Nuclear DNA Markers to Distinguish between Salix caprea, Salix gracilistyla, and Their Interspecific Hybrids

The Salix genus of the Salicaceae family has advantages such as rapid initial growth, a high germination rate, and asexual reproduction; therefore, it is used as a short-rotation energy crop for biomass production. The National Institute of Forest Science created new interspecific hybrid cultivars with superior biomass production by artificially interbreeding Salix caprea L. and Salix gracilistyla Miq. Identifying these hybrids during the seedling stage is challenging because their separate reproductive and vegetative growth stages necessitate prolonged observation of their morphological characteristics. Consequently, a reliable identification method is required to overcome these limitations. This study aimed to develop nuclear DNA markers to distinguish between S. caprea , S. gracilistyla , and their interspecific hybrids. An evaluation of 35 nuclear simple-sequence repeat (nSSR) markers in the Salix genus revealed two markers that distinguish these parent species and their hybrids. A sequence analysis confirmed the presence of insertion-deletion (InDel) regions within the nSSR markers that differed between S. caprea and S. gracilistyla . To effectively identify hybrids, a primer set comprising the InDel region, which exhibited only interspecies differences and no intraspecies differences, was developed. The results of this study will facilitate the genetic resource management of interspecific hybrids between S. caprea and S. gracilistyla , thus allowing for early identification and improved management of hybrids.