Thioredoxins (Trxs) are widely distributed, small proteins that function in redox regulation in a broad spectrum of cellular reactions. Experimental work with barley, wheat and a legume (Medicago truncatula) has established thioredoxin h (Trx h) as a central regulatory protein in seeds, reducing disulphide (S–S) groups of diverse seed proteins, including storage proteins, enzymes and enzyme inhibitors. Trxs appear to be particularly important in plants, as a large number of genes are present compared to mammalian organisms. In Arabidopsis thaliana there are approximately 20 different genes for classical Trxs, and large Trx gene families have also been found in cereals, such as rice, barley, wheat and sorghum. Extensive evidence indicates that adding Trx, reduced nicotinamide adenine dinucleotide phosphate (NADPH) and NADP-thioredoxin reductase (NTR) to cereal flour or seed preparations reduces disulphide (S–S) linkages of storage proteins. The early in vitro studies have been complemented with transgenic barley seed, overexpressing Trx h in protein bodies of the barley endosperm, which showed accelerated germination and early or enhanced expression of associated enzymes, i.e., α-amylase and pullulanase. Overexpression of Trx h levels in wheat was subsequently shown to (1) enhance protein solubility and digestibility, (2) reduce allergenicity of wheat gliadins, and (3) improve dough quality from poor-quality wheat flour. Most recently, we have demonstrated that down-regulation in wheat of Trx h9, a unique thioredoxin, leads to a reduction in the incidence of pre-harvest sprouting, demonstrated in several varieties over multiple generations with field-grown material. Yield and starch content were increased while baking quality in the high-gluten variety remained unchanged. These observations led to the intriguing question of how changes in the endosperm are communicated to the embryo. Studies of Trx h9, a membrane-associated Trx h that can move from cell to cell, provide suggestive evidence for a role of Trx h9 in intercellular communication of redox state.

This paper describes the exploratory use of isothermal micro-calorimetry (IMC) to measure directly the heat flow produced as seeds age. Heat flow was recorded in primed and non-primed (control) seeds of Ranunculus sceleratus L., aged in a micro-calorimeter at 35°C at three different seed water contents [c. 0.12, 0.075 and 0.045 g H2O (g dw)−1]. The rate of heat flow and total heat generated (an indicator of extent of reaction) were generally greater in control seeds, which aged at a faster rate, than in primed seeds. Total heat generated over a given period also increased with increasing water content. The power–time curves did not indicate first- or second-order rate kinetics, consistent with the probability that seed ageing is complex and involves a number of reactions. Even after the capacity to germinate had ceased, there was a residual power signal. As a method, IMC gave consistent results using independent samples at different times. Therefore, short-term experiments at relatively high water contents and/or temperatures may have the potential to predict the relative longevity of seed-lots, at least within a species.

Coastal desert vegetation of the Arabian Peninsula is almost entirely dominated by halophytes. Natural populations provide a genetic resource for ecological remediation and may also have direct economic value. High intrapopulation variation of seed traits is presumed to increase population persistence in the unpredictable climatic conditions of this hyper-arid desert. We investigated whether intrapopulation variation of seed mass, dormancy and germinability of four species was attributable to maternal individuals. Arthrocnemum macrostachyum, Halothamnus iraquensis, Haloxylon salicornicum and Seidlitzia rosmarinus are commonly distributed Arabian halophytes with differing seed weight variation. All species exhibited a higher germination when exposed daily to 12 h light, compared to seeds in darkness. A higher germination was correlated with a shorter germination time. For H. iraquensis and S. rosmarinus, a shorter germination time was negatively correlated with germination synchrony. H. salicornicum showed the highest intrapopulation variation of seed traits, followed by A. macrostachyum, S. rosmarinus and H. iraqensis. We found that individuals within populations of all the studied species showed variability in germination but the extent of variation was species-specific. The variation in seed mass and germination among the individuals of the studied species may facilitate a temporal distribution of germination, which may reduce the risk of seed bank exhaustion. The results of this study could assist conservation and management by improving the efficiency of seed collection from wild populations of these species.

