Oilseed crops are major sources of oils for human nutrition, and an increasing proportion is also being utilized for industrial purposes. Recent advances in our understanding of the basic biochemistry of seed oil biosynthesis, coupled with identification of genes for oilseed modification, have set the stage for the genetic engineering of oilseed crops that produce ‘designer’ plant seed oils tailored for specific applications. In this review we provide an overview of seed oil biosynthesis and highlight the enzymatic steps that have already been targeted for genetic manipulation, with the end goal of producing seed oils containing desired amounts of fatty acid components. Furthermore, we describe the identification of genes from various wild plant species that are capable of producing structurally diverse fatty acids, and how these advances open the door to the production of entirely novel oils in conventional oilseed crops. Transgenic oilseeds producing high amounts of these novel fatty acids represent renewable sources of raw materials that may compete with, and eventually replace, some petrochemicals that are derived from non-renewable crude oil.

This paper discusses how field and laboratory experiments, using a variety of genetic material, can be combined to investigate the genetic basis of germination under realistic ecological conditions, and it reviews some of our recent work on germination phenology of Arabidopsis thaliana in the field. Our results indicate that the genetic basis of germination depends on the environment. In particular, the conditions during seed maturation interact with post-dispersal environmental factors to determine germination phenology, and these interactions have a genetic basis. Therefore genetic studies of germination need to consider carefully the environment – both during seed maturation and after dispersal – in which the experiments are conducted in order to characterize genetic pathways involved with germination in the field. Laboratory studies that explicitly manipulate ecologically relevant environmental factors can be combined with manipulative field studies. These studies can identify the particular environmental cues to which seeds respond in the field and characterize the genetic basis of germination responses to those cues. In addition, a variety of genetic material – including mutant and transgenic lines, intact natural genotypes, recombinant genotypes, and near isogenic lines – can be used in field studies as tools to characterize genetic pathways involved in germination schedules under natural ecological conditions.

Higher plants have several strategies to perpetuate themselves under adequate ecophysiological conditions. The production of heterogeneous seeds is one such strategy. That is, to ensure the survival of the next generation, an individual plant might produce seeds that are heterogeneous with respect to the extent of dormancy, dispersion and persistence within the seed bank. Heterogeneity can affect not only certain physiological and molecular properties related to seed germination, but also such characteristics as colour, size and shape, parameters commonly used to differentiate morphs within a heterogeneous seed population. In heterogeneous seeds, the above features determine seed behaviour and alter their mechanism of germination. In this work, emphasis is placed on the existence of seed mutants having major alterations in characteristics of the testa and hormonal response. These mutants constitute a valuable tool for elucidating the mechanism of dormancy, germination and perpetuation of seeds. Finally, ontogeny and heterogeneity are reviewed, providing the first data related to the possible hormonal control of heterogeneity in seeds. These results raise the hypothesis that one of the factors triggering differences in germination among heterogeneous seeds may be an alteration in the signalling and action mechanism of ethylene and abscisic acid (ABA).

Seeds of different species are believed to have characteristic shelf lives, although data confirming this are scarce, and a mechanistic understanding of why this should be remains elusive. We have quantified storage performance of c. 42,000 seed accessions, representing 276 species, within the USDA National Plant Germplasm System (NPGS) collection, as well as a smaller experiment of 207 cultivars from 42 species. Accessions from the NPGS collection were harvested between 1934 and 1975, and had relatively high initial germination percentages that decreased at a variable rate during storage at both 5 and –18°C. Germination time courses, which represent the average performance of the species, were fitted to Avrami kinetics, to calculate the time at which germination characteristically declined to 50% (P50). These P50 values correlated with other longevity surveys reported in the literature for seeds stored under controlled conditions, but there was no correlation among these studies and seed persistence observed in the classic buried seed experiment by Duvel. Some plant families had characteristically short-lived (e.g. Apiaceae and Brassicaceae) or long-lived (e.g. Malvaceae and Chenopodiaceae) seeds. Also, seeds from species that originated from particular localities had characteristically short (e.g. Europe) or long (e.g. South Asia and Australia) shelf lives. However, there appeared to be no correlation between longevity and dry matter reserves, soluble carbohydrates and parameters relating to soil persistence or resource allocation. Although data from this survey support the hypothesis that some species tend to survive longer than others in a genebank environment, there is little information on the attributes of the seed that affect its storage performance.

