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For many species, seed storage protocols are still being improved to provide viable seeds of the highest quality. Seed storage is extremely problematic for short-lived seeds categorized as recalcitrant, including pedunculate oak (Quercus robur L.), for which the optimal seed storage protocol involves a temperature of –3 °C and 40% acorn moisture content recommendations. The sensitivity of pedunculated oak seeds to temperature manipulations under preparation for long-term storage has been poorly investigated, particularly in terms of the production of reactive oxygen species (ROS), which are assumed to be determinants of seed longevity. Thermotherapy, the pathogen elimination procedure, did not increase the level of three types of ROS: hydrogen peroxide (H2O2), superoxide anion radical and hydroxyl radical (•OH). The temporal heat stress of thermotherapy resulted in slightly reduced levels of H2O2, indicating activation of the antioxidant systems in acorn preparation for storage. The effect of constant storage temperatures (−3, −5, −7 °C) and their combinations (−3 → −5 °C or −3 → −5 →−7 °C) on ROS levels and seed viability was investigated in three provenances. The highest ROS levels were detected in acorns stored at −7 °C, whereas three-step cold acclimation was beneficial for reducing ROS levels. Interestingly, the levels of H2O2 were not affected by temperature in thermotherapized acorns. In contrast, decreasing storage temperature caused a linear increase in •OH levels in all provenances. The effect of heat stress and cold stress on ROS levels in relation to long-term seed storage of pedunculate oak is discussed here in relation to the seed viability evidenced via germination rates, seedling emergence and electrolyte leakage. Thermotherapy and cold acclimation of acorns can improve their viability after storage by decreasing ROS levels.

Thermotherapy and Storage Temperature Manipulations Limit the Production of Reactive Oxygen Species in Stored Pedunculate Oak Acorns