Recent advances in agricultural biotechnology have produced many new crop varieties with valuable transgenic traits. These varieties are being, and will continue to be, marketed alongside conventional non-transgenic varieties. As a result, seed purity in commercial seed lots is of particular importance to both seed consumers and seed producers. A key step in the seed production process is the design of sampling and testing procedures used to evaluate seed lot purity. However, due to uncertainties in such methods, there is always a risk of incorrectly rejecting or accepting a seed lot. This paper discusses factors that should be considered when designing and implementing seed purity testing procedures to manage this misclassification risk – especially with regard to the presence or absence of transgenic traits. Many sources of uncertainty in both seed lot sampling and in the assay methods are described, and recommendations for reducing their impact are provided. This paper also explains the statistical concepts of misclassification risk as it affects seed producers and seed consumers. Sampling plans and formulas for determining the sample sizes necessary to control these misclassification errors when accepting or rejecting seed lots are also provided. Both simple, single-stage testing plans and the, often more efficient, double-stage testing plans are described. Testing seed pools rather than individual seeds is introduced as another way of adding efficiency to the testing process. Formulas are given for determining, from a seed sample, the confidence limits for the actual purity level of a commercial lot.

Peptide transport plays a major role in the nitrogen nutrition of the cereal embryo during germination. During germination, enzymatic hydrolysis of storage proteins in the cereal grain endosperm forms a reservoir of small peptides and amino acids which are translocated across the scutellum to supply organic nitrogen to the growing embryo. Uptake of these solutes by the scutellum, a modified cotyledon which functions in nutrient transport, is mediated by carriers localized to the plasma membrane of the scutellar epithelium. To date the peptide transporter HvPTR1 is by far the best characterized example of a nutrient transporter involved in reserve mobilization during germination. Peptide transport in the barley scutellum has been relatively well-characterized biochemically, and in recent years the barley scutellar peptide transporter HvPTR1 has been cloned and characterized at the molecular level. Here, we review the physiological role and importance of peptide transport in germination, focusing on recent characterization of the barley peptide transporter HvPTR1. In barley, the uptake of small peptides by the scutellum appears to be mediated by a proton-coupled peptide transport system capable of handling peptides 2–4 amino acid residues in length.

Raffinose family oligosaccharides (RFOs) are of almost ubiquitous occurrence in plant seeds. They accumulate during seed development and disappear rapidly during germination. The biosynthesis of raffinose, the first member of the series, proceeds by addition of a galactosyl unit to sucrose. Galactinol, a galactosyl derivative of myo-inositol, acts as a galactosyl donor. It is synthesized from UDP-D-GALACTOSE AND MYO-INOSITOL. STACHYOSE, VERBASCOSE AND AJUGOSE, THE NEXT HIGHER RFOS, ARE EITHER SYNTHESIZED BY GALACTINOL-DEPENDENT GALACTOSYLTRANSFERASES OR BY TRANSFER OF GALACTOSYL UNITS BETWEEN TWO RFO MOLECULES. IN SEEDS, THE METABOLISM OF METHYLATED INOSITOLS, SUCH AS D-ononitol and D-pinitol, is linked with the RFO pathway. In contrast to myo-inositol, these cyclitols are galactosylated by transfer of galactosyl residues from galactinol and not from UDP-D-galactose. However, the resulting galactosyl cyclitols can replace galactinol as galactosyl donors for the biosynthesis of stachyose. These recently discovered branches of the RFO pathway are active in seeds of a range of crop species, especially in legumes. We focus here on the biochemistry and molecular biology of the enzymes of RFO and galactosyl cyclitol biosynthesis. The metabolic control and hormonal regulation of the pathway during seed development and germination is discussed. The controversial role of α-galactosidases, which are believed to hydrolyse RFOs during germination, is reviewed critically.

