The relationship between endosperm cap weakening and endo-β-mannanase activity during priming and the time to germination after priming was studied in tomato (Lycopersicon esculentum) seeds. During priming of seeds in −0.4 MPa PEG, the mechanical restraint of the endosperm cap decreased while the endo-β-mannanase activity in the endosperm cap increased. There was no decrease in required puncture force and no increase in endo-β-mannanase activity in seeds during priming in −1.0 MPa PEG. Two classes of seeds could be distinguished during priming in −0.7 MPa PEG: one with decreased required puncture force and one without. A strong correlation was found between the lowering of the mechanical restraint and endo-β-mannanase activity. It was concluded that individual seeds have to cross a threshold water potential in order to develop enzyme activity and lower their mechanical restraint. A decrease in required puncture force and increase in endo-β-mannanase activity correlated with ice crystal-induced porosity in the endosperm cap cell walls in scanning micrographs. It was presumed that ice crystal-induced porosity reflects cell-wall hydrolysis. Germination time after priming correlated positively with required puncture force during priming, depending on the osmotic potential. Seeds in −1.0 MPa PEG improved their time to germination, without a decrease in the required puncture force. Therefore it was concluded that lowering of the endosperm restraint during priming positively affects the time to germination of primed seeds but is not a prerequisite for rapid germination.

Morphological changes in tomato (Lycopersicon esculentum Mill. cv. Moneymaker) seed during osmopriming and imbibition (‘hydropriming’) were followed using X-ray photographs. Embryo, endosperm and free space areas were measured. Both osmopriming and hydropriming resulted in free space development (+ 8.1% and + 10.8% of the whole seed planar area, respectively), almost all at the cost of the endosperm area. Planar dimensions of whole seeds were relatively constant and the dimension perpendicular to the planar surface, the thickness, could account for the volume increase of primed seeds reported in the literature. In dead seeds, only a small amount of free space developed while the planar area of the seed remained the same. In the imbibing viable seeds no deterioration of endosperm could be detected until the moment of root protrusion.

Seeds which were osmoprimed directly after harvest, i.e. in the fresh state, did not showthe induction of any free space, while free space developed normally after dehydration and a second priming treatment. Apparently, a dehydration step prior to the priming treatments is required for the development of free space in osmo- or hydroprimed seed.

X-ray photographs of hydroprimed seeds showed that the radicle tip adhered tightly to theendosperm cap. This results in various forms of damage to the root tips upon redrying. Priming did not introduce cotyledon abnormalities.

A high level of genetic purity in crop varieties must be achieved and maintained for agronomic performance as well as to encourage investment and innovation in plant breeding and to ensure that the improvements in productivity and quality imparted by breeders are delivered to the farmer and, ultimately, to the consumer. Traditionally, morphological comparisons have formed the basis for genetic purity evaluations. However, replicated field observations are time-consuming, expensive and unreliable. Morphology cannot provide information on the purity of specific genetic attributes that relate to grain quality or to pest or herbicide resistance bred into varieties. Biochemical assays, including isozymes, can distinguish varieties within several species. Isozymes have been routinely used in checking seed-lot purity in maize (Zea mays L.) for the past 20 years. Newer DNA-based technologies such as restriction fragment length polymorphisms and more recently developed methods that use the polymerase chain reaction can allow even more discriminative and faster identification of varieties. However, none of the DNA methods have replaced biochemical methods for seed purity assays, other than in a relatively select group of crops with very high seed value, due to their high datapoint cost. It will require further miniaturization, automation and enhanced capabilities to process numerous samples simultaneously before newly developed methods supplant biochemical methods for routine usage in purity testing. New varieties that have major genes for herbicide or insect resistance incorporated within them require purity assays during product development and following seed production of the commercial variety. Immunological or DNA sequence assays can be developed and automated systems are required to process hundreds of thousands of individuals. Ultra-high, micro-array technologies and single-molecule detection systems are now under development. These technologies offer the promise that adequate distinction and high sample throughput will be combined. New methods may eclipse the capabilities of biochemical methodologies, thereby potentially raising genetic purity standards and enabling farmers and consumers better to utilize and benefit from increasingly productive varieties that are bred from a more diverse base of genetic resources.

