High-throughput method for analyzing methylation of CpGs in targeted genomic regions

Abstract

A unique microarray-based method for determining the extent of DNA methylation has been developed. It relies on a selective enrichment of the regions to be assayed by target amplification by capture and ligation (mTACL). The assay is quantitatively accurate, relatively precise, and lends itself to high-throughput determination using nanogram amounts of DNA. The measurements using mTACLs are highly reproducible and in excellent agreement with those obtained by sequencing (r = 0.94). In the present work, the methylation status of >145,000 CpGs from 5,472 promoters in 221 samples was measured. The methylation levels of nearby CpGs are correlated, but the correlation falls off dramatically over several hundred base pairs. In some instances, nearby CpGs have very different levels of methylation. Comparison of normal and tumor samples indicates that in tumors, the promoter regions of genes involved in differentiation and signaling are preferentially hypermethylated, whereas those of housekeeping genes remain hypomethylated. mTACL is a platform for profiling the state of methylation of a large number of CpG in many samples in a cost-effective fashion, and is capable of scaling to much larger numbers of CpGs than those collected here.array | technology | tumor

Nasal Cell DNA Methylation, Inflammation, Lung Function and Wheezing in Children with Asthma

“…%5mC levels of individual CpG dinucleotides were averaged to obtain a mean measure for each of the assays. The basis for using the average was the observation that methylation values in adjacent CpG sites over a short amplicon such as that used in pyrosequencing analysis are usually highly correlated [36]. In our previous work, we observed that averages of DNA methylation within each amplicon provide more robust methylation measures compared with analyses of individual CpGs [37].…”

Epigenetic regulation of theglucocorticoid receptorpromoter 17in adult rats

“…In genome wide studies in both normal and cancerous colon tissue, irrespective of the genomic locus, Nautiyal et al observed a very high correlation in methylation levels that decreased as the distance between dinucleotides increased. 43 Over the scale of a typical promoter, they observed a linear decrease in the Pearsons' r from > 0.8 for dinucleotide pairs less than 50 bp apart to < 0.4 for pairs 1 kb apart. Chronic stress increased methylation levels significantly in the pituitary and adrenal glands, and in both tissues a promoter-wide regulation was observed.…”

Birthweight is associated with DNA promoter methylation of the glucocorticoid receptor in human placenta

“…Moreover, the correlation in methylation between CpG sites appears to be a function of the distance between sites, consistent with the findings of Nautiyal and colleagues. 23 This relatively high degree of correlation suggests that use of the mean may be a reliable marker of the extent of methylation across this region.…”