The karyotype of the house mouse, Mus musculus domesticus, was examined in 282 specimens from 44 localities, in an effort to gain better understanding of the Robertsonian (Rb) variation known to exist in Greece. We consider that an Rb system exists in Peloponnisos, southern Greece, distributed in an area that is substantially larger than previously known. It consists of at least three Rb races with 2n = 30, 2n = 24 and 2n = 28, respectively, the last being reported for the first time in this paper and carrying Rb(3.6), Rb(8.12), Rb(10.14), Rb(13.15), Rb(9.16) and Rb(11.17) in a homozygous state. Additional instances of variation in this Rb system include individuals with 2n = 31 and 32 of variable Rb constitution and hybrids between the Rb races with 2n = 30 and 2n = 24. In southern Peloponnisos, Rb(10.14) was found in either a homozygous or a heterozygous state (2n = 38 or 39). The relationships among the Rb populations of Peloponnisos are discussed and hypotheses for their evolution are proposed. Rb variation was also recorded in two new locations of eastern Sterea Ellas (2n = 28 and 29) and one in Ipiros, north-west Greece (2n = 38). These findings corroborate the existence of two separate Rb systems in those two areas. Moreover, among a number of islands surveyed, Rb variation was only found in Kythira island, with Rb(10.14) in a heterozygous state (2n = 39). Finally, the typical all-acrocentric karyotype (2n = 40) was found in 51 of the animals studied from 13 localities. © 2005 The Linnean Society of London, Biological Journal of the Linnean Society, 2005, 84, 503–513.