This study addressed the difficulty of germinating drupelets (hereafter seeds) in the multi-seeded stony dispersal units (drupes) of Leptecophylla tameiameiae (Ericaceae). Embryos in fresh seeds were 77% the length of the endosperm, and seeds inside the intact drupes imbibed water. We monitored germination at 15/6, 20/10 and 25/15°C for 162 weeks, after which each drupe was cut open and ungerminated seeds counted. Drupes contained 1–6 seeds, and the total number of seeds in all treatments and controls was 1977, with 20, 29, 25, 18, 7 and <1% of them occurring in one-, two-, three-, four-, five- and six-seeded drupes, respectively. The percentage of seeds germinating in one-, two-, three-, four-, five- and six-seeded drupes was 74, 66, 65, 72, 56 and 0, respectively. Neither warm nor cold stratification for 6 or 12 weeks significantly increased germination percentages, compared to controls incubated continuously at 25/15°C for 162 weeks, where 72% of the seeds in the drupes germinated. At 25/15°C, 24–49 weeks were required for 20% of the seeds to germinate. Warm followed by cold stratification did not promote germination, and there was no widening of the temperature range for germination. Like seeds of other species known to have deep physiological dormancy (PD), those of L. tameiameiae required extended periods of time (16 to ≥162 weeks) to come out of dormancy and germinate, gibberellic acid (GA3) did not promote germination and excised embryos failed to grow. Thus, we conclude that seeds of L. tameiameiae have deep PD. However, unlike seeds of other species with deep PD, those of L. tameiameiae required an extensive period of warm rather than of cold stratification to come out of dormancy. It is suggested that a subtype a (seeds require a long period of cold stratification to come out of dormancy) and a subtype b (seeds require a long period of exposure to warm stratification to come out of dormancy) of deep PD be recognized in the Nikolaeva formula system for classifying seed dormancy.

Endo-β-mannanase (EC 3.2.1.78), an enzyme that mobilizes the endosperm cell walls of the lettuce (Lactuca sativa L.) seed, increases in activity in the micropylar and lateral regions of this tissue following the completion of germination. Its complementary DNA (cDNA) sequence (LsMan1) was determined using the polymerase chain reaction (PCR) with degenerate primers. The 3‘-end of the cDNA sequence was obtained by 3‘-end rapid amplification of cDNA ends (RACE), and the 5‘-end sequence by genome walking and 5‘-end RACE. The predicted amino acid sequence from the cDNA has a high identity with endo-β-mannanases present in other species (e.g. 67% identity with coffee β-1,4-mannan endohydrolase, 62% identity with tomato fruit endo-β-mannanase). Southern blot analysis suggests the presence of several members of an endo-β-mannanase gene family in the lettuce genome. Several isoforms of the enzyme, including three major ones, were detected by isoelectric focusing. Based on Northern blot analysis, accumulation of the endo-β-mannanase mRNA occurred only after lettuce seeds had germinated, and increased thereafter, although enzyme activity persisted after transcription declined.

Germinating seeds of many species, especially those of tropical or subtropical origin, suffer chilling injury when exposed to low but non-freezing temperatures, resulting in poor seedling establishment and reduction in yield. The severity of injury depends upon several factors such as (i) the species or the cultivars involved, (ii) the initial water content of the seed, (iii) the temperature to which seed is exposed, (iv) the duration of chilling exposure and (v) the period during the course of germination when the chilling exposure takes place. The period of imbibition by seeds is the period of sensitivity to the stresses but especially to chilling temperatures. Imbibitional chilling injury is defined as sensitivity to a combination of low seed-water content and imbibition at cold temperature. The mechanism of chilling injury in seeds is different from chilling injury of hydrated tissues and is discussed in relation to the involvement of membranes, respiration, rate of hydration and the concept of discreet moisture levels. Furthermore, the mechanistic and applied aspects of pre-sowing seed treatments for improved germination performance under chilling stress are discussed with an emphasis on hydration-dehydration, osmotic priming and growth regulator treatments.

