Question

Problem in using picard.jar

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5.4 years ago

Raheleh ▴ 260

Hello every one,

It's about a month that I stuck with picard. Every command that I try to run with picard failed. And for all of them I get the same error:

ERROR: Invalid argument '-I' , '-O'

When I try this argument I= or R=

I get this:

NOTE: Picard's command line syntax is changing.

* For more information, please see: * https://github.com/broadinstitute/picard/wiki/Command-Line-Syntax-Transition-For-Users-(Pre-Transition)

* The command line looks like this in the new syntax: * MergeVcfs -I sample1.vcf -I sample2.vcf.vcf -O Output.vcf.gz

can anyone tell me if I am doing sth wrong or it's a bug in picard?

I really appreciate any help!

picard • 7.9k views

ADD COMMENT • link updated 5.4 years ago by Pierre Lindenbaum 161k • written 5.4 years ago by Raheleh ▴ 260

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Please be as complete as possible and include the commands you tried, together with the version of Picard you are using.

ADD REPLY • link 5.4 years ago by WouterDeCoster 47k

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I'm using the latest version of picard: 2.18.20

This are the tools which I try to run by picard, but failed:

NormalizeFasta MarkDuplicates MergeVcfs CreateSequenceDictionary

I'm doing the exact command given from the website.

for example for MergVcfs, this is the command:

java -jar picard.jar MergeVcfs I=Germline.hc.fpfilterPassed.vcf  I=Somatic.hc.fpfilterPassed.vcf O=Output.vcf.gz

When I run the command I got this:

NOTE: Picard's command line syntax is changing.

** For more information, please see: ** https://github.com/broadinstitute/picard/wiki/Command-Line-Syntax-Transition-For-Users-(Pre-Transition)

** The command line looks like this in the new syntax:

** MergeVcfs -I Germline.hc.fpfilterPassed.vcf -I Somatic.hc.fpfilterPassed.vcf -O Output.vcf.gz

It returns me nothing. I tried this command:

java -jar picard.jar MergeVcfs -I Germline.hc.fpfilterPassed.vcf  -I Somatic.hc.fpfilterPassed.vcf -O Output.vcf.gz

But I got this error:

ERROR: Invalid argument '-I'

Thanks!

ADD REPLY • link 5.4 years ago by Raheleh ▴ 260

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Does removing the excessive space between Germline.hc.fpfilterPassed.vcf -I make a difference?

ADD REPLY • link 5.4 years ago by WouterDeCoster 47k

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Hello WouterDeCoster,

No, it doesn't.

ADD REPLY • link 5.4 years ago by Raheleh ▴ 260

WouterDeCoster · Accepted Answer · 2018-12-16

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5.4 years ago

Pierre Lindenbaum 161k

works on my machine using https://github.com/broadinstitute/picard/releases/download/2.18.20/picard.jar

java -jar /path/to/picard.jar MergeVcfs I=/path/to/S1.vcf.gz I=/path/to/S1.vcf.gz O=/path/to/out.vcf.gz
(...)
[Sun Dec 16 14:57:17 CET 2018] picard.vcf.MergeVcfs done. Elapsed time: 0.01 minutes.
Runtime.totalMemory()=40632320

yes there is the message NOTE: Picard's command line syntax is changing. but that's just an information.

ADD COMMENT • link 5.4 years ago by Pierre Lindenbaum 161k

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Dear Pierre, thanks for reply. I downloaded your machine and put all the files in the same directory and ran your command: java -jar picard.jar MergeVcfs I=Germline.hc.fpfilterPassed.vcf.gz I=Somatic.hc.fpfilterPassed.vcf.gz O=out.vcf.gz

It returned nothing and gave this exception:

Exception in thread "main" java.lang.IllegalArgumentException: A sequence dictionary must be available (either through the input file or by setting it explicitly).

