The Amaranthaceae-Chenopodiaceae alliance has been the focus of several phylogenetic studies, but major questions concerning the internal relationships of Amaranthaceae still remain unanswered. This study aims to test the monophyly of the subfamily Gomphrenoideae and to examine the generic relationships within this group. Parsimony and Bayesian analyses of trnL-F and rpl16 show that the subfamily Gomphrenoideae is monophyletic. The clade of Gomphrenoideae represents a large and diverse radiation of the Amaranthaceae in the New World. Unilocular anthers are a morphological synapomorphy for Gomphrenoideae that was derived from bilocular anthers. Three major clades are resolved: the Gomphrenoids, the Alternantheroids, and the Iresinoids, which are largely supported by pollen morphology. The Iresinoid clade is sister to Alternantheroids plus Gomphrenoids, rendering metareticulate pollen as the synapomorphy for the latter two clades. Tribes and subtribes delimited by androecium and inflorescence characters are poly- or paraphyletic. Several genera are monophyletic including the large genus Alternanthera, whereas Gomphrena is polyphyletic. Irenella and Woehleria are resolved within Iresine whereas Blutaparon and Lithophila fall within the polyphyletic Gomphrena. The trnL-F and rpl16 sequence data are the most variable chloroplast regions examined to date for the family and are highly effective in resolving relationships in Amaranthaceae.
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