Expansins are a multi-gene family of proteins involved in changes in cell wall properties. In seeds where the embryo is completely surrounded by the endosperm, dormancy breakage requires weakening of the micropylar endosperm, increased embryo growth potential or both. Cell wall alterations are fundamental components of these processes, and expansins are thought to participate in them. Here, we explore the possible involvement of expansins in the control of germination by phytochrome in Datura ferox L. seeds. Based on the conserved sequences of known expansin genes, corresponding primers were designed to investigate the expression of expansin mRNAs by reverse transcriptase polymerase chain reaction (RT-PCR). One expansin mRNA was detected in micropylar endosperm of D. ferox, while two mRNAs were present in the embryo prior to radicle emergence. Expansin transcript content is promoted by red (R) light, both in the micropylar endosperm and the embryo; these effects of R are far-red light (FR) reversible, displaying a typical low-fluence response (LFR) in a way fully consistent with the photocontrol of germination. On the other hand, when the promotion of germination by the LFR is antagonized by exposing the seeds to continuous FR through a high-irradiance response (HIR) or by ABA, the inhibitory treatments do not affect expansin mRNA abundance. The results support the participation of expansins in the promotion of germination by LFR of phytochromes, and suggest that inhibition of germination by HIR or ABA does not include a reduction in the transcription of expansin genes.

Seed storage proteins were analysed in 27 varieties of alfalfa (Medicago sativaL.); these included five subspecies (glomerata, caerulea, falcata, hemicycla, praefalcata), seven of the nine sources of Medicago germplasm introduced into North America and a sample of additional cultivars. The protein patterns were remarkably consistent for all of these taxa. One-dimensional and two-dimensional gel electrophoresis revealed only minor differences in polypeptide composition within each of the three major classes of storage protein (7S globulin, 11S globulin, 2S albumin). The slight variations that were found provided no information on either parentage or evolutionary relationships amongst these particular taxa. Nonetheless, persistent and reproducible heterogeneity of some minor polypeptides of 11S globulin (medicagin) may be useful under other circumstances for cultivar identification in alfalfa. Both subfamilies (I and II) of the 11S globulin were strongly expressed in all of the cultivars and subspecies examined. It was concluded that this structural divergence within the 11S storage protein family predated the evolution of the M. sativa L. species complex. Most of the variability in storage proteins was quantitative. However, even this variability was reduced when data were standardized with respect to seed dry weights. The consistent similarities in qualitative and quantitative expression of seed storage proteins amongst all of these taxa suggest a high degree of uniformity in both seed physiology and genetics within the alfalfa species complex.

The forest seed bank has been demonstrated to vary spatially at scales from 2 to 10 m. To our knowledge, the fine-scale spatial structure, i.e. < 2 m, has not been studied before. This study aims to make a thorough investigation of fine-scale spatial structure. Soil samples (128) were collected from each of five 2.1 m × 2.1 m plots, using a combined systematic (64) and random design (64). This allowed investigation of the fine-scale spatial structure of individual species–plot combinations using indicator-variograms. Our results indicated that over half of all species recorded in a particular plot were spatially structured. Remarkably, the presence of spatial structure seemed independent of species frequency. Visualization of the spatial structure showed an irregular spatial pattern, i.e. seed clusters that were randomly distributed in space. Spatial dependence occurred over small distances, possibly suggesting that a significant proportion of seeds was deposited near the mother plant. We conclude by presenting the relevance and implications of small-scale spatial seed-bank patterning for seed-bank sampling.

On the basis of previous research, we predict that Mediterranean grasslands should show larger-seeded annuals in: (1) more-arid grasslands; (2) more-fertile soils; (3) less-grazed grasslands; and (4) grasslands with lower intensities of seed predation by ants. To test these predictions, we set 29 sampling units of 50 m × 50 m in a 1000 km2 grassland area in Central Spain, and characterized them according to the former factors. We then recorded annual vegetation using ten quadrats of 20 cm × 20 cm in each sampling unit. Seed size at the community level was described using six variables: (1) mean seed mass; (2) standard deviation of seed mass; (3) weighted mean seed mass (by species frequencies); (4) proportion of small-seeded annuals; (5) proportion of medium-seeded annuals; and (6) proportion of large-seeded annuals. Most climate variables (mean annual temperature, length of the summer drought, water balance and mean annual precipitation) correlated with seed-size descriptors, showing that large-seeded annuals increase in warmer and more-arid communities. Mean seed size was modelled as a function of mean annual temperature and grazing pressure. According to this model, warmer and less-grazed communities tend to show a smaller mean seed size. These results confirm the importance of seed-size descriptors at the community level in Mediterranean grasslands, and the role of climate and grazing as major drivers in these communities. Conversely, hypotheses about soil fertility and seed predation by ants were not supported by our results.