It is well documented that the mother plant has much more influence than the father on seed dormancy/germination, especially of the F1 offspring, primarily by providing all material (maternally derived tissue) to the diaspore coat(s); by maternal environmental effects and provisioning of nutrient resources, mRNA transcripts, protein, the hormone abscisic acid and nitrate to the seed during its development; and by determining progeny environment via dispersal and phenology. There is some evidence that the paternal influence on seed dormancy/germination of the offspring (seeds) can be mediated through multiple paternity (including mate number and diversity), non-nuclear (cytoplasmic) and nuclear (genotypic) inheritance and paternal environmental effects. Our primary aim was to determine via a literature review the influence (or not) of the paternal parent on seed germination. Altogether, 37 of 59 studies (62.7%) indicated a positive influence of the father on seed germination, although not all of them were statistically significant. In general, however, results of studies reported in the literature do not offer strong support for the paternal parent having a major role in seed germination (or seed size) of his F1 offspring.

We have been studying a seed aspartic proteinase, termed AtAP, from Arabidopsis thaliana. In previous work, we purified the proteinase, analysed its activity and isolated the cDNA sequence. In this paper, the expression of the mRNA for the aspartic proteinase was analysed in seed tissues both by Northern blots for overall regulation and by in situ hybridization to follow cell-specific localization of message. We found a 1.9 kb aspartic proteinase message in dry seeds and seed pods. This message was expressed in many different cell types of the mature dry seed. The localization of the protein within these cells was also determined. Antibodies were raised against the AtAP and purified using affinity chromatography on an AtAP–immobilized-pepstatin A–agarose column. This purified antibody recognized several AtAP peptides in seeds. To localize the enzyme in cells, we isolated protein bodies from the dry seeds of Arabidopsis using a non-aqueous isolation method. The AtAP activity and antigenic peptides were found to be highest in the protein body fraction and co-localized with the seed storage protein 2S albumin and the vacuolar marker enzyme α-mannosidase. This protein body localization of the AtAP was confirmed with immunocytochemical localization by electron microscopy and shows that the protein is not secreted by these cells.

Seeds of Cenchrus ciliaris L. were produced under different hydro–photo–thermal environments with and without fertilizer. Dormancy loss of spikelets and extracted caryopses was tested during dry after-ripening at 40°C and 43% equilibrium relative humidity. Caryopses had higher initial germination and lost their dormancy faster than spikelets. Dormancy of both caryopses and spikelets generally decreased with an increase of maturation temperature and fertility, whereas dormancy increased if water stress was imposed during maturation. The latter effect was smaller when the mother plants were exposed to water stress after caryopses were fully formed than when water stress cycles were applied throughout maturation. Daylength extension (to 14 or 18 h d-1) by artificial light increased dormancy of both caryopses and spikelets. The effect of long days declined when plants were exposed to natural daylight for more than 10 h d-1. The after-ripening curves were consistent with the hypothesis that dormancy periods of individual seeds are normally distributed within each seed lot. Rates of loss of dormancy were quantified by the slopes of these curves. In a given experiment, these rates were identical for caryopses but not always for spikelets that matured in diverse environments. Even for caryopses, however, the slopes varied between experiments. Therefore, the results do not support the hypothesis that a dormancy model can be applied universally to all seed lots of Cenchrus ciliaris. Methods of predicting the period of after-ripening required to achieve desired levels of dormancy for reseeding degraded rangelands are discussed.

Many studies have claimed that fire acts as the chief ecological factor cueing dormancy break in seeds with a water-impermeable seed coat, i.e. physical dormancy (PY), in Mediterranean ecosystems. However, a proposal is made that seasonal temperature changes must be viewed as more meaningful dormancy-breaking cues because: (1) fire is erratic and may break PY in seasons during which seedlings cannot complete their life cycle; (2) fire may not occur for long periods, thereby only providing an opportunity for dormancy break and germination once in every several years; and (3) if fire opens the specialized anatomical structures called ‘water gaps’, in seconds, their evolutionary role of detecting environmental conditions becomes irrational. Although fire breaks dormancy in a proportion of seeds, given the risk of seed mortality and the post-fire environment providing cues for dormancy break, it is suggested that fire might possibly be an exaptation.