An Escherichia coli phytase gene was introduced into Arabidopsisplants using an embryo-specific promoter and a signal peptide for vacuolar targeting. Three independent transgenic lines were analysed. Phytase activity in dry seeds was observed in transgenic lines, whereas no activity was detected in control, untransformed seeds. Transgenic seeds expressing the phytase gene had lower levels of phytic acid than the controls. Concomitant with the decrease in phytic acid was an increase in free phosphate. These results indicated that embryo-expressed phytase can reduce the levels of phytic acid stored during development.

Lipoxygenases are widely distributed in the animal and plant kingdoms. These enzymes catalyse the hydroperoxidation of polyunsaturated fatty acids containing cis,cis-1,4-pentadiene moieties. Multiple isoenzymes, with different biochemical properties and tissue distribution, have been described for many plants. Lipoxygenases occur in vegetative tissues, but also accumulate in various seeds, and especially in leguminous seeds. Although several functions have been proposed for vegetative lipoxygenases, the roles of seed lipoxygenases remain enigmatic. In this review we discuss whether physiological functions assigned to vegetative lipoxygenases can be extended to seed isoforms.

The phenomenon of dormancy extends beyond the boundaries of the plant kingdom. While plant biologists typically associate dormancy-breaking treatments only with seeds, buds or tubers, these chemicals and environmental stimuli have much broader activity as general terminators of developmental arrest in other, non-plant species. The activation of growth by these treatments is associated with signal transduction processes, metabolic upregulation and changes in gene expression, in addition to other events that may or may not be species specific. The study of both the classic and current developmental arrest literature beyond the boundaries of plant biology may be helpful in generating useful ideas and analogies for meaningful experimental progress towards understanding seed dormancy.

Correlative evidence indicates that sucrose and α-galactosyl-sucrose oligosaccharides (raffinose family oligosaccharides; RFOs) may be involved in seed longevity. Priming treatments (hydration in water or osmotic solutions followed by drying) can improve short-term seed performance but often result in reduced seed longevity. As RFOs are metabolized quickly following seed imbibition, loss of RFOs during priming could lead to more rapid deterioration in dry storage. This hypothesis was tested by measuring sucrose and oligosaccharide contents and potential longevity of primed seeds. Raffinose contents of whole lettuce (Lactuca sativa L.) seeds declined during hydration and priming and were correlated with decreased median potential viability (p50). However, this relationship was less significant when only the embryonic axes were analysed. In tomato (Lycopersicon esculentum Mill.) and impatiens (Impatiens balsamina L.) seeds, planteose was the major galactosyl-sucrose oligosaccharide and only small quantities of RFOs were present. Planteose contents declined during priming in seeds of both species, while sucrose contents increased or remained constant. Post-priming treatments that restored longevity in primed impatiens and tomato seeds were not accompanied by consistent changes in RFO or planteose contents. Our data do not rule out a role for oligosaccharides in seed longevity, but they make it unlikely that changes in oligosaccharide contents alone are responsible for the reduction in longevity due to priming or its restoration by post-priming treatments.

