Seeds are increasingly being stored in seed banks for ex situ conservation, food security, and restoration reserves. Although it is hypothesized that seeds of many species can survive the dry, freezer conditions (−20°C) of seed banks for many years, shelf life is unknown for many species. In particular, changes to dormancy and germination requirements following freezer storage have not yet been studied extensively. To improve our understanding of how seed dormancy and germination respond to seed banking conditions, we evaluated seed of six restoration-priority species in the southwestern United States, which is a region of increasing restoration need. Germination tendencies of seeds that were and were not exposed to a −20°C treatment for 4–6 months were evaluated under 22 cold-moist stratification and incubation treatments to broadly assess changes to dormancy and germination requirements and speed. Direct or indirect (via interactions with stratification and/or incubation treatments) differences in dormancy breaking and germination were observed in seeds for four of the six species studied. Specifically, storage temperature accounted for differences in the final germination proportion, germination during stratification, and/or rate of germination in five of the six species. Notably, seeds of the one species that exhibit a combination of physiological and physical dormancy showed significant differences in all germination measures. However, while significant differences were found between seeds exposed to short-term seed bank storage and those that were not, these differences were small and may not be biologically meaningful, indicating that seed banking these species should not change how they are used for restoration or reintroduction purposes.

Seed longevity is a complex process of key ecological and agronomic importance. DNA damage is a significant factor affecting seed ageing. Likewise, epigenetic changes can control gene expression and, therefore, seed response to ageing. The aim of the present work was to investigate the effect of ageing on nucleic acid stability and to identify reliable molecular markers that might help to monitor epigenetic changes within plant genetic resources during conservation. DNA profiles, evaluated by RAPD (random amplified polymorphic DNA), and methylation patterns, obtained by MSAP (methylation-sensitive amplification polymorphism), were compared in non-aged and aged Mentha aquatica seeds and plants produced by them. Germination decreased to 50% by storing seeds at 35°C and 12% wc for 28 days. RAPD profiles were 99% similar in these aged seeds compared to non-aged seeds. However, seedlings produced from the aged seeds showed a 13% dissimilarity compared to seedlings produced from the non-aged seeds. About 8% difference in the MSAP epigenetic profile was detected in seeds after storage and 16% difference was detected in the seedlings produced from them. This indicates that stress from high temperature and humidity during storage induced changes on the methylation state of seeds, and that changes were also detectable in the regenerated plants. Our results suggest that DNA integrity was compromised in seeds during ageing, and on seedlings produced by aged seeds. Genotype screening techniques such as RAPD and MSAP have the potential as markers of nucleic acid stability during seed ageing.

UK trees require increased conservation efforts due to sparse and fragmented populations. Ex situ conservation, including seed banking, can be used to better manage these issues. We conducted accelerated ageing tests on seeds of 22 UK native woody species, in order to assess their likely longevity and optimize their conservation in a seed bank. Germination at four ageing time points was determined to construct survival curves, and it was concluded that multiple samples within a species showed comparable responses for most species tested, except for Fraxinus excelsior. Of all species studied, one could be classified as very short-lived, four as short-lived and 17 as medium, with none exceeding the medium category. The most important finding of this manuscript is that although some taxonomic trends were observed, the results indicate the need for caution when making broad conclusions on potential seed storage life at a species, genus or family level. Longevity predictions were compared to actual performance of older collections held in long-term storage at the Millennium Seed Bank, Kew. Although most collections remain high in viability in storage after more than 20 years, for short-lived species at least, there is some indication that accelerated ageing predicts longevity in seed bank conditions. For species with reduced potential longevity, such as Fagus sylvatica and Ulmus glabra, additional storage options are recommended for long-term gene banking.