This work presents the results of physiological studies developed to understand modifications linked to the reduction of seed dormancy provoked by domestication processes. The experiments performed compared wild and domesticated Cucurbita subspecies and their hybrids developed by reciprocal crossings. Seeds of two accessions of the wild subspecies presented dormancy, but it was largely reduced in seeds from the domesticated genotype, and partially reverted in hybrids, especially in those obtained when the domesticated genotype was used as the mother plant. In addition, naked embryos of all subspecies did not display dormancy when incubation was performed at 28°C, but embryo germination was progressively reduced only in the wild genotype under decreasing incubation temperatures (22 and 16°C). In the embryos, abscisic acid (ABA) concentrations were similar in both domesticated and wild subspecies, whereas in the seed coat, it was threefold higher in the wild subspecies. The naked embryos from the wild subspecies were far more responsive to ABA than those from the domesticated subspecies. These results indicate that dormancy in the wild subspecies is imposed by the seed coat tissues and that this effect is mediated by their high ABA content and the sensitivity of the embryos to ABA. These physiological aspects were apparently removed by domestication along with the temperature-dependent response for germination.

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.

We report the determination of abscisic acid (ABA) and its metabolites, phaseic acid (PA), dihydrophaseic acid (DPA) and ABA glucose ester (ABA-GE), in non-dormant dry and imbibed seeds of tomato (Solanum lycopersicum Mill.) cv. Moneymaker (wild type), and its tss1, tss2 and tos1 mutants. High ABA in dry seeds may originate from ABA accumulation in the sheath tissue, which was in contact with an ABA-containing medium, the endocarpus. The highest germination percentages at 72 h, observed in tss1 and tss2, coincided with minimal ABA content. Wild-type and mutant seeds showed different ABA and catabolic patterns, and these were correlated with their sensitivity to abiotic stress. Whereas dry seeds showed a high basal ABA, imbibed seeds showed higher ABA metabolite content, particularly DPA. The dramatic decrease of ABA following seed imbibition suggests an activation of ABA catabolism during the early stages of the germination process. The observed variation of ABA metabolites among dry and imbibed seeds of Solanum lycopersicum cv. Moneymaker and its tss1, tss2 and tos1 mutants shows that ABA metabolism is differentially regulated in these genotypes.

Investigations of physiological functions of seed lipoxygenase during germination and early seedling growth are complicated by the presence of multiple isoforms with different product specificity. The combination of isoelectric focusing, lipoxygenase in-gel activity staining and mass spectrometry showed that only two seed lipoxygenases, LOX-2 and LOX-3, were active at neutral pH in protein extracts from dry and germinating pea seeds (Pisum sativum L.), with LOX-3 more abundant than LOX-2. Each active seed LOX appeared at multiple pIs, probably resulting from post-translational modifications. The abundance of pea seed lipoxygenases was lower in embryonic shoots than in other parts of the axis. After radicle emergence, seed LOXs were degraded, as revealed by Western blotting using antibodies specific to pea seed LOXs. Two new vegetative isoforms, LOX-N3 and LOX-g, appeared in the shoot and gradually replaced the pea seed lipoxygenases as the principal contributors to LOX enzyme activity during early seedling growth. Activities of LOX-N3 and LOX-g were abundant in shoot tips, but not root tips, in pea seedlings. Thus, during germination, there is a shift of LOX activity from radicles to shoots that accompanies the transition from seed LOXs to vegetative LOXs. These results confirmed that identification of pea LOXs based solely on pI may be misleading; however, mass spectrometry enabled more accurate identification of these highly homologous proteins to be made.

Protein composition of sunflower seeds (Helianthus annuus L.) was analysed to identify a possible marker for germination capacity and to evaluate seed viability precociously after harvest and after natural ageing during storage. Electrophoresis on 2-D gels was used to compare total soluble and in vivo synthesized proteins in two seed lots showing different germination capacities. Differences were observed to a greater extent in in vivo protein synthesis profiles than in total soluble proteins. Inhibition of [35S]methionine uptake and incorporation in seeds with low germinability was also observed. The precise relationship between protein composition and prediction of germinability could not be determined. Other aspects such as membrane damage, capacity for protein synthesis and seed lot homogeneity in relation to germination are discussed.