A commercially available smoke-water solution (Seed Starter®) stimulated the germination of caryopses and intact florets of Avena fatuaL. The solution was most effective when diluted (5–50%) and presented to intact or dehulled grain that had received a short period of dry after-ripening. It was less effective when applied at full strength or to grains that had been freshly harvested. The same stimulatory effect was observed in partly after-ripened caryopses of nine different wild oat biotypes obtained from three different cropping regions of the world. When freshly harvested caryopses were re-tested with the commercial solution (100%) for just 7 days prior to placement on to distilled water, a much higher germination percentage was possible than seen with continuous smoke-water incubation. The stimulatory ability of smoke water was more closely matched to that of gibberellic acid than to potassium nitrate, which had little or no effect on freshly harvested caryopses. The smoke-water solution (5–100%) was tested on the germination of 18 other cool temperate arable weed species. All monocotyledonous species tested (viz. Avena sterilis ssp. ludoviciana L., Alopecurus myosuroides, Sorghum halepense, Phalaris paradoxa) responded positively, while those of the dicotyledonous species were either strongly stimulated (≥40%stimulation Malva neglecta), moderately stimulated (≥20% stimulation Galium aparine, Veronica persica), slightly stimulated (Polygonum persicaria, P. pennsylvanicum, Fallopia convolvulus), unaffected (P. aviculare, Sinapis arvensis, Heracleum sphondylium, Angelica sylvestris, Mercurialis annua, Veronica hederifolia) or inhibited (Lamium purpureum). The optimal concentrations required to stimulate germination of the monocotyledonous species were similar to those observed for A. fatua (5–10%). However, for the dicotyledonous species slightly stronger solutions were required (10–20%). When the unaffected species were retested using a 10-day pre-chilling treatment, smoke water showed a small promotive response in three (S. arvensis, P. aviculare and V. hederifolia) of the six species. When four different smoke-water solutions (Seed Starter®, ®, charred-wood solution and wheat-straw solution) were tested on two representative species (A. fatua and M. neglecta), three formulations were effective in promoting the germination of both species, while the fourth (charred-wood solution) was only active on A. fatua. The active concentrations were different for the four solutions. Three solutions were active in the 2–20% dilution range, while the fourth (®) was only active in the 1–2% dilution range and was inhibitory at higher concentrations. These observations are discussed in the context that smoke may play an important ecological role in the management and control of introduced weeds in native and arable communities.

The mature seeds of durian (Durio zibethinus L.) are recalcitrant, with the cotyledons storing both starch (amylose, 78% dry weight) and some protein (7%), but having few triglyceride reserves (<1%). Reserve accumulation occurs during the latter third of the 110-d seed developmental period prior to fruit shed. The storage proteins of durian seeds consist of two immunologically related unglycosylated albumins, light and heavy zibethinin (20.2, 21.4 kDa), that account for 37–46% of the total protein during development. Light zibethinin is a single polypeptide, most likely having internal sulphydryl linkages, whereas heavy zibethenin is composed of two sulphydryl-linked subunits. The zibethinins are deposited in swellings of the endoplasmic reticulum (ER) which then either expand or coalesce to form protein storage vacuoles. This mode of protein body formation in durian varies from both the ER–Golgi complex–vesicle–vacuole pathway characteristic of many dicotyledonous seeds and the direct ER to protein storage vacuole pathway involved in deposition of certain prolamins in cereal endosperm cells. Protein storage vacuoles in durian seed storage parenchyma never become fully filled, but contain only diffuse peripheral clumps of zibethinin protein. There is a 15% loss in starch, a precipitous loss of total protein, and a concomitant loss of the zibethinin storage proteins during the 5 d prior to fruit shed, suggesting that both reserve mobilization and germination has begun.

A common feature of desiccation-tolerant organisms, such as orthodox seeds, is the presence of large quantities of sugars, especially di- and oligosaccharides. These sugars may be one component of the suite of adaptations that allow anhydrobiotes to survive the loss of most of their cellular water. This paper describes the physical effects of dehydration on cellular ultrastructure, with particular emphasis on membranes, and explains quantitatively how sugars and other solutes can influence these physical effects. As a result of dehydration, the surfaces of membranes are brought into close approach, which causes physical stresses that can lead to a variety of effects, including demixing of membrane components and fluid-to-gel phase transitions of membrane lipids. The presence of small solutes, such as sugars, between membranes can limit their close approach and, thereby, diminish the physical stresses that cause lipid fluid-to-gel phase transitions to occur during dehydration. Thus, in the presence of intermembrane sugars, the lipid fluid-to-gel phase transition temperature (Tm) does not increase as much as it does in the absence of sugars. Vitrification of the intermembrane sugar solution has the additional effect of adding a mechanical resistance to the lipid phase transition; therefore, when sugars vitrify between fluid phase bilayers, Tm is depressed below its fully hydrated value (To). These effects occur only for solutes small enough to remain in very narrow spaces between membranes at low hydration. Large solutes, such as polymers, may be excluded from such regions and, therefore, do not diminish the physical forces that lead to membrane changes at low hydration.

