After-ripening and phytochrome action in seeds of dormant lines of wild oat (Avena fatua)

The effects of a short exposure to red, far-red or alternate red/far-red light on the germination of seeds after-ripened for different periods of time were studied in dormant lines of wild oat ( Avena fatua L.). Three stages were distinguishable in the after-ripening period in the response of germination to light. Seeds stayed dormant and showed no response to light during stage I. Phytochrome-mediated germination was observed in seeds during stage II. The phytochrome action disappeared during stage III, i.e. seeds fully germinated following treatments of all light qualities. When the seeds were imbibed in polyethylene glycol solutions, dark germination was reduced and phytochrome again had an effect, which suggested the involvement of phytochrome in water uptake of the seed.     

ATP Synthesis during Water Imbibition in Caryopses of Genetically Dormant and Non-dormant Lines of Wild Oat (Avena fatua L.)

Levels of ATP in dry caryopses of wild oats (Avena fatua L.)were much lower than in imbibed seeds of the seven geneticallypure lines surveyed. The ATP content of the lines with highgenetic dormancy was consistently lower than the ATP contentof genetically non-dormant lines, but no significant correlationwith depth of dormancy was found apart from this. Massive increasesin ATP content occurred within 30 min of water uptake by caryopsesof both dormant and non-dormant lines. The synthetic pathwaystudied utilized inorganic phosphate with great avidity to formATP. The ability to form ATP upon imbibition was present inboth embryo and de-embryonated caryopsis. The ATP levels attainedin imbibing caryopses appeared sufficient to support considerablesynthetic activity, and this reduced the possibility that adeficiency in ATP was responsible for the maintenance of dormancyin such imbibed seeds. The low levels of inorganic phosphatein the embryos of genetically dormant lines of wild oat couldrepresent a limiting factor, if the active formation of ATPupon water imbibition resulted in a scarcity of phosphate forother reactions essential to germination. Key words: Avena fatua, ATP synthesis, Inorganic phosphorus, Seed dormancy, Germination, Water uptake     

Embryo Dormancy and Light Requirements in the Germination of Helipterum craspedioides (Compositae), an Arid-zone Annual

Seeds of Helipterum craspedioides require light for germination,and this may be given while the seed is not fully imbibed. Removalof the seed coat, or leaching, does not bring about germinationin the dark. Gibberellin enhances dark germination to some extent,whether or not the seed coat is removed. The light requirementtherefore resides in the embryo. Fresh seed is dormant in lightor dark. Excised embryos of these dormant seeds grow slowly,rarely giving mature plants, but a larger number of normal plantsis produced after gibberellin treatment. Dormancy is lost afterhigh temperature storage, but not stratification.     

Role of Endogenous Growth Regulators in Seed Dormancy of Avena fatua: I. Short Chain Fatty Acids

The hypothesis that endogenous short chain fatty acids (C 6-C 10) are important in maintaining seeds of wild oat (Avena fatua L.) in the dormant state by acting as natural germination inhibitors (Berrie, Buller, Don, Parker, 1979 Plant Physiol 63: 758-764) was investigated. When germination of nondormant seeds was inhibited by treatment with short chain fatty acids, the seeds did not revert to a similar biochemical and physiological state as exhibited by dormant seeds. First, nonanoic acid-induced inhibition of seed germination was not reversed by hormone treatments which normally break dormancy in wild oat seeds. Second, nondormant seeds treated with short chain fatty acids maintained similar relative proportions of the pentose phosphate pathway and the Embden-Meyerhoff-Parnas pathway for respiratory glucose metabolism as that found in the nondormant controls. Seeds imbibed in the presence of nonanoic acid lost more amino acids and proteins into the imbibition solution than did the untreated controls, suggesting membrane damage had occurred. Inasmuch as increasing concentrations of nonanoic acid also progressively reduced the growth of the coleoptile and roots of intact seedlings until all growth ceased and no germination occurred, the inhibition of seed germination could be due to a nonspecific inhibition of growth of the embryo, perhaps because of disruption of membrane structure and function. Finally, no correlation between endogenous levels of short chain fatty acids in seeds or isolated embryonic axes and seed dormancy could be demonstrated.     

