Can anyone help me with normal-only calling for panel of normals with MuTect2.
The code below does not spit any error as well as variants in vcf, except vcf header.
Here is the code:
java -Xmx45g -XX:+AggressiveOpts -jar /nexus/GenomeAnalysisTK-3.5/GenomeAnalysisTK.jar \
-T MuTect2 \
-R /home/ngs/data/ucsc.hg19.fasta \
-I:tumor /nexus/normal.bam \
--dbsnp /home/ngs/data/dbsnp_137.hg19.vcf \
--cosmic /nexus/rshahi/data/CosmicCodingMuts.vcf5 \
--artifact_detection_mode \
--intervals /nexus/NimbV3_target.gatk.interval_list \
-o $HOME/jobout/normal_only.vcf
Thanx,
Rajendra
Brussels
5 answers
The most likely source of error is your intervals list. If this is incomplete for instance, or not matching up with the regions you have data for, you would get no variants and also no error messages (unless the intervals list file was actually malformed). Try running the same command but without passing it an intervals file and see what happens.
You should also probably check your BAM file to make sure it is actually containing aligned data as expected. Visualize it with IGV for instance and check some genomic coordinates of your favourite genes as a quick check.
Maybe try changing your java calling? It's possible something is going on with the JVM. You are passing 45G of RAM and Aggressive Opts to Java. If the BAM file is for a single exome or targeted resequencing experiment (assuming since you're passing a Nimblegen targets list) then 45G is about 10x-20x what is actually needed. If GATK runs out of memory it will throw an error, but if the problem is earlier on with the JVM then it could potentially die silently, especially with the -XX:+AggressiveOpts also being passed. That will turn on a whole suite of options and I'm not sure what effect that may have as I have never used it.
If neither of those seem to be the case it is hard to diagnose without trying on your same BAM file (or a portion of the BAM file) to diagnose. There doesn't seem to be anything wrong with the command line. When you look at the BAM file in IGV or something can you find obvious variants bye eye that should be called? If you throw the BAM at FreeBayes or another variant caller do you get results then?
@Dan,hanx for your effort. I will update you with some tweaks from Geraldine_VdAuwera (GATK). Cheers!
Don t worry your command is correct, it s not your fault. There is a bug in MuTect2 when try to create a PON
I am facing the same issue.
Some people claim that the problem was solved but I am still getting an empty vcf file.
Try the nightly build
It looks like the --artifact_detection_mode flag is for HaplotypeCaller (the GATK recommendation is to use HaplotypeCaller for generating the panel of normals): https://www.broadinstitute.org/gatk/guide/tooldocs/org_broadinstitute_gatk_tools_walkers_cancer_m2_MuTect2.php
Hi, I think maybe you can find GATK4 to have a try, I heard it has been published.
•
written
by
winter_li •
Log in to answer this question.