I tried to create sequence dictionary using this command:

java -jar picard.jar CreateSequenceDictionary R=hg38.fa O=hg38.fa.dic

It returned an empty file and gave this error:

[Mon Dec 17 08:20:31 IRST 2018] CreateSequenceDictionary OUTPUT=hg38.fa.dic REFERENCE=hg38.fa    TRUNCATE_NAMES_AT_WHITESPACE=true NUM_SEQUENCES=2147483647 VERBOSITY=INFO QUIET=false VALIDATION_STRINGENCY=STRICT COMPRESSION_LEVEL=5 MAX_RECORDS_IN_RAM=500000 CREATE_INDEX=false CREATE_MD5_FILE=false GA4GH_CLIENT_SECRETS=client_secrets.json USE_JDK_DEFLATER=false USE_JDK_INFLATER=false [Mon Dec 17 08:20:31 IRST 2018] Executing as mohammadi@user-Z10PE-D16-WS on Linux
4.13.0-16-generic amd64; OpenJDK 64-Bit Server VM 1.8.0_144-8u144-b01-2-b01; Deflater: Intel; Inflater: Intel; Provider GCS is not available; Picard version: 2.18.20-SNAPSHOT [Mon Dec 17 08:20:31 IRST 2018] picard.sam.CreateSequenceDictionary done. Elapsed time: 0.00 minutes. Runtime.totalMemory()=1011351552 To get help, see http://broadinstitute.github.io/picard/index.html#GettingHelp Exception in thread "main" htsjdk.samtools.SAMException: Error opening file: hg38.fa   at htsjdk.samtools.util.IOUtil.openFileForReading(IOUtil.java:637)  at htsjdk.samtools.reference.FastaSequenceFile.<init>(FastaSequenceFile.java:64)    at htsjdk.samtools.reference.ReferenceSequenceFileFactory.getReferenceSequenceFile(ReferenceSequenceFileFactory.java:140)   at htsjdk.samtools.reference.ReferenceSequenceFileFactory.getReferenceSequenceFile(ReferenceSequenceFileFactory.java:96)    at htsjdk.samtools.reference.ReferenceSequenceFileFactory.getReferenceSequenceFile(ReferenceSequenceFileFactory.java:84)    at picard.sam.CreateSequenceDictionary.doWork(CreateSequenceDictionary.java:220)    at picard.cmdline.CommandLineProgram.instanceMain(CommandLineProgram.java:295)  at picard.cmdline.PicardCommandLine.instanceMain(PicardCommandLine.java:103)    at picard.cmdline.PicardCommandLine.main(PicardCommandLine.java:113) Caused by: java.nio.file.NoSuchFileException: hg38.fa  at sun.nio.fs.UnixException.translateToIOException(UnixException.java:86)   at sun.nio.fs.UnixException.rethrowAsIOException(UnixException.java:102)    at sun.nio.fs.UnixException.rethrowAsIOException(UnixException.java:107)    at sun.nio.fs.UnixFileSystemProvider.newByteChannel(UnixFileSystemProvider.java:214)    at java.nio.file.Files.newByteChannel(Files.java:361)

Any help? Many thanks!

ADD REPLY • link updated 5.4 years ago by WouterDeCoster 47k • written 5.4 years ago by Raheleh ▴ 260

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htsjdk.samtools.SAMException: Error opening file: hg38.fa at

If your shell/terminal is not within the same folder as hg38.fa you have to specify the full path to it.

O=hg38.fa.dic

The file extension should be .dict. This is what picard tools are looking for. And it has to be in the same folder as hg38.fa.

fin swimmer

ADD REPLY • link 5.4 years ago by finswimmer 16k

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A sequence dictionary must be available (either through the input file or by setting it explicitly).

a vcf should contain the ##contig lines. After CreateSequenceDictionary, you should add it with https://broadinstitute.github.io/picard/command-line-overview.html#UpdateVcfSequenceDictionary

ADD REPLY • link 5.4 years ago by Pierre Lindenbaum 161k

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Dear fin and Pierre, many thanks for your help!