The physical and chemical seed characteristics of 11 dipterocarp species (four Dipterocarpus, two Dryobalanops and five Shorea species) were quantified to examine variations among species. We measured dry weight, pericarp thickness and concentrations of nitrogen, total phenol, condensed tannin, lignin, lipid, starch and total sugar. Although there was low intraspecific variation among parent trees, each species had unique characteristics. We found significant correlations between chemical defensive compounds (i.e. lignin and total phenol) and nitrogen concentration. However, these relationships were no longer significant when an outlier was removed. Fruiting frequency was not significantly correlated with any chemical compound. Seeds of Shorea species had a higher concentration of lipid than seeds of Dipterocarpus. The implications of variation in the physical and chemical characteristics of dipterocarp seeds are discussed.

Pretreatment of moist seeds at certain temperatures can reduce seed dormancy, but such observations represent the net effect of pretreatment. Seeds of Sitka spruce (Picea sitchensis [Bong.] Carr.) were raised to 30% moisture content and pretreated at five different temperatures between 5 and 30°C for up to 24 weeks (168 d). Subsequent ability to germinate at 10°C and viability were then determined in order to investigate the effects of pretreatment on seed dormancy and survival. There was a curvilinear, negative semi-logarithmic relationship between seed longevity and pretreatment temperature, such that Q10 for loss in viability increased from 2.6 between 10 and 20°C to 2.8 between 20 and 30°C. Simple multiplicative models combining cumulative normal frequency distributions for each of loss in viability and loss in dormancy were able to describe the changes in ability to germinate at 10°C, after pretreatment at 5, 10 and 30°C. However, in order to quantify the changes in ability to germinate observed at 10°C after pretreatment at 15°C, it was necessary also to invoke a model of dormancy reimposition, while for the results at 20°C it was necessary to postulate both dormancy reimposition and the further loss of this reimposed dormancy. It is concluded that moist seeds of Sitka spruce held at 15 and 20°C cycle between the dormant and non-dormant condition.

There are a number of studies describing the gross range of morpho-anatomical variability in epiphytic Tillandsia species, but the interspecific variation in seed traits remain largely unexplored, although these play an important role in determining dispersal and establishment success. In order to evaluate interspecific variation in seed morphology, anatomy and germination, we sampled six Tillandsia species from the Yucatan peninsula, Mexico, distributed along a precipitation gradient. We studied morpho-anatomical traits (seed length, seed mass, ratio of coma to seed, ratio of embryo to endosperm), seed terminal velocity in still air, and performed histochemical analyses and germination trials under controlled conditions. Tillandsia recurvata differs from the other five species in the structure of the plumose coma; it was the only species lacking an endosperm and showed distinct seedling development. Among the species, bigger seeds were related to longer comas, and had higher germinability. Overall, seed terminal velocity was invariably slow, compared with reports of other anemochorous species, suggesting a high dispersal potential. Taxonomical and ecological implications of our results are discussed.

Somatic embryo quality is an important aspect of artificial or synthetic seed production and is dependent on proper morphological development and deposition of sufficient storage reserves to support early growth of the seedling. The abilities of a somatic embryo to survive desiccation and to establish a rapidly growing seedling were used as estimates of quality and were dependent on a number of factors during the elongation and maturation phases of somatic embryo development. Embryo quality was significantly enhanced when the sucrose content of the elongation and maturation media was increased to 50 g I−1 from 30 g I−1. This increased dry weight to >2 mg embryo−1 suggesting greater deposition of storage reserves. Higher sucrose concentrations were inhibitory and reduced survival after desiccation and seedling vigour. An optimum number of embryos can be supported on one Petri plate; the number was controlled by the plating density of the embryogenic cell clusters spread on the elongation medium. This optimum was dependent to some extent on the sucrose content of the medium, indicating that sucrose had a nutritive effect on embryo development and not solely an osmotic effect. Light intensity was another critical factor. Intensities > 75 μmol m−2 s−1 photosynthetic photon flux density (PPFD) at the elongation stage of development blocked globular-stage somatic embryo development. Maturing somatic embryos were far less sensitive to high light intensities and development was more rapid at 150 μmol m−2 s−1 PPFD. Light quality was manipulated by covering plates with filters that selectively transmitted certain regions of the visible spectrum. All filter treatments reduced embryo yield and quality compared with embryos developing under fullspectrum light; varying the light spectrum does not seem to be a viable method of enhancing somatic embryo quality.

