The Importance of Genetically Determined Seed Coat Characteristics to Seed Quality in Grain Legumes

Comparisons of five pairs of isogenk lines of peas, differingonly in the A gene for seed coat colour showed that white seeds(genotype aa) imbibed more rapidly than coloured seeds (AA),suffered greater imbibition damage revealed by dead tissue onthe cotyledons, and higher solute leakage. Seed-coat pigmentationwas closely associated with slow water uptake, since when expressionof the A gene was suppressed by the recessive pollens gene,the resulting white seeds {palpal AA) imbibed rapidly. The slowwater uptake by coloured seeds was not due to the restrictionof water entry by the seed coat since the differences in imbibitionrate were maintained when a portion of the seed coat was removedand seeds were imbibed with the exposed cotyledon in contactwith moist filter paper. Imbibition of similarly treated seedsby immersion in polyethylene glycol solutions (1–4%) whichincreased the seed/solution wettability, had little effect onthe water uptake of coloured seeds compared to imbibition inwater whereas that of white seeds increased in the first 10mins imbibition. Poor wettability of the inner surface of colouredseed coats did not therefore explain the slow imbibition ofthese seeds. The white seed coats loosened rapidly during imbibitionwhilst the coloured seed coats remained closely associated withthe cotyledons suggesting that the adherence of the seed coatto the cotyledons and therefore the ease of access of waterbetween the testa and cotyledons determines the rate of imbibition.The rapid water uptake by white-coated seeds and the subsequentimbibition damage may explain the high incidence of infectionof these seeds by the soil-bome fungus Pythhan after 2 d insoil. Improved seed quality and emergence may therefore be achievedby breeding for seed coat characteristics leading to reducedrates of imbibition Pisum sativum, isogenic lines, A gene, seed coat colour, imbibition, imbibition damage, wettability, pollens gene, seed quality, grain legumes     

Identification of defence proteins from the seed exudates of Cicer arietinum L. and its effect on the growth of Fusarium oxysporum f.sp. ciceri

Germinating seeds tend to release a variety of proteins into their surrounding surfaces; some of which have an inhibitory action against plant pathogens. The aim of this study was to investigate and identify defence proteins present in the exudates from water-imbibed and chitosan-imbibed (0.1% w/v) seeds of chickpea (Cicer arietinum L). Chickpea seeds imbibed in chitosan released a higher amount of proteins in the exudate when compared to the seeds imbibed in water. The obtained exudates were analysed in regard to specific protein activities by enzymatic assays and SDS-PAGE analysis. Results showed that the exude obtained from chickpea seeds imbibed in chitosan solution exhibited a new isoform of chitinase, chitosanase and protease inhibitors. These exudates also have an “in vitro” inhibitory effect on the growth of the fungus, Fusarium oxysporum f.sp. ciceri. Our results suggest that seed exudates protect seeds during their germination from soil pathogens.     

The Damaging Effect of Water on Dry Pea Embryos During Imbibition

When pea seeds were imbibed in water without their seed coats,vital staining revealed that cells on the abaxial surface ofthe cotyledons were dead. No damage occurred on the surfaceof cotyledons when the seeds were imbibed intact, or beneaththe testa when only half of the testa was removed. Cell deathoccurred as a result of rapid water uptake within the first2 min of imbibition, since reducing the rate of imbibition insolutions of Carbowax 4000 lessened the damage. Cell death wasrestricted to the outer layers of the cotyledons; inner tissuesremained alive. These observations supported the hypothesisthat rapid early leakage during imbibition of dry embryos resultedfrom the death of cells caused by the physical disruption ofmembranes. Imbibition damage resulted in reduced respirationand germination, a decline in the rate of food reserve transferfrom the cotyledons to the growing axis, and a lower growthrate in the seedlings produced. Greater sensitivity of embryosto imbibition damage at low temperature, and similarities betweenfeatures of imbibition damage and chilling injury led to thesuggestion that so-called chilling injury is the result of imbibitiondamage rather than the effects of low temperature.     

