Poor storability of recalcitrant seeds is due to their inability to tolerate low moisture content. Understanding the processes underlying their recalcitrance is a prerequisite to developing a maintenance strategy and prolonging their lifespan. Multiple studies have investigated the differences between orthodox (desiccation-tolerant) and recalcitrant (desiccation-sensitive) seeds. Information on epigenetic regulation, however, is lacking and thus limits our understanding of the processes defining the physiology of seeds. In the present comparative study, changes in the global levels of 5-methylcytosine (m5C) in orthodox and recalcitrant seeds of Acer platanoides L. and Acer pseudoplatanus L. were characterized during progressive stages of severe drying. Concomitant with their differential sensitivity to desiccation stress, we demonstrate variation in the response of embryonic axes and cotyledons to water deficit at the level of DNA methylation. Results indicate that desiccation-induced changes in m5C are both tissue- and seed category-specific and are highly correlated with recalcitrant seed viability. Moreover, we demonstrate that m5C global changes in response to desiccation are not retained in DNA isolated from seedlings, except in seedlings that are derived from strongly desiccated orthodox seeds (moisture content of 3.5%). Finally, the potential utilization of m5C status as a universal seed viability marker is discussed.

Black poplar (Populus nigra L.) is a widely distributed species that plays a crucial role in riparian forest ecosystems. Due to a reduction in its natural habitats and hybridization with introduced poplar species clones, its genetic pool is decreasing and efforts are required to preserve this species. Seeds of black poplar are short-lived and quickly lose viability during conventional storage in gene banks. Therefore, in order to preserve ex situ the genetic diversity of this species, the feasibility of seed cryopreservation in liquid nitrogen (at -196°C, LN) for periods of 24 h and 2 years was investigated. Seeds were harvested from three individual trees (two provenances) and desiccated to different levels of water content (WC) in the range of 0.02-0.35 g g -1 (g H 2 O/g dry mass, g g -1 ) prior to immersion in LN. Seed germination was assessed after each treatment. P. nigra seeds tolerated desiccation to WC 0.07 g g -1 but after severe desiccation to WC\0.05 g g -1 exhibited a significant reduction in germination. Results indicated that all black poplar seeds, regardless of origin, year of harvesting and seed quality, could be cryopreserved for 24 h when their WC was in the range of 0.11-0.17 g g -1 . Physiology of P. nigra seeds showed in this paper is consistent with attributes of intermediate seed storage behavior. This study provides a foundation for using cryopreservation for the ex situ conservation of P. nigra seeds.

) is an alluvial forest tree species whose genetic pool is decreasing in Europe. Poplar trees produce short-lived seeds that do not store well.