The genus Nolana (Solanaceae) comprises numerous species endemic to the coastal Atacama Desert of Chile and Peru of high ornamental potential and conservation value. The environments in which these species have evolved and are present today correspond to particular conditions in the midst of a hyper-arid habitat, so the study of their germination requirements and characterisation of seed dormancy becomes important in terms of conservation but also for ecological and evolutionary purposes. Different treatments were performed on mericarps of 12 species of Nolana: control (intact seeds imbibed in distilled water), scarification in funicular plug and distilled water and scarification in funicular plug and addition of GA3 (500 ppm); their permeability to water was also tested. It was determined that the species did not present physical dormancy, as had been previously reported, but rather physiological dormancy (PD). Germination results after treatments were not homogeneous among all 12 species, indicating differences in their dormancy levels. Also, the important role of the endosperm in the prevention of germination for the studied Nolana species was highlighted. Regarding the relationship between the level of PD (expressed as the percentage of germination for the most successful treatment) and the latitudinal distribution of the species or their phylogenetic closeness, it was determined that, for the studied species, their proximity in terms of clades was more relevant than their latitudinal distribution.

The endosperm is the storage tissue of seeds and is an important source of nutrients for humans and animals. In the previous work, the gene expression was characterized at 3 and 24 h after pollination (AP). The results suggested that eATP would act as a signalling molecule at the beginning of endosperm development and that sucrose metabolism could be related to EBN insensitivity. In addition, differentially expressed transcripts derived fragments (DETDFs) were related to the failure of fusion of the polar nuclei and the accumulation of storage products in seeds of Arabidopsis thaliana. The objective of the present study was to identify genes related to endosperm development in apomictic and sexual ovaries of Paspalum notatum 48 h AP, a stage at which development is prior to post-zygotic collapse. The cDNA-AFLP analysis was carried out to analyse different crosses and DETDFs categorized according to their function. The main cellular functions at 48 h AP were metabolism and signal transduction. Fourteen out of 39 DETDFs with relevant functional information were found in crosses for which normal endosperm development was expected. Three DETDFs were found in crosses where viable and unviable seeds were predicted and presented similarity with a casein kinase II (CK2), an enzyme that governs the accumulation of storage proteins in seeds of A. thaliana and Zea mays. The results obtained at 3, 24 and 48 h AP suggest that CK2 is involved in early endosperm development in P. notatum.

In higher plants, the seed precursor (ovule primordia) is composed of three parts: funiculus, nucellus and chalaza, generating the latter one (II) or two (OI and II) protective maternal integuments (seed coat, SC). The appearance of a viable seed requires the coordinate growth and development of the preceding three compartments. Integuments are essentials for seed life as they nourish, protect and facilitate seed dispersion. Endosperm and integument growth and development are tightly coupled. Gymnosperm and angiosperm ovules are commonly unitegmic and bitegmic, respectively. Unusually, ategmy and threetegmy (OI, II and aril) also exist. The expression of the INO, ATS and ETT genes, involved in integument development, seems to have demonstrated that the fusion of OI and II leads to the appearance of unitegmy in higher plants. Likewise, INO expression also manifests the conservation of OI during evolution. The molecular control of SC development is constituted by a signalling network with still a multitude of gaps. The fertilization-independent development of the ovule is repressed by the FERTILIZATION INDEPENDENT SEED (FIS), a Polycomb-Repressive-Complex-2 (PRC2). Both endosperm and SC development are tightly linked to PRC2 function. As in many other developmental processes, auxin plays an essential role during ovule and SC development. Auxin transport from the endosperm to the integuments is regulated by AGL62 (AGAMOUS-LIKE 62), the encoding gene of which is specifically expressed in the endosperm to suppress its cellularization. In the absence of AGL62 (i.e. agl62 mutants), auxin remains trapped in the endosperm and the SC fails to develop (i.e. seed abortion). This update shows that auxin biosynthesis, transport and signalling play a predominant role and seem to be absolutely required in the pathway(s) that lead to SC formation, most likely not as a unique hormonal component.

