Investigating 5-hydroxymethylcytosine (5hmC): the state of the art

The discovery of 5-hydroxymethylcytosine (5hmC) as an abundant base in mammalian genomes has excited the field of epigenetics, and stimulated an intense period of research activity aimed at decoding its biological significance. However, initial research efforts were hampered by a lack of assays capable of specifically detecting 5hmC. Consequently, the last 3 years have seen the development of a plethora of new techniques designed to detect both global levels and locus-specific profiles of 5hmC in mammalian genomes. This research effort has culminated in the recent publication of two complementary techniques for quantitative, base-resolution mapping of 5hmC in mammalian genomes, the first true mammalian hydroxymethylomes. Here, we review the techniques currently available to researchers studying 5hmC, discuss their advantages and disadvantages, and explore the technical hurdles which remain to be overcome.