Juncaginaceae are a small monocot family of mostly coastal and wetland herbs of almost cosmopolitan distribution. A phylogenetic analysis of the family and members of Alismatales was conducted to clarify the circumscription of Juncaginaceae and to understand intrafamilial relationships. For the first time, all genera associated with the family in the past were analysed together. Two plastid (rbcL and matK) and one mitochondrial gene (atpA) were sequenced. The separate and combined analysis of the three markers showed that Juncaginaceae are not monophyletic in their current circumscription. The family is re-circumscribed to exclude Maundia which is proposed to belong to a separate family Maundiaceae. In the new classification Juncaginaceae comprise three genera: Tetroncium, Cycnogeton, and Triglochin. Tetroncium is weakly supported as sister to the rest of the family. The reinstated Cycnogeton (formerly included in Triglochin) is highly supported as sister to Triglochin s.s. The enigmatic Lilaea is nested within Triglochin s.s. and highly supported as sister to the T. bulbosa complex. The results of the molecular analysis are discussed in combination with morphological characters. A key to the genera of the family is presented, and six new combinations are proposed: Cycnogeton alcockiae, Cycnogeton dubium, Cycnogeton microtuberosum, Cycnogeton multifructum, Cycnogeton rheophilum, and Triglochin scilloides.