Subtribe Gonolobinae of the cosmopolitan family Apocynaceae is diverse and represents the largest radiation of milkweeds in the New World (~500 species). The largest genus, Matelea, is an amalgam that was united largely by a perceived lack of discrete morphological variation, particularly in floral structures. Using plastid and nuclear DNA sequences, we estimate a phylogeny of Gonolobinae using four chloroplast loci and three cloned nuclear regions. We investigate monophyly of Matelea and other established and provisional genera via a series of hypothesis tests. We find variable support and considerable conflict among the gene trees. Despite this, our data confirm the paraphyly of Matelea while Gonolobus and other segregate genera receive some support as monophyletic assemblages. We attribute conflicting signal in our data primarily to incomplete lineage sorting, a likely result of rapid radiation. Ancestral character reconstructions of growth form, fruit, and corolla morphology reveal a woody twining growth form to be plesiomorphic with one to two transitions to an herbaceous, non-twining growth form followed by one putative reversal to the woody twining growth form in the subtribe. Plesiomorphic states for fruit ornamentation and corolla shape are less clear and these characters exhibit high levels of homoplasy. This study is one of the most comprehensive phylogenies of gonoloboid milkweeds to date and the first to sample multiple nuclear loci and include allelic data.