Convallariamajalis has double dormancy and hypogeal germination, but no information is available on embryo growth or on the effects of light and gibberellic acid (GA3) on germination in this genus. Therefore, we investigated embryo growth and other germination features in seeds of C. keiskei and compared the data with those of Trillium camschatcense in another study. Until now, in seeds with double dormancy, embryo growth and germination (epigeal) have been studied in detail only for seeds of T. camschatcense. Phenology of embryo growth and emergence of cotyledonary petiole/root (hereafter root) and shoot in seeds of C. keiskei were monitored outdoors. Effects of temperature, light and GA3 on embryo growth and root and shoot emergence were tested under laboratory conditions. Roots emerged the first spring following seed dispersal in autumn. The embryo grew soon after root emergence, and germination was hypogeal. Seeds with an emerged root formed buds from which a shoot (leaf) emerged above ground during the second spring. Alternating temperatures and light had negative effects on root emergence, and GA3 did not substitute for cold stratification in root emergence. Seeds of C. keiskei have double dormancy, but it differs from that in T. camschatcense. Based on differences in embryo growth before (T. camschatcense) versus after (C. keiskei) root emergence, and on epigeal (T. camschatcense) versus hypogeal (C. keiskei) germination, we suggest that two types of deep simple double morphophysiological dormancy (MPD) be recognized. Since embryo growth in C. keiskei does not fit the standard definition of MPD, we propose to expand this definition.

Nothofagus alpina, N. obliqua, N. glauca, N. leonii, N. dombeyi and N. pumilio seeds exhibited consistent, albeit slight, sensitivity to extreme desiccation, but nevertheless maintained viability at low moisture contents and cool temperatures (–10° to –20°C) over 2 years. Nothofagus alpina, N. obliqua, N. glauca, N. leonii and N. dombeyi conformed to the seed viability equation of Ellis and Roberts; sensitivity of longevity to temperature was quantitatively similar to that of crop seeds, sensitivity to moisture was somewhat less, and a low-moisture-content limit to the equation was detected at 4.8% moisture content in hermetic storage at 65 °C, and possibly similar moisture contents at 30–40°C. These five species show orthodox seed storage behaviour. Therefore, ex-situ conservation of these Nothofagus species in seed banks is possible, but the quality of seed lots collected requires attention. Seed storage behaviour was not defined in N. pumilio: initial seed quality was poor and loss of viability was detected over 2 years at 0°, –10° and –20°C at 2.7% moisture content, but not at 5.2%. The results confirm that the economy of nature in seed storage physiology extends to forest tree seeds, but the repeated observation of reduced sensitivity of longevity to moisture in forest tree seeds requires further investigation.

In this study we developed a sample size procedure for estimating the proportion of genetically modified plants (adventitious presence of unwanted transgenic plants, AP) under inverse negative binomial group testing sampling, which guarantees that exactly r positive pools will be present in the sample. To achieve this aim, pools are drawn one by one until the sample contains r positive pools. The use of group testing produces significant savings because groups that contain several units (plants) are analysed without having to inspect individual plants. However, when using group testing we need to consider an appropriate pool size (k) because if the k individuals that form a pool are mixed and homogenized, the AP will be diluted. This effect increases with the size of the pool; it may also decrease the AP concentration in the pool below the laboratory test detection limit (d), thereby increasing the number of false negatives. The method proposed in this study determines the required sample size considering the dilution effect and guarantees narrow confidence intervals. In addition, we derived the maximum likelihood estimator of p and an exact confidence interval (CI) under negative binomial pool testing considering the detection limit of the laboratory test, d, and the concentration of AP per unit (c). Simulated data were created and tables presented showing different potential scenarios that a researcher may encounter. We also provide an R program that can be used to create other scenarios.

Amaranthus tuberculatus is a weed species that has shifted emergence patterns over the past few years, presumably due to changes in seed dormancy in response to selection in agricultural fields. Although it is recognized that the seed dormancy phenotype is greatly affected by the environment, it is also acknowledged that the genotype plays a significant role. However, the importance of the genotype in determining intra-population seed dormancy variability, and the effect on emergence patterns, is not well understood. The objective of the present study was to determine the importance of the genotype on deep dormancy and the stratification-mediated dormancy alleviation in A. tuberculatus. Wild populations differing in seed dormancy were crossed and F2 families were generated. These families were used to determine narrow sense heritability of dormancy and stratification-mediated dormancy alleviation at the individual (hi2) and family (hf2) levels. hi2 ranged from 0.13 to 0.4 and 0.04 to 0.06 for the dormancy and stratification response, respectively. In the case of hf2, the values ranged from 0.76 to 0.91 for deep dormancy and from 0.33 to 0.58 for the stratification response. The genetic correlation between these two traits was below 0.075, indicating that different genes control them. High temperature strengthened the dormancy of deeply dormant seeds, making them less sensitive to stratification. However, high temperature promoted the germination of non-deeply dormant seeds. It is proposed that delayed weed emergence can be generated by selecting genes that control stratification response, and not necessarily only the genes that are directly responsible for deep dormancy.