Phytochrome-induced softening of the micropylar endosperm of Datura ferox seeds is followed by radicle protrusion and preceded by a significant increase in the extractable activity of β-mannanase and β-mannosidase (6 and 3 times over the far-red light controls, respectively) and a decrease in the mannan content of the micropylar endosperm cell walls. This relationship between phytochrome-induced germination, endosperm softening and mannan degradation was studied in Datura ferox seeds subjected to a range of water potentials during the presence of Pfr. Low water potential inhibited the decrease in mannose-rich cell-wall insoluble polysaccharides, the increase in activity of β-mannosidase and endosperm softening. A good correlation was found between activity of β-mannosidase measured at 45 h after exposure to a red light pulse and germination counted 24 h later when, with different external water potentials, germination varied between 0 and 90%. In contrast, no inhibitory effect of low water potential on in vitro β-mannanase activity was detected. We suggest that the in vivo action of β-mannanase might be limited by accumulation of manno-oligosaccharides caused by inhibition of β-mannosidase. These data are consistent with the concept of a link between mannan mobilization and endosperm softening. Part of the effect of low water potential on Pfr-induced germination may be mediated by inhibition of β-mannosidase activity, with consequences for mannan hydrolysis and endosperm softening.

Reversible phosphorylation is catalysed by protein kinases that transfer the γ-phosphate from ATP to amino acid residues of proteins. The process can be reversed by protein phosphatases. Phosphorylation can dramatically activate or inhibit enzymes and affect protein-protein interaction. Through phosphorylation protein kinases can amplify and propagate cellular signals. In plants and now seeds, protein kinases involved in hormone, defence and environmental stress responses are being identified. Increasingly, these protein kinases are being cloned and characterized, demonstrating the major role of reversible protein phosphorylation in seeds.

Seed dispersal effectiveness (SDE) is a useful framework to explore the evolutionary and ecological consequences of seed dispersal to plant fitness. However, SDE is poorly studied in tropical open grasslands. Here, we studied both quantitative and qualitative components of SDE in two species of Miconia (Melastomataceae) from ferruginous campo rupestre, a vegetation highly threatened by mining activities. We determined fruit traits and fruit availability and found that fruits of both species are produced in times of resource scarcity at the study site. Based on the number of visits and the number of fruits removed per visit, we calculated the quantitative component of SDE for both species. Finally, we explored the qualitative component of SDE by means of a controlled experiment that simulated the effects of gut passage on seed germination. Bird species differed strongly in the quantitative component of SDE. Gut passage did not affect germination compared with hand-extracted seeds, except for a minor negative effect on germination time in M. pepericarpa. However, seeds within intact fruits showed lower germination percentages compared with hand-extracted seeds. Our data indicate that Miconia species from ferruginous campo rupestre are visited by a diverse assemblage of generalist birds that differ in quantitative, but not qualitative, seed dispersal effectiveness. We argue that planting Miconia species can overcome seed limitation in degraded areas and thus assist ecological restoration after mining abandonment.

Cumulative oxidative damage from the unavoidable formation of reactive oxygen species (ROS) contributes to seed ageing. Low-molecular-weight (LMW) antioxidants, such as water-soluble glutathione (GSH) and lipid-soluble tocochromanols, can prevent ROS from causing damage, especially when antioxidant enzymes are inactive due to desiccation. However, loss of tocochromanols does not always accompany seed ageing, such as during accelerated ageing or controlled deterioration, despite the presence of oxygen and prevalent loss of GSH. To assess relationships between total germination (TG) and antioxidant changes under storage conditions with practical relevance, commercial seeds of Cucumis sativus, Daucus carota, Helianthus cucumerifolius, Latuca sativa, Lepidium sativum, Phaseolus vulgaris and Raphinus sativus of the same cultivar were obtained over multiple years and stored under ambient conditions (21.9 ± 2.1°C; 36.8 ± 6.6% relative humidity). Sigmoidal fitting of TG revealed time to when 50% of seeds had lost viability, which ranged from <5 years (D. carota) to >15 years (C. sativus). Cellular redox states were quantified via the half-cell reduction potential of LMW thiol/disulphide couples. These negatively correlated with TG (i.e. cell redox states were more oxidized in lots with lower TG), with an average R2 value of 0.62 for the most abundant thiol (GSH, or γ-glutamyl-cysteine in P. vulgaris). Concentrations of tocochromanols positively correlated with TG, with an average R2 value of 0.50 for the most abundant tocochromanol (γ or α in L. sativa and H. cucumerifolius). Therefore, during viability loss under ambient ageing conditions leading to the cytoplasm having a glassy state, the lipid domain in all species experienced oxidative damage.

