Iris subgenus Xiphium is a small group of taxa that occur in the Mediterranean Basin, a long-recognized biodiversity hotspot. Phylogenetic relationships among these Iris were reconstructed based on sequence data from 110 nuclear markers (coding regions) and plastomes using Bayesian inference and maximum likelihood methods. Best trees based on plastome and combined datasets resolved Iris subgenus Xiphium and I. xiphium as not monophyletic while nuclear and all-gene (combined nuclear and plastid coding regions) datasets resolved the subgenus as monophyletic but I. xiphium as not monophyletic. Topology tests indicated the alternative hypothesis of a monophyletic subgenus is not rejected by the combined dataset while a monophyletic I. xiphium is rejected. Topology tests also showed the hypothesis of a polyphyletic subgenus is rejected by the nuclear and all-gene datasets, however, a monophyletic I. xiphium is not rejected by these two datasets. We hypothesize that the subgenus is monophyletic based on these analyses, morphology, and biogeography, and that uneven patterns of missing data is a likely reason for topological incongruence among datasets. A previously suggested informal group within the subgenus was supported. Patterns of relationships among species suggest multiple exchanges between the African and European continents but also the importance of the Strait of Gibraltar as a barrier to genetic exchange.