Total DNA was extracted from 41 species (20 genera) of the subfamily Nepetoideae (family Labiatae). Using rbcL-specific primers, the rbcL gene wsa amplified by polymerase chain reaction (PCR) and sequenced directly. RbcL sequences were evaluated with character state (maximum parsimony; PAUP) and distance methods (neighbour-joining; MEGA). In agreement with classical systematics all taxa studied cluster within the Nepetoideae and are clearly distinguished from members of the subfamily Lamioideae. A number of distinctive clades are apparent within the Nepetoideae: I-Collinsonia, II-Lavandula, III-Agastache, Glechoma, IV-Satureja, Hyssopus, Dracocephalum, V-Nepeta, VI-Hormium, VII-Prunella, VIII-Melissa, Ocimum, IX-Monarda, Mentha, X-Origanum, Thymus, XI-Salvia, XII-Rosmarinus, and XIII-Perovskia. At least five main branches representing the clades I, II, III to VII, VIII, and IX to XIII respectively, can be distinguished within the Nepetoideae studied. They might be considered representing the tribes (according to Cantino, 1992) Elsholtzieae (I). Lavanduleae (II), and Mentheae (III-XIII). The tribe Mentheae needs to be subdivided into at least three main groups (clades III-VII, VIII and IX-XIII). Majorana hortensis which is often classified as Origanum hortensis does not cluster with Origanum and deserves a genetic status of its own.