Human Genomic Epidemiology Asian Course 2022

Genotyping data for GWAS

1. PLINK binary data (Illumina ASA array)
2. 1000Genomes data (for PCA plot) converted from vcf files from 1000 Genomes

Simulation

1. Genotypes of Chr1-22 and X for 1000 samples simulated from haplotypes of 1000 Geomes CHB and CHS samples using HAPGEN2
2. ChrY genotypes (with random/missing genotypes for females) simulated based on MAF of in-house Chinese samples
3. Binary (497 cases + 501 controls, 6 missing) phenotype simulated using GCTA with hsq=0.8 and K=0.5
4. Quantitative phenotypes simulated using GCTA with hsq=0.5

Software

• PLINK 1.9 beta

Download

• Raw genotyping data
  • Simulated SNP array data with reference to Illumina ASA array
  • 1000 Genomes data for SNPs included in Illumina ASA array
• Data passed QC

Simulated samples with varying QC problems (n=12)

1. Missingness
2. Sex mismatch (id2_300,id2_301,id2_500,id2_501)
3. Heterozygosity (CHSHet002, CHSHet01)
4. Non-East asian ancestry / admixture (BEB-BEB_1, CHS-BEB_1, CHS-CEU_1, CHS-ITU_1)
5. Relatedness (CHSQUAD)
6. CNV (CHSTri21: trimsomy 21, CHSDel: deletion of part of Chr1)

Testing commands

1. Missingness

~/Programs/plink --bfile chrAll.ASA --missing --out QC/chrAll.ASA

sort --key 6 -gr  QC/chrAll.ASA.imiss | head
#        FID         IID MISS_PHENO   N_MISS   N_GENO   F_MISS
#    id1_382     id2_382          N    92781   481283   0.1928
#    id1_944     id2_944          N    71370   481283   0.1483
#    id1_671     id2_671          N    61587   482088   0.1278
#    id1_563     id2_563          N    48433   481283   0.1006
#    id1_875     id2_875          N    36187   482088  0.07506
#    id1_220     id2_220          N    27181   481283  0.05648
#    id1_500     id2_500          N     5957   482088  0.01236
#    id1_300     id2_300          N     5836   482088  0.01211
#    id1_826     id2_826          N     2782   481283  0.00578
#  CHSHet002   CHSHet002          N     1861   482088  0.00386
  
sort --key 5 -gr  QC/chrAll.ASA.lmiss | head
# CHR                        SNP   N_MISS   N_GENO   F_MISS
#  19                19:41642811      484     1012   0.4783
#  16                16:73793954      451     1012   0.4457
#  10                kgp21558165      446     1012   0.4407
#  12                12:78152680      416     1012   0.4111
#   4                 rs77925406      393     1012   0.3883
#  17                  rs2367537      387     1012   0.3824
#   2                rs144130077      383     1012   0.3785
#  11                 rs76693355      344     1012   0.3399
#  11                  rs1580873      336     1012    0.332
#  15                 kgp8838271      333     1012   0.3291

2. Sex mismatch

~/Programs/plink --bfile chrAll.ASA --check-sex ycount 0.2 0.8 40 --out QC/chrAll.ASA

egrep PROBLEM QC/chrAll.ASA.sexcheck
#        FID         IID       PEDSEX       SNPSEX       STATUS            F   YCOUNT
#  CHSHet002   CHSHet002            1            0      PROBLEM       0.6763      804
#    id1_300     id2_300            1            2      PROBLEM      0.03288        2
#    id1_301     id2_301            2            1      PROBLEM            1      802
#    id1_500     id2_500            1            2      PROBLEM     0.008734        2
#    id1_501     id2_501            2            1      PROBLEM            1      805