Recalcitrant seeds of Trichilia dregeana were stored at 16 or 25°C, either at the water content at which they were shed or partially dried. Although having been exposed to a short period (approx. 6 h) at temperatures up to 30°C prior to storage, seeds at the original water content maintained viability for several weeks at 16°C. However, storage of undried seeds at 25°C was deleterious within 8 d, indicating a chemical basis for degeneration of hydrated recalcitrant seeds. Seeds that had been mildly dehydrated to the relatively high axis level of 1.68 g H2O g—1 dry mass, while maintaining full germinability immediately after drying, exhibited only 4% viability after 8 d in storage at 16°C and had completely lost viability after the same storage period at 25°C. Ultrastructural features characterizing hydrated seeds included indications of enhanced activity associated with initial exposure to the elevated temperature as well as some signs of stress. However, over an effective 15 d storage period at 16°C, ultrastructural features showed the cells to have retained little damage and to have been in an enhanced state of activity commensurate with ongoing development towards germination. After a longer storage period, however, signs of damage, including indirect evidence for disarray of the cytoskeleton in some axis cells, became apparent in line with the declining seed viability. Immediately following dehydration from an average axis water content of 1.97 to 1.68 H2O g g—1(the sub-imbibed condition), some ultrastructural abnormalities were apparent, but the seeds remained 100% germinable. However, within the 8 d storage period in this sub-imbibed condition, a spectrum of severe ultrastructural degeneration, including indirect evidence of the collapse of the nucleoskeleton and extensive cell lysis, accompanied viability decline of the seeds to 4%.

The understanding that many aspects of the spatial and temporal patterns of epiphyte communities may be explained by the comportment of early life stages has given rise to a considerable number of germination studies in recent years. Unfortunately, protocols frequently use unproven assumptions and arbitrary experimental conditions. To make future studies as ecologically meaningful as possible we address a number of potential pitfalls with a series of experiments with seeds from a total of 16 species. We show that it is safe to collect capsules for experiments before natural dehiscence – there is afterripening even in the case of very early collections. The application of fluctuating temperatures is not imperative, because there is no consistent difference in the germination response under constant versus fluctuating temperatures. The effects of different osmotic potentials and intermittent drought of varying intensity on germination are qualitatively, but not quantitatively, comparable. Due to the greater ecological realism, we encourage the use of the latter. However, care must be taken to use realistic temperatures – the impact of intermittent drought on germination is modulated by temperature. This highlights the need for data on the in situ temperature regimes during germination as an important prerequisite towards more realistic experiments in the field of germination ecology of vascular epiphytes.

In 2009, the bicentenary of Charles Darwin's birth on 12 February 1809 is being celebrated. For seed scientists, celebrations of the contributions of the great biologist should also mark his involvement with seeds. Darwin was interested in seeds, particularly in their role in dispersal and distribution of plant species over long distances. His studies of seeds, laid down in several books and articles, contributed to the development of his ideas on evolution and the distribution of living organisms on the planet. In this review, the place of seeds in Darwin's work is surveyed and it is shown how he referred to them to support and illustrate some of his most important ideas.

Fleshy fruits fall on to the ground together with cleaned seeds previously ingested by primary dispersers, offering a wide range of fruits and seeds to the ground foragers. Although nutritional properties strongly differ between fruits and seeds, this different seed presentation (cleaned seeds versus seeds within the pulp) has not been addressed in seed removal studies. This study reports on the removal of fruits versus their seeds in five fleshy-fruited species in a temperate forest. We found that rodents removed most of the seeds and partially consumed most of the fruits, preferring seeds to fruits. Rodents bit the fruits to extract the seeds, leaving most of the pulp. We found a preference ranking for the seeds (Sorbus aucuparia>Ilex aquifolium>Sorbus aria>Rosa canina>Crataegus monogyna) but no preferences were found for the fruits, probably due to their similarities in pulp constituents. Seed and fruit choice were affected by chemical and physical properties and not by their size. The presence of alternative and preferred seeds (nuts) delayed the encounter of the fruits and seeds and diminished their removal rates. We found that higher rodent abundance is not necessarily associated with higher removal rates of fleshy fruits. Rodent abundance, fruit size and seed size are minor factors in the removal of fleshy fruits and their seeds. This study underlines that scatter-hoarding rodents are important removers of fleshy fruits and their seeds, producing a differential seed removal depending on the seed presentation (with or without pulp), the nutritional properties of the seeds (but not of the fruits) and the presence of alternative food.

