Stomatal morphological variation contributes to global ecological adaptation and diversification of Brassica napus

Stomatal density and guard cell length of 274 global core germplasms of rapeseed reveal that the stomatal morphological variation contributes to global ecological adaptation and diversification of Brassica napus. Stomata are microscopic structures of plants for the regulation of CO₂ assimilation and transpiration. Stomatal morphology has changed substantially in the adaptation to the external environment during land plant evolution. Brassica napus is a major crop to produce oil, livestock feed and biofuel in the world. However, there are few studies on the regulatory genes controlling stomatal development and their interaction with environmental factors as well as the genetic mechanism of adaptive variation in B. napus. Here, we characterized stomatal density (SD) and guard cell length (GL) of 274 global core germplasms at seedling stage. It was found that among the significant phenotypic variation, European germplasms are mostly winter rapeseed with high stomatal density and small guard cell length. However, the germplasms from Asia (especially China) are semi-winter rapeseed, which is characterized by low stomatal density and large guard cell length. Through selective sweep analysis and homology comparison, we identified several candidate genes related to stomatal density and guard cell length, including Epidermal Patterning Factor2 (EPF2; BnaA09g23140D), Epidermal Patterning Factor Like4 (EPFL4; BnaC01g22890D) and Suppressor of LLP1 (SOL1 BnaC01g22810D). Haplotype and phylogenetic analysis showed that natural variation in EPF2, EPFL4 and SOL1 is closely associated with the winter, spring, and semi-winter rapeseed ecotypes. In summary, this study demonstrated for the first time the relation between stomatal phenotypic variation and ecological adaptation in rapeseed, which is useful for future molecular breeding of rapeseed in the context of evolution and domestication of key stomatal traits and global climate change.