Simultaneous sequencing of hundreds of nuclear loci for phylogenomic analyses across Rosaceae: A next generation sequencing approach

Phylogenetic reconstruction using organelles or small numbers of nuclear loci often fails to accurately reconstruct organismal relationships, especially in groups with high rates of speciation and/or reticulate evolution, such as Rosaceae. The apple, peach and strawberry genomes have been sequenced, and these genera represent 3 of the 4 lineages of the family. To simultaneously obtain sequence data from hundreds of nuclear loci, probes were designed for solution hybridization-based target enrichment followed by sequencing on the Illumina HiSeq platform (Hyb-Seq). A total of 1419 conserved orthologous exons (<15% sequence divergence in pairwise comparisons across the three genomes and >10% divergence among loci in a genome) were targeted from 257 single-copy nuclear loci. The analysis included 24 species representing the 12 recognized Rosaceae tribes and 4 unplaced genera. A mean of 1342 exons and 392,000 base pairs were obtained per species. In addition, nearly complete plastomes were assembled via genome skimming. Maximum likelihood phylogenetic analysis was conducted using nucleotide and amino acid sequences with concatenation and gene tree approaches. The robust phylogenetic resolution of all inter-tribal relationships provides a strong foundation for studies of gene family evolution, biogeography and character evolution in Rosaceae. The probes are available for phylogenetic analysis within genera of Rosaceae, and are currently being used in Fragaria, Rubus, Crataegus and others. These studies highlight their utility for the resolution of species delimitation, interspecific hybridization and polyploid origins.