Computational protocol: The renal urate transporter SLC17A1 locus: confirmation of association with gout

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Protocol publication

[…] Using CHB HapMap data as the most closely related and available population to Polynesia, Haploview software (Broad Institute, Cambridge, MA, USA) was used to define four haplotype blocks using the Gabriel confidence interval method covering SLC17A1 (defined by rs4712972 (25.772 Mb) to rs12192635 (25.881 Mb)). Variants tagging major haplotypes were selected: rs9358890 in block 1, rs3799344 in block 2, rs1183201 in block 3 (previously associated with control of serum urate concentration) [] and rs12664474 in block 4. The haplotype blocks extended into flanking genes (SLC17A4 and SLC17A3). In Centre d'Etude du Polymorphisme Humain (CEU) Caucasian population and the CHB population, rs1183201 and rs3799344 exhibited some intermarker linkage disequilibrium (LD) (r2 = 0.77 and 0.50, respectively) and rs9358890 and rs12664474 also exhibited LD (r2 = 0.35 and 0.62, respectively). SNP rs9358890 is in SLC17A4, and rs12664474 is in SLC17A3. SNP rs1165196 (SLC17A1) was also selected, in strong LD with rs1183201 (r2 = 0.87 in CEU and 0.91 in CHB) (Figure ).Genotyping was done by TaqMan® SNP genotyping assays (Applied Biosystems, Foster City, CA, USA) using a Lightcycler® 480 Real-Time PCR System (Roche, Indianapolis, IN, USA) for four SNPs: rs1183201 (assay ID: C_1911034_10), rs9358890 (assay ID: C_25595118_10), rs3799344 (assay ID: C_194536_10) and rs12664474 (assay ID: C_11189653_10). SNP rs1165196 was genotyped using Sequenom technology (Sequenom, Inc. San Diego, CA, USA). SNPs rs9358890 and rs12664474 had been genotyped over 590 of the Caucasian controls on the Affymetrix 6 SNP array (Affymetrix, Santa Clara, CA, USA) [] - genotypes were imputed for rs3799344 and rs1183201with IMPUTE2, using HapMap3 CEU (NCBI Build 36 (db126b)) as reference haplotypes. [...] ORs were calculated using PLINK software []. Because the case-control sample sets were not matched for sex, association analysis also included sex as a possible confounder. Analysis of association of haplotypes was also performed using PLINK. Meta-analysis was carried out using Rmeta software (within STATA 8.0, Stata, College Station, TX, USA) to calculate the combined Mantel-Haenszel OR using a fixed-effects model and the Breslow-Day test for heterogeneity.The Māori population of NZ is admixed, primarily Caucasian. This leads to genetic stratification, which is a confounding factor for case-control genetic studies, especially when the prevalence of disease differs between the interbreeding populations. The prevalence of gout in NZ Māori is approximately double that in NZ Caucasian []. Given this, it is not surprising that the EP case sample set, which is predominantly NZ Māori (78% of cases, 93% of controls), has a significantly greater proportion of self-reported EP grandparents than does the control sample set (average of 3.1 EP grandparents in cases vs. 2.5 in controls, P = 3.1 × 10-14 by t test). Sixty-seven biallelic genomic control markers (see Supplemental Table in Additional file ) were genotyped in the EP sample set, and STRUCTURE software [] was used to estimate the individual proportion of EP ancestry. This estimation was performed using the following parameters: number of populations assumed to be two, 30,000 burn-in period, and 100,000 Markov chain Monte Carlo replications after burn-in. Caucasian control individuals genotyped for the 67 markers were included as representative of the ancestral Caucasian population to aid in population clustering, although we were unable to include EP ancestral representatives. Plots of self-reported ancestry versus STRUCTURE estimated ancestry are shown in Supplemental Figure in Additional file . For association analysis we created two datasets matched for EP ancestry - EP/N with estimated EP ancestry > 0.65 (the geometric mean; 236 cases and 192 controls), and EP/Z with estimated EP ancestry ≤ 0.65 (48 cases and 157 controls). The estimated average proportion of EP ancestry in the EP/N sample set was 0.90 in cases and 0.88 in controls, and for the EP/Z group was 0.41 in cases and 0.40 in controls. […]

Pipeline specifications

Software tools Haploview, IMPUTE, PLINK, rmeta
Application GWAS
Diseases Rheumatic Diseases
Chemicals Sodium, Uric Acid