Maturing yellow lupin seeds were desiccation tolerant. Glucose, sucrose and cyclitols (mainly D-pinitol, D-chiro-inositol and myo-inositol) were predominant at the early stages of seed growth. Accumulation of the raffinose family oligosaccharides (RFOs) and the galactosyl cyclitols including galactinol, digalactosyl myo-inositol, galactopinitol A, galactopinitol B, trigalactopinitol A, ciceritol, fagopyritol B1 and fagopyritol B2 appeared during seed maturation; their increase correlated with seed germinability after desiccation. The loss of desiccation tolerance after seed germination was also studied. For the desiccation tolerance test, intact seedlings were dried rapidly or slowly followed by rehydration. Soluble carbohydrates were assayed before and after drying. Root tissues were more sensitive to desiccation than hypocotyl tissues and completely lost desiccation tolerance within 36 h of imbibition after both fast and slow-drying treatments. Survival of hypocotyls decreased gradually up to 96 h after imbibition. Loss of RFOs and galactosyl cyclitols in axis tissues preceded visible germination. Loss of desiccation tolerance was accompanied by loss of RFOs and galactosyl cyclitols and an increase in reducing sugars in cotyledon, hypocotyl and radicle tissues. Drying did not induce the accumulation of RFOs and galactosyl cyclitols in seedling tissues.

Germination over 3 years was followed in 25 soil samples of 31 × 31 × 5 cm each from primary and secondary stands in Białowieża forest, Poland. The number of seedlings emerging and the number of species declined over the 3 years. In year 1 the mean densities of emerging seedlings for the primary and secondary stands were similar (2142 m−2 and 2488 m−2 respectively); in year 3 the difference was greater (248 vs 498 m−2) but not statistically significant. The trend reflected the greater longevity of the seed bank species characteristic of the secondary forest. Two functional groups of herb and shrub species were distinguished in the seed bank based on (a) frequency as adults in the present vegetation, and (b) relative abundance in the seed banks of primary and secondary forest. Group A species need large-scale gaps in the tree canopy; most have very small seeds (<0.15 mg mean dry mass) and are very persistent in the soil. Group B species benefit from single-treefall gaps in the tree canopy, but appear to be suited primarily to gaps in the canopy of the herb layer and in the root layer made mainly by animals; their seeds are not as small (mostly >0.15 mg) and not as persistent in the soil. Group B species made up 86% of the seedlings emerging over 3 years in soil from the primary stand, and 56% of those from the secondary stand. In the laboratory all five Group B species tested showed some inhibition of germination in darkness, but only one showed unequivocal inhibition by far-red-enrichment of low irradiance light.

The fruits of Quercus robur are shed containing seeds at high moisture contents which remain desiccation sensitive and exhibit recalcitrant storage behaviour. Little is known of the control of seed development in these and other recalcitrant seeds. In the present work the changing concentrations of four plant growth regulators during seed reserve accumulation was studied over five years on seeds from the same tree. The pattern of reserve accumulation and changing ABA content in seeds differed between years. Although ABA content in the cotyledons increased in line with increasing dry weight to different contents at shedding, similar concentrations existed in cotyledons in each year. Thus ABA did not appear to be influencing dry weight accumulation. However, unlike orthodox seeds the decline in ABA concentration prior to shedding was limited and consistent with a continuing role for ABA in preventing precocious germination. An earlier peak in ABA concentration was associated with greater desiccation tolerance at shedding across years. The concentrations of zeatin and zeatin riboside in cotyledons were similar in each year and declined during reserve accumulation in a similar fashion to that reported for orthodox seeds. By contrast, IAA concentration increased in both the cotyledons and axes in the latter stages of seed development, opposite to that reported for orthodox seeds. It is possible that the increasing IAA concentration in cotyledons and axes and the stable concentration of zeatin and zeatin riboside throughout the latter stages of development in the axes of Q. robur are linked to the maintenance of active metabolism for the rapid initiation of germination upon shedding observed in seeds of this species.