Embryonic axes of tea (desiccation sensitive) and pea (desiccation tolerant) were dried at different rates or stored at different water contents to distinguish between damage associated with the immediate effects of water loss and the longer-term effects of a partially hydrated state. No loss of viability was observed if pea axes were dried sufficiently rapidly (from 1.8 to 0.1 g H2O g-1dry mass (g/g) within 5 d). However, viability was lost in tea axes dried below 0.5 g/g (approximately -15 MPa) even if axes were dried within 1 h. Death in tea axes dried to moisture contents less than 0.5 g/g probably resulted from the removal of water necessary for cellular structural integrity (i.e. desiccation damage sensu stricto). When axes of both species were dried at slower rates, viability losses were observed at water potentials between about -3 and -15 MPa. The timing for this type of damage was species dependent, occurring within 2 d for tea and after 5 d for pea, and may be explained by higher oxidative activity in tea compared to pea. Embryos of both species with water potentials below -3 MPa were lethally damaged if oxygen consumption exceeded 1000–5000 μmol O2 g-1dry mass. Recalcitrant seeds are different than orthodox seeds because the former do not survive drying below a critical water content, regardless of the drying rate. Rapid drying is required for accurate assessment of the critical water content. Slow drying leads to metabolic imbalance and artefactual assessment of the critical water content for desiccation damage. Both tea and pea seeds were susceptible to damage from metabolic imbalances, suggesting that the predominant stress from slow drying is ageing.

Homologues of the pea SBP65, a late embryogenesis abundant (LEA) biotinylated protein that behaves as a putative sink for the free vitamin biotin during embryo development, were characterized biochemically in various plant species, including soybean, lentil, peanut, rape, cabbage, carrot and sugarbeet. Based on sequence homologies, the genome of Arabidopsis thaliana contains a gene putatively encoding a homologue of pea SBP65. These proteins exhibit two remarkable features. First, they only accumulate in seeds, particularly during late stages of embryo development. The results strongly suggest that these seed-specific biotinylated proteins belong to the class of plant proteins called seed maturation proteins, which are presumed to play major roles in embryo development. Secondly, covalent attachment of biotin occurs at a lysine residue within a conserved motif of (V/M)GKF, which shows no resemblance to the highly conserved AMKM tetrapeptide that houses the target lysine residue in the well-characterized biotin-dependent carboxylases and decarboxylases. These findings highlight novel structural features for protein biotinylation.

Common buckwheat (Fagopyrum esculentum Moench) plants produce many flowers, but fewer seeds. Seed set is highly variable among years, between plants, and during the period of flowering within a plant or raceme. Seasonal variations suggest that temperature and water-deficit stresses are important factors for seed set. The effects of mild temperature and water-deficit stresses on seed set and seed filling were determined in planta and in vitro. An in vitro method to culture matched sets of racemes from individual plants was used for precise comparisons between experimental treatments. Buckwheat racemes form new flowers continuously during several weeks in planta. Seed set resulting in yield occurs during the first 2–3 weeks of flowering and then rapidly declines in planta independently of mild stresses. Plants grown at 18°C have 40% increased seed set, set seeds over a longer duration, and produce 40% more dry matter per seed than plants grown at 25°C. Similar patterns occurred in vitro. A 3-day water-deficit stress during the first week of flowering reduced the number of seeds by 50%without a reduction in seed size and dry weight, or the number of flowers formed in planta or in vitro. The effect of water-deficit stress continued after rewatering and subsequently was expressed as a reduction in fertility in newly formed flowers, both in planta and in vitro. Mild temperature and water-deficit stresses affected both female and male components of seed set in common buckwheat, resulting in a persistent but non-additive reduction in sink strength.

