Maddenia (Rosaceae) was long recognized as a distinct genus closely related to Prunus and was recently merged with the latter. The Maddenia clade of Prunus has been revised taxonomically with four species from eastern Asia recognized. Recent molecular studies have provided evidence that this group is nested within Prunus, having a close relationship with the Padus-Laurocerasus complex. To further test the phylogenetic position of Maddenia, this study investigates pollen morphology of all former Maddenia species and 28 other Prunus species, using scanning electron microscopy and light microscopy. The Prunus species sampled here represent major subgenera: Amygdalus, Cerasus, Laurocerasus, Padus, and Prunus s.s. We found that pollen grains of these groups are all monad and tricolporate with striate or rugulate sculpturing. The pollen evidence supports a close relationship between Maddenia and species of Padus and Laurocerasus subgroups (excluding the Pygeum group), all of which have elongated, thicker, and less directional muri. Pollen of the subgenera of Cerasus, Amygdalus, and Prunus has generally thinner, highly directional, and predominantly parallel muri. The pollen grains of the Pygeum subgroup are quite distinct from the other groups of Prunus. Pygeum pollen has rugulate exines with much shorter and rod-shaped muri.