Three closely linked X‐chromosomal genes potentially control sex determination in <i>Cannabis sativa</i>

Summary Sex determination mechanisms in dioecious plants remain poorly understood yet offer an excellent model system to study genetic changes underlying morphological evolution. We investigated the genetic basis of sex determination in Cannabis sativa , combining quantitative trait locus mapping in a segregating population, comparative transcriptomics between monoecious and dioecious cultivars, and a genomic analysis of X–Y chromosome divergence. Quantitative trait locus mapping identified Monoecy1 , a locus on the X chromosome putatively controlling the monoecy–dioecy trait. This locus resides in the most ancient and diverged region of the sex chromosomes and contains three genes within 60,000 bp ( CsREM16 , lncREM16 and CsKAN4 ) with distinct sex‐specific and monoecy‐specific expression patterns. Monoecy1 harbors genes for male–female as well as monoecious–dioecious sex determination. We propose that the combinatorial interaction of CSREM16 , lncREM16 and CsKAN4 provides a unifying genetic framework for understanding male–female and monoecious–dioecious sex determination in C. sativa .