The genus Microtus presents several species with extremely large sex chromosomes that contain large blocks of constitutive heterochromatin. Several cytogenetic and molecular studies of the repetitive sequences in species of the genus Microtus have demonstrated that the heterochromatin is highly heterogeneous. We have cloned and characterized a family of repetitive DNA sequences from M. cabrerae, a species with large heterochromatic blocks on the giant sex chromosomes. These repetitive sequences are 65.84% A-T rich, organized in tandem, with a 161-bp unit and are located on the centromeric region of autosomes and the X chromosome. In addition, this repetitive DNA is located throughout the entire heterochromatic block of the X chromosome and on three interstitial bands in the heterochromatic block of the Y chromosome. Comparative analysis of this family of repetitive sequences from three Microtus species revealed that the development of these sequences has occurred by concerted evolution. Our results support the hypothesis that the heterochromatic blocks from the sex chromosomes of different species are evolving independently and they probably have the genetic capacity to amplify and retain different satellite DNAs. For a topic related to the location of these repetitive DNA sequences on the Y chromosome of M. cabrerae, we propose a model to explain the origin of a length polymorphism previously described for this chromosome.