Storage proteins (2S, 7S and 11S) are synthesized at the mid- to late stages of development in alfalfa seeds, mainly within the embryos. Mature dry seeds cannot synthesize these proteins upon subsequent germination and growth. When embryos were isolated from the seed during development (stages VII and VIII) and placed on water or nutrient medium, they germinated. They exhibited a pattern of protein synthesis which was identifiable as germinative/post-germinative, and was identical to the pattern synthesized in embryos of germinated dry seeds, and of embryos from seeds that were subjected to drying prematurely at stages VII and VIII. When, after 48 h from the start of imbibition, abscisic acid or osmoticum was introduced to germinated embryos isolated at stage VII and not desiccated, the synthesis of the 11S storage protein was restored to a rate comparable to that of stage-VII embryos before isolation. This was accompanied by an increase in transcription of the 11S storage protein gene. Transfer of germinated stage-VII embryos, which had first been desiccated, to abscisic acid or osmoticum 48 h after imbibition started did not result in any restoration of storage protein synthesis. Thus, desiccation, prematurely or at maturation, off-regulates storage protein synthesis and within the embryos this process is no longer responsive to abscisic acid or osmoticum.

Inorganic P, total P and phytic acid-P leakage from imbibing whole soybean, bean, and pea seeds, as well as whole maize and barley grains were measured. The amounts of P and phytic acid leaked during a 6-h soaking period varied for the five species. Soybeans leaked the most P and phytic acid of the species examined. Peas, beans and barley leaked little or no phytic acid. Based on reported levels of total P and phytic acid in the dry seeds/grains that were studied, total P leakage ranged from 0.5 to 1.2%, while phytic acid leakage ranged from 0.01 to 0.5%. No relationship was found between initial levels of total P or phytic acid in the dry seeds/grains and the amount of total P or phytic acid leaked by the imbibed seed/grain. Similarly, the amounts of phytic acid leaked by the seeds/grains of this study did not appear to be related to the water solubility of the phytic acid in the dry seeds/grains.

A novel method has been devised to extract poly (A+)RNA from seed tissues. The advantages of this technique include the speed of extraction and the elimination of the hazardous phenol:chloroform extraction step which usually precedes oligo (dT)-cellulose column chromatography. Instead, the crude supernatant, containing a high concentration of SDS and salts to effectively inhibit RNase activity, is passed directly over the oligo (dT)-cellulose column. The resultant isolated poly (A+)RNA is of high yield and purity and is suitable for in vitro translation and Northern blot analysis.

Dynamics of abscisic acid (ABA) and indole-3-acetic acid (IAA) contents were followed in developing barley caryopses of the ABA-deficient mutant AZ34 and its parental cultivar Steptoe. Distribution of these hormones and HvPIP2 aquaporins (AQPs) was studied with the help of immunohistochemical methods in the roots and coleorhiza of developing embryos. In Steptoe, maturation of the caryopsis was accompanied by vast accumulation of ABA, while this hormone accumulated more slowly in the caryopsis of AZ34 and its content was lower than in Steptoe. Accumulation of ABA was accompanied by a decline in IAA level in the developing caryopsis, the process being delayed in AZ34 in accordance with the slower accumulation of ABA. ABA accumulated to high levels in the coleorhiza cells of Steptoe, while the effect was absent in AZ34. The high level of ABA was likely to be important for maintaining the barrier function of the coleorhiza, preventing germination of seminal roots and enabling seed dormancy, while the absence of ABA accumulation in coleorhiza of AZ34 may be responsible for the initiation of root germination inside the caryopsis. The abundance of HvPIP2 AQPs in the seminal roots was higher at the beginning of maturation of Steptoe caryopsis and declined afterwards, while the levels of APQs increased later in AZ34 in accordance with the delay in ABA accumulation. These results suggest the importance of ABA accumulation in coleorhiza for preventing precocious growth of seminal roots, and suggest regulation of IAA and aquaporin levels by this hormone during maturation of embryos.