Release and Modification of nod-Gene-Inducing Flavonoids from Alfalfa Seeds

Traces of luteolin, an important rhizobial nod gene inducer in Rhizobium meliloti, are released by alfalfa (Medicago sativa L.) seeds, but most luteolin in the seed exudate is conjugated as luteolin-7-O-glucoside (L7G). Processes affecting the production of luteolin from L7G in seed exudate are poorly understood. Results from this study establish that (a) seed coats are the primary source of flavonoids, including L7G, in seed exudate; (b) these flavonoids exist in seeds before imbibition; and (c) both the host plant and the symbiotic R. meliloti probably can hydrolyze L7G to luteolin. Glycolytic cleavage of L7G is promoted by glucosidase activity released from sterile seeds during the first 4 hours of imbibition. Thus, L7G from imbibing alfalfa seeds may serve as a source of the nod-gene-inducing luteolin and thereby facilitate root nodulation by R. meliloti.     

Deep Sequencing of Maize Small RNAs Reveals a Diverse Set of MicroRNA in Dry and Imbibed Seeds

Seed germination plays a pivotal role during the life cycle of plants. As dry seeds imbibe water, the resumption of energy metabolism and cellular repair occur and miRNA-mediated gene expression regulation is involved in the reactivation events. This research was aimed at understanding the role of miRNA in the molecular control during seed imbibition process. Small RNA libraries constructed from dry and imbibed maize seed embryos were sequenced using the Illumina platform. Twenty-four conserved miRNA families were identified in both libraries. Sixteen of them showed significant expression differences between dry and imbibed seeds. Twelve miRNA families, miR156, miR159, miR164, miR166, miR167, miR168, miR169, miR172, miR319, miR393, miR394 and miR397, were significantly down-regulated; while four families, miR398, miR408, miR528 and miR529, were significantly up-regulated in imbibed seeds compared to that in dry seeds. Furthermore, putative novel maize miRNAs and their target genes were predicted. Target gene GO analysis was performed for novel miRNAs that were sequenced more than 50 times in the normalized libraries. The result showed that carbohydrate catabolic related genes were specifically enriched in the dry seed, while in imbibed seed target gene enrichment covered a broad range of functional categories including genes in amino acid biosynthesis, isomerase activity, ligase activity and others. The sequencing results were partially validated by quantitative RT-PCR for both conserved and novel miRNAs and the predicted target genes. Our data suggested that diverse and complex miRNAs are involved in the seed imbibition process. That miRNA are involved in plant hormone regulation may play important roles during the dry-imbibed seed transition.     

Protein synthesis in the embryo ofPinus thunbergii seed III.

Dissociability of the monomer ribosomes prepared from dry and imbibed pine (Pinus thunbergii) seed embryos was analyzed in sucrose density gradient containing a high salt buffer. Abnormal dissociation into the subunits was observed with the ribosome preparation from dry seed embryos when compared with that from imbibed seed embryos, i.e. each subunit peak was broader and localized at a lower site in sucrose density gradient. This indicates some change(s) in ribosomes during imbibition of seeds. These ribosomal changes also progressedin vitro. That is, after incubation of ribosome preparation from dry seed embryos in a high salt buffer for 5 min at 30 C or in a low salt buffer for 15 hr at 0 C, complete dissociation into the normal subunits was observed. No difference was found between polyacrylamide gel electrophoresis patterns of ribosomal RNA from dry and imbibed seed embryos. These results suggest some alteration in the protein components of ribosome during imbibition of pine seeds. This paper is dedicated to Prof. Shyogo Sawamura, Utsunomiya University on his retirement in March, 1979.     

聚乙烯醇预处理提高大豆种子活力和抗冷能力的作用

聚乙烯醇预处理大豆种子和聚乙二醇一样,能明显地提高种子的活力、萌发率和抗冷能力。其主要作用可能是:在整个吸胀过程中减少了吸水速度,使膜的修复作用有足够的时间;启动和改善种子内部的生理代谢过程.保证了生长胚根有更多的物质供应。     

Allelopathy due to purine alkaloids in tea seeds during germination

Summary During imbibition of whole tea seeds (6 days) two purine alkaloids, caffeine and theobromine, did not decrease in the seed coats and there was no increase in the seeds. In parallel with and after the breaking of seed coats there was a gradual release of caffeine from coats of germinating seeds. By contrast, when the seed was freed from the outer seed coat and soaked, imbibition of the seed required only 2 days and simultaneously caffeine was released from the inner seed coat. In such seeds, but not in whole seeds, growth of embryonic tissues (roots and shoots) was inhibited after the breaking of the inner seed coats. Nevertheless, caffeine increased more in such roots of the seedlings of decoated seeds than in roots of normal seedlings.     

