Infinium human methylation 450k bead chip technology

To extract the whole methylation of the OGG1 gene, a genome-wide methylation analysis was conducted. To do this, high-quality genomic DNA samples (500 ng) in the adipose tissue from healthy participants (N = 25) and patients with CRC (N = 29) were treated with bisulfite reagent using the EZ-96 DNA Methylation kit (Zymo Research, Irvine, CA, USA). Subsequently, DNA methylation was analyzed by microarray assays using Infinium Human Methylation 450K bead chip technology (Illumina, San Diego, CA, USA). After obtaining data from the OGG1 gene, DNA quality was checked, normalized, and filtered using the minfi package [47 (link)]. After that, DNA methylation for each CpG site was represented by beta and M values ranging from 0 to 1, corresponding to fully unmethylated and fully methylated, respectively. Specific differentially methylated positions from the OGG1 gene were extracted from both the promoter region and the whole gene.

Venous blood samples were drawn on EDTA tubes. Genomic DNA was extracted from PWBCs using the MasterPure^TM DNA Purification kit (Epicenter, Madison, WI), whose quality was assessed with the Pico Green dsDNA Quantitation Reagent (Invitrogen, Carlsbad, CA). High-quality DNA samples (500 ng) were treated with bisulfite using the EZ-96 DNA Methylation Kit (Zymo Research Corporation, Irvine, CA) according to the manufacturer’s instructions, converting cytosine into uracil. DNA methylation levels were measured by microarray with the Infinium Human Methylation 450 K bead chip technology (Illumina, San Diego, CA, USA) in all the cohorts, except OBEKIT, which was performed with Infinium MethylationEPIC beadchip (Illumina). This analysis was conducted in the Unidad de Genotipado y Diagnóstico Genético from Fundación Investigación Clínico de Valencia, as detailed elsewhere^{80 (link)}.