Universal primers based on plant, fungal, and animal sequences were designed and used to amplify the intergenic spacer (IGS) of 18S-26S rDNA and to sequence the 3' end of the external transcribed spacer (ETS) in Compositae. Based on these sequences, an internal ETS primer useful across Heliantheae was designed and used to amplify and sequence directly a 3' region of the ETS in 21 representatives of Calycadenia, Osmadenia, and two outgroups that were previously studied phylogenetically using rDNA ITS sequences. Size variation in the amplified ETS region varied across taxa of Heliantheae from approximately 350 bp to 700 bp, in part attributable to an approximately 200 bp tandom duplication in a common ancestor of Calycadenia and Osmadenia. Phylogenetic analysis of the repeat region alone and examination of apomorphic changes in the repeat region demonstrate that the 200 bp repeats in Calycadenia/Osmadenia have evolved divergently. Phylogenetic analyses of the entire amplified ETS region yielded a highly resolved strict consensus tree that is nearly identical in topology to the ITS tree, with strong bootstrap and decay support on most branches of the ETS tree. Combined parsimony analyses of ETS and ITS data yielded a strict consensus tree that is better resolved and generally better supported than trees based on either data set analyzed separately. We calculated a 1.3 to 1.4 fold higher rate of sequence evolution by nucleotide substitution in the ETS region studied than in ITS-1 + ITS-2. A similar disparity in the proportion of variable (1.3 ETS: 1 ITS) and potentially informative (1.5 ETS: 1 ITS) sites was observed for the ingroup. Levels of homoplasy are similar in the ETS and ITS data. We conclude that the ETS holds great promise for augmenting ITS data for phylogenetic studies of young lineages.

Key words: Calycadenia, Compositae, ETS, external transcribed spacer, phylogeny, rDNA