Mixing paired-end and single-end Exomes

BerntPoppBerntPopp Posts: 47Member
edited October 2012 in Ask the GATK team

Hello,

I am currently working on a Exome sequencing projekt with older single-end SOLiD exomes and new paired-end exomes.
In a first attempt (GATK 1.7 and best practices v3 back then) i tried calling and recalibrating all exomes together (at that time 120) without selecting for paired/single-end. As I already had validatet many variants I could check the quality of the calls and got very bad results, especially for InDels (previously called, true positive variants missing).
My idee is that the UnifiedGenotyper has Problems mixing paired-end with single-end exomes.

Is there any official recommendation for this problem?
My solution right now is to group the exomes in batches (40-50 Exomes) which ran on the same technology.

Also a second Problem/Question:
For some individuals exomes where sequenced twice, and for some of these the first run was single-end and the second one was paired.
The best practices mentions one should se all available reads for a individual for calling. Do you have any experience on how to handle these cases?

Any help is greatly appreciated!

Bernt

Post edited by BerntPopp on

Best Answer

  • ebanksebanks Broad InstitutePosts: 684Member, Administrator, GATK Developer, Broadie, Moderator, DSDE Member, GP Member admin
    Answer ✓

    You can freely mix single and paired end sequencing data with no problem in the GATK. The problem is probably with the data itself, not the genotyper.

    Eric Banks, PhD -- Senior Group Leader, MPG Analysis, Broad Institute of Harvard and MIT

Answers

  • ebanksebanks Broad InstitutePosts: 684Member, Administrator, GATK Developer, Broadie, Moderator, DSDE Member, GP Member admin
    Answer ✓

    You can freely mix single and paired end sequencing data with no problem in the GATK. The problem is probably with the data itself, not the genotyper.

    Eric Banks, PhD -- Senior Group Leader, MPG Analysis, Broad Institute of Harvard and MIT

  • BerntPoppBerntPopp Posts: 47Member

    Ok,

    the problem is that i ran the exact same analysis for the single-end runs (GATK 1.5) before and found said CNVs. After adding the paired-end exomes the CNVs are missing (GATK 1.7).
    VCF files are checked and comple, log files showed no errors and the BAM files have been processed according to the best practices and checked with FastQC and Picards ValidateSam.
    ValidateSam seems to show the only remaining error source, see also:
    http://gatkforums.broadinstitute.org/discussion/1562/need-to-run-a-step-with-fixmateinformation-after-realignment-step

    Maybe it was a version problem. I will redo the Analysis with all exomes and the new GATK and repost upon having the resulting calls.

    Thank you Eric!

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