苦马豆和披针叶黄华种子硬实特性与活力关系

将苦马豆和披针叶黄华种子在恒温25℃下吸胀,每24 h取出吸胀种子,16 d后未吸胀的种子为硬实种子（H）,硬实种子用硫酸处理后恒温吸胀24 h,与非硬实种子进行发芽试验和各项活力指标测定。结果显示每日内吸胀的种子数量随时间推移以一定比列下降,苦马豆非硬实种子第3天后吸胀率下降到1%,第13～16天突然上升后又下降到1%,披针叶黄华非硬实种子第3天后下降到1%,第9、10天突然上升后又下降到1%。两种豆类都显示出硬实种子发芽率、发芽指数、活力指数、脱氢酶活性、呼吸速率和超氧化物歧化酶（SOD）活性均高于非硬实种子,而电导率、浸出液可溶性糖和丙二醛（MDA）含量低于非硬实种子,缓慢吸胀的硬实种子活力指标高于快速吸胀的硬实种子,这表明硬实种子活力高于非硬实种子,硬实种子吸胀过程中存在吸胀损伤。而在非硬实种子中,根据以上活力指标判断,晚吸胀的种子比早吸胀的种子活力高。     

Imbibition of Swietenia macrophylla (Meliaceae) seeds: the role of stomata

BACKGROUND AND AIMS: The occurrence of stomata in seed coats is uncommon and there is limited information about their function(s). The aim of this study was to verify the distribution of stomata in seed coats of Swietenia macrophylla and to relate it to the imbibition process and aspects of the structure of the outer integument layers. METHODS: For the structural and ultrastructural studies, the seeds were processed using the usual techniques and studied under light and scanning electron microscopes. Histochemical tests were employed to identify the cell wall composition in the different seed coat portions. To assess the role of the stomata in the imbibition, non-impervious seeds were compared with partially impervious ones, in which only the embryo, median or hilar regions were left free. Further, the apoplastic pathway marker was employed to confirm the role of the stomata as sites of water passage during imbibition. KEY RESULTS: A positive relationship was observed between seed coat thickness and stomata density. The stomata were devoid of movement, with a large pore. They occurred in large numbers in the embryo region and extended with lower frequency towards the wing. Imbibition rates were related to stomata density, suggesting that the stomata act as preferential sites for water entry in the S. macrophylla seeds. CONCLUSIONS: At maturity, the stomata in the seed coat play a significant role in seed imbibition. The data may also infer that these permanently opened stomata have an important role in gas exchange during seed development, aiding embryo respiration.     

Translocation of iron from soybean cotyledons

Soybean seeds, Glycine max L. Merrill, were produced by plants treated from anthesis to seed maturity with ⁵⁹Fe supplied as ferric ethylenediaminedi (o-hydroxyphenylacetate). Seed coats accounted for 7.4% of dry seed weight and had Fe concentrations 5 times greater than the embryos. After germinating 2 days, cotyledons contained 69.6% and radicles 5.0% of original seed Fe. Fractions of seed Fe unavailable to seedlings were: 19.8% in seed coats, 1.7% in germination paper, 0.1% in the water under germinating seeds, and 3.8% unaccounted for. Every 3 days seedlings received nutrient solution without Fe or with 10 μm ferric ethylenediaminedi (o-hydroxyphenylacetate) and developed as deficient Fe or normal Fe plants. The deficient Fe cotyledons on day 18 retained 13% of the labeled Fe originally present. Cotyledons of normal Fe plants retained 50 to 70% of their original Fe. Moreover, cotyledons of the normal Fe plants accumulated externally supplied Fe and finally contained twice the quantity of Fe originally present. Stem exudate collected above cotyledons of deficient Fe plants contained 5.3 μm⁵⁹Fe. Electrophoresis of exudate showed that most of the ⁵⁹Fe migrated anodically as a single band and was in the position of ferric citrate.     