Localization of Glucose-6-phosphate Dehydrogenase Activity in Seeds and its Possible Involvement in Dormancy Breakage

A quantitative cytochemical analysis of glucose-6-phosphatedehydrogenase activity of deeply dormant seeds of Avena fatuashowed that although the enzyme activity is present in mostcell types of the embryo and seed, it is only in the embryothat activity is increased on treatment with GA₃ to break dormancy.This would appear to happen prior to any measurable embryonicaxis growth, and supports the idea that activation of the pentosephosphate pathway is an early event in dormancy break. A similar,though less marked, change occurred in less dormant seeds ofA. fatua, but could not be detected in dormant seeds of Lactucasaliva. Dry seeds of L. sativa and weakly dormant A. fatua containedtwice the activity seen in seeds imbibed with either water orGA₃, indicating that this might be a marker of low levels ofdormancy. Avena fatua, Lactuca sativa, seeds, dormancy, pentose phosphate pathway, cytochemistry, glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase     

Germination stimulation in wild oats (Avena fatua L.) by synthetic strigol analogs and gibberellic acid

At concentrations of 0.01–1 mM, five synthetic multiring analogs of strigol were effective germination stimulants of intact and dehulled wild oat (Avena fatua L.) seeds. The effect was concentration-dependent and equaled or exceeded that produced by equimolar gibberellic acid (GA₃). The most effective strigol analog treatments induced 55–80% germination within 7 days in intact wild oat seeds and resulted in 63–86% germination and normal seedling growth over 14 days. Intact wild oat controls germinated 14% after 14 days. The stimulation of wild oat germination by these synthetic strigol analogs demonstrates that these compounds, initially developed as germination stimulants for the seeds of the parasitic weed, witchweed (Striga asiatica L. Kuntz.), have bioregulatory activity in dormant seeds of monocots, as well as dicots. None of the compounds tested significantly affected the germination of nondormant cultivated oat seeds (Avena sativa L.). The commonly used dispersal agent, Tween 20 (0.1%), was found to inhibit germination of cultivated oats, alone and in the presence of 2% acetone.     

The Role of Germination Inhibitors and Oxygen in the Dormancy of the Light-densitive Seed of Betula spp.

Although unchilled, intact seeds of Betula pubescens and B.verrucosa require light for germination, isolated embryos germinateequally well in both light and darkness. An aqueous extract of these seeds has germination-inhibitoryproperties correlated with the presence of a non-fluorescent,single substance. The light requirement of isolated embryosis restored by the inhibitor. When intact seeds are leachedwith water to remove some inhibitor, it is found that the lightrequirement is reduced, short days and single light periodsthen eliciting greater germination than in unleached seeds. It has been found that scratching, pricking, and cutting theseed coat increases the germination of intact seeds in darkness,and that this is probably due to enhanced oxygen entry. Further,it has been found that germination in short days is increasedin oxygen-enriched atmospheres. It has been found that although the inhibitory effect of theseed coat in intact seeds is partially due to the reductionof the oxygen supply to the embryo, a low oxygen concentrationdoes not prevent germination of isolated embryos. Experimentalresults suggest that the inhibitor in the seed coat increasesthe oxygen requirement of the embryo.     

A Seed Coat Bedding Assay to Genetically Explore In Vitro How the Endosperm Controls Seed Germination in Arabidopsis thaliana

The Arabidopsis endosperm consists of a single cell layer surrounding the mature embryo and playing an essential role to prevent the germination of dormant seeds or that of nondormant seeds irradiated by a far red (FR) light pulse. In order to further gain insight into the molecular genetic mechanisms underlying the germination repressive activity exerted by the endosperm, a "seed coat bedding" assay (SCBA) was devised. The SCBA is a dissection procedure physically separating seed coats and embryos from seeds, which allows monitoring the growth of embryos on an underlying layer of seed coats. Remarkably, the SCBA reconstitutes the germination repressive activities of the seed coat in the context of seed dormancy and FR-dependent control of seed germination. Since the SCBA allows the combinatorial use of dormant, nondormant and genetically modified seed coat and embryonic materials, the genetic pathways controlling germination and specifically operating in the endosperm and embryo can be dissected. Here we detail the procedure to assemble a SCBA.     

