PREMISE OF THE STUDY: Hybridization has played an important role in the evolution and ecological adaptation in diploid and polyploid plants. Artemisia tridentata (Asteraceae) tetraploids are extremely widespread and of great ecological importance. These tetraploids are often taxonomically identified as A. tridentata ssp. wyomingensis, or as autotetraploids of diploid subspecies tridentata and vaseyana. Few details are available as to how these tetraploids are formed or how they are related to diploid subspecies. METHODS: We used amplicon sequencing to assess phylogenetic relationships among three recognized subspecies: tridentata, vaseyana and wyomingensis. DNA sequence data from putative genes were pyrosequenced and assembled from 329 samples. Nucleotide diversity and putative haplotypes were estimated from the high-read coverage. Phylogenies were constructed from Bayesian coalescence and neighbor-net network analyses. KEY RESULTS: Analyses support distinct diploid subspecies of tridentata and vaseyana in spite of known hybridization in ecotones. Nucleotide diversity estimates of populations compared to the total diversity indicate the relationships are predominately driven by a small proportion of the amplicons. Tetraploids, including subspecies wyomingenesis, are polyphyletic occurring within and between diploid subspecies groups. CONCLUSIONS: Artemisia tridentata is a species comprised of phylogenetically distinct diploid progenitors and a tetraploid complex with varying degrees of phylogenetic and morphological affinities to the diploid subspecies. These analyses suggest tetraploids are formed locally or regionally from diploid tridentata and vaseyana populations via autotetraploidy, followed by introgression between tetraploid groups. Understanding the phylogenetic versus ecological relationships of A. tridentata subspecies will have bearing on how to restore these desert ecosystems.