Xenia is the direct effect of pollen genotype on the development and characteristics of the seed and fruit in the period that spans from fertilization to seed germination. Xenia effects cause phenotypic variations in the seed and fruit, which have importance for seed and fruit production but are not heritable to the progeny. Two hypotheses have been proposed as a mechanism for xenia: the hormonal hypothesis and the mobile mRNAs hypothesis. Although xenia effects have been studied in seeds and fruits in many crops, its effects and mechanism have been poorly studied at the molecular level. The aim of this work was to perform an initial screening of the xenia effect in the hybrid endosperm at the molecular level by differential gene expression among different pollen genotype sources from Paspalum notatum Flüggé. In order to characterize xenia effects and mechanisms, crosses were made between an emasculated mother plant with donors from two different pollen genotypes. RNA was isolated from ovaries 3 h after pollination. Some of the 24 differentially expressed transcript-derived fragments (DETDFs) provided relevant information. Four of those DETDFs were related to germination, pollen tube growth and pollen–pistil interaction. Seven DETDFs were associated with seed development and production. Finally, four DETDFs were predicted to encode for mobile mRNAs. These DETDFs might be involved in xenia effects and mechanisms in P. notatum.

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.

Two experiments were conducted to examine woolly cupgrass seed reserve utilization and the effect of germination depth on seed reserve utilization, tiller number, and timing of tiller emergence. The first experiment included germination in light with water, in light with Hoagland's solution, and in darkness with water; the second included five planting depths of 1, 3, 5, 7, and 9 cm. Endosperm utilization and shoot and root growth over time were recorded. Endosperms lost weight rapidly until 6 d after germination (DAG), when weight loss slowed down. Endosperm weight loss ceased 8 or 10 DAG. Endosperm weight loss was fastest for germination in light with Hoagland's solution. Deep planting significantly decreased emergence rate and increased the time to emergence. Differences in residual endosperms may have affected the early development of woolly cupgrass seedlings. Germination from a depth of 9 cm depleted the endosperm by the time of emergence. Deep planting also decreased shoot dry weight and tiller number 15 d after planting (DAP). Because the first tiller of woolly cupgrass usually appeared 10 to 12 DAP, and the endosperm had been depleted before that, there was no endosperm to support the new tillers.

Iron (Fe) and zinc (Zn) contents in hexaploid wheat are very low and are further reduced because of the removal of micronutrient-rich bran of wheat grains during milling and processing. Therefore, hexaploid wheat, its wild species and wheat–Aegilops kotschyi substitution lines were evaluated to identify the genome(s) carrying gene(s) for high Fe and Zn concentrations in bran and endosperm fractions of grains. It is reflected from the results that Triticum monococcum (acc. W463) may serve as a promising donor for biofortification of Fe, and Aegilops speltoides (acc. 3804) may serve as a promising donor for biofortification of Zn in the endosperm of cultivated wheat. Further, among the three wheat–Ae. kotschyi substitution lines, the higher concentration of Fe and Zn in endosperm fraction was observed in BC2F4 63-2-13-1. The work on precise transfer of useful gene(s) from 7Uk chromosome of this line is in progress to reduce linkage drag.

Recently, a list has been published of angiosperms capable of germinating in less than 24 h (‘very fast germination’). Here, we add three families and 23 species to that list. The main extra families complement ecological groups already recognized – the Cactaceae into the aridity-adapted group and the Tamaricaceae into the floodplain-adapted group. These amended findings on very fast germination (VFG) are integrated into the recent work on the functional ecology of embryo size. They confirm the important connection between germination speed and embryo to seed ratio. Plotting the plant orders containing VFG species on a phylogenetic tree shows that VFG has evolved multiple times throughout angiosperm history, including at least three times within the Caryophyllales. The fact that species with VFG are mainly restricted to advanced clades shows that VFG is a derived trait that evolved as an adaptation to either arid, saline or floodplain habitats.

The seed consists of three distinct regions, the zygotic embryo and endosperm and the maternal seed coat. Development of the seed requires co-ordinated growth and differentiation within each of these regions. Although the major events that occur during seed development have been studied extensively at the anatomical, physiological and molecular level, a comprehensive understanding of gene activity in each seed region has been lacking. DNA microarrays and extensive sequencing of cDNA libraries permit genome-wide analyses of mRNA populations of biological samples. We review recent studies of genome-wide gene activity that illustrate approaches used to provide new insights into the processes that occur during seed development.

