A novel set of single-copy nuclear gene markers in white oak and implications for species delimitation

Oaks have often been the focus of research on plant evolution owing to their propensity to intercross and their important role in ecology and economy. Compared with traditional molecular markers, such as amplified fragment length polymorphisms (AFLPs) and simple sequence repeats (SSRs), multiple single-copy nuclear genes (SCNGs) are of greater utility in inferring evolutionary processes in oaks. Nineteen primer pairs were developed from expressed sequence tags (ESTs) of Quercus mongolica and Q. robur that could produce orthologous products in Chinese white oaks (section Quercus). These SCNG markers showed a moderate to high level of nucleotide polymorphism in 42 individuals of two closely related white oaks, Q. mongolica and Q. liaotungensis, and demonstrated high transferability across seven white oaks, four oaks from section Cerris, and one oak from section Lobatae. A phylogenetic tree based on these SCNGs provided resolution at deep nodes and robust support for delimiting populations of Q. mongolica and Q. liaotungensis; Bayesian analysis clustered individuals into their respective species with high probability and no admixture. When the same individuals were used, Bayesian clusters based on either 194 AFLPs or 19 SSRs gave comparable results, but one or several individuals respectively were identified as having admixed ancestry. This indicates that the hybridization rate between these two oaks may have been overestimated using SSR markers due to the occurrence of homoplasy. The SCNGs are powerful for species delimitation of white oaks, and these markers could be useful for future phylogenetics and phylogeography research in white oaks.