Seed is a fertilized mature ovule, which possesses an embryonic plant. When the dry, mature seeds are subjected to imbibition, they release a wide range of organic substances, which include low molecular weight carbonyl compounds (gases and volatiles) and water-soluble organic substances (enzymes and polysaccharides). The volatile organic compounds (VOCs) are molecules of low molecular weight (300 g mol−1) and high vapour pressure (0.01 kPa at 20°C) and include diverse chemical compounds. The nature and emission kinetics of volatiles produced from seeds vary, depending on the moisture content of the seeds. Orthodox seeds stored at ‘low seed moisture content’ undergo seed deterioration, predominantly due to lipid peroxidation, initiated by autoxidation or enzymatic oxidation of unsaturated or polyunsaturated fatty acids. This peroxidation leads to emission of volatile compounds. The quantity of VOCs emitted is positively correlated with the advancement of seed deterioration. With respect to the seed germination process, exposure of seeds to ‘high moisture conditions’ leads to increased respiration, triggers glycolysis and mobilization of storage reserves, resulting in the emission of volatile metabolic products. The quantity of VOCs emitted on commencement of metabolic activity in germinating seeds depends on (1) vigour status and (2) amount of storage reserves. Since it has been established that there is a significant difference between high and low vigour seeds with respect to quantity and profile of VOCs emitted, there is great potential for utilizing the VOC profile to obtain a quick and reproducible test of vigour status of crop seeds. In order to harness the VOC profile for quick assessment of vigour status of seeds, research has to be taken up to develop standard protocols for fingerprinting of VOCs for the purpose of seed vigour assessment and to fix the standard volatile biomarker(s) specific to crop and vigour status of seeds.

Recent evidence points to relationships between intra-specific seed mass variation and viability loss in response to ageing stress. However, little is known about how seed quality may change temporally in response to such stress. Here we examined seed–water relations of mass-separated Rudbeckia mollis seeds to better understand physiological status among mass classes. We then evaluated seed viability and vigour changes in response to various storage conditions or post-storage vigour tests (a 41°C, 75% RH stress for up to 45 d). We found similar pre-storage physiology among mass classes. However, seeds of lower mass deteriorated up to 1.5-fold faster than heavier seeds under certain conditions. Stressing seeds after storage resulted in distinct vigour differences among mass classes. For example, vigour in lower mass seeds tended to decline more compared to heavier seeds following storage in a climate-controlled room. Alternatively, vigour loss varied among mass classes following storage in a non-climate-controlled shed. Our results highlight the importance of distinguishing between pre-sowing storage and post-storage vigour effects when quantifying relative levels of viability loss among seeds of different mass. Furthermore, differential responses to storage and ageing stress among mass classes may have important implications for post-storage regeneration and subsequent population dynamics.

Vitamin E is known to scavenge lipid peroxy radicals and has a purported role in preventing seed deterioration during storage. In our previous studies using 20 rice varieties from different variety groups, the specific ratio of vitamin E homologues rather than total vitamin E content was associated with seed longevity. To validate this result, we extended the experiment to a rice panel composed of 185 Aus (semi-wild rice) varieties. Seed longevity values were determined through storage experiments at 45°C and 10.9% seed moisture content (MC). Eight types of vitamin E homologues (α-, β-, γ- and δ-tocopherol/tocotrienol) were quantified by ultra-performance liquid chromatography. The theoretical initial viability in NED, Ki, was positively correlated with γ- and δ-tocopherols and negatively correlated with α-tocotrienol. The time for viability to fall to 50% during storage at elevated temperature and relative humidity, p50, was positively correlated with δ-tocopherol. The harvest MC was negatively correlated with all seed longevity traits. Taking this factor into account in a genome-wide association (GWA) analysis, we were able to correct false positives. A consistent major peak on chromosome 4 associated with −σ−1 was detected with a mixed linear analysis. Based on rice genome annotation and gene network ontology databases, we suggest that RNA modification, oxidation–reduction, protein–protein interactions and abscisic acid signal transduction play roles in seed longevity extension of Aus rice. Although major GWA regions were not overlapped across traits, three genetic markers, on chromosomes 1, 3 and 4, were associated with both δ-tocopherol and Ki and two markers on chromosome 1 and 8 were associated with both δ-tocopherol and p50.

