Mitochondrial (12SrRNA and cyt b, 1086 bp) and nuclear (c-mos, 374 bp) DNA sequences were used to investigate relationships and biogeography of 24 living and extinct taxa of Phelsuma geckos. Monophyly of Phelsuma and sister relationship to the SW African Rhoptropella is corroborated. Phelsuma originated on Madagascar and made multiple long-distance invasions of oceanic islands including the Mascarenes, Aldabra, Comores, Seychelles, Andamans, and perhaps Pemba. The Mascarenes were probably colonised once, about 4.2-5.1Ma, and here Phelsuma rosagularis and Phelsuma inexpectata are newly recognised as species, as are three lineages of Phelsuma cepediana. Mascarene relationships are: Phelsuma edwardnewtoni, Phelsuma gigas (Phelsuma guentheri ((((P. cepediana A (P. cepediana B, C)) P. rosagularis) Phelsuma borbonica) ((Phelsuma ornata, P. inexpectata) Phelsuma guimbeaui))). The two recently extinct species, P. edwardnewtoni and the giant secondarily nocturnal Phelsuma gigas, differentiated on Rodrigues while on Mauritius the large nocturnal P. guentheri separated from a small diurnal form that radiated into six species, a likely result of volcanic activity. Two small-bodied lineages from Mauritius invaded the more recent island of Réunion producing two more species. Outside the Mascarenes, two mainly Madagascan assemblages are substantiated: Phelsuma serraticauda (Phelsuma lineata, Phelsuma laticauda, Phelsuma quadriocellata); (Phelsuma m. kochi (Phelsuma m. grandis, Phelsuma abboti)) (Phelsuma astriata, Phelsuma sundbergi). Their relationships to the Mascarene clade, and to Phelsuma mutabilis, Phelsuma standingi and Phelsuma andamanensis are unresolved.