Data from three chloroplast genes (rps4 trnS spacer, trnL spacer; 1350 base pairs) for 27 of the recognized segregates show the Thelypteridaceae to be monophyletic and sister to an unresolved alliance of blechnoid, athyrioid, onocleoid, and woodsioid ferns. The family comprises two primary lineages, one phegopteroid, the other thelypteroid (including cyclosoroid). The phegopteroid lineage (Macrothelypteris, Pseudophegopteris, and Phegopteris) includes those elements that are the most dissected, lack adaxial grooves on the frond axes, and are generally morphologically the most distinct elements within the family. Within the thelypteroid-cyclosoroid lineage, three predominantly north-temperate subgroups, including Thelypteris s.s., form a free-veined clade that is in turn sister to the rest of the family. All segregates possessing x=36 (Cyclosorus sensu Smith, with predominantly anastomosing veins) form a strongly supported clade. Those groups with dysploid base chromosome numbers (x=27, 29, 30, 31, 32, 33, 34, 35) form a series of smaller clades basal to Cyclosorus s.l. Although our sampling is not yet sufficient to favor one classification over another, recognition of an intermediate number of genera may be the most reasonable taxonomic course.