Unveiling the Identity of Wenwan Walnuts and Phylogenetic Relationships of Asian Species Using Restriction Site-Associated DNA-Sequencing.

species have considerable ecological and economic value worldwide. In China, Wenwan walnuts have been collected by aristocrats and noblemen for more than 2000 years. As a diversity center of Asian , five species are widely distributed in China. The most famous of these is Mahetao (), which is an uncharacterized species that is mostly cultivated. Wild individuals are very rare and are endemic to Hebei Province. Because of the minimal variations in previously used molecular markers and the heterogeneity between chloroplast and nuclear genomes, determining the phylogenetic relationships among the species has been challenging, and has hindered subsequent evolutionary inferences. In this study, we collected enough materials for both cultivated and wild Mahetao to construct well-resolved phylogenetic trees for Asian species. We used a high-throughput genome-wide restriction site-associated DNA sequencing method. Consequently, the identity of has been clearly resolved. Our results indicate that is a hybrid of and . However, and should be considered as a single species from section . Additionally, , and likely represent one species from section according to morphological and molecular studies. These results are supported by population structure analysis and morphological comparison. We propose that trees growing in the wild should be conserved because of the economic value of their nuts. These trees may be of particular importance to impoverished communities. Furthermore, they may serve as a valuable genetic resource relevant for enhancing the production of edible walnuts. The 2b-RAD method is a viable option for future phylogenetic studies of species as well as other plant species.