The objective of this study was to fit a hydrothermal germination model to germination data for a seedlot of radiata pine (Pinus radiata D. Don). Seeds were incubated for 50 d at constant temperatures and water potentials (T = 12.5–32.5°C, Ψ = 0 to − 1.2 MPa). Most seeds completed germination within 50 d, but for low Ψ and/or non-optimal temperatures (T < 17.5°C, T>25°C) many seeds did not complete germination. In general, germination data conformed to the hydrothermal model. Departures from the model were encountered for slow-germinating seeds at suboptimal temperatures (T ≤ 20°C). To account for these departures, two alternative hydrothermal models were fitted with an additional term for an upwards shift in seed base water potential with increasing time to germination. The alternative models more correctly predicted germination time than the original model. Similarly, reduced percentage germination at supra-optimal temperatures (T>20°C) was explained by including a term in the hydrothermal model which shifted the base water potential of seeds upwards towards zero, which in turn reduced the predicted rate that hydrothermal time would be accumulated by seeds. The rate of this upwards shift in base water potential was dependent on time to complete germination and ambient water potential as well as supra-optimal temperature.

The most common way to test seed quality is to use a simple and reliable but time- and space-consuming germination test. In this paper we present a fast and simple method to analyse cabbage seed deterioration by measuring ethanol production from partially imbibed seeds. The method uses a modified breath analyser and is simple compared to gas chromatographic or enzymatic procedures. A modified method using elevated temperatures (40°C instead of 20°C) shortened the assay time and improved its sensitivity. The analysis showed an inverse correlation between ethanol production and seed quality (e.g. the final percentages or speed of germination and the number of normal seedlings). The increase in ethanol production was observed when cabbage seeds were deteriorated by storage under ambient conditions or hot water treatments, both of which reduced the number of normal seedlings. Premature seeds produced more ethanol upon imbibition than mature seeds. Ethanol production occurred simultaneously with oxygen consumption, indicating that lack of oxygen is not the major trigger for ethanol production.

Each taxon is characterized by a temperature range over which seed germination is possible and this may vary in space and time in relation to climate and ecological conditions. We used thermal modelling to test the hypothesis that thermal thresholds for seed germination can predict germination timing of Mediterranean species along an altitudinal and environmental gradient. Seeds of 18 species were collected in Sardinia from sea level to 1810 m above sea level, and germination tests were carried out at a range of constant (5 to 25°C) and alternating (25/10°C) temperatures. Different dormancy-breaking treatments [gibberellic acid (GA3), cold (C) and warm (W) stratifications and dry after ripening (DAR)] were applied. The annual pattern of soil temperatures was recorded using 24 data-loggers buried close to the study species. The logged soil temperatures distinguished ‘Mediterranean lowland’ from ‘Mediterranean mountain’ species. Although germination was >50% in untreated seeds of most species, GA3 had a positive effect in all species. C either inhibited or had a neutral effect on germination, W did not enhanced seed germination, while DAR had a positive effect only in species from coastal environments. The thermal time constant (S) for 50% germination ranged from 22 to 357°Cd (degree days) above base temperatures (Tb) of –9 to 9°C, depending on species and treatments. Mediterranean lowland species had lower Tb values compared with upland species. This study revealed significant differences in germination thresholds of Mediterranean lowland and mountain species in relation to Tb and S that probably have an impact on germination timing in the field and niche competitiveness.

The Millennium Ecosystem Assessment estimates that between 60,000 and 100,000 plant species are threatened with extinction–equivalent to around one-quarter of the total number of known plant species. Why should we care? There are a number of reasons. The first is that these plants may be useful to us in unknown ways. Secondly, ecology has taught us that resilience is found in diversity. Thirdly, we should be saving plant species from extinction because we can–there is no technological reason why any plant species should become extinct. Where we can't protect and manage plant diversity in situ, we should be employing ex situ conservation techniques, ranging from seed banks to habitat restoration. The Millennium Ecosystem Assessment describes such interventions as ‘techno-gardening’. This is not an abstract concept–it is already a reality in the majority of man-managed landscapes. In this context the perception of ex situ conservation as simply a back-up strategy for in situ conservation is mistaken. We are all involved in ex situ conservation to some degree, from cultivating our back gardens, to farming, to management of protected areas. Ex situ conservation should be seen as a complementary approach to in situ conservation and on the same spectrum. Kew's Millennium Seed Bank Partnership, comprising more than 120 plant science institutions in 50 countries, epitomizes this philosophy in action. We work actively on every seed collection we bank, finding out how useful it is and how we can grow it to enable human innovation, adaptation and resilience. Challenges remain at the policy level; for example, the need to factor-in the value of natural capital to development decision making, and better defining a role for public-sector science. At the technical level, also, there is much to do. Perhaps the greatest technical challenges relate to the restoration and management of complex, self-sustaining habitats or species assemblages. If we are to techno-garden effectively, in order to maintain ecosystem services and sustain biodiversity, then a multidisciplinary approach will be required. Many plant science institutions have recognized this and are becoming engaged increasingly in restoration activities and in situ management. Ultimately, humanity's ability to innovate and adapt is dependent on our having access to the full range of plant species and the alleles they contain.

