DNA methylation occurring on the 5 position of the pyrimidine ring of cytosines in the context of the dinucleotide sequence CpG forms one of the multiple layers of epigenetic mechanisms controlling and modulating gene expression through chromatin structure. It closely interacts with histone modifications and chromatin remodeling complexes to form the genomic chromatin landscape. DNA methylation is essential for proper mammalian development, crucial for imprinting and plays a role in maintaining genomic stability as well as in dosage compensation. DNA methylation patterns are susceptible to change in response to environmental stimuli such as diet or toxins, whereby the epigenome seems to be most vulnerable during early in utero development. Aberrant DNA methylation changes have been detected in several diseases, particularly cancer where genome-wide hypomethylation coincides with genespecific hypermethylation. DNA methylation patterns can be used to detect cancer at very early stages, to classify tumors as well as predict and monitor the response to antineoplastic treatment. As a stable nucleic-acid-based modification with limited dynamic range that is technically easy to handle, DNA methylation is a promising biomarker for many applications.