As seed dormancy is released within a seed population, both the rate and percentage of germination increase progressively with increasing dose of a dormancy-breaking treatment or condition. Population-based models can account for this behaviour on the basis of shifting response thresholds as dormancy is alleviated. In particular, hydrothermal time analysis of germination sensitivity to water potential (Ψ) and temperature (T) can describe these features of seed behaviour. We used the hydrothermal time model to analyse the effects of dormancy-breaking treatments on germination of dormant true (botanical) potato (Solanum tuberosum L.) seeds (TPS). After-ripening (37°C and 4% seed moisture content) of TPS for 7 or 30 days partially or fully alleviated primary dormancy. The median base water potential required to prevent germination [Ψb(50)] decreased from –0.25 MPa in control seeds to –0.87 MPa and –1.83 MPa after 7 and 30 days of after-ripening, respectively. In contrast, the base temperature for germination (Tb) was relatively unaffected (0–3.3°C). Fluridone (50 μM), an inhibitor of abscisic acid (ABA) biosynthesis, also promoted germination of dormant TPS and lowered Ψb(50), indicating a role for de novo synthesis of ABA during dormancy maintenance. Moist chilling (3 days at 4°C) or gibberellin (100 μM) alleviated secondary dormancy and lowered Ψb(50) values from –0.08 MPa to –0.36 and –0.87 MPa, respectively. The hydrothermal time model allows quantification of dormancy levels and explains why changes in germination speed and percentage are closely correlated during dormancy alleviation.

Although viable seeds of Mediterranean dry-fruited shrubs are found in herbivore dung, the ecological importance of this observation is still not well understood. We analysed seed retrieval percentages, defecation time and germinability after sheep gut passage for the five most common shrub species of an area in central Spain (Retama sphaerocarpa, Cytisus scoparius, Halimium umbellatum subsp. viscosum, Cistus ladanifer and Lavandula stoechas subsp. pedunculata). Five ewes were fed seeds, and their dung was collected regularly during the following week. Seeds were hand-sorted from dung subsamples and tested for germinability. The defecated seeds were clustered in time, with a majority retrieved in the 24–40 h period, although over 1% of the seeds were retained in the gut for more than 72 h. Data suggested a possible link between seed size and retrieval, with medium-sized seeds less damaged (16–23%) than larger and smaller seeds (10–12%), although only a small number of species were studied. Germination results showed an increased percentage of germination after gut passage for H. umbellatum (x2 test, P<0.05) and a marginally significant difference for C. scoparius (P<0.1). Soft-seeded L. stoechas did not germinate after gut passage. The results indicate a potential role of herbivore endozoochory for the long-distance dispersal of dry-fruited shrubs and their potential colonization of distant sites.

The most often used time-line for distinguishing a transient seed bank from a persistent seed bank is one calendar year. Thus, species whose seeds live in or on the soil for <1 year have a transient seed bank, whereas those whose seeds live for ≥1 year have a persistent seed bank. However, dormancy cycling of seeds buried in soil has not been given due consideration in these models. When dormancy cycling is considered, it is shown that seeds of both autumn-germinators and spring-germinators are in the dormant state when they are 1 year old. Thus, unless the seeds live until at least the second germination season (i.e. usually 16–18 months following dispersal), they are, in effect, part of a transient seed bank, having lived through only one germination season. We propose that for seeds of such species to be considered part of a short-term persistent seed bank, they should remain viable and germinable until at least the second germination season, and to be part of a long-term persistent seed bank, until at least the sixth germination season. Our definitions are applicable to seeds with physiological, physical or morphophysiological dormancy, which often require >1 year after maturity to come out of dormancy in nature. We discuss modifications of the seedling emergence method for detection of a soil seed bank, so that they correspond to our definitions of seed-bank strategies.