A characteristic mechanism of gene expression regulation during seed germination is the selective translation of mRNAs. Previous findings indicate that the two cap-binding complexes eIF4F (with eIF4E and eIF4G subunits) and eIF(iso)4F [with eIF(iso)4E and eIF(iso)4G subunits] are differentially expressed during maize seed germination. In addition, several studies in vitro have suggested that these factors may participate in selective mRNA translation. The translational activities of eIF4E and eIF(iso)4E were tested in vitro using transcripts from two different sets: dry (0 h) and 24-h-imbibed maize embryonic axes. In vitro translation of these mRNA pools in the presence of the recombinant eIF4E or eIF(iso)4E, and the native cap-binding complexes from dry- or 24-h-imbibed axes, produced different profiles of proteins which were visualized by two-dimensional protein gels and autoradiography. The data indicated that eIF(iso)4E was particularly required for translation of the stored mRNAs from dry seeds, and that eIF4E was unable to fully replace the eIF(iso)4E activity. In addition, the dry seed mRNA pool was translated by the cap-binding complex isolated from dry seeds better than by the complex isolated from 24-h-imbibed seeds, whereas the translational efficiency of the mRNA pool from 24-h-imbibed seeds was similar between the cap-binding complexes from these two stages. Interestingly, eIF(iso)4E was more abundant than eIF4E in dry seeds, while both cap-binding proteins were present at similar levels in 24-h-imbibed seeds. These results suggest that the ratio of eIF(iso)4E to eIF4E in the corresponding eIF4F complex is critical for the mechanisms of translational control during maize germination.

Soil salinity is one of the major abiotic stresses affecting seed germination, crop growth and productivity. In this study, seeds of three wheat (Triticum aestiveum L.) and three barley (Hordeum vulgare L.) cultivars were treated with different concentrations of NaCl to investigate the effect of salt on seed germination physiology and metabolism through the characterization of seed germination pattern, gamma-aminobutyric acid (GABA) shunt metabolite accumulation [GABA, glutamate (Glu) and alanine (Ala)] and glutamate decarboxylase (GAD) expression using RT-PCR. A trend of decreasing germination percentage with increasing NaCl concentrations was observed. Under all salt stress treatments, data showed significant increase with positive correlation (r = 0.50–0.99) between abundance of GABA shunt metabolites and salt concentration in all wheat and barley cultivars for 5 days. Increased GABA content was associated with a small but significant increase in Ala and Glu content in all cultivars. In all NaCl treatments, the transcription of GAD in terms of RNA abundance showed a significant increase in all cultivars with positive correlation (r = 0.50–0.98). Data showed significant association between GAD RNA transcription and the response of germinating seeds to salt stress in terms of GABA shunt metabolite accumulation. The elevated expression of GAD under salinity suggests the need for elevated activity of the GAD-mediated conversion of Glu to GABA during seed germination, which provides alternative metabolic routes to the respiratory machinery, balancing carbon and nitrogen metabolism and osmolyte synthesis in germinating seeds of wheat and barley under salt stress.

Development of the rudimentary embryo in mature carrot (Daucus carota) seed during imbibition was characterized. The small embryo in the carrot seed, located in the micropylar region, elongated into the lateral part during imbibition and attained about two-thirds the length of the seed before radicle protrusion. Developing embryos excised from imbibed seeds were only capable of germinating in both water and Murashige–Skoog (MS) medium when they reached maximum size. The corrosion cavity into which the embryo grew enlarged concomitantly with endosperm degradation. The expression of endo-β-mannanase (EC 3.2.1.78), which is assumed to be involved in endosperm degradation, was characterized. A cDNA encoding an endo-β-mannanase was obtained by reverse transcription polymerase chain reaction (RT-PCR) using total RNA extracted from 24-h-imbibed carrot seeds. The full-length cDNA (DcMAN1) exhibited 64% deduced amino acid sequence identity with tomato (Lycopersicon esculentum) seed germination-associated mannanase (LeMAN2). DcMAN1 mRNA and endo-β-mannanase activity were first detected in the micropylar-half seed and then in the lateral-half seed. The timing of the appearance of DcMAN1 mRNA and endo-β-mannanase activity in the lateral-half seed corresponded with that of embryo development into this region. These results suggest that the expression of DcMAN1 and endo-β-mannanase activity in imbibed carrot seeds is associated with the enlargement of the corrosion cavity, which accompanies embryogenesis.

