Question: Rna-Seqc Generates Error
0
gravatar for federico.gaiti
6.7 years ago by
Brisbane
federico.gaiti70 wrote:

Hi all,

I was running RNA-SeQC software in command line. I tried withto follow the example sets provided on RNA-SeQC website and following the instructions as stated.

The command was:

"java -jar /hox/u/uqfgaiti/Tools/picard-tools-1.84/RNA-SeQC_v1.1.7.jar -r ampQue1.fasta -s "TestID|CEL-Seq.final.sorted.bam|TestDesc" -t 2507_Intergenic+Intronic.gtf -n 1000 -BWArRNA rRNA.fasta -o RNA-SeQC_out"

However I ran into the below error, and I am trying to see if anyone else faced the same error or knows the fix:

RNA-SeQC v1.1.7 05/14/12

Retriving contig names from reference

contig names in reference: 13397

Loading GTF for Read Counting

Converting to refGene

Transcript objects to RefGen format: 2 s

Running IntronicExpressionReadBlock Walker ....

Arguments: [-T, IntronicExpressionReadBlock, --outfile_metrics, RNA-SeQC_out/TestID/TestID.metrics.tmp.txt, -R, ampQue1.fasta, -I, CEL-Seq.final.sorted.bam, -refseq, RNA-SeQC_out/refGene.txt, -l, ERROR]

net.sf.samtools.util.RuntimeEOFException: Premature EOF; BinaryCodec in readmode; streamed file (filename not available)

at net.sf.samtools.util.BinaryCodec.readBytes(BinaryCodec.java:373)
at net.sf.samtools.util.BinaryCodec.readByteBuffer(BinaryCodec.java:480)
at net.sf.samtools.util.BinaryCodec.readInt(BinaryCodec.java:491)
at net.sf.samtools.BAMFileReader.readSequenceRecord(BAMFileReader.java:429)
at net.sf.samtools.BAMFileReader.readHeader(BAMFileReader.java:403)
at net.sf.samtools.BAMFileReader.<init>(BAMFileReader.java:144)
at net.sf.samtools.BAMFileReader.<init>(BAMFileReader.java:114)
at net.sf.samtools.SAMFileReader.init(SAMFileReader.java:514)
at net.sf.samtools.SAMFileReader.<init>(SAMFileReader.java:167)
at org.broadinstitute.sting.gatk.datasources.reads.SAMDataSource$ReaderInitializer.call(SAMDataSource.java:927)
at org.broadinstitute.sting.gatk.datasources.reads.SAMDataSource$SAMReaders.<init>(SAMDataSource.java:788)
at org.broadinstitute.sting.gatk.datasources.reads.SAMDataSource$SAMResourcePool.createNewResource(SAMDataSource.java:747)
at org.broadinstitute.sting.gatk.datasources.reads.SAMDataSource$SAMResourcePool.getAvailableReaders(SAMDataSource.java:718)
at org.broadinstitute.sting.gatk.datasources.reads.SAMDataSource.<init>(SAMDataSource.java:261)
at org.broadinstitute.sting.gatk.GenomeAnalysisEngine.createReadsDataSource(GenomeAnalysisEngine.java:755)
at org.broadinstitute.sting.gatk.GenomeAnalysisEngine.initializeDataSources(GenomeAnalysisEngine.java:666)
at org.broadinstitute.sting.gatk.GenomeAnalysisEngine.execute(GenomeAnalysisEngine.java:227)
at org.broadinstitute.sting.gatk.CommandLineExecutable.execute(CommandLineExecutable.java:113)
at org.broadinstitute.sting.commandline.CommandLineProgram.start(CommandLineProgram.java:236)
at org.broadinstitute.sting.commandline.CommandLineProgram.start(CommandLineProgram.java:146)
at org.broadinstitute.cga.rnaseq.gatk.GATKTools.runIntronReadCount(GATKTools.java:226)
at org.broadinstitute.cga.rnaseq.ReadCountMetrics.runRegionCounting(ReadCountMetrics.java:243)
at org.broadinstitute.cga.rnaseq.ReadCountMetrics.runReadCountMetrics(ReadCountMetrics.java:58)
at org.broadinstitute.cga.rnaseq.RNASeqMetrics.runMetrics(RNASeqMetrics.java:220)
at org.broadinstitute.cga.rnaseq.RNASeqMetrics.execute(RNASeqMetrics.java:166)
at org.broadinstitute.cga.rnaseq.RNASeqMetrics.main(RNASeqMetrics.java:135)

RNA-SeQC Total Runtime: 0 min

Any comments are highly appreciated.

Thanks Federico

bioinformatics java picard • 4.5k views
ADD COMMENTlink modified 4.9 years ago by Biostar ♦♦ 20 • written 6.7 years ago by federico.gaiti70
2

In all such cases you need to make sure your bam file is exists, it is indexed and that it does contain the samples.

Include a few lines of the command to invoke and the results of doing a samtools view

ADD REPLYlink written 6.7 years ago by Istvan Albert ♦♦ 81k
3
gravatar for federico.gaiti
6.7 years ago by
Brisbane
federico.gaiti70 wrote:

I re-did everything from the beginning and it seems the problem was in the command "AddOrReplaceReadGroups".

Now I have all the required input files:

  • an indexed, coordinate-sorted BAM file with read group information

  • .BAI of the BAM file in the same folder

  • my FASTA ref sequence

  • .FAI of the ref sequence

  • .DICT of the ref sequence

  • GTF annotation file

Now it's running and it's not giving me any sort of error.

Let you know if I'll be stuck again

Thanks for the comments

ADD COMMENTlink written 6.7 years ago by federico.gaiti70
1

thanks for following up

ADD REPLYlink written 6.7 years ago by Istvan Albert ♦♦ 81k
0
gravatar for Chris Miller
6.7 years ago by
Chris Miller21k
Washington University in St. Louis, MO
Chris Miller21k wrote:

The relevant part of this error seems to be: "filename not available"

Are you sure your bam path is correct and that the software can access it (permissions are set properly, etc)?

ADD COMMENTlink written 6.7 years ago by Chris Miller21k
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