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impute17q21

A protocol to impute 17q21.32 haplotypes from surrounding genotypes computed from genotype array or whole genome sequence data using the reference panel from:

Boettger L., McCarroll S., et al. Structural haplotypes and recent
evolution of the human 17q21.31 region. Nat. Genet. 44, 881–885 (2012)

This reference panel for HapMap samples was generated using droplet digital PCR as explained here. For any feedback, send an email to giulio.genovese@gmail.com or mccarroll@genetics.med.harvard.edu

Installation

Install basic tools (Debian/Ubuntu specific):

sudo apt install wget gzip samtools bcftools plink1.9 openjdk-11-jre-headless

Preparation steps

mkdir -p $HOME/res

Download Beagle binary

wget -P $HOME/res/ https://faculty.washington.edu/browning/beagle/beagle.25Nov19.28d.jar

Download reference panels

wget -P $HOME/res/ https://personal.broadinstitute.org/giulio/panels/chr17q21_haplotypes_{hm3,1kg}.GRCh3{7,8}.vcf.gz

Run 17q21.32 imputation from an input VCF

Run imputation using Beagle

vcf="..."
out="..."
sfx="1kg" # sfx="hm3"
build=38 # build=37
declare -A reg=( ["37"]="17:43165384-45781599" ["38"]="chr17:45088016-47704233" )

bcftools view --no-version "$vcf" -r ${reg[$build]} | \
  java -Xmx8g -jar $HOME/res/beagle.25Nov19.28d.jar gt=/dev/stdin \
  ref=$HOME/res/chr17q21_haplotypes_$sfx.GRCh$build.vcf.gz out="$out" \
  map=<(bcftools query -f "%CHROM\t%POS\n" $HOME/res/chr17q21_haplotypes_$sfx.GRCh$build.vcf.gz | \
  awk '{print $1"\t.\t"$2/1e7"\t"$2}')

Extract imputed 17q21.32 alleles into a table

out="..."
build=38 # build=37
declare -A reg=( ["37"]="17:44166101-44166101" ["38"]="chr17:46088735-46088735" )

bcftools index -ft "$out.vcf.gz" && \
bcftools query -f "[%SAMPLE\t%ALT\t%GT\n]" "$out.vcf.gz" -r ${reg[$build]} | tr -d '[<>]' | \
  awk -F"\t" -v OFS="\t" '{split($2,a,","); a["0"]="NA"; split($3,b,"|"); \
  print $1,a[b[1]],a[b[2]]}' > "$out.tsv"

Build the reference panels yourself

This section is only in case you want to build the reference panels yourself, you can skip it otherwise

Download GRCh37 human genome reference

wget -O- ftp://ftp.1000genomes.ebi.ac.uk/vol1/ftp/technical/reference/human_g1k_v37.fasta.gz | \
  gzip -d > $HOME/res/human_g1k_v37.fasta
samtools faidx $HOME/res/human_g1k_v37.fasta

Download GRCh38 human genome reference

wget -O- ftp://ftp.ncbi.nlm.nih.gov/genomes/all/GCA/000/001/405/GCA_000001405.15_GRCh38/seqs_for_alignment_pipelines.ucsc_ids/GCA_000001405.15_GRCh38_no_alt_analysis_set.fna.gz | \
  gzip -d > $HOME/res/GCA_000001405.15_GRCh38_no_alt_analysis_set.fna
samtools faidx $HOME/res/GCA_000001405.15_GRCh38_no_alt_analysis_set.fna

Download 17q21.32 reference panel in Beagle 3 format and an additional file with marker positions for the GRCh37 human genome reference

wget -P $HOME/res/ https://raw.githubusercontent.com/freeseek/impute17q21/master/imputation_cnv_{panel_{hm3,1kg}.bgl,haps_encoded}.txt

Liftover marker positions for the GRCh38 human genome reference

wget http://hgdownload.cse.ucsc.edu/admin/exe/linux.x86_64/liftOver
chmod a+x liftOver
wget http://hgdownload.cse.ucsc.edu/goldenPath/hg19/liftOver/hg19ToHg38.over.chain.gz
for sfx in hm3 1kg; do
  tail -n+2 $HOME/res/imputation_cnv_panel_$sfx.bgl.txt | grep -v 17:44166 | \
    awk '{print "17\t"substr($2,4)}' > $HOME/res/imputation_cnv_panel_$sfx.GRCh37
  tail -n+2 $HOME/res/imputation_cnv_panel_$sfx.bgl.txt | grep -v 17:44166 | \
    awk '{print "chr17\t"substr($2,4)-1"\t"substr($2,4)}' | \
    ./liftOver /dev/stdin hg19ToHg38.over.chain.gz /dev/stdout /dev/stderr | \
    awk '{print $1"\t"$3}' > $HOME/res/imputation_cnv_panel_$sfx.GRCh38
done