Seeds of Medicago truncatula, a genomic model species for legumes, exhibit physiological and physical dormancy. Here, the factors influencing the germination behaviour of freshly harvested and stored seeds were investigated using several genotypes. Hardseededness is promoted when mature seeds are equilibrated at relative humidities (RH) below 75%. The release of physical dormancy during imbibition was dependent on the initial water content/RH that the seeds were dried to: the drier the seeds, the longer the imbibition time needed to break physical dormancy. The kinetics of physical dormancy release was slower than that of physiological dormancy, making it possible to uncouple both phenomena. Freshly harvested embryos without seed coverings germinated at the same speed as afterripened seeds. The depth of dormancy varied between different M. truncatula genotypes, from more to less dormant: DZA315.16>A17 (Jemalong)>R108>DZA45.5. This difference was eliminated by removing the endosperm. Collectively, these observations indicate that the endosperm is likely the main factor in the reduced germination of freshly harvested seeds. White light decreased germination speed of dormant seeds whereas it had no effect on non-dormant seeds. Recently harvested seeds were most dormant at temperatures above 17°C, whereas afterripened seeds germinated over a wider range of temperature. Fluridone could efficiently break dormancy, reinforcing the role of abscisic acid (ABA) synthesis. However, dormancy was not affected by gibberellic acid (100 μM GA3) or nitrate. The particular dormancy features unravelled here for M. truncatula, combined with the available genomic resources, make it a new, useful model for genetic and molecular studies which can complement those developed for Arabidopsis.

The quality of a seed (germination and vigour) is established during its development and maturation, but can be improved by post-harvest processing and pre-sowing treatments. During commercial seed production, maturity is usually estimated visually, relying on experience of the growers, but seed researchers are working to find molecular markers that can be applied easily to help in establishing optimal harvest time. One marker is cell cycle activity expressed as DNA replication in the seeds, analysed by flow cytometry. This fast and accurate method for the estimation of DNA content in plant nuclei allows detection of nuclei at different replication stages in different seed tissues and thus makes it possible to follow changes in the physiological state of a seed. DNA replication, as a late event during germination, can also be used to mark completion of germination and transition to early seedling growth. This information can be useful in the evaluation of seed quality and for following the advancement of priming. Flow cytometric analysis of ploidy can be also used as a basis for control of purity of some polyploid species seed lots.

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.

This review focuses on the possible evolutionary status and functional significance of recalcitrance and certain associated ovule/seed characters, within the framework of modern systems of angiosperm classification. The presence of recalcitrant seed viability in 45 dicotyledonous families is significantly associated with bitegmic and crassinucellate ovules and with nuclear endosperm development, all considered ancestral (plesiomorphic) character states of the ovule; as well as with greater seed size, woody habit and tropical habitat, also regarded as ancestral character states in the dicotyledons. In many species with recalcitrant seeds, the predominant storage reserve is carbohydrate. Recalcitrance is significantly associated with the exalbuminous type of reserve storage. It is proposed that in large recalcitrant seeds, the transfer of the main storage function from endosperm to embryo was probably an early development. In many species with recalcitrance, the ovules/seeds are characterized by extensive vascularization of the integument(s)/seed coat or by a pachychalaza. Pachychalazy is proposed to be a significant functional adaptation for a more efficient transfer of nutrients to the embryo/seed. Recalcitrance and some of the other character states proposed to be plesiomorphic in dicotyledons are also present in some gymnosperms, including presumed sister groups of the dicotyledons. In relatively advanced dicotyledonous families, mostly with orthodox seeds, recalcitrance probably persisted only in isolated relict members. Determination of character polarity is particularly problematic at lower taxonomic levels, because there is always the possibility that, in some taxa, a character state such as recalcitrance may have arisen secondarily as a reversal. Available evidence supports our view that seed recalcitrance can be regarded as a relatively ancestral character state in dicotyledons.

Galactomannan hydrolysis in isolated endosperms of fenugreek seeds is promoted by the presence of the embryo. Incubation in a large volume in the absence of the axis also results in endosperm mobilization and an increase in endo-β-mannanase activity. This is prevented when endosperms are incubated in a small volume, or when abscisic acid is present in the large volume. Fenugreek endosperms contain abscisic acid which is present in greater concentrations when these tissues have been incubated in a small volume rather than in a large volume. Antibodies prepared against purified tomato endo-β-mannanase detect the fenugreek enzyme on Western blots. Endosperms contain equal amounts of this enzyme whether they are incubated in a large volume, or in a small volume, or in the presence of abscisic acid. Thus, increased enzymic activity may not be related to increased synthesis, but rather to activation or release from the cells into the galactomannan-containing cell walls.