The unique feature of the seed of tetraploid cotton (Gossypium hirsutum and Gossypium barbadense) is that about 30% of the seed coat epidermal cells develop into cellulose-enriched fibres, while the embryos synthesize oils and proteins. Hence, both the maternal and filial tissues of the cotton seed are of significant economic value. After initiation from the ovule epidermis at or just before anthesis, the single-celled fibres elongate to 2.5–6.0 cm long in the tetraploid species before they switch to intensive secondary cell wall cellulose synthesis. Thus, apart from its agronomic importance, the cotton fibre represents a model single-cell system to study the control of cell differentiation and elongation, carbon partitioning to cellulose synthesis and also the interaction between maternal (fibre) and embryonic tissues in seeds. Over the past decade or so, significant effort has been made to understand the cellular and molecular basis of cotton fibre development and oil biosynthesis in the embryo. Metabolic engineering of the oil biosynthetic pathway in cotton seed has successfully produced healthier and stable oils. A number of candidate genes and cellular processes that potentially regulate various aspects of fibre development have been identified. Further elucidation of the in vivo functions of those candidate genes could significantly deepen our understanding of fibre development and offer potential for improvement of fibre quality through genetic engineering or marker-assisted breeding approaches.

When the laboratory methods employed are qualitative, the statistical methodologies used in testing for the adventitious presence (AP) of transgenic material in conventional seed and grain lots are well defined. However, when the response from the method used by the laboratory is quantitative (e.g. percent transgenic DNA), the statistical methodologies developed for qualitative laboratory methods are not fully appropriate. In this paper, we present the details of procedures specific to quantitative laboratory methods. In particular we consider: (1) the assessment of quantitative laboratory method errors using linear modelling; and (2) the process of deciding whether or not a lot meets pre-specified purity standards, including the development of probability calculations needed to develop operating characteristic curves and estimate consumer and producer risks for a given lower quality limit (LQL), acceptable quality limit (AQL) and testing plan. We also describe implementation of this approach in a useful spreadsheet application.

Seeds of Carduus acanthoides, Cynara cardunculus, Cirsium vulgare, Brassica campestris, and Sisymbrium altissimum were incubated at a range of decreasing osmotic potentials (Ψo) under fluctuating temperatures or the median temperature of the fluctuation cycle. Fluctuating temperatures promoted total seed germination in water and at reduced osmotic potential. Total germination was reduced as the Ψo decreased. However, this trend was smallest under fluctuating temperatures, signalling a higher tolerance of seeds to reduced osmotic potential. Effects of osmoticum and temperature were modelled with the hydrotime model. The parameters estimated from the model, the hydrotime constant (θH), the mean base water potential Ψb(50) and its standard deviation (σΨb) gave good descriptions of germination time courses. For all species, incubation under fluctuating temperatures shifted Ψb(50) values downwards without modifying their distribution substantially. This accounted for the greater tolerance of germination to reduced Ψo under fluctuating temperatures. To confirm that these effects were mediated by temperature fluctuations per se, the behaviour of C. acanthoides and C. cardunculus incubated at the minimum, the mean and the maximum temperature of the fluctuation cycle was also analysed. Constant maximum and minimum temperatures of the cycle did not stimulate germination, nor did they shift Ψb(50) towards more negative values. The hydrotime model provides a physiologically based quantitative description for germination promotion due to fluctuating temperature.

This paper describes the dispersal–germination characteristics of seeds of 77 native tree species in a seasonal monsoon forest in Uttaranchal state, northern India. Results indicate that 50% of the species dispersed in the hot, dry summer months, 18% during the rainy season, 23% during the cold season and the remainder in late spring. Germination tests on fresh and laboratory-stored seeds revealed a relationship between morphological features of the fruit and both germination percentage and mean germination time (MGT). Highest mean germination (50%) was for dry-dehiscent fruits with winged wind-dispersed seeds, followed by dry-dehiscent fruits with non-winged seeds (38%) and seeds of dry-indehiscent fruits (37%). Lowest germination (29%) was for seeds from fleshy or pulpy fruits. MGT followed the reverse course. Germination data for seeds stored dry in the laboratory during one seeding cycle indicated six patterns of seed germination: (1) average germination percentage of fresh seeds lower than that of stored seeds, indicating an after-ripening requirement; (2) initial high germination percentage followed by low values, indicating a steep to moderate decline in viability following harvest; (3) no germination after 1 or 2 months of seed storage, due to complete loss of viability, indicating short seed longevity; (4) fresh seed germination in some species equalled the average germination value of stored seeds, indicating constant germination for one whole seeding cycle; (5) germination of both fresh and stored seeds remained consistently low throughout the season, indicating a requirement for some kind of pre-treatment or having poor quality of seeds; (6) initial high germination followed by decline and again increase, showing a seasonal rhythm of germination.