Evidence is provided to show that frozen, hydrated seeds and seed tissues can be observed by low temperature field emission scanning electron microscopy. This technique allows preparation and observation of seed samples that contain 13–60% water without altering their moisture content. The technique provides information about the surface structure of seeds and also allows specimens to be fractured to reveal internal features of tissues. Futhermore, both halves of fractured specimens can be retained, examined and photographed either as two-dimensional micrographs or as stereo pairs when three-dimensional observation (stereology) or quantitative measurements (stereometry) are desired. Use of this technique avoids artefacts associated with chemical fixation, dehydration, and critical point drying—procedures that are usually required to prepare seed tissues for scanning electron microscope examination. This technique does not affect the degree of hydration in specimens; it can be used to localize water in tissues to determine their degree of hydration. This technique should find wide application in developmental studies of seeds, especially during maturation, imbibition and germination.

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

Temperate wetland species often require light and warm temperatures for seed germination. However, recent studies indicate that species which specialize on permanently saturated wetlands that are maintained by groundwater discharge (fens, seeps and mountain springs), rather than wetlands with surface-water-driven hydrologic regimes, diverge from the typical wetland germination niche by germinating at cool temperatures and lacking photoblastic seeds. We conducted laboratory experiments that manipulated stratification conditions (non-stratified versus cold stratification in light and darkness), thermal regime (15/6, 25/15 and 35/20°C), and light (14 h photoperiod versus continuous darkness) to test whether seeds of the North American calcareous fen specialist Parnassia grandifolia diverged from the typical temperate wetland germination niche. After 30 d, fresh seeds were conditionally dormant and could only germinate to high percentages in light at 25/15°C. During 16 weeks of incubation, non-stratified seeds germinated to low percentages ( < 40%) at all thermal regimes in darkness. In contrast, cold-stratified seeds germinated to high percentages in both light and darkness at all thermal regimes, although germination was incomplete (no cotyledon emergence) at 35/20°C. Further, seeds did not require light during cold stratification to germinate to high percentages when incubated in light or darkness. Thus, seeds diverged from the typical temperate wetland germination syndrome in lacking a light and warm temperature requirement for germination. Our results reinforce previous work from European fens and Mediterranean wetlands. This indicates that multiple germination strategies are found in fen wetlands that are maintained by the continuous or near-continuous discharge of cool groundwater.

Plant community-level studies on seed dormancy traits are important to understand and determine the significance of seed dormancy in different ecosystems. Hence, we studied seed dormancy and other related seed biological traits of 42 selected tropical lowland rainforest tree species from Sri Lanka, aiming to address seed dormancy class(es) for a biodiverse tropical lowland wet zone forest community and the relationship between dormancy classes, forest strata and seed dispersal mechanisms. Seed germination, imbibition, embryo length:seed length ratio, embryo morphology and the effect of gibberrelic acid on seed germination were determined. Sixty-two percent of the species with T50 < 30 days were identified as having fast-germinating seeds and the remaining 38% with T50 > 30 possessed slow-germinating seeds. Seeds of 33 species had fully developed embryos, while nine species had underdeveloped embryos; three had morphological dormancy (MD) and six morphophysiological dormancy (MPD). Treatment with gibberellic acid revealed physiological dormancy (PD) in seeds of six species, and the response to manual scarification confirmed physical dormancy (PY) in seeds of Pericopsis moonina. The majority of tropical lowland rainforests had non-dormant (ND) species (62%), and 14.3, 14.3, 7 and 2.3% of the species had MPD, PD, MD, and PY, respectively. Non-dormancy decreased for taxa from the upper strata to the lower strata of the forest. ND seeds were dispersed during the rainy season. Thus, non-dormancy seems to be the most dominant germination behaviour among the tree species in the lowland rainforest of Sri Lanka with the class of dormancy related to forest strata and dispersal time.