The rice (Oryza sativa L.) cv. Taichung 65, a japonica subspecies, was used to characterize the isoform, protein and gene expression patterns of endo-β-mannanase during and after seed germination. Activity assays and isoform analyses of whole grains or seed parts (scutellum, aleurone layer and starchy endosperm) revealed that seeds began to express endo-β-mannanase activity at 48 h from the start of imbibition at 25°C, after the completion of germination of most seeds. Three isoforms of endo-β-mannanase (pI 8.86, pI 8.92 and pI 8.98) were detected in the aleurone layer and starchy endosperm, but only two (pI 8.86 and pI 8.92) were present in the scutellum. The endo-β-mannanase in the starchy endosperm was mainly from the aleurone layer. Western blot analysis, using a tomato anti-endo-β-mannanase antibody, indicated that an endo-β-mannanase protein was present in an inactive form in dry grains. The amount of this protein decreased in the scutellum, but increased in the aleurone layer during and after germination. Thus, the increase in endo-β-mannanase activity in rice grains may be due to the activation of extant proteins and/or the de novo synthesis of the enzyme. Northern blot analysis showed that four putative rice endo-β-mannanase genes (OsMAN1, OsMAN2, OsMAN6 and OsMANP) were expressed in germinating and germinated rice grains. However, OsMANP was not expressed in the scutellum. The amount of OsMAN6 mRNA decreased after the completion of germination and paralleled the decline in endo-β-mannanase protein. In the aleurone layer, the increase of OsMAN2, OsMAN6 and OsMANP mRNA was prior to the increase of endo-β-mannanase protein.

Australia's native grass species contain a diverse array of wild cereal relatives which are adapted to a broader range of environmental conditions than current commercial cereals and may contain novel alleles which have utility in commercial production systems. Characterizing the available variation in endosperm morphology is one of the first steps towards in planta manipulation of endosperm by either the introgression of novel alleles or bioengineering cereal starch and protein. The endosperm of 19 crop wild relatives (CWR) was examined using scanning electron microscopy (SEM). Mature caryopses were fixed, dehydrated, critical-point dried and then snap fractured transversely through the grain. Wild relatives exhibited similar types of starch granules to that of their respective cultivated species, though in general the wild species retained a greater proportion of the endosperm cell wall at maturity. The two species examined with no closely related cultivated species exhibited a rice-like endosperm. Wild sorghum relatives exhibited an abundance of endosperm variations described as variations in starch granule size, shape and surface morphology, and the distribution of protein bodies. This is particularly important because the grain of Sorghum bicolor has inherently low starch and protein digestibility. These variations within the wild relatives of commercial cereals may provide novel sources of genetic diversity for future grain improvement programmes.

The fate of cells in the endosperm of developing wheat kernels was investigated under normal conditions and upon premature slow drying on the cut ear. To follow the changes in membrane integrity and cellular ultrastructure, an electron spin resonance (ESR) spin probe technique and low temperature scanning electron microscopy (LTSEM) were used. ESR data indicated that during development, the relative amount of cells with intact membranes decreased and became almost zero at the stage of mass maturity, i.e. several days before the onset of maturation drying. This suggests that the death of starchy endosperm cells is a developmental phenomenon rather than one induced by water loss. Even at 8 days after anthesis (daa), early in the differentiation phase, a considerable proportion of endosperm cells had already lost plasma membrane integrity. Comparison of ESR data with LTSEM micrographs revealed that the loss of plasma membrane integrity occurred early in the starch accumulation process, from which it was concluded that the programmes of starch accumulation and developmental death are simultaneously switched on. When the differentiation into starchy endosperm was arrested by premature slow drying of kernels on the cut ear, meristematic cells (aleurone initials) acquired desiccation tolerance as assessed by the combination of the spin probe technique and LTSEM. In contrast, fast drying caused immediate death of these cells. Thus, meristematic endosperm cells have the competence to acquire desiccation tolerance. This occurs upon premature slow drying or during normal differentiation into aleurone cells, but is lost when the cells differentiate into starchy endosperm.

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.

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.

The structural components and physical characteristics of sorghum kernels were studied as factors of resistance against Sitotroga cerealella (Oliv.) and Sitophilus oryzae (L.). Specifically, the effects of kernel glumes, weight, size and hardness on larval entry site, oviposition site, emergence site, progeny production, and kernel damage and weight loss, were investigated. The larvae of S. cerealella entered kernels primarily in the germ end and its periphery. Females of S. oryzae oviposited eggsmainly in the endosperm portion of kernels. Larval entry into kernels, and subsequent development of S. cerealella was enhanced in kernels that were enclosed by glumes. In addition, S. cerealella caused greater damage and weight loss in kernels with glumes. In contrast, kernels with glumes yielded less adult progeny of S. oryzae, and sustained less kernel damage and weight loss. The emergence holes of S. cerealella were located mainly in the crown end, whereas those of S. oryzae were mostly in the endosperm portion of kernels. The heavier and larger kernels produced heavier and larger adult progeny of both species of insect.