Interchangeable effects of temperature, moisture content and oxygen on seed longevity have been mostly examined to estimate seed viability during long-term dry storage, whereas few experiments have studied seed viability under near-natural conditions to evaluate seed persistence in the soil. To this end, we artificially aged seeds of Ranunculus baudotii, a hydrophyte widely distributed in temporary ponds constituting an abundant soil seed bank. Seeds were exposed to controlled ageing at three different relative humidities (RH) under both aerobic and anoxic conditions. Their viability, water content, membrane damage, oxidative stress and anti-oxidant enzymatic defence activity were evaluated. Seed survival was longer at higher relative humidity (97% RH), and lowest at a relative humidity (90% RH) simulating moist but not waterlogged soils. Anoxic conditions showed a protective role on viability at lower moisture contents (70% RH). Seed viability was negatively associated with hydrogen peroxide content and correlated with anti-oxidant enzyme activities, but not with membrane damage. Altogether, these results suggest negative roles for moist soils and anoxia in determining seed persistence in the field, but at higher moisture contents the negative effects of anaerobia diminished. The anti-oxidant system activation, even under unfavourable conditions, might recover seeds once all protective processes can operate, pointing out the plasticity of mechanisms involved in seed loss viability.

The understanding of the genetic basis of grain dormancy in wheat has rapidly improved in the last few years, and a number of genes have been identified related to that trait. We recently identified the wheat genes TaPM19-A1 and -A2 and we have now taken the first step towards understanding the role of this class of genes in seeds. By investigating the Arabidopsis homologous PM19-Like 1 (PM19L1) we have found that it has a seed-specific expression pattern and, while its expression is higher in dormant than in non-dormant seeds, knock-out mutations produced seeds with increased dormancy. Not only primary dormancy, but also secondary dormancy in response to high temperature was increased by the loss-of-function. We have also examined the function of PM19L1 by localizing the PM19 protein primarily to the cotyledon cells in seeds, possibly in membranes. By investigating the co-expression network of this gene we have found that it is connected to a small group of abscisic acid (ABA)-induced seed maturation and storage-related genes. The function of PM19L1 represents a good opportunity to explore the interactions of key factors that can influence seed dormancy such as ABA, temperature and membrane properties.

To improve understanding of how a rare endemic species of Centaurium adapts to a specialized ecological niche, we studied the germination ecology of the mountain spring specialist, C. somedanum, a perennial species restricted to an unusual habitat for this genus. We conducted laboratory experiments with fresh seeds collected from two populations for three consecutive years, to investigate: (1) the effect of temperature and light on germination; (2) the existence of seed dormancy; and (3) inter-population and inter-annual variation in germinability. Germination occurred only in the light and at relatively low temperatures (15–22°C) with no differences between constant and alternating regimes, and a significant decrease at high temperatures (25°C and 30°C). We found non-deep simple morphophysiological dormancy and variation in seed germinability depending on the year of seed collection. C. somedanum diverged from the common germination characteristics of the genus in: (1) its germination at lower temperatures, which contrasts with what is generally expected in wetland species but could be adaptive in the spring habitat; and (2) its morphophysiological dormancy, which we report here for the first time in the genus and which could be an adaptation to its mountain habitat.

Time of seedling emergence is an important step in the life cycle of annual plants because it may determine subsequent performance and success. Timing of emergence is especially critical to plant performance in habitats like arable fields which are subject to frequent disturbances. Within-season variation in timing of germination in the range of only a few days is typical for many arable weeds. However, since it is unclear whether such small deviations in germination date translate into fitness differences in the course of the life cycle, the aim of this paper was to quantify the effects of short germination delays on plant performance. We conducted two generalized randomized block experiments in an unheated greenhouse to study the impact of delayed germination (1, 2, 3 and 7 d) with and without competition, respectively, on the fitness of the arable weed species Agrostemma githago (L.). We expected that delayed germination significantly reduces fitness in terms of several life-history traits, and that the decrease of fitness is higher in the presence of competition. Under realistic conditions with competition through barley, Agrostemma plants with delayed germination of 7 d produced 54% fewer shoots, 57% less biomass, 52% fewer flowers, 36% lighter seeds and were 23% shorter as compared to control plants without delayed germination. Without additional stress through competition with barley this pattern was less pronounced. Thus, in the situation of interspecific competition, early emerging seedlings have biologically significant fitness advantages over later emerging seedlings of the same species.