A novel variant near PIK3CG associated with mean platelet ...



Supplementary information for

A genome-wide meta-analysis identifies 22 loci associated with eight hematological parameters in the HaemGen consortium

Corresponding author:

Nicole Soranzo, PhD

Human and Medical Genetics Department

Wellcome Trust Sanger Institute,

Hinxton

Cambridge CB10 1HH

UK

Phone: +44 1223 492364

Fax:  +44 1223 494919

Email: ns6@sanger.ac.uk

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SUPPLEMENTARY NOTE

POPULATION SAMPLES

UK Blood Services Donor Panel 1 (UKBS-CC1)

Sample. The UKBS collection of Common Controls is an anonymised collection of DNA samples from 3,000 healthy blood donors. The collection has been established by the three British Blood Services of England, Scotland and Wales as part of the Wellcome Trust Case Control Consortium study and 1,500 of the samples (panel 1) served as shared controls in this study 1.

Genotyping, imputation and data analysis. 1,500 samples from the UKBS panel 1 were genotyped using the Affymetrix 500K Gene Chip as part of the Wellcome Trust Case Control consortium 1. The following QC criteria were applied to the data: HWE test P ≥ 1x10-6 (11,407 markers excluded), per-SNP missingness ≥ 0.05 (19,272 markers excluded) and MAF ≥ 0.05 (116,025 SNPs excluded) leaving 361,352 SNPs in the analysis. Imputation was carried out using HapMap2 CEU phased data (Build 36, v22, ) using the software IMPUTE (v0.4) 2. Association analyses of 2,619,497 imputed and genotyped SNPs (additive models) were carried out using the software SNPTEST (v1.1.5)2 adjusting for age, sex and geographical region. The average genomic inflation lambda for the eight hematological parameters was 1.01.

KORA F3 500K

Sample. The study population for the KORA F3 500K GWAS () was recruited from the KORA S3 survey (4,856 subjects, response 75%), an independent population-based sample from the general population living in the region of Augsburg, Southern Germany, examined in the years 1994/95 (KORA S3). The standardized examinations have been described in detail elsewhere 3. A total of 3,006 subjects participated in a follow-up examination of S3 in 2004/05 (KORA F3). For KORA F3 500K we selected 1,644 subjects of these participants then aged 35 to 79 years. The study has been approved by the local ethical committee and all participants gave signed informed consent.

Genotyping, imputation and data analysis. Genotyping of KORA F3 samples was performed using Affymetrix 500K Array Set as described in 4. Prior to imputation, the following QC filters were applied: sample call rate ≥ 93% and SNP call rate ≥ 90%. For fine scale analysis genotypes were imputed for all polymorphic HapMap SNPs (CEU, Build 35, v21) using a Hidden Markov Model as programmed in MACH (v1.0.9)5. For association testing of the 2,374,873 imputed and genotyped SNPs, we used the program MACH2QTL (v1.0.4), which uses an expected dosage value (0.0-2.0) as predictor in a linear regression framework. Analyses were adjusted for age and sex. The average genomic inflation lambda for the eight hematological parameters for KORA F3 was 1.03.

TwinsUK

Sample. The TwinsUK cohort (KCL, twinsuk.ac.uk) is an adult twin British registry shown to be representative of the UK singleton population 6. A total of 1,763 twins (100% females) were available with FBCs, of which 1,050 had data for MPV. Ethics approval was obtained from the Guy’s and St. Thomas’ Hospital Ethics Committee. Written informed consent was obtained from every participant to the study.

Genotyping, imputation and data analysis. All samples were typed with Infinium assay (Illumina, San Diego, USA) as described in 6. Imputation of genotypes was carried out using the software IMPUTE 2. The following QC filters were applied prior to imputation: call rate > 95% (130 SNPs excluded), heterozygosity 37% (46), MAF ≥ 1% (9489 SNPs excluded), HWE P ≥ 10-6 (1555 SNPs excluded) and per SNP call rate ≥ 95% (777 SNPs excluded). Genotypes with imputation posterior probability on individual genotype calls ≤ 0.9 were discarded. Imputation was carried out using HapMap2 CEU phased data (Build 36, v22) using the software IMPUTE (v0.4) 2. A total of 2,436,605 imputed and genotyped SNPs were retained for analysis. Association analyses (additive models) were carried out using the software MERLIN 7 adjusting for age, instrument type and family structure. The average genomic inflation lambda for the eight hematological parameters was 1.01.

KORA F4

Sample. The KORA S4 survey, an independent population-based sample from the general population living in the region of Augsburg, Southern Germany, was conducted in 1999/2001. The standardized examinations applied in the survey (4261 participants, response 67%) have been described in detail elsewhere 3. A total of 3,080 subjects participated in a follow-up examination of S4 in 2006/08 (KORA F4). For KORA F4 GWAS we selected 1,814 subjects of these participants. The study has been approved by the local ethical committee and all participants gave signed informed consent. The KORA S3 and S4 samples do not overlap.

Genotyping, imputation and data analysis. Genotyping of the KORA F4 GWAS sample was performed using Affymetrix Human SNP Array 6.0. Prior to imputation, the following QC filters were applied: sample call rate ≥ 93% and SNP call rate ≥ 90%. For fine scale analysis genotypes were imputed for all polymorphic HapMap SNPs using IMPUTE (v0.4.2) 2. For association testing of imputed genotypes, we used the software SNPTEST (v1.1.5) which uses posterior probabilities for genotypes 2. Analyses were adjusted for age and sex. The average genomic inflation lambda for the eight hematological parameters was 1.02.

SHIP

Sample. The Study of Health in Pomerania (SHIP) is a longitudinal population-based cohort study conducted in West Pomerania, the north-east area of Germany 8. For the baseline examinations, a sample of 6,267 eligible subjects aged 20 to 79 years was drawn from population registries. Only individuals with German citizenship and main residency in the study area were included. Selected persons received a maximum of three written invitations. In case of non-response, letters were followed by a phone call or by home visits if contact by phone was not possible. The final SHIP sample comprised 4,310 participants (response 68.8%), of which 4,291 are part of this study. Baseline examinations were collected between 1997 and 2001.

Genotyping, imputation and data analysis. Genotyping was performed using the Affymetrix Human SNP Array 6.0 platform. A per-sample call rate ≥ 93% filtering threshold was used prior to imputation. Imputation was carried out using HapMap2 CEU phased data (Build 36, v22) using the software IMPUTE (v0.5) 2. A total of 2,823,129 imputed and genotyped SNPs were retained for analysis. Association analyses (additive models) were carried out using the software SNPTEST (v1.1.5) 2 adjusting for age, sex and study centre.

Cambridge Bioresource (CBR)

Sample. The Cambridge BioResource (CBR) is a collection of pseudo-anonymised DNA samples from 4,000 healthy blood donors that has been established by the Cambridge Biomedical Research Centre in collaboration with NHS Blood and Transplant for use in genotype-phenotype association studies.

CAD/MI CASE-CONTROL SERIES

WTCCC-CAD

Details of the WTCCC-CAD study can be found elsewhere 1,9. Cases were recruited from families of European descent with a strong familial basis of CAD, where cases (one per family) had a validated history of either MI or coronary revascularisation (coronary artery bypass surgery (CABG) or percutaneous coronary angioplasty (PCI)) before their 66th birthday. Recruitment was carried out on a national basis in the UK 10. Controls were ascertained from two UK collections, namely the UKBS collection of Common Controls Panel 1 already described, and the 1958 Birth Cohort 11.

GerMIFS I

The German MI Family Study (GerMIFS) comprises unrelated German MI patients (age of onset ................
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