Germination stimulation in wild oats (Avena fatua L.) by synthetic strigol analogs and gibberellic acid

At concentrations of 0.01–1 mM, five synthetic multiring analogs of strigol were effective germination stimulants of intact and dehulled wild oat (Avena fatua L.) seeds. The effect was concentration-dependent and equaled or exceeded that produced by equimolar gibberellic acid (GA₃). The most effective strigol analog treatments induced 55–80% germination within 7 days in intact wild oat seeds and resulted in 63–86% germination and normal seedling growth over 14 days. Intact wild oat controls germinated 14% after 14 days. The stimulation of wild oat germination by these synthetic strigol analogs demonstrates that these compounds, initially developed as germination stimulants for the seeds of the parasitic weed, witchweed (Striga asiatica L. Kuntz.), have bioregulatory activity in dormant seeds of monocots, as well as dicots. None of the compounds tested significantly affected the germination of nondormant cultivated oat seeds (Avena sativa L.). The commonly used dispersal agent, Tween 20 (0.1%), was found to inhibit germination of cultivated oats, alone and in the presence of 2% acetone.Names of companies or commercial products are given solely for the purpose of providing specific information; their mention does not imply recommendation or endorsement by the U.S. Department of Agriculture over others not mentioned.     

Nonstructural carbohydrates in dormant and afterripened wild oat caryopses

Nonstructural carbohydrates were determined in both embryo and endosperm of dormant (nongerminating) and afterripened (germinating) intact caryopses of wild oat ( Avena fatua L.). No changes in endosperm starch or soluble sugar were observed at the onset of germination (18 h). No changes in glucose, fructose, sucrose or starch within dormant or afterripened embryos correlated with onset of visual germination. In afterripened embryos, depletion of raffinose (18 h), stachyose (18 h) and galactose (24 h) was correlated with germination. In contrast, raffinose-family oligosaccharide levels in dormant embryos remained constant for 7 days following imbibition. Germination of isolated dormant embryos on 88 m M galactose-containing media was accompanied by decreased endogenous levels of raffinose and stachyose. Isolated embryos from dormant caryopses incorporated ¹⁴C from ¹⁴C-fructose into both raffinose and stachyose during 24 h of imbibition. In contrast, no ¹⁴C incorporation into stachyose was observed in embryos from afterripened caryopses. No ¹⁴C incorporation into raffinose was observed at 18 and 24 h. When in vitro activities of α galactosidase were measured, no temporal differences between dormant or afterripened caryopses were detected in either embryo or endosperm tissue. Although the mechanism associated with differences in utilization of raffinose and stachyose is yet unidentified, alterations in raffinose-family oligosaccharide metabolism in the embryo appear to be a unique prerequisite for afterripening-induced germination.     

Control of Seed Germination by Abscisic Acid: I. Time Course of Action in Sinapis alba L

The germination process of mustard seeds (Sinapis alba L.) has been characterized by the time courses of water uptake, rupturing of the seed coat (12 hours after sowing), onset of axis growth (18 hours after sowing), and the point of no return, where the seeds lose the ability to survive redesiccation (12 to 24 hours after sowing, depending on embryo part). Abscisic acid (ABA) reversibly arrests embryo development at the brink of radicle growth initiation, inhibiting the water uptake which accompanies embryo growth. Seeds which have been kept dormant by ABA for several days will, after removal of the hormone, rapidly take up water and continue the germination process. Seeds which have been preincubated in water lose the sensitivity to be arrested by ABA after about 12 hours after sowing. This escape from ABA-mediated dormancy is not due to an inactivation of the hormone but to a loss of competence to respond to ABA during the course of germination. The sensitivity to ABA can be restored in these seeds by redrying. It is concluded that a primary action of ABA in inhibiting seed germination is the control of water uptake of the embryo tissues rather than the control of DNA, RNA, or protein syntheses.     

Seed Coat Dormancy in Two Species of Grevillea(Proteaceae)

The role played by the seed coat in seed dormancy of Grevillealinearifolia(Cav.) Druce and G. wilsonii(A. Cunn.) was testedby a series of manipulations in which the seed coat was dissectedand removed, dissected and returned to the decoated seed, ordissected, removed and given a heat shock, and returned to thedecoated seed. Germination of intact seeds of both species wasalso examined after exposure to heat shock, smoke, or heat shockand smoke combined. Water permeability of the seed coat wasinvestigated by examining imbibition. For intact seeds, virtuallyno germination occurred under any treatment (G. wilsonii), orgermination was increased by exposure to either heat or smoke(G. linearifolia). Removal of the seed coat led to germinationof all decoated seeds for G. linearifolia, or a proportion ofdecoated seeds for G. wilsonii. Inclusion of smoked water inthe incubation medium led to a higher proportion of decoatedseeds germinating for G. wilsonii. Returning the seed coat,either with or without heat shock to the seed coat, did notsignificantly affect germination in either species. Seed coatswere permeable to water in both species. For the two Grevilleaspecies, there were different dormancy mechanisms that werecontrolled by the seed coat (G. linearifolia) or by both theseed coat and embryo (G. wilsonii). Copyright 2000 Annals ofBotany Company Grevillea linearifolia, Grevillea wilsonii, dormancy, seed coat dormancy, seed coat permeability, smoke, heat shock, germination     