Among 14 species of herbaceous Fabaceae, all six winter annuals show a marked non-deep physiological dormancy of the embryo in addition to physical dormancy. This physiological dormancy is apparent at 23°C, but not at lower temperatures of 10°C and 5°C and disappears after 3 months of dry storage. These results corroborate the hypothesis that combinational dormancy is a double safety mechanism for delaying germination during summer: physical dormancy postpones germination, and even in early softened seeds germination is prevented by physiological dormancy of the embryo. Softened, swollen seeds of Medicago arabica tolerate a subsequent desiccation and remain viable even after five cycles of dehydration and rehydration. The rate of natural softening of M. arabica seeds increases exponentially at higher temperatures, with a Q10 between 3.4 and 5.1, and obeys the Arrhenius equation. This indicates that a chemical reaction might be involved in breakdown of physical dormancy. Winter annuals with hard seeds show similar properties as winter annuals with permeable seeds: the need for afterripening and requirement of lower temperatures delay germination until autumn. Only one species, Vicia sativa, loses physical dormancy during dry storage. Drying during summer might be a supplementary cue for germination in autumn.

Rouya polygama (Apiaceae) is an endangered Mediterranean species of great phytogeographical and ecological interest, growing on coastal sandy dunes. Intraspecific variability in the responses to constant temperatures (5–25°C) and an alternating temperature regime (25/10°C), salt stress (0–600 mM NaCl) and recovery of seed germination was evaluated among six populations from Sardinia and Corsica. Seeds were non-dormant and germination percentages ranged from 10 to 83%, depending on temperature and population. Differences in germination percentages were mainly due to different seed mortality among seed lots. R. polygama seeds germinated in salt concentrations up to 200 mM NaCl, whereas higher salt concentrations totally inhibited germination. Salt affected seed viability, and the recovery response decreased with increasing salinity and temperature. Inter-population variability and different sensitivity to NaCl in seed germination were detected. Our results are consistent with field germination in a period from autumn to spring, when water is available in the soil and temperatures are not prohibitive for seedling establishment, representing an advantageous ecological adaptation for seedling establishment to the unpredictable Mediterranean rainfall pattern. Further studies on R. polygama are needed to investigate germination requirements at temperatures higher than 25°C and its germination in the field, and to clarify genetic inter-population variability, considering a higher number of populations and possibly extending to North African populations.

Priming in polyethylene glycol (PEG, −1.5 MPa) for 7 d followed by drying, led to a 4–5 fold increase in mean longevity when achenes (seeds) of Ranunculus sceleratus L. were subsequently stored at 35°C and 9.2% moisture content on a fresh weight basis (67.1% equilibrium relative humidity). The increase in longevity was due to an increase in the standard deviation of the frequency distribution of individual seed lifespans (decrease in the slope of transformed survival curves) and to a lesser extent to an increase in the intercept of survival curves. Priming for 1 d resulted in a smaller but significant increase in longevity independent of whether seeds were primed in PEG, distilled water or a saturated atmosphere (100% RH).

The effects of priming were not due to the prevention of imbibition injury, and there was no evidence that the water relations of whole achenes was affected by priming. The effects of a 7 d priming treatment were dependent on the temperature and nature of the priming medium. However, no differences were recorded between corresponding treatments which were primed in PEG or distilled water. Moreover, the survival and responsivity to priming of R. sceleratus seeds was not related to dormancy status. The effects of priming on the longevity of R. sceleratus seeds are apparently unrelated to seed quality as the response of seeds previously aged for 8 d at 35°C and 7.9% moisture content was similar to that for unaged seeds.

Priming followed by drying did not increase seed longevity in the related species R. acris L. The promotive effects of priming on seed survival appear to be species specific and may be related to ecological factors.