Abstract We have studied genetic divergence and phylogenetic relationships of Alpine, Spanish and Abruzzo chamois (genus Rupicapra) by sequencing a region of 330 nucleotides within the mitochondrial DNA cytochrome b gene (mtDNA cyt b). These sequences were aligned with additional homologous sequences of Caprinae: Japanese serow, Chinese goral, Canadian mountain goat, Mishmi takin, muskox, Sardinian mouflon and domestic goat. Results suggest that, using representatives of the Bovini as outgroups, the Caprinae constitute a monophyletic clade. However, inferred phylogenetic relationships among and within tribes of Caprinae were poorly defined and did not reflect current evolutionary and taxonomical views. In fact, the Asian Rupicaprini goral and serow constituted a strongly supported clade, which included the muskox, while the takin grouped with Ovis. Therefore, the monophyly of Ovibovini was not supported by cyt b sequences. Species of Rupicapra joined a strongly supported monophyletic clade, which was distantly related to the Asian rupicaprins and Oreamnos. Therefore, the monophyly of the Rupicaprini was not supported by these cyt b sequences. There were sister species relationships within Rupicapra, Spanish and Alpine chamois and the Abruzzo chamois (Rupicapra pyrenaica ornata) was strictly related to the Spanish chamois (Rupicapra pyrenaica parva), as previously suggested by allozyme data and biogeographic reconstructions.