Monophyly, origin and systematics of the subfamily Chloridoideae have long been debated. We present evidence from the plastid matK gene sequences of 74 species representing 55 genera and all tribes, and from extensive outgroup survey to address these questions. The monophyly of the Chloridoideae and its origin from an arundinoid group with C4 photosynthesis are unequivocally ascertained by this study. Rooted with the arundinoid Centropodia, a basal polytomy was observed among Triraphis schlecteri and three strongly supported clades labeled A, B, and C. Clade A comprises the Pappophoreae, Uniolinae, and three Eragrostideae genera; clade B includes most representatives of subtribe Sporobolinae, Spartina, Zoysia, and some Eragrostis; and clade C encompasses the remaining taxa. The two largest tribes Eragrostideae and Chloridoideae are polyphyletic. Uniolinae, Orcuttieae and Triodiinae form strongly supported monophyletic entities. The taxonomic status of other tribes is discussed. Two large genera, Chloris and Eragrostis, do not appear monophyletic. A repeated pattern of segregation of Old World from New World taxa is evident. The results provide a framework for further detailed studies at the tribal and generic levels to reach a comprehensive systematic treatment of the Chloridoideae.

Key words: Chloridoideae, matK, molecular, phylogeny, Poaceae, systematics