Kids First DRC Joint Genotyping Workflow

Kids First Data Resource Center Joint Genotyping Workflow (cram-to-deNovoGVCF). Cohort sample variant calling and genotype refinement. Note: The DNA annotation has been significantly upgraded since v2.2.3, if you'd like to use the old version, revert to that release.

Using existing gVCFs, likely from GATK Haplotype Caller, we follow this workflow: Germline short variant discovery (SNPs + Indels), to create family joint calling and joint trios (typically mother-father-child) variant calls. Peddy is run to raise any potential issues in family relation definitions and sex assignment.

If you would like to run this workflow using the cavatica public app, a basic primer on running public apps can be found here. Alternatively, if you'd like to run it locally using cwltool, a basic primer on that can be found here and combined with app-specific info from the readme below. This workflow is the current production workflow, equivalent to this Cavatica public app.

Runtime Estimates

• Trio of 6 GB gVCFs Input: 540 Minutes & ~$6.95

Tips To Run:

1. inputs vcf files are the gVCF files from GATK Haplotype Caller, need to have the index .tbi files copy to the same project too.
2. If you are experiencing issues with Variant Recalibration either in VariantRecalibrator or ApplyVQSR, consider adjusting the max_gaussians. If a dataset gives fewer variants than the expected scale, the number of Gaussians for training should be turned down. Lowering the max-Gaussians forces the program to group variants into a smaller number of clusters, which results in more variants per cluster.
3. ped file in the input shows the family relationship between samples, the format should be the same as in GATK website link, the Individual ID, Paternal ID and Maternal ID must be the same as in the inputs vcf files header.
4. Here we recommend to use GRCh38 as reference genome to do the analysis, positions in gVCF should be GRCh38 too.
5. Reference locations:
   • https://console.cloud.google.com/storage/browser/genomics-public-data/resources/broad/hg38/v0/
   • kfdrc bucket: s3://kids-first-seq-data/broad-references/
   • cavatica: https://cavatica.sbgenomics.com/u/kfdrc-harmonization/kf-references/
6. Suggested inputs:
   • Axiom_Exome_Plus.genotypes.all_populations.poly.hg38.vcf.gz
   • Homo_sapiens_assembly38.dbsnp138.vcf
   • hapmap_3.3.hg38.vcf.gz
   • Mills_and_1000G_gold_standard.indels.hg38.vcf.gz
   • 1000G_omni2.5.hg38.vcf.gz
   • 1000G_phase1.snps.high_confidence.hg38.vcf.gz
   • Homo_sapiens_assembly38.dict
   • Homo_sapiens_assembly38.fasta.fai
   • Homo_sapiens_assembly38.fasta
   • 1000G_phase3_v4_20130502.sites.hg38.vcf
   • hg38.even.handcurated.20k.intervals
   • homo_sapiens_merged_vep_105_GRCh38.tar.gz, from ftp://ftp.ensembl.org/pub/release-105/variation/indexed_vep_cache/ - variant effect predictor cache.
   • wgs_evaluation_regions.hg38.interval_list
   • dbNSFP4.3a_grch38.gz
     • dbNSFP4.3a_grch38.gz.tbi
     • dbNSFP4.3a_grch38.readme.txt
   • CADDv1.6-38-gnomad.genomes.r3.0.indel.tsv.gz
   • CADDv1.6-38-whole_genome_SNVs.tsv.gz
   • gnomad_3.1.1.vwb_subset.vcf.gz
   • clinvar_20220507_chr.vcf.gz

Import info on cloning the git repo

This repo takes advantage of the git submodule feature. The germline annotation workflow is maintained in a different repo. Therefore, in order to get the rest of the code after cloning, you need to run: git submodule init and git submodule update. Currently this workflow uses tools from v0.4.2 of the germline workflow. If that is updated, submodule should be as well. More info on how this worked here

Annotation sub workflow

Information of default annotation performed can be found in the Kids First DRC Germline SNV Annotation Workflow docs

Other Resources

• dockerfiles: https://github.com/d3b-center/bixtools