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ocanela
Member
Hello,
I have a BAM from RNA Seq experiment. The reads are aligned with tophat and then I filtered secondary reads, fixed mate pairs, marked duplicates with picard and realigned with gatk. However, when I try to use BaseRecalibrator function,
-T BaseRecalibrator -nct 12 -R ./SusGenome10v69/Sus_scrofa.Sscrofa10.2.69.dna.toplevel.fa -I ./7_15.corrected.ordered.sremoved.marked.fixed.realigned.fixed.corrected.ordered.sremoved.marked.fixed.realigned.fixed.bam -knownSites ./SusGenome10v69/Sus_scrofa.vcf -o ./7_15.RecalData.grp
It stops with the error:
org.broadinstitute.sting.utils.exceptions.ReviewedStingException: Trying to clip before the start or after the end of a read
at org.broadinstitute.sting.utils.clipping.ReadClipper.hardClipByReferenceCoordinates(ReadClipper.java:534)
at org.broadinstitute.sting.utils.clipping.ReadClipper.hardClipByReferenceCoordinatesLeftTail(ReadClipper.java:176)
at org.broadinstitute.sting.utils.clipping.ReadClipper.hardClipAdaptorSequence(ReadClipper.java:389)
at org.broadinstitute.sting.utils.clipping.ReadClipper.hardClipAdaptorSequence(ReadClipper.java:392)
at org.broadinstitute.sting.gatk.walkers.bqsr.BaseRecalibrator.map(BaseRecalibrator.java:244)
at org.broadinstitute.sting.gatk.walkers.bqsr.BaseRecalibrator.map(BaseRecalibrator.java:131)
at org.broadinstitute.sting.gatk.traversals.TraverseReadsNano$TraverseReadsMap.apply(TraverseReadsNano.java:230)
at org.broadinstitute.sting.gatk.traversals.TraverseReadsNano$TraverseReadsMap.apply(TraverseReadsNano.java:218)
at org.broadinstitute.sting.utils.nanoScheduler.NanoScheduler$ReadMapReduceJob.run(NanoScheduler.java:471)
at java.util.concurrent.Executors$RunnableAdapter.call(Executors.java:471)
at java.util.concurrent.FutureTask$Sync.innerRun(FutureTask.java:334)
at java.util.concurrent.FutureTask.run(FutureTask.java:166)
at java.util.concurrent.ThreadPoolExecutor.runWorker(ThreadPoolExecutor.java:1146)
at java.util.concurrent.ThreadPoolExecutor$Worker.run(ThreadPoolExecutor.java:615)
at java.lang.Thread.run(Thread.java:679)
The bam file validates properly with the Picard's ValidateSamFile function. I tried with gatk versions, 2.3-9 and 2.4-7 obtaining the same results.
I used the same procedure with other samples without any problem. However, I am unable to recalibrate this sample. I tried to realign the reads with different tophat versions, applied different filters and different procedures, however i am unable to use the BaseRecalibrator function with this sample.
Could you help me?
Thanks!
ebanks
Broad InstituteMember, Broadie, Dev
Thanks for the very helpful test files. We were able to implement a fix for this problem.
The official patch will be available in version 2.5 (although if you cannot wait then you can access our unsupported nightly builds).
Answers
Hi there,
This sounds like a bug; we'll need to reproduce this error locally. Could you please upload a BAM snippet to our server? Detailed instructions here: http://www.broadinstitute.org/gatk/guide/article?id=1894
Be sure to also upload your reference since you're using a custom one.
Hi,
I uploaded the requested files in your ftp server (including the reference).
The name of the compressed file is: baseracalibrator.bug.ocanela.tar.gz
Thanks,
Thanks for the very helpful test files. We were able to implement a fix for this problem.
The official patch will be available in version 2.5 (although if you cannot wait then you can access our unsupported nightly builds).
Ok, thank you for the software support!
Hi,
I have the exact same error with my BAM file even though I used the latest version 2.4.9-g532efad. Would you please let me know where I can download version 2.5 with the patch?
Thanks,
Huy
Hi @huyvuong, version 2.5 is not yet available, but you can try the latest nightly build (look on the Downloads page).
Thank you very much. I tried the latest nightly build and it worked now.
The version 2.5-2 fixed the problem with my sample.
Thank you for the fix!