cDNAs for the major cysteine endopeptidase (CEpase) of mung bean (Vigna radiata [L.] Wilczek) seedling cotyledons have been cloned using gene-specific primers with the polymerase chain reaction (PCR) in a 3'-RACE system. A cDNA clone for CEpase, pKL042, that is 1221 bp long excluding the poly A tail was isolated. It appears to contain the entire coding sequence for a 362-residue-long polypeptide. The N-terminal sequence for the mature CEpase begins at position 128 of the putative translation product, suggesting removal of an N-terminal hydrophobic signal peptide and additional sequences to produce the mature protease. Northern blot hybridization with CEpase cDNA pKL042 as probe indicates that CEpase transcript is not detectable in the cotyledons or the embryonic axis of dry seeds, but is first detectable in the day 1 cotyledons and in the day 3 axis. The level of CEpase mRNA in both cotyledons and axis increases as growth proceeds. A decline of protease activity, however, is observed after day 3 in the cotyledons, even though the level of protease transcripts continues to increase until day 8. Detachment of the axis from the cotyledons before day 3 results in the prevention of the normal increase in both protease activity and CEpase mRNA.

Regardless of whether or not seeds of the geographically-widespread Solidago altissima and S. nemoralis were exposed to light in autumn, those ‘dispersed’ in autumn (15/°C) or winter (5°C) required 12 weeks of light in winter to germinate to ≥80% in darkness in spring (2 weeks at 20/10°C). On the other hand, seeds of the narrow-endemic S. shortii dispersed in autumn and exposed to ≥2 weeks of light in early winter germinated to ≥77% in darkness in spring, and those dispersed in winter and exposed to ≥6 weeks of light germinated to ≥82%. S. altissima and S. nemoralis seeds not exposed to light during any season germinated to only 0–1% in darkness in spring, whereas S. shortii seeds germinated to 45–56%. Seeds of S. altissima and S. nemoralis kept in darkness in autumn and winter needed a 1-day (14-h photoperiod) light exposure in spring to germinate to ≥75% in darkness, whereas those of S. shortii required only one 5-s exposure. Cold-stratified (nondormant) seeds of S. altissima, S. nemoralis and S. shortii exposed to light with a high far-red/red ratio germinated to significantly higher percentages than dark controls and freshly-matured and lab-stored seeds. Results of this study suggest that a soil seed bank of S. shortii should be smaller and be depleted at a faster rate than those of S. altissima and S. nemoralis, and portions of the seeds of the three species can germinate in the far-red-enriched light under plant canopies.

The LEA-like protein previously isolated from a homogenate of pea (Pisum sativum L.) embryonic axes heated at 80°C for 10 min (Russouw et al., 1995) was purified without exposure to heat. Peptides produced by trypsin digestion were separated by HPLC and sequenced. The protein was identified as a member of the LEA group I family. The conformation of the protein was compared before and after heat treatment by antibody affinity, circular dichroism spectroscopy, fluorescence spectroscopy and 8-anilino-1-naphthalenesulfonic acid binding. No differences could be detected, demonstrating that the protein was not irreversibly denatured by exposure to high temperature.

The viability of Norway maple seeds harvested at mass maturity (57.3% moisture content) was reduced from 100% to only 38% if dried rapidly (at 15% RH and 15°C for 3 days) to 4.7% moisture content. In contrast, slow drying for 32 days (the optimum duration of several investigated) to 29.9% moisture content enabled 93% of the seeds to survive subsequent rapid desiccation to 3.5% moisture content. This is similar to the 95% viability shown by seeds harvested 40 days after mass maturity and then dried rapidly to 4.4% moisture content. However, fruits or seeds harvested at mass maturity and then held moist for 21 days also showed 94 and 91% viability after subsequent rapid desiccation to 3.8 and 3.3% moisture content, respectively. Thus a post-ovule-abscission programme is required before Norway maple seeds are able to tolerate rapid enforced desiccation to low moisture contents, but loss in moisture during this period is not essential to the development of desiccation tolerance.

Biotin is an important vitamin. It is biologically active as a protein prosthetic group, where it functions in enzymatically catalysed carboxylation reactions. It has previously been shown that the ability to synthesize biotin is not necessary for germination of Arabidopsis thaliana seeds, but that this process is required for early seedling growth. This research was conducted to determine if changes in the detection of biotinylated proteins could be observed that reflect changes in the need for biotin-mediated enzyme reactions observed during early soybean seedling growth. A seed specific 75-kDa biotinylated protein present in the embryonic axes and the cotyledons was lost during the first 3 d of germination. Seed specificity, and pattern of expression during germination suggest that this protein is a homologue of the seed specific 65-kDa biotinylated protein previously identified in pea (Pisum sativum). If samples were not treated with 2-mercaptoethanol, three equally spaced proteins at approx. 85 kDa were visible. In the presence of 2-ME these proteins appeared as a single 85-kDa band. This triplet was distinct only in the embryonic axes of dry seeds and not in imbibed seeds or in other plant parts. This demonstrates that imbibitional changes do occur in the pool of biotinylated proteins present in dry soybean seeds, and that 2-ME treatment can inhibit complete identification of the biotinylated proteins present in seed tissues.