This review focuses mainly on eudicot seeds, and on the interactions between abscisic acid (ABA), gibberellins (GA), ethylene, brassinosteroids (BR), auxin and cytokinins in regulating the interconnected molecular processes that control dormancy release and germination. Signal transduction pathways, mediated by environmental and hormonal signals, regulate gene expression in seeds. Seed dormancy release and germination of species with coat dormancy is determined by the balance of forces between the growth potential of the embryo and the constraint exerted by the covering layers, e.g. testa and endosperm. Recent progress in the field of seed biology has been greatly aided by molecular approaches utilizing mutant and transgenic seeds of Arabidopsis thaliana and the Solanaceae model systems, tomato and tobacco, which are altered in hormone biology. ABA is a positive regulator of dormancy induction and most likely also maintenance, while it is a negative regulator of germination. GA releases dormancy, promotes germination and counteracts ABA effects. Ethylene and BR promote seed germination and also counteract ABA effects. We present an integrated view of the molecular genetics, physiology and biochemistry used to unravel how hormones control seed dormancy release and germination.

Direct sowing is the simplest method of plant establishment for restoration and remediation purposes, but relatively few plants can establish under high salinity conditions. In this study, the ability of different seashore plants and grass cultivars to germinate in different dilutions of seawater (0–400 mM NaCl) was tested. Highest germination was found in distilled water or seawater dilutions up to 100 mM NaCl. When seawater concentrations were increased from 100 to 200 mM NaCl, a strong decline in germination percentage and rate was observed in less salt-tolerant species, such as Matricaria maritima and Achillea millefolium. The more salt-tolerant species, Plantago maritima, Juncus gerardii, Artemisia vulgaris, Agrostis spp. and Rumex spp., had a threshold salinity, where germination was significantly decreased in seawater dilutions between 200 and 400 mM NaCl. Even among the salt-tolerant species, only two, Agrostis stolonifera and Artemisia vulgaris, germinated at 400 mM. Variation in salinity response was observed among populations of Artemisia vulgaris and among cultivars of Festuca spp. Increasing salinity to 200 mM NaCl delayed germination in most species. Ungerminated seeds of most salinity-tolerant species were still viable after 21 d at the highest salinity (400 mM), and showed a rapid and high germination when transferred to distilled water. These species would be able to survive high salinity and germinate when the salinity of the sediments decreases through dilution or leaching of salts. The experiment revealed species and cultivars that will be of interest in further testing for restoration and remediation in saline habitats.

We undertook a comparative study of molecular changes during development of seed coats in the wild-type and a recessive hull-less mutant of pumpkin (Cucurbita pepo L.), with the goal of identifying key genes involved in secondary cell wall development in the testa. The mature mutant testa has reduced amounts of cellulose and lignin as compared to the wild type. The expression patterns of several genes involved in secondary cell wall biosynthesis during the development of the testa are described. These genes are: CELLULOSE SYNTHASE, PHENYLALANINE AMMONIA-LYASE, 4-COUMARATE-CoA LIGASE, and CINNAMOYL-CoA REDUCTASE. Additionally, the expression patterns of a few genes that were differentially expressed in the two genotypes during testa development (GLUTATHIONE REDUCTASE, ABSCISIC ACID RESPONSE PROTEIN E, a SERINE-THREONINE KINASE, and a β-UREIDOPROPIONASE) are presented. The results show a coordinated expression of several genes involved in cellulose and lignin biosynthesis, as well as marked differences in the level of their expression between the two genotypes during testa development. There is generally a higher expression of genes involved in cellulose and lignin biosynthesis in the wild-type testa as compared to the mutant. The molecular data presented here are consistent with anatomical and biochemical differences between the wild-type and the mutant testae. An understanding of the genes involved in cell wall development in the testa will facilitate the manipulation of seed coat development in Cucurbita and other species for diverse commercial applications.

