Cinnamomum kanehirae Hayata and C. canephora (L.) Presl is important to tree species in eastern Asia. The wood of C. kanehirae is in increasing demand for culturing Antrodia cinnamomea, a medicinal fungus that naturally grows inside the trunk of C. kanehirae. Putative hybrids between C. kanehirae and C. canephora were previously reported but with no scientific evidence, leading to confusion or misplanting. First, to identify the female parent of putative hybrids, the maternal inheritance InDel (insertion/deletion) markers were developed by using low-coverage sequencing. SNPs were developed by using the genotyping-by-sequencing (GBS) approach in C. kanehirae, C. camphora, and putative hybrids. The results indicated that the female parent of the studied hybrids was C. camphora. Eight hundred and forty of the 529,006 high-density SNPs were selected and used for analysis. Hybrids were classified as F1 (C. kanehirae × C. camphora), F2, and backcrosses. Hybridization has occurred in the human-developed area of eastern and southwestern Taiwan, and the introgression was bidirectional. For producing pure wood, buffer zones should be established around seed orchards to avoid cross-species pollination and to preserve the genetic purity of C. kanehirae. The DNA markers developed in this study will also be valuable for further wood identification, breeding, and evolutionary research.