Water Relations of Tomato Seeds During Imbibition and Osmotic Priming

Water uptake of tomato (Lycopersicon esculentum Mill. cv. Moneymaker) seeds during germination was obviously triphasic. The completion of the first phase of water uptake by whole seed could not be realized until 10~12 h later after sowing though varies in different parts of seed. The mechanical resistance of endosperm and seed coat restricted water uptake of the embryo envoleped by the endosperm. Water potential of the intact embryo was still 0. 6~0. 9 Mpa lower than the whole seed when the equilibrium between seed and imbibing solution was established. GA and ABA had no direct effects on the water uptake of tomato seeds. The water potential of embryo was positively correlated with its moisture content. The osmotic potential of tomato embryos decreased slowly during imbibition in water and osmotic solution as well.     

Differentiation of a Functional Aleurone Layer Within the Seed Coat of Sinapis alba L.

The hitherto unresolved ontogenetic origin of the aleurone layerin mustard (Sinapis alba L.) seeds was investigated with lightand electron microscopy. Contrary to previous views, this layerof storage cells is neither derived from the endosperm nor fromthe nucellus, but from a particular cell layer within the innerintegument of the seed coat. These cells differentiate and becomefilled with storage protein and fat concurrently with the maturationof the embryo. They survive seed desiccation and become depletedof storage materials during seed germination. Temporally correlatedwith the germinating embryo, the aleurone cells produce microbodyenzymes, which are controlled by light in a similar fashionin both tissues. Sinapis alba L., mustard, aleurone layer, seed coat, seed formation, germination     

Photoinhibition of radish (Raphanus sativus L) seed germination: control of growth potential by cell-wall yielding in the embryo

The biophysical mechanism underlying photoinhibition of radish (Raphanus sativus L.) seed germination was investigated using three cultivars differing in sensitivity to continuous irradiation with far-red light (high-irradiance reaction of phytochrome). Sensitivity of germination to the inhibitory action of light was assessed by probing germination under osmotic stress (incubation in media of low water potentials adjusted with polyethylene glycol 6000) and expressed in terms of ‘germination potential’ (positive value of the water potential at which germination is inhibited by 50%). Far-red light decreases the germination potential to various degrees in the different cultivars, reflecting the light-sensitivity of germination in water. Removal of the seed coat increases the germination potential by a constant amount in darkness and light. It is concluded that germination depends on the expansive force of the embryo which can be drastically diminished by far-red light. Seed-coat constraint and expansive force of the embryo interact additively on the level of the germination potential. Photoinhibition of germination was accompanied by an inhibition of water uptake into the seed. Analysis of seed water relations showed that osmotic pressure and turgor assumed higher levels in photoinhibited seeds, compared to seeds germinating in darkness, while the water potential was close to zero under both conditions. Far-red light produced a shift (to less negative values) in the curve relating water-uptake rate to external water potential, i.e. a reduction in the driving force for water uptake. It is concluded that photoinhibition of germination results from the maintenance of a high threshold of cell-wall extensibility in the embryo.     

Development of seed coat-imposed dormancy during seed maturation in Cynoglossum officinale

The relationship between seed phenolics and appearance of seed coat–imposed dormancy during seed development in Cynoglossum officinale L. was studied. Up to 24 days after anthesis, seeds failed to germinate upon imbibition in Petri dishes at 25°C. At 44 days after anthesis, seeds were fully germinable; removal of seed coats did not improve their germination or O₂ uptake. At 72 days after anthesis, mature seeds at the base of the cyme did not germinate unless their coats were removed. Removal of seed coat also stimulated O₂ uptake at this harvest date. The methanol-soluble phenolic content of the seeds increased during the early stages of seed development, in both the seed coat and the embryo. As seed development continued, the methanol-soluble phenolic content of the embryo stabilized, but that of the seed coat declined. This decline was associated with an increase in the thioglycolic acid–soluble phenolics, presumably lignins, in the seed coat. These results suggest that polymerization of methanol–soluble phenolics into lignins in the seed coat during later stages of seed development renders the seed coat of C. officinale impermeable to 03, and thus keeps the seed dormant.     

