO  FastaAlternateReferenceMaker - ------------------------------------------------------------
14:15:30.431 INFO  FastaAlternateReferenceMaker - HTSJDK Version: 2.18.2
14:15:30.432 INFO  FastaAlternateReferenceMaker - Picard Version: 2.18.25
14:15:30.432 INFO  FastaAlternateReferenceMaker - HTSJDK Defaults.COMPRESSION_LEVEL : 2
14:15:30.432 INFO  FastaAlternateReferenceMaker - HTSJDK Defaults.USE_ASYNC_IO_READ_FOR_SAMTOOLS : false
14:15:30.432 INFO  FastaAlternateReferenceMaker - HTSJDK Defaults.USE_ASYNC_IO_WRITE_FOR_SAMTOOLS : true
14:15:30.432 INFO  FastaAlternateReferenceMaker - HTSJDK Defaults.USE_ASYNC_IO_WRITE_FOR_TRIBBLE : false
14:15:30.432 INFO  FastaAlternateReferenceMaker - Deflater: IntelDeflater
14:15:30.432 INFO  FastaAlternateReferenceMaker - Inflater: IntelInflater
14:15:30.433 INFO  FastaAlternateReferenceMaker - GCS max retries/reopens: 20
14:15:30.433 INFO  FastaAlternateReferenceMaker - Requester pays: disabled
14:15:30.433 INFO  FastaAlternateReferenceMaker - Initializing engine
14:15:30.853 INFO  FeatureManager - Using codec VCFCodec to read file file:///my_file.vcf
14:15:30.859 INFO  FastaAlternateReferenceMaker - Shutting down engine
[November 3, 2020 2:15:30 PM CET] org.broadinstitute.hellbender.tools.walkers.fasta.FastaAlternateReferenceMaker done. Elapsed time: 0.12 minutes.
Runtime.totalMemory()=623378432
org.broadinstitute.hellbender.exceptions.GATKException: Error initializing feature reader for path my_file.vcf
        at org.broadinstitute.hellbender.engine.FeatureDataSource.getTribbleFeatureReader(FeatureDataSource.java:353)
        at org.broadinstitute.hellbender.engine.FeatureDataSource.getFeatureReader(FeatureDataSource.java:305)
        at org.broadinstitute.hellbender.engine.FeatureDataSource.<init>(FeatureDataSource.java:256)
        at org.broadinstitute.hellbender.engine.FeatureManager.addToFeatureSources(FeatureManager.java:234)
        at org.broadinstitute.hellbender.engine.FeatureManager.initializeFeatureSources(FeatureManager.java:208)
        at org.broadinstitute.hellbender.engine.FeatureManager.<init>(FeatureManager.java:155)
        at org.broadinstitute.hellbender.engine.GATKTool.initializeFeatures(GATKTool.java:417)
        at org.broadinstitute.hellbender.engine.GATKTool.onStartup(GATKTool.java:638)
        at org.broadinstitute.hellbender.engine.ReferenceWalker.onStartup(ReferenceWalker.java:36)
        at org.broadinstitute.hellbender.cmdline.CommandLineProgram.runTool(CommandLineProgram.java:136)
        at org.broadinstitute.hellbender.cmdline.CommandLineProgram.instanceMainPostParseArgs(CommandLineProgram.java:191)
        at org.broadinstitute.hellbender.cmdline.CommandLineProgram.instanceMain(CommandLineProgram.java:210)
        at org.broadinstitute.hellbender.Main.runCommandLineProgram(Main.java:162)
        at org.broadinstitute.hellbender.Main.mainEntry(Main.java:205)
        at org.broadinstitute.hellbender.Main.main(Main.java:291)
Caused by: htsjdk.tribble.TribbleException$MalformedFeatureFile: Unable to parse header with error: Your input file has a malformed header: The FORMAT field was provided but there is no genotype/sample data, for input source: my_file.vcf
        at htsjdk.tribble.TribbleIndexedFeatureReader.readHeader(TribbleIndexedFeatureReader.java:263)
        at htsjdk.tribble.TribbleIndexedFeatureReader.<init>(TribbleIndexedFeatureReader.java:102)
        at htsjdk.tribble.TribbleIndexedFeatureReader.<init>(TribbleIndexedFeatureReader.java:127)
        at htsjdk.tribble.AbstractFeatureReader.getFeatureReader(AbstractFeatureReader.java:120)
        at org.broadinstitute.hellbender.engine.FeatureDataSource.getTribbleFeatureReader(FeatureDataSource.java:350)
        ... 14 more
Caused by: htsjdk.tribble.TribbleException$InvalidHeader: Your input file has a malformed header: The FORMAT field was provided but there is no genotype/sample data
        at htsjdk.variant.vcf.AbstractVCFCodec.parseHeaderFromLines(AbstractVCFCodec.java:185)
        at htsjdk.variant.vcf.VCFCodec.readActualHeader(VCFCodec.java:111)
        at htsjdk.tribble.AsciiFeatureCodec.readHeader(AsciiFeatureCodec.java:79)
        at htsjdk.tribble.AsciiFeatureCodec.readHeader(AsciiFeatureCodec.java:37)
        at htsjdk.tribble.TribbleIndexedFeatureReader.readHeader(TribbleIndexedFeatureReader.java:261)
        ... 18 more

I show the header of my vcf file, which looks good :

##fileformat=VCFv4.2
##INFO=<ID=DP,Number=1,Type=Integer,Description="Total Depth">
##INFO=<ID=AF,Number=A,Type=Float,Description="Allele Frequency">
##FORMAT=<ID=GT,Number=1,Type=String,Description="Genotype">
##FORMAT=<ID=HQ,Number=2,Type=Integer,Description="Haplotype Quality">
#CHROM  POS     ID      REF     ALT     QUAL    FILTER  INFO    FORMAT
Super-Scaffold_100001   672149  .       C       T       .       PASS    DP=210;AF=0.51  GT:HQ
Super-Scaffold_100001   862122  .       A       T       .       PASS    DP=305;AF=0.5   GT:HQ
Super-Scaffold_100001   931168  .       C       A       .       PASS    DP=127;AF=0.5   GT:HQ
Super-Scaffold_100001   967240  .       C       T       .       PASS    DP=127;AF=0.5   GT:HQ

Any idea why I get Your input file has a malformed header: The FORMAT field was provided but there is no genotype/sample data ?

Bests

[GenoMax]

I think the main problem is here:

Error initializing feature reader for path my_file.vcf

Is that vcf file in the right location?

[pablo]

Yes, the vcf file is in the right location. Even if I use the full path, it does not work.

[Pierre Lindenbaum]

The FORMAT field was provided but there is no genotype/sample data, for input source: my_file.vcf

there is a FORMAT column but there is no associated genotype in your vcf.