Seeds of the narrow-endemic Solidago shortii and of the geographically-widespread S. altissima and S. nemoralis buried in December 1993 were exhumed in June 1995 and given 10 cycles of 1 day wet/5 days dry, 9 of 2/5, 8 of 3/5, 7 of 4/5 and 6 of 5/5 in light at 30/15°C; the control was kept continuously wet during the experiment. Seeds of the three species incubated on wet substrate for 3, 4 or 5 days germinated to ≥47% during the first cycle. On the other hand, seeds kept moist 2 days germinated to only 4–26% in the first cycle, and none kept moist for 1 day germinated. Cumulative germination percentages of seeds of all three species at the end of the final cycle of the 1/5 treatment were 0–4%. In the 2/5 treatment, cumulative germination percentages of S. altissima and S. shortii seeds at the end of the final cycle were 50 and 41%, respectively, but that of S. nemoralis was only 4%. For all three species, cumulative germination percentages were ≥55% at the end of the final cycle of the 3/5, 4/5 and 5/5 treatments. Control seeds of the three species germinated to 85–99% after 2 weeks, and no additional seeds germinated during the remainder of the experiment. High percentages of seeds were viable in the treatments and control at the end of the experiment; however, some seeds of S. nemoralis and S. shortii given 1/5, 2/5 and 3/5 treatments became dormant. The ecological implication of this study is that seeds of the three species will not germinate on the soil surface after brief rainfall events in summer. The germination response of the narrow endemic is similar to that of its two geographically-widespread congeners.

Solidago shortii is endemic to a small area in northcentral Kentucky (USA), whereas two of its sympatric congeners, S. altissima and S. nemoralis, are geographically widespread. Seeds (achenes) of S. shortii (0.370 mg) are significantly larger (PLSD, P=0.05) than those of S. altissima (0.070 mg) and S. nemoralis (0.068 mg). Germination percentages of freshly-matured seeds of the three Solidago species collected in November 1991, 1992 and 1994 were 0–2% in light at 15/6°C, 1–37% at 20/10°C, 9–56% at 25/15°C and 10–85% at 30/15 and 35/20°C. Stratification increased the percentage and rate of germination and decreased the time to the onset of germination (measured by Timson's index only at 20/10°C in light) in the three species. Following 12 weeks of cold stratification in light, seeds of the three species germinated to 72–100% in the light and to 22–100% in darkness over the range of thermoperiods; those cold-stratified in darkness germinated to 39–100% in light. Freshly-matured seeds of S. altissima and of S. nemoralis germinated to 0–4% in darkness, whereas those cold-stratified for 12 weeks in darkness germinated to 0–28% in darkness. On the other hand, freshly-matured and cold-stratified (in darkness) seeds of S. shortii germinated to 0–13 and 13–73%, respectively, in darkness. Under near-natural temperatures in a glasshouse without temperature control, germination of the three species peaked in March. Thus, the primary difference in dormancy-breaking and germination requirements of the three species is that the endemic germinates to a much higher percentage in darkness than its two congeners. Seeds of S. shortii do not have any special dormancy-breaking or germination requirements that could not be fulfilled outside its present-day geographic range.

