The increase in biological diversity with decreasing latitude is widely appreciated but the cause of the pattern is unknown. This pattern reflects latitudinal variation in both the origin of new species (cladogenesis) and the number of species that coexist. Here we address latitudinal variation in species origination, by examining population genetic processes that influence speciation. Previous data suggest a greater number of speciation events at lower latitudes. If speciation events occur more frequently at lower latitudes, we predicted that genetic divergence among populations within species, an important component of cladogenesis, should be greater among lower latitude populations. We tested this prediction using within-species patterns of mtDNA variation across 60 vertebrate species that collectively spanned six continents, two oceans, and 119 degrees latitude. We found greater genetic divergence of populations, controlling for geographic distance, at lower latitudes within species. This pattern remained statistically significant after removing populations that occur in localities previously covered by continental glaciers during the last glaciation. Results suggest that lower latitude populations within species exhibit greater evolutionary independence, increasing the likelihood that mutation, recombination, selection, and/or drift will lead to divergence of traits important for reproductive isolation and speciation. Results are consistent with a greater influence of seasonality, reduced energy, and/or glacial (Milankovitch) cycles acting on higher latitude populations, and represent one of the few tests of predictions of latitudinal variation in speciation rates using population genetic data.