The thickened cell walls of endosperm cells of asparagus (Asparagus officinalis) seeds are composed of glucomannans. During development, the extensive thickening of the cell wall is specific to the lateral region of the endosperm and not to the micropylar region. The micropylar endosperm may thus be predisposed to facilitate germination. Following germination there is a progressive mobilization of the reserves in the cells of the lateral endosperm in a wave-like manner away from the haustorial cotyledon, accompanied by the loss of cytoplasm and the crushing of the lateral endosperm cells. Although the internal reserves are mobilized from the micropylar endosperm cells, the cells themselves remain intact throughout the period of lateral endosperm mobilization and may form a living barrier to prevent the loss of soluble hydrolysis products of the lateral endosperm to the surrounding environment. The location and timing of endo-β-mannanase production and the increase in activity of β-mannoside mannohydrolase in seeds of germinated asparagus were followed. Endo-β-mannanase activity increases greatly in the endosperm until the mid-point of mobilization, and is about 45 times higher than in the embryo on a per seed part basis. Unlike endo-β-mannanase, which is extractable in low-salt buffer, β-mannoside mannohydrolase requires high salt (0.5 M NaCl) for extraction. This enzyme continually increases in activity in both the endosperm and embryo following germination, with the majority of the activity being concentrated in the embryo when considered on a per seed part basis.

To determine whether seeds of the weedy shrub Wigandia urens, from the Valley of Mexico, undergo natural priming when buried in soil, comparative experiments were performed with seeds: (1) harvested directly from the plants; (2) buried in three natural habitat conditions; and (3) laboratory primed with polyethylene glycol. Seeds were sown in a growth chamber and in a shade house. Final germination percentages, emergence, germination and emergence rates, survival and initial growth were determined. Burial and priming enhanced the germination and emergence parameters evaluated in the laboratory and in the shade house. Effects of treatments on survival were not significantly different. Nevertheless, burial improved emergence and mean survival, and induced differences in specific leaf area of seedlings that could have ecological significance. Heat-stable proteins were extracted and electrophoresed. Proteins formed in W. urens seeds during burial had molecular weights (14–21 kDa) similar to those reported for late embryogenesis abundant (LEA) proteins induced by priming in other species. Nevertheless, the presence and abundance of proteins expressed (14–23, 36 and more than 45 kDa) differed among control, primed and buried seeds. During soil burial, molecular and physiological responses were induced that were similar to the effects of priming.

This study examined the hypothesis that desiccation sensitivities of recalcitrant and intermediate seeds can be categorized into discrete levels of critical water potential. The equilibrium dehydration method was used to determine the critical water potential (CWP) below which desiccation damage started to occur. The CWP values of Bruguiera cylindrica, Lansium domesticum, Litchi chinensis and Lumitzera racemosa are approximately –4 MPa. The CWP values of Andira inermis, Avicennia alba, Castanea sinensis (from New Zealand), Citrus aurantifolia, Ginkgo biloba, Nephelium lappaceum and Theobroma cacao (immature axis) are approximately –8 MPa. The CWP values of Acer pseudoplatanus, Castanea sinensis (from China), Quercus rubra and Theobroma cacao(mature axis) are approximately –12 MPa. The CWP values of Artocarpus heterophyllus and Hevea brasiliensis are approximately -23 MPa, while the CWP values of Acer platanoides, Azadirachta indica, Carica papaya and Coffea arabica are approximately –73 MPa. Together with data available in earlier literature, these CWP values suggest that there are five discrete levels of critical water potential among desiccation-sensitive seed tissues. These data support the hypothesis that discrete levels of desiccation sensitivity occur among recalcitrant and intermediate seeds, and suggest that specific damaging and protective mechanisms exist at certain hydration levels.

