Hi there,

I'm following the genotype refinement workflow (https://gatk.broadinstitute.org/hc/en-us/articles/360035531432-Genotype-Refinement-workflow-for-germline-short-variants) and doing CalculateGenotypePosteriors. I got four questions

1. I have a big vcf file of 109 trio families. To do CalculateGenotypePosteriors, I first split my big vcf into109 trio vcf files by using selectvariant, and then with each trio vcf file, I did  CalculateGenotypePosteriors with .ped as the family prior. I'm wondering - for such analysis with only 3 samples in a trio vcf, would you recommend to supply population prior as well? I saw 'By default, priors will be applied to each variant separately, provided each variant features data from at least 10 called samples (no-calls do not count)' in the document (https://gatk.broadinstitute.org/hc/en-us/articles/360037226592-CalculateGenotypePosteriors). I'm confused in my case of only 3 called samples in a vcf, this default condition may not hold and thus may lead to some false positives? 

2. This statement - By default, priors will be applied to each variant separately, provided each variant features data from at least 10 called samples - did make me confused... does it mean I CANNOT do CalculateGenotypePosteriors for a trio vcf of 3 samples only? in what condition/scenario does this 'at least 10 called samples' hold?

3. right now I do CalculateGenotypePosteriors for each trio vcf individually, so I have to do CalculateGenotypePosteriors 109 times (I have 109 trio vcf files in total).  I'm wondering is it okay to do CalculateGenotypePosteriors for my big vcf with all samples from 109 families and supply a .ped file for all the 109 families without splitting into single trio vcf files? In such case I just need to do CalculateGenotypePosteriors once. Would the posterior be better if I do all the 109 families all together in one big vcf compared to do one family at a time?

4. I know CalculateGenotypePosteriors is for diploid only so I have to excl. regions on chrX & chrY, which I used haploid mode in haplotypecaller. However, I'm wondering are you thinking of developing a similar tool for haploid or mixed ploidy? sex chromosomes also play an important role in many diseases and it would be great if I can still consider them in running my workflow. 

Thank you so much for your help!

REQUIRED for all errors and issues:
a) GATK version used: gatk/4.2.1.0 

b) Exact command used:

gatk --java-options '-Xmx40g -Xms32g -XX:ParallelGCThreads=2' \

    CalculateGenotypePosteriors \

    -V $per_fam_vcf \

    -ped $per_fam_ped \

    -XL chrX \

    -XL chrY \

    -O ${outdir_per_fam}/Chaklab_posterior_GT_${famID}.vcf.gz \

    --skip-population-priors

c) Entire program log:

23:33:41.682 INFO  CalculateGenotypePosteriors - ------------------------------------------------------------

23:33:41.683 INFO  CalculateGenotypePosteriors - HTSJDK Version: 2.19.0

23:33:41.683 INFO  CalculateGenotypePosteriors - Picard Version: 2.19.0

23:33:41.683 INFO  CalculateGenotypePosteriors - HTSJDK Defaults.COMPRESSION_LEVEL : 2

23:33:41.683 INFO  CalculateGenotypePosteriors - HTSJDK Defaults.USE_ASYNC_IO_READ_FOR_SAMTOOLS : false

23:33:41.683 INFO  CalculateGenotypePosteriors - HTSJDK Defaults.USE_ASYNC_IO_WRITE_FOR_SAMTOOLS : true

23:33:41.683 INFO  CalculateGenotypePosteriors - HTSJDK Defaults.USE_ASYNC_IO_WRITE_FOR_TRIBBLE : false

23:33:41.683 INFO  CalculateGenotypePosteriors - Deflater: IntelDeflater

23:33:41.683 INFO  CalculateGenotypePosteriors - Inflater: IntelInflater

23:33:41.683 INFO  CalculateGenotypePosteriors - GCS max retries/reopens: 20

23:33:41.683 INFO  CalculateGenotypePosteriors - Requester pays: disabled

23:33:41.683 INFO  CalculateGenotypePosteriors - Initializing engine

23:33:42.000 INFO  FeatureManager - Using codec VCFCodec to read file file:///gpfs/data/chaklab/data/HSCR_WES_2020_MF/Psomagen_broad_Macrogen_all/combined_gVCF_Psomagen_broad_Macrogen_1KG_union/statistical_analysis/family_based/DNM/per_fam_vcf/438/Chaklab_438.vcf.gz

23:33:42.306 INFO  IntervalArgumentCollection - Initial include intervals span 3217346917 loci; exclude intervals span 213268310 loci

23:33:42.307 INFO  IntervalArgumentCollection - Excluding 213268310 loci from original intervals (6.63% reduction)

23:33:42.308 INFO  IntervalArgumentCollection - Processing 3004078607 bp from intervals

23:33:42.342 INFO  CalculateGenotypePosteriors - Done initializing engine

23:33:42.379 INFO  PedReader - Reading PED file /gpfs/data/chaklab/data/HSCR_WES_2020_MF/Psomagen_broad_Macrogen_all/combined_gVCF_Psomagen_broad_Macrogen_1KG_union/statistical_analysis/family_based/DNM/input_file/per_fam_ped/438.ped with missing fields: []

23:33:42.382 INFO  PedReader - Phenotype is other? false

23:33:42.475 INFO  ProgressMeter - Starting traversal

23:33:42.475 INFO  ProgressMeter -        Current Locus  Elapsed Minutes    Variants Processed  Variants/Minute

23:33:51.692 INFO  ProgressMeter -       chr22:38107840              0.2                 55532         361497.2

23:33:51.693 INFO  ProgressMeter - Traversal complete. Processed 55532 total variants in 0.2 minutes.

23:33:51.761 INFO  CalculateGenotypePosteriors - Shutting down engine

[April 14, 2022 11:33:51 PM EDT] org.broadinstitute.hellbender.tools.walkers.variantutils.CalculateGenotypePosteriors done. Elapsed time: 0.20 minutes.

Runtime.totalMemory()=35113664512

Using GATK jar /gpfs/share/apps/gatk/4.1.2.0/gatk-4.1.2.0/gatk-package-4.1.2.0-local.jar

Running:

    java -Dsamjdk.use_async_io_read_samtools=false -Dsamjdk.use_async_io_write_samtools=true -Dsamjdk.use_async_io_write_tribble=false -Dsamjdk.compression_level=2 -Xmx40g -Xms32g -XX:ParallelGCThreads=2 -jar /gpfs/share/apps/gatk/4.1.2.0/gatk-4.1.2.0/gatk-package-4.1.2.0-local.jar CalculateGenotypePosteriors -V /gpfs/data/chaklab/data/HSCR_WES_2020_MF/Psomagen_broad_Macrogen_all/combined_gVCF_Psomagen_broad_Macrogen_1KG_union/statistical_analysis/family_based/DNM/per_fam_vcf/438/Chaklab_438.vcf.gz -ped /gpfs/data/chaklab/data/HSCR_WES_2020_MF/Psomagen_broad_Macrogen_all/combined_gVCF_Psomagen_broad_Macrogen_1KG_union/statistical_analysis/family_based/DNM/input_file/per_fam_ped/438.ped -XL chrX -XL chrY -O /gpfs/data/chaklab/data/HSCR_WES_2020_MF/Psomagen_broad_Macrogen_all/combined_gVCF_Psomagen_broad_Macrogen_1KG_union/statistical_analysis/family_based/DNM/per_fam_posterior_GT/438/Chaklab_posterior_GT_438.vcf.gz --skip-population-priors

_ESTATUS_ [ CalculateGenotypePosteriors]: 0

_END_ Thu Apr 14 23:33:51 EDT 2022

 

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