Desiccation-tolerant (DT) plant germplasm (i.e. seeds, pollen and spores) survive drying to low moisture contents, when cytoplasm solidifies, forming a glass, and chemical reactions are slowed. DT germplasm may survive for long periods in this state, though inter-specific and intra-specific variation occurs and is not currently explained. Such variability has consequences for agriculture, forestry and biodiversity conservation. Longevity was previously considered in the context of morphological features, cellular constituents or habitat characteristics. We suggest, however, that a biophysical perspective, which considers the molecular organization – or structure – within dried cytoplasm, can provide a more integrated understanding of the fundamental mechanisms that control ageing rates, hence the variation of longevity among species and cell types. Based on biochemical composition and physical–chemical properties of dried materials, we explore three types of the interplay between structural conformations of dried cytoplasm and ageing: (1) cells that lack chlorophyll and contain few storage lipids may exhibit long shelf life, with ageing probably occurring through slow autoxidative processes within the glassy matrix as it relaxes; (2) cells with active chlorophyll may die quickly, possibly because they are prone to oxidative stress promoted by the photosynthetic pigments in the absence of metabolic water and (3) cells that lack chloroplasts but contain high storage lipids may die quickly during storage at −20°C, possibly because lipids crystallize and destabilize the glassy matrix. Understanding the complex variation in structural conformation in space and time may help to design strategies that increase longevity in germplasm with generally poor shelf life.

Inga vera subsp. affinis (Fabaceae) is a tree species native to riparian forests in Southeast Brazil and is key for the restoration of deforested areas. The species produce seeds that are highly recalcitrant. Extreme sensitivity to desiccation as well as vivipary are commonly observed in mature seeds, which also tend towards polyembryony. Past research has shown that typical strategies to store seeds are inapplicable to Inga vera as viability is completely lost when seeds are either dried to around 28% water content (wet basis) or stored at 5°C for a few weeks. Here, we examine the feasibility of storing the seeds under hydrated conditions but at reduced water potential. Freshly collected seeds were kept under conventional storage conditions (plastic bags in cold chamber, 5°C) and in polyethylene glycol (PEG) solutions (−1.6 and −2.4 MPa) at 10°C. Seed germination was assessed after various intervals of time, until all seeds had lost viability. Before storage, seeds attained 100% germination and produced an average of 1.8 normal seedlings per seed (due to polyembryony). Storage in PEG at −1.6 MPa maintained 90% germination (radicle protrusion) and one normal seedling per seed on average for more than 200 d. Osmotic storage likely slowed down metabolism within the seed and hence consumption of food reserves. The storage time achieved has practical applications for in situ restoration, but cannot address ex situ germplasm conservation. Extending shelf life for an additional 6 months allows tree nurseries to optimize the production of seedlings so that they can be planted during the wet season.

Characterizing non-lethal damage within dry seeds may allow us to detect early signs of ageing and accurately predict longevity. We compared RNA degradation and viability loss in seeds exposed to stressful conditions to quantify relationships between degradation rates and stress intensity or duration. We subjected recently harvested (‘fresh’) ‘Williams 82’ soya bean seeds to moisture, temperature and oxidative stresses, and measured time to 50% viability (P50) and rate of RNA degradation, the former using standard germination assays and the latter using RNA Integrity Number (RIN). RIN values from fresh seeds were also compared with those from accessions of the same cultivar harvested in the 1980s and 1990s and stored in the refrigerator (5°C), freezer (−18°C) or in vapour above liquid nitrogen (−176°C). Rates of viability loss (P50−1) and RNA degradation (RIN⋅d−1) were highly correlated in soya bean seeds that were exposed to a broad range of temperatures [holding relative humidity (RH) constant at about 30%]. However, the correlation weakened when fresh seeds were maintained at high RH (holding temperature constant at 35°C) or exposed to oxidizing agents. Both P50−1 and RIN⋅d−1 parameters exhibited breaks in Arrhenius behaviour near 50°C, suggesting that constrained molecular mobility regulates degradation kinetics of dry systems. We conclude that the kinetics of ageing reactions at RH near 30% can be simulated by temperatures up to 50°C and that RNA degradation can indicate ageing prior to and independent of seed death.