Premise The genus Gynoxys and relatives form a species‐rich lineage of Andean shrubs and trees with low genetic distances within the sunflower subtribe Tussilaginineae. Previous molecular phylogenetic investigations of the Tussilaginineae have included few, if any, representatives of this Gynoxoid group or reconstructed ambiguous patterns of relationships for it. Methods We sequenced complete plastid genomes of 21 species of the Gynoxoid group and related Tussilaginineae and conducted detailed comparisons of the phylogenetic relationships supported by the gene, intron, and intergenic spacer partitions of these genomes. We also evaluated the impact of manual, motif‐based adjustments of automatic DNA sequence alignments on phylogenetic tree inference. Results Our results indicate that the inclusion of all plastid genome partitions is needed to infer well‐supported phylogenetic trees of the Gynoxoid group. Whole plastome‐based tree inference suggests that the genera Gynoxys and Nordenstamia are polyphyletic and form the core clade of the Gynoxoid group. This clade is sister to a clade of Aequatorium and Paragynoxys and also includes some but not all representatives of Paracalia. Conclusions The concatenation and combined analysis of all plastid genome partitions and the construction of manually‐curated, motif‐based DNA sequence alignments are found to be instrumental in the recovery of well‐supported relationships of the Gynoxoid group. We demonstrate that the correct assessment of homology in genome‐level plastid sequence data sets is crucial for subsequent phylogeny reconstruction and that the manual post‐processing of multiple sequence alignments improves the reliability of such reconstructions amid low genetic distances between taxa.

PREMISEThe genus Gynoxys and relatives form a species-rich lineage of Andean shrubs and trees with low genetic distances within the sunflower subtribe Tussilaginineae. Previous molecular phylogenetic investigations of the Tussilaginineae have included few, if any, representatives of this Gynoxoid group or reconstructed ambiguous patterns of relationships for it.METHODSWe sequenced complete plastid genomes of 21 species of the Gynoxoid group and related Tussilaginineae and conducted detailed comparisons of the phylogenetic relationships supported by the gene, intron, and intergenic spacer partitions of these genomes. We also evaluated the impact of manual, motif-based adjustments of automatic DNA sequence alignments on phylogenetic tree inference.RESULTSOur results indicate that the inclusion of all plastid genome partitions is needed to infer fully resolved phylogenetic trees of the Gynoxoid group. Whole plastome-based tree inference suggests that the genera Gynoxys and Nordenstamia are polyphyletic and form the core clade of the Gynoxoid group. This clade is sister to a clade of Aequatorium and Paragynoxys and also includes some but not all representatives of Paracalia.CONCLUSIONSThe concatenation and combined analysis of all plastid genome partitions and the construction of manually curated, motif-based DNA sequence alignments are found to be instrumental in the recovery of strongly supported relationships of the Gynoxoid group. We demonstrate that the correct assessment of homology in genome-level plastid sequence datasets is crucial for subsequent phylogeny reconstruction and that the manual post-processing of multiple sequence alignments improves the reliability of such reconstructions amid low genetic distances between taxa.