We examined taxa from 13 of the 17 chiropteran families, using single-copy DNA hybridization. Five taxa that either represented points of controversy in systematics or were members of problematic families were included in the experiment. The resulting data were used to build phylogenetic trees of 14 and 19 taxa, and by combining this study's data with those from two previous studies, a supertree of 36 taxa was constructed. The trees based on the three different matrices are compared and contrasted, and a phylogenetic hypothesis supporting the association of the rhinolophoid and the pteropodid groups of bats is presented. On the basis of this study, we conclude that the phylogenetically correct placement of the family Nycteridae is in a clade that does not include their putative relatives, the Rhinolophoidea. Our results suggest that the Emballonuridae, while a monophyletic group, are well embedded within the Yangochiroptera, and do not comprise the sister taxon to all other microbats. This study supports earlier DNA-hybridization results with respect to the placement of Mystacinidae within the Noctilionoidea, replicating those earlier findings. Finally, we determine that Miniopterus may well warrant recognition as a family distinct from the Vespertilionidae in which it is usually placed.