This study investigates the karyotypes, genome sizc, constitutive heterochromatin, and NOR localization in three European brown frog species: Rana iberica, R. dalmatina and R. graeca. The three species all possess a karyotype of 26 chromosomes. The genome size is respectively 6.9, 9.7 and 11.3 pg/N in R. iberica, R. dalmatina and R. graeca. Constitutive heterochromatin is 1.7% in R. iberica, 3.1 % in R. dalmatina and 7.0% in R. graeca. The NOR is located on the long arm of the 10th chromosome in R. iberica and R. graeca, whereas it is pericentromerically localized on the short arm of the 3rd chromosome in R. dalmatina. A single NOR extrasite is found in R. graeca. Results suggest that European brown frogs have evolved independently from North American and Asian groups. Moreover, the variation observed in constitutive heterochromatin is informative of different amplification levels of satellite-like DNA. Data also suggest that in R. dalmatina NOR localization on the 3rd chromosome may be derivcd by amplification of a pre-existing NOR site.