3. Heterozygosity

~/Programs/plink --bfile chrAll.ASA --autosome --maf 0.05 --hwe 1e-5 --geno 0.02 --indep-pairwise 200 50 0.1 --out QC/chrAll.ASA
~/Programs/plink --bfile chrAll.ASA --extract QC/chrAll.ASA.prune.in --het --out QC/chrAll.ASA.pruned

sort --key 6 -gr  QC/chrAll.ASA.pruned.het | head
#        FID         IID       O(HOM)       E(HOM)        N(NM)            F
#     CHSDel      CHSDel        47030    4.592e+04        67195      0.05212
#  BEB-BEB_1   BEB-BEB_1        46796    4.592e+04        67195      0.04112
#    id1_430     id2_430        46315    4.591e+04        67183      0.01886
#    id1_510     id2_510        46268    4.591e+04        67179       0.0168
 
sort --key 6 -gr  QC/chrAll.ASA.pruned.het | tail
#        FID         IID       O(HOM)       E(HOM)        N(NM)            F
#   id1_583     id2_583        45486    4.591e+04        67183     -0.02004
#   CHSHet01    CHSHet01        44968    4.592e+04        67195     -0.04481
#  CHS-ITU_1   CHS-ITU_1        44825    4.592e+04        67195     -0.05153
#  CHS-BEB_1   CHS-BEB_1        44817    4.592e+04        67195     -0.05191
#  CHS-CEU_1   CHS-CEU_1        44550    4.592e+04        67195     -0.06446
#  CHSHet002   CHSHet002        41372    4.592e+04        67195      -0.2138

4. Relatedness

~/Programs/plink --bfile chrAll.ASA --extract QC/chrAll.ASA.prune.in --genome --out QC/chrAll.ASA.pruned

sort --key 12 -gr QC/chrAll.ASA.pruned.genome | head
#       FID1       IID1       FID2       IID2 RT    EZ      Z0      Z1      Z2  PI_HAT PHE       DST     PPC   RATIO
#    CHSQUAD         C1    CHSQUAD         C2 OT     0  0.2423  0.4298  0.3279  0.5428  -1  0.870124  1.0000 11.0616
#    CHSQUAD         C2    CHSQUAD         F1 OT     0  0.0000  0.9851  0.0149  0.5075  -1  0.843984  1.0000      NA
#    CHSQUAD         C1    CHSQUAD         F1 OT     0  0.0000  0.9875  0.0125  0.5062  -1  0.843597  1.0000      NA
#    CHSQUAD         C1    CHSQUAD         M1 OT     0  0.0000  0.9932  0.0068  0.5034  -1  0.842704  1.0000      NA
#    CHSQUAD         C2    CHSQUAD         M1 OT     0  0.0000  0.9946  0.0054  0.5027  -1  0.842481  1.0000      NA
#  BEB-BEB_1  BEB-BEB_1  CHS-BEB_1  CHS-BEB_1 UN    NA  0.0000  1.0000  0.0000  0.5000  -1  0.834296  1.0000      NA
#    id1_245    id2_245    id1_834    id2_834 UN    NA  0.9537  0.0341  0.0122  0.0293  -1  0.751325  0.9736  2.1466

5. Validation of ancestry

• PCA plot for unrelated, sex-matched samples with <2% missingess
  • legend: square (contaminated), diamond (admixed), circle (1000G BEB)

~/Programs/plink --bfile chrAll.ASA --update-name ASA.1000G.to-update-name.snp --make-bed --out chrAll.ASA.id-1000G
~/Programs/plink --bfile chrAll.ASA.id-1000G --bmerge chrAll.ASA.1000GP-All --geno 0.05 --make-bed --out merged.chrAll.ASA.1000G
~/Programs/plink --bfile merged.chrAll.ASA.1000G --maf 0.05 --hwe 1e-5 --geno 0.05 --indep-pairwise 200 50 0.1 --out merged.chrAll.ASA.1000G
~/Programs/plink --bfile merged.chrAll.ASA.1000G --extract merged.chrAll.ASA.1000G.prune.in --pca 3 --out merged.chrAll.ASA.1000G.pruned

• PCA plot for final QC+ samples

6. Association

~/Programs/plink --bfile chrAll.ASA --remove QCneg.to-remove.indiv --maf 0.01 --geno 0.05 --hwe 1e-5 --assoc --adjust --out assoc/chrAll.ASA.QCpos.binary