Patterns of seed mass variation in the perennial spurge Euphorbia characias(L.) (Euphorbiaceae) in three populations from north-east Spain were examined. Mature inflorescences have a distinctive structure, with a distal umbrella of rays subtending dichotomously grown cyathia. Under the umbrella, cyathia are loosely arranged on rays growing from the bases of leaves. Seed characteristics were studied, especially in relation to position along the inflorescence. Maximum inflorescence height had no effect on seed set or seed mass. Seed set did not differ between populations. Flower position along the inflorescence had a marked effect on seed set and seed mass. Seed set was higher in the distal (upper) portion of the inflorescence. Seeds from the distal area had a greater mass than those from lower parts of the inflorescence. Seeds were planted in the field and observed under natural weather conditions. The bigger seeds from the higher position in the inflorescence produced seedlings with a greater percentage emergence, which, in addition, had an increased percentage establishment than seedlings originating from the smaller seeds from lower areas of the inflorescence. For the three populations considered together, seeds from the basal (lower) portion of the inflorescence were reduced in germination by 51% and establishment by 59%. No differences in seedling dry mass were observed. Seed dispersal-related structures (elaiosome and elaiosome/load ratio) did not vary in relation to position or population. Positional effects on female performance – seed quantity and quality – may be a very strong selective pressure for the peculiar flower disposition in the architecture of E. characias.

Cucurbita maxima Duch. cv. Warted Hubbard, which produces large fruits with many seeds, was used as a test system to study changes in several mineral nutrients during seed formation. Developing embryos increased markedly in weight, size and oil content. Energy dispersive X-ray (EDX) analysis was carried out on mineral nutrient storage globoids in developing Cucurbita cotyledons from 4 to 9 weeks after pollination. Phosphorus peak-to-background (p/b) and potassium p/b ratios increased slightly after the fourth week, while magnesium p/b ratios remained unchanged. The calcium p/b ratio was statistically higher at 4 weeks than in the subsequent weeks. Traces of iron and zinc were found in most of the globoids at all fruit ages. The zinc p/b ratio was higher in globoids at 4 weeks than at 9 weeks. Traces of manganese were detectable in only 18% of the globoids at 4 weeks and not detectable in any globoids at 9 weeks. The frequency of the small globoids of 0.25 μm in diameter was the highest at 4 weeks and was zero at 9 weeks. The frequency of the larger globoids increased progressively from 4 to 9 weeks. As the spherical globoids increased in size and hence in volume, there was a 6.4-fold increase in the embryo phytic acid, which is the main component of globoids. On a whole embryo basis, P content increased 4.5-fold, K 3.3-fold, Mg 5.4-fold and Ca 3.2-fold from the fourth to the ninth week.

At shedding, the moisture content (MC) of Avicennia marina (Forssk.) Vierh. propagules was 65% (fresh mass basis), and there was no significant difference in the MC of four tissues (hypocotyl, cotyledons, plumule and root primordia). Viability declined as the propagules were dried below 60% MC, so that only 40%of seeds were capable of germination at 54% MC. At 47% MC all the seeds had died. The four tissues dried at the same constant rate of 0.02 g water g dwt-1 h–1 throughout this range of MCs. There was no significant depletion of the free-radical-quenching mechanisms measured. In each tissue an organic free-radical was detected by electron paramagnetic resonance (EPR). In the plumule the amplitude of the signal increased by a further 50% at MCs where viability was lost, but there was no increase in the other tissues. There was a concurrent increase in the amount of tocopherol and the activity of superoxide dismutase in the plumule. Lipid peroxidation, assessed by the amount of thiobarbituric acid-reactive substances, also increased in advance of viability loss, suggesting that propagules were experiencing oxidative stress. However, lipid peroxidation decreased at 54–57% MC, where most seeds lost viability. The results presented are consistent with a propagule reacting to oxidative stress, but overtaken by more catastrophic physical damage.