DNA was extracted from blood or saliva using standard DNA isolation products (Roche Diagnostics, Germany; QIAGEN, USA; DNA Genotek, USA). The four SNP assays (ADH1B: rs1229984, rs1229982, rs1159918; and ADH1C: rs698) were designed for the Sequenom MassArray system (Sequenom, USA). Genotyping was done using a modified single nucleotide extension reaction (iPLEX assays) with allele detection by mass spectrometry (Sequenom MassArray System). For quality control, assays were tested on two sets of 40 unrelated individuals from the Coriell European and African-American samples (Coriell Institute for Medical Research, USA), and Hardy-Weinberg equilibrium (HWE) was confirmed in both sets. A panel of 32 ancestry informative short tandem repeat markers (AIMs) to assess population substructure (and shown to be adequate to this purpose; (Listman et al., 2010) was genotyped on a subset of the sample (N=1,096) as well as other related samples (Listman et al., 2010). Briefly, STRUCTURE 2.2 (Pritchard et al., 2000) was used to identify four parental populations, with two main subpopulations reflecting the Northern-to-Southern European cline (average proportions of contribution to present sample: 47.3% Northern Europe, 48.3% Southern Europe) with only minor contributions from “African” (1.7%) and “Asian” (2.7%) ancestral populations (Listman et al., 2010).