Generate 17q21.32 reference panels in VCF format for both the GRCh37 and GRCh38 human genome references

declare -A fasta=( ["37"]="$HOME/res/human_g1k_v37.fasta" ["38"]="$HOME/res/GCA_000001405.15_GRCh38_no_alt_analysis_set.fna" )
declare -A chr=( ["37"]="17" ["38"]="chr17" )
declare -A pos=( ["37"]="44166101" ["38"]="46088735" )

for sfx in hm3 1kg; do
  grep 17:44166 $HOME/res/imputation_cnv_panel_$sfx.bgl.txt | \
    awk 'NR==FNR {if (!($3 in y)) {y[$3]=i; x[i++]=$3} z[$4$5$6$7$8$9$10$11$12$13$14$15]=y[$3]}
    NR>FNR {for (j=3; j<=NF; j++) w[j]=w[j]$j}
    END {for (i in x) if (i==1) printf "<"x[i]">"; else if (i>0) printf ",<"x[i]">";
    printf "\t.\t.\t.\tGT";
    for (i in w) printf "\t"z[w[i]]}' $HOME/res/imputation_cnv_haps_encoded.txt - | \
    sed 's/\t\([1-9]\)\t\([1-9]\)/\t\1|\2/g' > tmp
  for build in 37 38; do
    tail -n+2 $HOME/res/imputation_cnv_panel_$sfx.bgl.txt | grep -v 17:44166 | tr ' ' '\t' | cut -f3- | \
      paste $HOME/res/imputation_cnv_panel_$sfx.GRCh$build - | \
      sed 's/?/-/g;s/\t\([ACGT-]\)\t\([ACGT-]\)/\t\1\2/g' | \
      bcftools convert --no-version --tsv2vcf /dev/stdin -c CHROM,POS,AA -f ${fasta[$build]} --samples \
      $(head -n1 $HOME/res/imputation_cnv_panel_$sfx.bgl.txt | tr ' ' '\n' | \
      tail -n+3 | uniq | tr '\n' ',' | sed 's/,$//') | tr '/' '|' | \
      awk -v chr=${chr[$build]} -v pos=${pos[$build]} 'NR==FNR {line=$0}
      NR>FNR {if (line && $0!~"^#" && $2>pos) {print chr"\t"pos"\t.\tT\t"line; line=""} print}' tmp - | \
      bcftools view --no-version -Oz \
      -o $HOME/res/chr17q21_haplotypes_$sfx.GRCh$build.vcf.gz
  done
  /bin/rm tmp
done

Check for consistency of the 17q21.32 reference panel

Convert the 17q21.32 and 1000 Genomes project reference panels to plink and then merge to compute consistency

url="ftp://ftp.1000genomes.ebi.ac.uk/vol1/ftp/phase1/analysis_results/integrated_call_sets/ALL.chr17.integrated_phase1_v3.20101123.snps_indels_svs.genotypes.vcf.gz"
bcftools view --no-version $url -r 17:43165384-45781599 | \
  awk 'NF==2 {print "##contig=<ID=17,length=81195210>"} {print}' | \
  bcftools view --no-version -v snps | \
  bcftools annotate --no-version -x ID -I +'%CHROM:%POS:%REF:%ALT' | \
  $HOME/bin/plink --vcf /dev/stdin --keep-allele-order --const-fid --make-bed \
  --out ALL.chr17.integrated_phase1_v3.20101123.snps_indels_svs.genotypes

for sfx in hm3 1kg; do
  bcftools annotate --no-version -x ID -I +'%CHROM:%POS:%REF:%ALT' \
    $HOME/res/chr17q21_haplotypes_$sfx.GRCh37.vcf.gz | \
    $HOME/bin/plink --vcf /dev/stdin --biallelic-only --keep-allele-order --const-fid --make-bed \
    --out chr17q21_haplotypes_$sfx.GRCh37

  plink --bfile chr17q21_haplotypes_$sfx.GRCh37 \
    --bmerge ALL.chr17.integrated_phase1_v3.20101123.snps_indels_svs.genotypes --merge-mode 6
done

You should get the following results

217071 overlapping calls, 216606 nonmissing in both filesets.
215812 concordant, for a concordance rate of 0.996334.
...
2747672 overlapping calls, 2747672 nonmissing in both filesets.
2741694 concordant, for a concordance rate of 0.997824.

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A protocol to impute 17q21.32 alleles from 17q21.32 genotypes

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