The Effect of pH on the Action of Respiratory Inhibitors in Avena fatua Seeds

The effects of pH on the action of sodium azide, a cytochome-oxidaseinhibitor, and salicylhydroxamate (SHAM), an alternative respirationinhibitor, on the respiration of dormant seeds of wild oat (Avenafatua L.; line AN-51) were studied. While pH had little effecton seed respiration in controls, it strongly affected the activityof azide. One mM azide inhibited seed respiration at pH5, butstimulated it at pH 7. SHAM (10 mM) completely inhibited thestimulation of respiration by 1 mM azide in an unbuffered medium,but failed to do so when the medium was buffered at pH 7. Inunbuffered medium, 10 mM SHAM actually augmented the stimulationof respiration by 0.25 mM azide to the same degree as when theazide solution was acidified to mimic the pH change incurredwith dissolution of 10 mM SHAM. These results suggest that theinhibitory effect of SHAM on the action of azide in an unbufferedsystem may in part be due to its acidification of the incubationmedium rather than by the inhibition of alternative oxidase.Lower pH favours the formation of the undissociated hydrazoatemolecules causing greater inhibition of cytochrome-oxidase andother azide-sensitive enzymes. Avena fatua L, wild oat, seed dormancy, azide, salicylhydroxamate     

Water Uptake and Protein Synthesis in Germinating Wheat Embryos under the Osmotic Stress of Polyethylene Glycol

The effect of osmotic stress on wheat-seed germination was testedby imbibition in aqueous polyethylene glycol solutions at differentconcentrations. The experiments were designed to allow blockingand the subsequent recovery of germination by 12 h or 24 h pre-imbibitionof seeds in osmoticum, followed by transfer to water. Seedswere alternatively presoaked in water for 12 or 24 h, then transferredto polyethylene-glycol solutions to study the induced blockingof germination. Water content and [³H]leucine incorporationinto embryo tissues (as a measure of in vivo protein synthesis)were determined over a 48-h imbibition period. A close relationshipwas established overall between hydration status and proteinsynthesis rate. Osmotic stress seems to have a strong influenceupon the quantitative synthesis of proteins, suggesting thatthis biochemical activity is associated with the regulationof the germination process. Triticum durum, embryo, osmotic stress, water uptake, protein synthesis     

Strophiole of seeds of the black locust acts as a water gap

In Japan the black locust (Robinia pseudoacacia L.) is undergoing rapid habitat expansion, which has an adverse effect on native vegetation. It is therefore a priority to clarify the regeneration characteristics of the black locust and establish adequate management of this invasive species in Japan. To determine the germination characteristics of physically dormant black locust seeds, we observed anatomical features of the seed coat and identified the water gap that acts as a signal detector. Our microscopic observations showed that seed coats of this species had hilum, micropyle and strophiole. The anatomical features of these regions correspond to the general characteristics of papilionoid legume seeds. Based on our microscopic observations, water absorption blocking experiments and a dye tracking experiment, we identified the strophiole as a water gap in black locust seeds. Our results suggest that the opening of the strophiole is important for water uptake to the embryo and subsequent germination of black locust seeds under natural conditions.     

Induction and release of secondary seed dormancy in genetically pure lines of Avena fatua

Induction and release of secondary dormancy in genetically pure dormant (AN-51, Mont 73) and non-dormant (CS-40, SH-430) lines of wild oat ( Avena fatua L.) were studied. These lines differed with regard to the optimal period of anaerobiosis necessary for induction of dormancy, and/or the degree (% of seeds acquiring dormancy) and duration of the dormancy induced. Secondary dormancy could be induced more effectively in the after-ripened seeds of dormant lines than in the non-dormant lines, where only a short-term dormancy could be induced (in 5–7 week-old-seeds). Higher anaerobiosis temperatures were more effective in inducing dormancy in all lines studied. Thus, as with primary dormancy, wild oat biotypes exhibit genetic variability in their secondary dormancy behaviour and factors like temperature can modify the expression of this trait.
The germination stimulants kinetin, isopentenyl adenine, sodium azide, potassium nitrate, ethanol and substituted phthalimides, which break primary dormancy in wild oats, stimulated germination of secondarily dormant seeds (line AN-51). Since these chemicals are structurally diverse, primary and secondary dormancies appear to be similar in part in their regulation.
Salicylhydroxamic acid, an inhibitor of cyanide-insensitive (alternative) respiration, did not inhibit: 1, spontaneous release of secondary dormancy in the line SH-430; and 2, stimulation of germination of secondarily dormant AN-51 seeds by various chemicals (except azide), suggesting that this respiratory pathway is not necessary for the release of induced dormancy.