Abstract Results of phylogenetic analyses of nuclear 18S–26S rDNA internal transcribed spacer (ITS) region sequences for representatives of most genera of helenioid Heliantheae and various members of Heliantheae s.s. (sensu stricto) and Eupatorieae help to clarify major lineages and relationships in the clade corresponding to Heliantheae s.l. (sensu lato) and Eupatorieae. Most subtribes of helenioid Heliantheae as circumscribed by Robinson (1981) correspond closely with ITS clades. Polygeneric subtribes of helenioid Heliantheae that appear to be monophyletic based on ITS data include Flaveriinae sensu Turner and Powell (1977), Madiinae sensu Carlquist (1959), and Peritylinae sensu Robinson (1981). Chaenactidinae sensu Robinson (1981) is polyphyletic, but most members of the group are encompassed within only four ITS clades. Based on the ITS data and results of combined analyses of ITS variation plus previously published morphological and chloroplast DNA data, we conclude that Heliantheae s.s. and Eupatori...