Unified Genotyper Error

LisaLattarulo

Hi. I was hoping you could help track down the source of this error. I haven't seen this exact error anywhere here in the forum. I am trying to run unified genotyper on 48 bam files at once. Each bam file has been aligned using bwa and processed through the best practices as mentioned on the website. They all pass validation with no error (I had to run picard tools, cleanSamFile and fixMateInformation, along with removing one read from a single file). The data is targeted sequencing. I am running GATKLite 2.3-5-gd738181.

The error message is: Somehow the requested coordinate is not covered by the read. Alignment 189665677 | 48M103S

The command line I am using is: java -Xmx4g -jar pathToGATK.jar -R standardRefFileHG37 -I (48 bam files) -o OutputFile.vcf -nt 4 -nct 4 -T UnifiedGenotyper --dbsnp standardRef -glm Both -metrics metric.file -debug_file debug.txt -A DepthOfCoverage -A AlleleBalance

Is there any way to see if it is a particular bam file that is causing the error? In the error message, what Alignment is it referring to? I am doing the processing on a machine that is not connected to the internet so it is harder for me to cut/copy paste information to here. Is there anything else that you would need to help me track down the source of this error?

In the meantime I am going to try to run it with a .bed file of targeted areas to see if that helps. Should I also try to process each file individually? Thank you for your input.

Lisa

Geraldine_VdAuwera

Hi Lisa, can you tell me if you reduced your bams with ReduceReads?

LisaLattarulo

I did not as the ReduceReads is not available with the GATKLite version. My coverage is anticipated to be at most 80X so I didn't think it was that important anyhow as my files aren't that big.

Geraldine_VdAuwera

OK, just checking -- reduced bams were throwing a similar error a little while ago and I wanted to rule that out.

I'm not sure what is going on, so let me know if you still get the same error when you run with the intervals file. FYI, for targeted sequencing you'll get better performance if you use the intervals file at all steps.

LisaLattarulo

Ok. It now finished without any error when using the targeted sequence file. I'm guessing there was probably just some sort of funny alignment in a file. Thank you.

LisaLattarulo

I just wanted to post some follow up with this in case other people may be having this error. I have found this error to be wholly unreproducible. I have run into it again when running different parameters. A re-start of the run often "fixes" it, i.e. it doesn't happen again and the run finishes through. I've also gotten an error when running something again that had finished the first time through with no issues. The Alignment number in the error message changes along with the text string after the pipe sign. Still don't know what is going on with it.

Geraldine_VdAuwera

Thanks for following up. It will be interesting to see if this bug still occurs after release 2.4 (coming really really soon!) as we added a lot of internal tests that should prevent subtle bugs from getting out.