Seed dormancy of Cardiospermum halicacabum (Sapindaceae) from three precipitation zones in Sri Lanka

• This study investigated seed germination of Cardiospermum halicacabum, a medicinally important invasive species.
• We compared mass, moisture content (MC), dormancy and dormancy‐breaking treatments and imbibition and germination of scarified and non‐scarified seeds of C. halicacabum from a low‐elevation dry zone (DZ), low‐elevation wet zone (WZ1) and mid‐elevation wet zone (WZ2) in Sri Lanka to test the hypothesis that the percentage of seeds with water‐impermeable seed coats (physical dormancy, PY) decreases with increased precipitation.
• Seed mass was higher in WZ2 than in DZ and WZ1, while seed MC did not vary among the zones. All scarified DZ, WZ1 and WZ2 and non‐scarified DZ and WZ1 seeds imbibed water, but only a few non‐scarified WZ2 seeds did so. When DZ and WZ1 seeds were desiccated, MC and percentage imbibition decreased, showing that these seeds have the ability to develop PY. GA₃ promoted germination of embryos excised from fresh DZ and WZ1 seeds and of scarified WZ2 seeds.
• At maturity, seeds from DZ and WZ1 had only physiological dormancy (PD), while those from WZ2 had combinational dormancy (PY+PD). Thus, our hypothesis was not supported. Since a high percentage of excised embryos developed into normal seedlings; this is a low‐cost method to produce C. halicacabum plants for medicinal and ornamental purposes.

     

Control of seed dormancy in Nicotiana plumbaginifolia: post-imbibition abscisic acid synthesis imposes dormancy maintenance

The physiological characteristics of seed dormancy in Nicotiana plumbaginifolia Viv. are described. The level of seed dormancy is defined by the delay in seed germination (i.e the time required prior to germination) under favourable environmental conditions. A wild-type line shows a clear primary dormancy, which is suppressed by afterripening, whereas an abscisic acid (ABA)-deficient mutant shows a non-dormant phenotype. We have investigated the role of ABA and gibberellic acid (GA₃) in the control of dormancy maintenance or breakage during imbibition in suitable conditions. It was found that fluridone, a carotenoid biosynthesis inhibitor, is almost as efficient as GA₃ in breaking dormancy. Dry dormant seeds contained more ABA than dry afterripened seeds and, during early imbibition, there was an accumulation of ABA in dormant seeds, but not in afterripened seeds. In addition, fluridone and exogenous GA₃ inhibited the accumulation of ABA in imbibed dormant seeds. This reveals an important role for ABA synthesis in dormancy maintenance in imbibed seeds. Received: 31 December 1998 / Accepted: 9 July 1999     

Is The Barley Endosperm a Water Reservoir for the Embryo When Germinating Seeds Are Dried?

The water content of germinating seeds fluctuates in response to water potential changes in the surrounding environment. We tested the hypothesis that the endosperm functions as a water reservoir when imbibed seeds experience drying, and we characterized water uptake and movement within barley (Hordeum vulgare cv. Triumph) caryopses (hereafter referred to as seeds). Water movement into and through germinating barley seeds during imbibition and drying was determined gravimetrically and with the fluorescent dye trisodium 8-hydroxy-1,3,6-pyrenetrisulfonate (PTS). During imbibition, embryo tissues hydrated more rapidly and reached a higher water content (g H20/g dry weight) than did the endosperm, although the endosperm eventually contained nine times as much total water. When barley seeds that had imbibed for 12 h were exposed to moderate (-4 MPa) drying, PTS solution moved from the endosperm into the shoot meristem, radicle, and scutellum, but not vice versa. Radicle emergence and elongation proceeded for up to 8 h. With harsh (-150 MPa) drying, PTS concentrated almost exclusively in the radicle. These data illustrate that the endosperm is at least a temporary water storage compartment external to the embryo itself. We speculate that water supplied by the endosperm may be important in reducing the harmful effects of drying during the critical transition period when a germinating seed changes from a desiccation-tolerant to a desiccation-intolerant organism.     

