Abstract
BackgroundThe Illumina Infinium HumanMethylation450 BeadChip is an array-based technology for analysing DNA methylation at approximately 475,000 differentially methylated cytosines across the human genome. Hitherto, the array has been used for case-control studies, where sample numbers can be sufficient to yield statistically robust data on a genome-wide basis. We recently reported an informatic pipeline capable of yielding statistically and biologically significant results using only five cases, which expanded the use of this technology to rare disease studies. However, the clinical application of these technologies requires the ability to perform robust analysis of individual patients.ResultsHere we report a novel informatic approach for methylation array analysis of single samples, using the Crawford-Howell t-test. We tested our approach on patients with ultra-rare imprinting disorders with aberrant DNA methylation at multiple locations across the genome, which was previously detected by targeted testing. However, array analysis outperformed targeted assays in three ways: it detected loci not normally analysed by targeted testing, detected methylation changes too subtle to detect by the targeted testing and reported broad and consistent methylation changes across genetic loci not captured by point testing.ConclusionsThis method has potential clinical utility for human disorders where DNA methylation change may be a biomarker of disease.Electronic supplementary materialThe online version of this article (doi:10.1186/s13148-015-0081-5) contains supplementary material, which is available to authorized users.
Highlights
The Illumina Infinium HumanMethylation450 BeadChip is an array-based technology for analysing DNA methylation at approximately 475,000 differentially methylated cytosines across the human genome
As mentioned earlier, we developed a pipeline for small sample size against large control groups, using patients with Transient Neonatal Diabetes (TND)-multi-locus methylation disorders (MLMD) and BeckwithWiedemann Syndrome (BWS)-MLMD and broadly similar patterns of methylation change as determined by targeted testing
The pipeline applied a linear model as the statistical method, and CpGs were selected where they were hypomethylated compared with controls, with an adjusted P value < 1.33 × 10− 7 and M values between −1 and +1 in normal controls, to enrich for the intermediate methylation consistent with the hemimethylation of genomic imprinting
Summary
The Illumina Infinium HumanMethylation450 BeadChip is an array-based technology for analysing DNA methylation at approximately 475,000 differentially methylated cytosines across the human genome. Recent advances in technologies have made it possible to study the epigenetic changes associated with these diseases using robust genome-wide technologies including the Infinium. The 450 k array measures the intensity of fluorescent signal from methylated and unmethylated probes at approximately 475,000 CpG dinucleotides across the genome, including CpG islands, promoters, gene bodies, intergenic regions and the majority of imprinted loci. These intensities are used to calculate DNA methylation levels, with advantageous throughput, cost, coverage and technical consistency
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