BaseRecalibrator error in a BAM validated with Picard

ocanela

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:

ERROR ------------------------------------------------------------------------------------------

ERROR stack trace

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)

ERROR ------------------------------------------------------------------------------------------

ERROR A GATK RUNTIME ERROR has occurred (version 2.4-7-g5e89f01):

ERROR

ERROR Please visit the wiki to see if this is a known problem

ERROR If not, please post the error, with stack trace, to the GATK forum

ERROR Visit our website and forum for extensive documentation and answers to

ERROR commonly asked questions http://www.broadinstitute.org/gatk

ERROR

ERROR MESSAGE: Trying to clip before the start or after the end of a read

ERROR ------------------------------------------------------------------------------------------

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

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).

Geraldine_VdAuwera

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.

ocanela

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,

ocanela

Ok, thank you for the software support!

huyvuong

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

Geraldine_VdAuwera

Hi @huyvuong, version 2.5 is not yet available, but you can try the latest nightly build (look on the Downloads page).

huyvuong

Thank you very much. I tried the latest nightly build and it worked now.

ocanela

The version 2.5-2 fixed the problem with my sample.

Thank you for the fix!