Changes in Some ATP-Dependent Activities in Seeds during Treatment with Polyethyleneglycol and during the Redrying Process

During the imbibition of seeds in polyethyleneglycol (PEG),increasing amounts of ATP accumulated up to 24 h. Similar amountsaccumulated in the seeds during 4–5 h of imbibition inwater. Radioactive amino acids were increasingly incorporatedin the acid-insoluble fraction up to 24 h imbibition in PEG,as well as in water, after which a sharp decrease occurred upto 5 d of imbibition. If seeds were imbibed in PEG or waterin the presence of radioactive acetate, water-insoluble radioactivityincreased linearly in seeds during 5 d of imbibition. The amountsof incorporated amino acids or acetate were about double inPEG-imbibed as compared with in water-imbibed seed. The incorporationof AMP into the acid-insoluble fraction in seeds imbibed inPEG in the presence of radioactive AMP levelled off after 24h followed by a sharp decrease of up to 10% of the peak 5 dafter the start of imbibition. In water-imbibed seeds the incorporationof AMP continued to increase during at least 5 d of imbibition.During redrying of PEG-treated seeds (24 h), at least 80% ofthe accumulated ATP decreased during 18 d. The total radioactiveamino acids and nucleotide decreased during 3 d of redryingby 20% and 60%, respectively. At that time, the acid-insolubleincorporates increased by 20% and 50%, respectively. Some ofthe AMP was released as CO₂. Key words: AMP, Germination, Nucleic acid synthesis, Osmoconditioning, PEG, Protein synthesis     

Importance of methionine biosynthesis for Arabidopsis seed germination and seedling growth

Proteomics of Arabidopsis seeds revealed the differential accumulation during germination of two housekeeping enzymes. The first corresponded to methionine synthase that catalyses the last step in the plant methionine biosynthetic pathway. This protein was present at low level in dry mature seeds, and its level was increased strongly at 1-day imbibition, prior to radicle emergence. Its level was not increased further at 2-day imbibition, coincident with radicle emergence. However, its level in 1-day imbibed seeds strongly decreased upon subsequent drying of the imbibed seeds back to the original water content of the dry mature seeds. The second enzyme corresponded to S -adenosylmethionine synthetase that catalyses the synthesis of S -adenosylmethionine from methionine and ATP. In this case, this enzyme was detected in the form of two isozymes with different p I and M r. Both proteins were absent in dry mature seeds and in 1-day imbibed seeds, but specifically accumulated at the moment of radicle protrusion. Arabidopsis seed germination was strongly delayed in the presence of dl -propargylglycine, a specific inhibitor of methionine synthesis. Furthermore, this compound totally inhibited seedling growth. These phenotypic effects were largely alleviated upon methionine supplementation in the germination medium. The results indicated that methionine synthase and S -adenosylmethionine synthetase are fundamental components controlling metabolism in the transition from a quiescent to a highly active state during seed germination. Moreover, the observed temporal patterns of accumulation of these proteins are consistent with an essential role of endogenous ethylene in Arabidopsis only after radicle protrusion.     

The effect soaking pea seeds with or without seedcoats has on seedling growth

Pea seeds (Pisum sativum L. `Alaska') with intact seedcoats (WC) and with seedcoats removed (WOC) were soaked in distilled water for 24 hours at 20°. The water, containing the pea diffusate, was decanted after the second, fourth, sixth, eighth, twelfth, and twenty-fourth hour and analyzed for total nitrogen, α-amino nitrogen, carbohydrate, and total solute dry weight. The seeds were germinated at 20° in a 16 hour photoperiod of 300 foot candles. Stem lengths and dry weights of roots, shoots and cotyledons were determined after 4, 11, and 18 days of growth. WOC seeds imbibed more water than WC seeds during the 24 hour imbibition period. Diffusates from WOC seeds always contained more solute than diffusates from WC seeds. Maltose, glucose, and fructose were not detected in the early diffusates from WOC seeds but were found in WC seed diffusates at all times. Seedlings from WC seeds had longer stems than those from WOC seeds. The dry weight of stems and roots of WC seedlings was greater than those from WOC seedlings. The dry weight of cotyledons from 18 day-old WC seedlings was less than from WOC seedlings. Water absorption by WC seeds was slower than by WOC seeds. Removal of the seedcoat allowed rapid imbibition resulting in seed injury presumably because of the loss of solutes which included monosaccharides, disaccharides, amino acids, and other nitrogen containing compounds. These results are consistent with the hypothesis that rapid imbibition disrupts membrane organization leading to reduction of seedling growth.