Now I have hg38.fa.dict file :-) However, when I run this command: java -jar picard.jar VcfFormatConverter I=Germline.hc.fpfilterPassed.vcf O=Germline.hc.fpfilterPassed.bcf REQUIRE_INDEX=true to update my vcf file with the new sequence dictionary, I get this error:

> [Mon Dec 17 11:03:47 IRST 2018] VcfFormatConverter
> INPUT=Germline.hc.fpfilterPassed.vcf
> OUTPUT=Germline.hc.fpfilterPassed.bcf REQUIRE_INDEX=true   
> VERBOSITY=INFO QUIET=false VALIDATION_STRINGENCY=STRICT
> COMPRESSION_LEVEL=5 MAX_RECORDS_IN_RAM=500000 CREATE_INDEX=true
> CREATE_MD5_FILE=false GA4GH_CLIENT_SECRETS=client_secrets.json
> USE_JDK_DEFLATER=false USE_JDK_INFLATER=false [Mon Dec 17 11:03:47
> IRST 2018] Executing as adi@user-Z10PE-D16-WS on Linux
> 4.13.0-16-generic amd64; OpenJDK 64-Bit Server VM 1.8.0_144-8u144-b01-2-b01; Deflater: Intel; Inflater: Intel; Provider GCS is not available; Picard version: 2.18.20-SNAPSHOT [Mon Dec 17
> 11:03:47 IRST 2018] picard.vcf.VcfFormatConverter done. Elapsed time:
> 0.00 minutes. Runtime.totalMemory()=1011351552 To get help, see http://broadinstitute.github.io/picard/index.html#GettingHelp
> Exception in thread "main" htsjdk.tribble.TribbleException: An index
> is required, but none found., for input source:
> file:///media/adi/DA9CC7989CC76D97/WES_project/apps/test/gatk-4.0.11.0/Germline.hc.fpfilterPassed.vcf
>   at
> htsjdk.tribble.TribbleIndexedFeatureReader.<init>(TribbleIndexedFeatureReader.java:135)
>   at
> htsjdk.tribble.AbstractFeatureReader.getFeatureReader(AbstractFeatureReader.java:113)
>   at
> htsjdk.tribble.AbstractFeatureReader.getFeatureReader(AbstractFeatureReader.java:77)
>   at htsjdk.variant.vcf.VCFFileReader.<init>(VCFFileReader.java:140)
>   at htsjdk.variant.vcf.VCFFileReader.<init>(VCFFileReader.java:92)   at
> picard.vcf.VcfFormatConverter.doWork(VcfFormatConverter.java:101)     at
> picard.cmdline.CommandLineProgram.instanceMain(CommandLineProgram.java:295)
>   at
> picard.cmdline.PicardCommandLine.instanceMain(PicardCommandLine.java:103)
>   at picard.cmdline.PicardCommandLine.main(PicardCommandLine.java:113)

Can you help me again out of this problem? Thanks, Raheleh

ADD REPLY • link updated 5.4 years ago by WouterDeCoster 47k • written 5.4 years ago by Raheleh ▴ 260

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I added code markup to your post for increased readability. You can do this by selecting the text and clicking the 101010 button. When you compose or edit a post that button is in your toolbar, see image below:

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ADD REPLY • link 5.4 years ago by WouterDeCoster 47k

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your vcfs are not indexed. bzip them and index with bcftools index -t your.vcf.gz

ADD REPLY • link 5.4 years ago by Pierre Lindenbaum 161k

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why do you use VcfFormatConverter ?

ADD REPLY • link 5.4 years ago by Pierre Lindenbaum 161k

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Oops, my bad. So sorry. I ran this command: java -jar picard.jar UpdateVcfSequenceDictionary I=Germline.hc.fpfilterPassed.vcf O=Germline.hc.fpfilterPassed_new.vcf SEQUENCE_DICTIONARY=hg38.fa.dict

and now everything is fine :-)

Many thanks!

ADD REPLY • link 5.4 years ago by Raheleh ▴ 260