Basic and translational research Figure 2 Structure of associated BLK haplotypes. (A) Haplotypes formed by single-nucleotide polymorphisms (SNPs) in each associated block B1, B3 and B5 as estimated with expectation maximisation algorithm (Haploview v.4.1). Blue and red squares represent major and minor alleles, respectively. To the right of each haplotype, its population frequency (cases and controls) and connections between blocks (thin lines if >1%, thick lines if >10%) are displayed. In crossing areas are the values of the multiallelic D′ which represent the level of recombination between two blocks. See table 1 for haplotype-speciﬁc ORs and p values. Haplotypes associated with increased risk of developing systemic lupus erythematosus are enclosed within rectangles (solid line if p<1×10−3; dashed line if p<0.05). B1 contains a risk haplotype tagged by rs998683-A(b) (strongest hit in this study) and its proxies including the rs13277113-A allele(a) previously described.1 Blocks B1 and B3 are highly correlated. B3 contains SNP rs9329246(c), which is shared by the two strongest four-SNP windows of the whole BLK locus. B5 contains the independent risk haplotype tagged by the mutant allele of the rs55758736/Ala71Thr non-synonymous SNP(d) and a set of highly correlated allelic variants (grey circles). The independence of B5 risk haplotype is further supported by (B) the linkage disequilibrium plot displaying r2 values between the representative variants of each block and (C) the relationship between their alleles. the strata argument, we performed a structured test of associ- ation for all SNPs. In a stepwise manner, the SNP with higher p value was identiﬁed in each step and then used as covariate for the next association step until no SNP remained associ- ated. GenABEL package was run using R version 2.9.2. Genome sequence track analysis for epigenomic marks and transcription factor binding sites Associated and non-associated SNPs within haplotype blocks were analysed for enrichment with eight epigenomic marks using GenomeRunner.23 For SNPs within each haploblock the same number of randomly selected nucleotides from the whole range of haploblocks within chr8:11374997-11475653 coordi- nates (NCBI36/hg18 genome assembly) was sampled over the course of 100 Monte-Carlo simulations. The average expected frequency of their association with a given genomic feature Ann Rheum Dis 2012;71:1219–1226. doi:10.1136/annrheumdis-2011-200987 (eg, nuclear factor κ B (NFkB) binding site, epigenomic marks) was compared with the observed frequency using a χ2 test. Speciﬁcally, SNPs within haploblocks were tested for enrich- ment against experimentally validated NFκBp65, IRF4 and PAX5 transcription factor binding sites annotated in the wgenco- deregtfbs clustered UCSC table of the genome browser.24 Other tables included histone acetylation and methylation marks (Regulation/Broad histone tables for Gm12878 lymphoblastoid cell line). p Values were –Log10 converted and a – was included for under-represented p values. The values are shown as a heat- map plotted using Matlab (Natick, Massachusetts, USA) (online ﬁgure S2). p Values are presented in the online table S3. We used SIFT25 to predict the status of rs55758736. SIFT pre- dicts whether an amino acid substitution affects protein function based on similarity to related sequences and physical properties of amino acids (http://sift.jcvi.org/). 1221