GATK Questions

[Jane M]

Hello everybody,

I would appreciate your help to discuss the files to be provided as known indels and known variants for realignment and recalibration steps.
I am using GATK v3.4.0.

1. For realignment, as discussed here:
https://www.biostars.org/p/96022/

http://seqanswers.com/forums/showthread.php?t=19600

it is possible to provide several known indels datasets.
The most widely used files seem to be:
ftp://ftp.broadinstitute.org/bundle/...9.sites.vcf.gz
and
ftp://ftp.broadinstitute.org/bundle/...9.sites.vcf.gz

I wonder if the second file does not include most of indels in the first file?

As mentionned by Rocketknight,

The Mills and 1000G gold standard indels are a very stringently curated list of indels. For the purposes of indel realignment in GATK you probably want one or more of the broader sets from the GATK bundle instead, though which ones I'm not completely sure.

Which file is less stringent? 1000G_phase1.indels.hg19.sites.vcf?

java -jar GenomeAnalysisTK.jar \ 
    -T RealignerTargetCreator \ 
    -R reference.fa \ 
    -I dedup_reads.bam \ 
    [I]-known 1000G_phase1.indels.hg19.sites.vcf \ [/I]   
    [I]-known Mills_and_1000G_gold_standard.indels.hg19.sites.vcf \ [/I]   
    -o realignment_targets.list

Should these datasets be provided in the next step too? If yes, why?

java -jar GenomeAnalysisTK.jar \
  -T IndelRealigner \
  -I dedup_reads.bam \
  -R reference.fa \
  -targetIntervals realignment_targets.list \
  -o realignedBam.bam \
  [I]-known 1000G_phase1.indels.hg19.sites.vcf \ [/I]   
  [I]-known Mills_and_1000G_gold_standard.indels.hg19.sites.vcf \ [/I]

Can java -jar GenomeAnalysisTK.jar -T IndelRealigner be multithreaded with -nt or -nct options?

2. For recalibration, both SNV and indels datasets should be provided.
Can I use both indels datasets previously used for realignment and the last dbSNP release for the SNV?

java -jar GenomeAnalysisTK.jar \ 
    -T BaseRecalibrator \
    -l INFO \
    -R reference.fa \ 
    -I realigned_reads.bam \ 
    -knownSites dbsnp144.vcf \ 
    -knownSites 1000G_phase1.indels.hg19.sites.vcf \ 
    -knownSites Mills_and_1000G_gold_standard.indels.hg19.sites.vcf \ 
    -cov ReadGroupCovariate -cov QualityScoreCovariate -cov CycleCovariate
    -o recal_data.table

java -jar GenomeAnalysisTK.jar \
    -l INFO \
    -T PrintReads \
    -R reference.fa \
    -I realigned_reads.bam \
    -BQSR recal_data.table \
    -o realigned.fixed.recal.bam

Can java -jar GenomeAnalysisTK.jar -T PrintReads be multithreaded with -nt or -nct options?

Thank you for your help
Jane