BACKGROUND Göttingen Minipigs (GMP) is the smallest commercially available minipig breed under a controlled breeding scheme and is globally bred in five isolated colonies. The genetic isolation harbors the risk of stratification which might compromise the identity of the breed and its usability as an animal model for biomedical and human disease. We conducted whole genome re-sequencing of two DNA-pools per colony to assess genomic differentiation within and between colonies. We added publicly available samples from 13 various pig breeds and discovered overall about 32 M loci, ~ 16 M. thereof variable in GMPs. Individual samples were virtually pooled breed-wise. FST between virtual and DNA pools, a phylogenetic tree, principal component analysis (PCA) and evaluation of functional SNP classes were conducted. An F-test was performed to reveal significantly differentiated allele frequencies between colonies. Variation within a colony was quantified as expected heterozygosity. RESULTS Phylogeny and PCA showed that the GMP is easily discriminable from all other breads, but that there is also differentiation between the GMP colonies. Dependent on the contrast between GMP colonies, 4 to 8% of all loci had significantly different allele frequencies. Functional annotation revealed that functionally non-neutral loci are less prone to differentiation. Annotation of highly differentiated loci revealed a couple of deleterious mutations in genes with putative effects in the GMPs . CONCLUSIONS Differentiation and annotation results suggest that the underlying mechanisms are rather drift events than directed selection and limited to neutral genome regions. Animal exchange seems not yet necessary. The Relliehausen colony appears to be the genetically most unique GMP sub-population and could be a valuable resource if animal exchange is required to maintain uniformity of the GMP.