The enhanced laboratory and field emergence characteristics of osmo- and halo-primed tomato seeds (cv. Pusa Ruby) were related to changes in hydration–dehydration kinetics, modified sorption properties and nuclear magnetic resonance (NMR) relaxation behaviour of humidified and imbibed seeds. Water sorption isotherms were constructed for primed and unprimed seeds by equilibrating to different water activities (aw) at 25°C. Analysis of the isotherms by the D'Arcy–Watt equation revealed that priming reduced the number of strong binding sites and the associated water content, and increased significantly the number of weak binding sites and the associated water content. This redistribution of water, which increased the availability of seed water, may be the reason for the higher speed of germination of primed seeds. The changes in transverse relaxation time (T2) of seed water and its components, measured in vivo using nuclear magnetic resonance spectroscopy, showed interesting differences between primed and unprimed seeds. With an increase in humidification time, the T2 of primed seeds could be resolved into three components with varying mobilities, while the control seeds had only two components until 10 d of humidification. During imbibition, the third component appeared after 2 and 6 h in primed and control seeds, respectively. This component disappeared after the germination process started in all treatments. The third fraction, with very low molecular mobility, which accounted for about 40% of the proton population, was assigned to hydration water of macromolecules. Hence, we propose that better performance of primed seeds may be attributed to the modifications of seed water-binding properties and reorganization of seed water during imbibition, so as to increase the macromolecular hydration water required for various metabolic activities related to the germination process.

Although jasmonates (JAs) are involved in germination and seedling development, the regulatory mechanism of JAs, and their relation with endogenous level modifications in these processes, is not well understood. We report here the detection of 12-oxo-phytodienoic acid (OPDA), jasmonic acid (JA), 11-hydroxyjasmonate (11-OH-JA), 12-hydroxyjasmonate (12-OH-JA) and methyljasmonate (JAME) in unimbibed seeds and seedlings of tomato Lycopersicon esculentum Mill cv. Moneymaker (wild type) and tss1, tss2, tos1 mutants. The main compounds in wild-type and tss1, tss2, tos1 seeds were the hydroxylate-JAs; 12-OH-JA was the major component in dry seeds of the wild type and in tss2 and tos1. The amounts of these derivatives were higher in seeds than in seedlings. Changes in JAs during wild-type and tss1 imbibition were analysed in seeds and the imbibition water. In wild-type imbibed seeds, 11-OH-JA content was higher than in tss1. 12-OH-JA showed a different tendency with respect to 11-OH-JA, with high levels in the wild type at early imbibition. In tss1, levels of 12-OH-JA rose from 24 to 48 h of imbibition. At 72 h of imbibition, when radicles had emerged, the amounts of both hydroxylates in wild-type and tss1 seeds were minimal. An important release of the hydroxylate forms was observed in the imbibition water. 11-OH-JA decreased in the imbibition water of wild-type seeds at 48 h. On the contrary, a high and sustained liberation of this compound was observed in tss1 after 24 h. 12-OH-JA increased in wild-type as well in tss1 until 24 h. Thereafter, a substantial reduction in the content of this compound was registered. NaCl-treated wild-type seedlings increased their 12-OH-JA, but tss1 seedlings increased their JA in response to salt treatment. In tss2 seedlings, NaCl caused a slight decrease in 11-OH-JA and JAME, whereas tos1 seedlings showed a dramatic OPDA and 12-OH-JA decrease in response to salt treatment. Under salt stress the mutant seedlings showed different patterns of JAs according to their differential hypersensitivity to abiotic stress. The JA-hydroxylate forms found, and the differential accumulation of JAs during germination, imbibition and seedling development, as well as their response to NaCl stress, provide new evidence about the control of many developmental processes by JA.

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.

