Large-scale validation of single nucleotide polymorphisms in gene regions

Abstract

Genome-wide association studies using large numbers of bi-allelic single nucleotide polymorphisms (SNPs) have been proposed as a potentially powerful method for identifying genes involved in common diseases. To assemble a SNP collection appropriate for large-scale association, we designed assays for 226,099 publicly available SNPs located primarily within known and predicted gene regions. Allele frequencies were estimated in a sample of 92 CEPH Caucasians using chip-based MALDI-TOF mass spectrometry with pooled DNA. Of the 204,200 designed assays that were functional, 125,799 SNPs were determined to be polymorphic (minor allele frequency > 0.02), of which 101,729 map uniquely to the human genome. Many of the commonly available RefSNP annotations were predictive of polymorphic status and could be used to improve the selection of SNPs from the public domain for genetic research. The set of uniquely mapping, polymorphic SNPs is located within 10 kb of 66% of known and predicted genes annotated in LocusLink, which could prove useful for large-scale disease association studies.

Figure 1

SNP allele frequency distribution. Minor allele frequency (MAF) distribution of 158,295 functional SNP assays that map uniquely to the human genome from this study (SQNM CEPH; histogram and solid line) compared to the distribution of 61,173 SNPs with Caucasian frequencies estimated by TSC (TSC CAUC; dashed line). The distributions are presented as densities, such that the area under each curve sums to one. The y-axis is truncated at 10, although the left-most histogram bar for SQNM CEPH extends to 17.

Figure 2

Correlation between frequencies from this study and matched Caucasian TSC SNP allele frequencies. The relationship between allele frequencies for 5,644 SNPs measured in the current study (SQNM CEPH) and matched Caucasian frequencies in the TSC collection (TSC CAUC) is compared with a bivariate contour plot. Density lines are plotted every 0.5 units, ranging from 0.5 for the outermost line to the highest density of 3.0 for the innermost line. The allele presented was arbitrarily standardized to the MassEXTEND high mass allele from each assay in this study. SNPs with minor allele frequencies <0.05 in both SQNM CEPH and TSC CAUC samples were excluded from this plot. Lower frequency SNPs are compared at the bottom of Table 1.