Computational protocol: Genome-wide pathway analysis identifies VEGF pathway association with oral ulceration in systemic lupus erythematosus

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Protocol publication

[…] In the discovery stage, the genome-wide genotyping of the 482 SLE patients was performed using the Illumina Quad610 Beadchips (Illumina, San Diego, CA, USA) at the Centro Nacional de Genotipado (CeGen, Madrid, Spain). The genotyping quality control analysis was performed using PLINK software (Additional file : Figure S1) []. To evaluate the presence of potential population stratification in the SLE patient cohorts, we used the principal component analysis (PCA) implemented in the EIGENSOFT (v4.2) software []. Using the first 10 PCs of variation over 10 iterations we identified 14 samples showing an outlier genetic background and were excluded from downstream analysis (Additional file : Figure S1). After the quality control analysis, a final dataset of 507,051 single nucleotide polymorphisms (SNPs) and 395 SLE patients was available for the GWPA.The validation of the two genetic pathways associated with SLE in the discovery stage required the genotyping and analysis of a total 1347 SNPs. Given the large number of variants to be tested and the utility of genome-wide data for accurate genetic ancestry identification, the 425 SLE patients in the validation cohort were genotyped using the same microarray platform. Genotyping for the validation stage was performed at the HudsonAlpha Institute for Biotechnology (Huntsville, AL, USA). The same quality control analysis as in the discovery stage was performed (Additional file ). A total of 394 SLE patients and all 1347 SNPs from the two genetic pathways passed the quality control and were available for the pathway-based analysis of the validation stage. [...] The statistical association analysis between the allele dosage of the established SLE risk SNPs and the SLE clinical phenotypes was performed using the logistic regression model implemented in the SNPTEST v2 software (Oxford, UK) []. In this model, the allele dosage was defined as follows:AlleleDosagei=∑g=02PrG=g*g Where g represents each genotype of a particular genetic variant i and Pr(G = i) is the marginal posterior probability obtained by imputation. The allele dosage takes values between 0 and 2. SLE patients without phenotypical data available for the phenotype analyzed were excluded from the association analysis. Finally, the P values obtained from the discovery and replication stages were combined using the METAL software []. […]

Pipeline specifications

Software tools PLINK, EIGENSOFT, SNPTEST
Applications Population genetic analysis, GWAS
Organisms Homo sapiens
Diseases Hematologic Diseases, Lupus Erythematosus, Systemic, Melanoma, Rheumatic Diseases
Chemicals Tretinoin