Physical dormancy (PY) was identified in six genera representative of Australian Rhamnaceae and subsequently was broken, based on identification of key seed dormancy characteristics: (1) isolation and classification of embryo features; (2) imbibition experiments to determine the rate and amount of water uptake in seeds; and (3) determination of optimum temperature regimes for germination. All six species had relatively large spatulate embryos. Imbibition studies showed all species possessed PY (i.e. a water-impervious seed coat) that was broken by a hot-water treatment. Alleviation of PY resulted in high germination (<70%) at 7/18°C, temperatures similar to winter in south-west Western Australia. Germination was suppressed at higher temperatures and in the presence of light. The study adds information to our knowledge of seed dormancy in Australian Rhamnaceae, and highlights the benefits of understanding dormancy states in seeds prior to evaluating dormancy-release mechanisms on wild species used in restoration ecology and horticulture.

Grains of indica rice (Oryza sativa cv. Peiza 67) exhibit an increase in endo-β-mannanase activity, mostly after the completion of germination. According to tissue blots, the initial increase occurs in association with the embryo, and possibly the scutellum, although the largest sustained increase in activity is in the peripheral regions of the endosperm. The aleurone layer, being the only living region of the endosperm, is presumably the site of synthesis and secretion of the enzyme into the non-living, starch-laden region. β-Mannosidase activity is low throughout germination and subsequent seedling growth, particularly in the endosperm regions. Its activity profile does not mimic that of endo-β-mannanase. In the intact grain, gibberellin (GA) causes a relatively small increase in endo-β-mannanase activity, while abscisic acid (ABA) causes a large decrease; this inhibition is overcome to a considerable extent when GA is supplied along with ABA. β-Mannosidase activity is little affected by either GA or ABA. Embryoless half-grains imbibed in water exhibit only a small increase in endo-β-mannanase activity with time of imbibition, showing the necessity for a stimulus from the embryo for this to occur. Incubating half-grains in the presence of GA results in a large increase in enzyme activity; ABA reduces the amount of activity compared to the water controls. GA is capable of reversing the inhibitory effect of ABA with respect to endo-β-mannanase activity. As in the intact grains, β-mannosidase activity in the half-grains is unaffected by either GA or ABA. It is concluded that the major site for the production of endo-β-mannanase activity is the aleurone layer, and this event is influenced by the presence of the embryo; in the absence of the latter, the increase in enzyme activity is stimulated by GA. β-Mannosidase activity is low throughout germination and post-germination, it is not influenced by GA and ABA, and thus its activity is not regulated in a coordinated manner with that of endo-β-mannanase.

MicroRNAs (miRNAs) play critical roles in the development of animals and plants. Characterizing the stage- and tissue-specific expression of miRNAs that potentially regulate target transcription factor expression is becoming more important for understanding the regulatory mechanisms of critical events during plant development. A simple method for purifying small RNAs from seeds is described, as well as an efficient non-radioactive labelling system for making miRNA probes. In Arabidopsis thaliana seed extracts, low molecular-weight (LMW) RNAs (e.g. 5S rRNA, tRNA and miRNA) were separated from high molecular-weight (HMW) nucleic acids (e.g. 28S and 18S rRNA, mRNA and genomic DNA) by fractionation using isopropanol. HMW RNAs precipitated in 20% isopropanol, while most LMW RNAs remained in the supernatant. The purified LMW RNAs were used successfully for RNA gel blotting to detect miRNAs expressed in Arabidopsis and tomato (Lycopersicon esculentum) seeds. To increase the detection sensitivity of the microRNA probes, additional digoxigenin-labelled uridine triphosphates (UTPs) were incorporated into the miRNA probes by designing template oligo DNAs with three extra adenines (A) at each end of their sequence. These DNA oligomers were used to make double-stranded DNA templates for miRNA probe synthesis. This probe (termed AAAPLUS) exhibited stronger signals than normal probes. A technique was also developed to quickly screen expressed miRNAs in seeds using a miniblot system, which enabled simultaneous examination with multiple miRNA probes. This method provides a simple alternative to microRNA microarrays to identify the major miRNAs expressed in seeds.