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Methylation of cytosine is a DNA modification associated with gene repression. Recently, a novel cytosine modification, 5-hydroxymethylcytosine (5-hmC) has been discovered. Here we examine 5-hmC distribution in cellular systems and during development and show that unlike 5-methylcytosine the presence of immunochemically detectable levels of 5-hmC in nuclei is lineage specific. 5-hmC correlates with a pluripotent cell state in cellular systems, The developmental dynamics of 5-hmC are different from those of 5-mC; in particular 5-hmC is enriched in embryonic contexts comparing to adult tissues. Our findings suggest that apart from the cells of neuronal lineages high levels of 5-hmC are an epigenetic feature of cellular pluri- and multi-lineage potency.