The developmental pattern of biotinylated proteins (BP) during embryogenesis and maturation of soybean seed was characterized. Detection of these BP was compared with the development of desiccation tolerance in seeds. Three groups of BP were detected in soybean seeds using a biotin-streptavidin detection method: the first group consisted of a set of three bands with a mean apparent MW of 85 kDa (called BP85), detected in crude extracts of embryonic axes (EA) from non-dehydrated seeds (NDS) and from artificially slow-dehydrated seeds (DS); the second one, BP75, was a single protein with an apparent MW of 75 kDa and was expressed in cotyledons (COT) and EA tissues of NDS and DS; the third group with a mean apparent MW of 35 kDa (BP35), was expressed at high levels only in COT of NDS. BP35 concentration was highest in the early stages of seed development (21 days after flowering — DAF) and decreased as seeds developed, being almost imperceptible after 47 DAF. Conversely, only traces of BP75 and BP85 extracted from EA and COT were detected at early stages of seed development (21–33 DAF). Maximum levels of accumulation of these proteins were expressed at 42–47 DAF and remained constant until harvest maturity. Desiccation-tolerant stage of the seeds was initiated at 47 DAF, which coincided with the stage of maximum accumulation of BP75 and BP85 in the seeds, however, appearance of these proteins could be stimulated by desiccation of immature seeds that had not achieved desiccation tolerance. Therefore changes in biotinylated proteins are coincident with, but not sufficient for, the development of desiccation tolerance.

Late embryogenesis abundant (LEA) proteins accumulate in developing seeds prior to maturation drying and are presumed to help protect embryos from desiccation stress. The unusual solubility properties of these proteins, such as resistance to heat coagulation, have led to suggestions that they alter the hydration properties of cellular constituents. Hydration characteristics and water potential range at which wheat heat-soluble LEA proteins were expressed have been determined. Levels of heatsoluble proteins decline in germinating seeds but can be induced by dehydration (ψ ≤ −0.5 MPa) in crown meristematic tissue that is desiccation tolerant. The heatsoluble extract from mature wheat embryos contained proteins and sugars at about a 1:1 (w/w) ratio. Only about half of the sugars could be removed by exhaustive dialysis; the rest appeared to be tightly associated with the proteins. The water sorption characteristics of undialysed and dialysed heat-soluble protein/sugar fractions were compared with other water-soluble proteins (bovine serum albumin, lysozyme or gluten) and with sucrose. At relative humidities greater than 50%, the amount of water absorbed by protein-sugar mixes was a function of the sugar content. For the same sugar content, the heat-soluble protein preparation absorbed 2–3 times more water than a lysozyme/sucrose preparation. The rate at which heat-soluble protein fractions dried was also different to desorption rates of lysozyme/sucrose mixes. While lysozyme/sucrose mixtures dried either very rapidly (within 20 min) or very slowly (about 2 months) depending on the sugar content, desorption rates of the heat soluble protein-sugar preparations were intermediate (2–10 days) and modulated by sugar concentration. Based on the presumption that the hydrophilic properties of LEA heat-soluble proteins are important to their function, it is suggested that these proteins function to control drying so that cells stay at critical water potentials for the proper time. In this respect, the heat-soluble LEA proteins do not prevent desiccation, but serve as hydration buffers.

The effect of dry storage under ambient laboratory conditions on after-ripening and survivorship was tested on seeds of the geographically-widespread Solidago altissima and S. nemoralis and the narrow-endemic S. shortii. Freshly-matured seeds of S. altissima collected in 1991 and in 1992 germinated to low or moderate percentages in light at 15/6, 20/10 and 25/15°C and to high percentages at 30/15 and 35/20°C, whereas those of S. nemoralis and S. shortii germinated to low percentages over the range of temperature regimes. After 0.8–1.8 years of storage, 1991 seeds of S. altissima incubated in light germinated to high percentages at 25/15, 30/15 and 35/20°C, those of S. nemoralis did so at 30/15 and 35/20°C and those of S. shortii at 20/10, 25/15, 30/15 and 35/20°C; 1992 seeds of all three species germinated to high percentages at 20/10, 25/15, 30/15 and 35/20°C. Freshly-matured 1991 and 1992 seeds of the three species germinated to low percentages in darkness over the range of temperature regimes, and only seeds of S. shortii germinated to high percentages after 0.8–1.8 years of storage. Compared with cold stratification, dry storage was only moderately effective in breaking dormancy in these three species. The primary difference in after-ripening of seeds of the three species was that seeds of the narrow endemic germinated to higher percentages in darkness than those of its two geographically-widespread congeners. Survivorship curves for 1991 and 1992 seeds of S. altissima and S. nemoralis and for 1992 seeds of S. shortii were of Deevey Type I; the survivorship curve for 1991 seeds of S. shortii was closest to Type II. Longevity of 1991 seeds of S. altissima, 1992 seeds of S. nemoralis and 1991 and 1992 seeds of S. shortii was <4.0 years, whereas that of 1991 seeds of S. nemoralis was <2.3 years; 5% of 1992 seeds of S. altissima were viable after 4.0 years.

