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At interphase, de-condensed chromosomes have a non-random three-dimensional architecture within the nucleus, however, little is known about the extent to which nuclear organisation might influence expression or vice versa. Here, using imprinting as a model, we use 3D RNA- and DNA-fluorescence-in-situ-hybridisation in normal and mutant mouse embryonic stem cell lines to assess the relationship between imprinting control, gene expression and allelic distance from the nuclear periphery. We compared the two parentally inherited imprinted domains at the Dlk1-Dio3 domain and find a small but reproducible trend for the maternally inherited domain to be further away from the periphery however we did not observe an enrichment of inactive alleles in the immediate vicinity of the nuclear envelope. Using Zfp57KO ES cells, which harbour a paternal to maternal epigenotype switch, we observe that expressed alleles are significantly further away from the nuclear periphery. However, within individual nuclei, alleles closer to the periphery are equally likely to be expressed as those further away. In other words, absolute position does not predict expression. Taken together, this suggests that whilst stochastic activation can cause subtle shifts in localisation for this locus, there is no dramatic relocation of alleles upon gene activation. Our results suggest that transcriptional activity, rather than the parent-of-origin, defines subnuclear localisation at an endogenous imprinted domain., Competing Interests: The authors have declared that no competing interests exist.