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The family Melanotaeniidae (rainbowfishes) represents the largest monophyletic group of freshwater fishes found in Australia and New Guinea. The family consists of seven genera and a total of 81 species, which are broadly distributed throughout the region. We conducted a phylogenetic analysis of Melanotaeniidae based on nearly complete taxonomic sampling of all species. We sequenced seven protein coding mitochondrial genes and the first two introns of the nuclear S7 gene, for a total of 6827 base pairs. Our goal was to use the phylogeny to infer the biogeographic history of rainbowfishes in this region, to provide a framework for the timing of divergence within the family, and to test for possible introgression between species. We found strong support for the monophyly of Melanotaeniidae. Three species-poor genera-Cairnsichthys, Rhadinocentrus and Iriatherina-were all resolved as early branching …