Water contents, desiccation tolerance, respiratory rates and subcellular characteristics of three contrasting seed types were studied during development. Avicennia marina (a tropical wetland species) and Aesculus hippocastanum (a temperate species) produce recalcitrant seeds and Phaseolus vulgaris produces orthodox seeds. During development, A. hippocastanum and P. vulgaris seeds showed a decline in water content and respiration rate with a concomitant increase in desiccation tolerance. These parameters did not change during the development of A. marina seeds once they had become germinable. There was a decrease in the degree of vacuolation and an increase in the deposition of insoluble reserves in A. hippocastanum and P. vulgaris seeds, while A. marina seeds remained highly vacuolated and did not accumulate insoluble reserves. Mitochondria and endomembranes degenerated during the development of A. hippocastanum and P. vulgaris seeds, but remained unchanged in A. marina seeds. The data are consistent with the hypothesis that extensive vacuolation and high metabolic rates contribute to desiccation sensitivity. However, the development of recalcitrant A. hippocastanum seeds is similar to that of orthodox P. vulgaris seeds. These data are in accord with the concept of seed recalcitrance being a consequence of truncated development. The results suggest that there may be three categories of seeds: orthodox seeds which develop desiccation tolerance, seeds which show similar development to orthodox seeds, but are shed before desiccation tolerance is well developed, and seeds which show no developmental trends giving rise to increased tolerance.

Protein synthesis in embryonic axes of Norway maple (Acer platanoides) was studied in seeds stratified at 3 or 15°C. At 3°C stratification, germination of seeds started between 6 and 7 weeks; at 15°C stratification germination did not occur. During imbibition of embryonic axes undergoing cold and warm stratification many new proteins became visible. Two of these proteins may be connected with the release from dormancy at 3°C. In seeds stratified at 15°C, these proteins occur in low amounts or are completely absent.

Seeds of wild rice (Zizania palustris var. interior) have been reported to show maximum survival when dehydrated at 25°C. It has also been reported that axis cells sustain least damage at this drying temperature. In the present study, a linear relationship between drying rate and dehydration temperature was established. Whereas the highest positive tetrazolium staining and lowest leakage were recorded for seeds dehydrated at 25°C, maximum germination was recorded for seeds dried at 20°C. A proportion of seeds showed glass formation irrespective of the dehydration temperatures used. Parameters of the glass to liquid transition, however, correlated with neither water content nor sugar profiles. The ratio of raffinose to sucrose was similar in all the treatments. A hydroperoxide test revealed a linear relationship between peroxide levels and drying temperature, although fatty acid amounts were not correlated with hydroperoxide amounts. Butanal amounts and total aldehydes evolved, on the other hand, showed a high negative correlation with peroxide amounts. Electron microscopy showed that the variability and relative abundance of peripheral membrane complexes (PMCs) was highest for embryonic axes dehydrated at 25°C and lowest for cells of embryonic axes of seeds dried at 10°C. Furthermore, intramembrane particles (IMPs) were evenly distributed in cells of axes dried at 25 or 37°C. In contrast, membranes of cells of axes dehydrated at 10°C showed large IMP-free areas. The relative abundance of IMPs was highest in cells of embryonic axes dried at 25°C and lowest in cells of axes dehydrated at 10°C. From these observations, it is suggested that membrane phase transitions, with the concomitant elimination of proteins, accompany dehydration of Z. palustris seeds at 10°C, whereas at 37°C peroxidation may predominate.

Osmotic priming in a polyethylene glycol solution (300 g/kg water) for 48 h resulted in a partial loss of desiccation tolerance for seeds of Vigna radiata (L.) Wilczek (mung bean). The percentage of germination began to decrease after primed seeds were dried down to water contents less than 0.06 g/g DW. As compared with control seeds, primed mung bean seeds also had poorer storage stability. The decline of storage stability after osmotic priming was correlated with the modifications of seed water sorption properties. Priming significantly increased the amount of water associated with the weak water-binding sites, and reduced the amount of water associated with the strong binding sites and multi-molecular binding sites in seed tissues. The enhancement of molecular mobility in seeds, as a result of such water redistribution, probably accelerates seed deterioration and decreases storage stability.