Current and Emerging Technologies for the Analysis of the Genome-Wide and Locus-Specific DNA Methylation Patterns

DNA methylation is the most studied epigenetic modification, and altered DNA methylation patterns have been identified in cancer and more recently also in many other complex diseases. Furthermore, DNA methylation is influenced by a variety of environmental factors, and the analysis of DNA methylation patterns might allow deciphering previous exposures. A number of techniques to study DNA methylation either genome-wide or at specific loci have been devised using a limited number of principles for differentiating the methylation state: (1) methylation-sensitive/dependent restriction enzymes, (2) antibody or methyl-binding protein-based enrichment, or (3) chemical or enzymatic conversion, (4) direct sequence readout. Second-generation sequencing has largely replaced microarrays as a readout platform and is also becoming more popular for locus-specific DNA methylation analysis. In this chapter, the currently used methods for both genome-wide and locus-specific analysis of 5-methylcytosine as well as its oxidative derivatives such as 5-hydroxymethylcytosine are reviewed in detail and advantages and limitations of each approach are discussed. Furthermore, emerging technologies avoiding PCR amplification and allowing a direct readout of DNA methylation are summarized, together with novel applications, such as the detection of DNA methylation in single cells or in circulating cell-free DNA.