Abscisic acid controls dormancy development and bulb formation in lily plantlets regenerated in vitro

Plantlets of lily regenerated in vitro from scale explants consist of scales and leaves from which the base of the petiole has swollen to a scale. Fluridone, an inhibitor of ABA-synthesis, applied during culture in vitro, inhibited the swelling of the petioles and promoted leaf formation. At high fluridone concentrations (10 or 33μ M ), swelling was completely blocked, and plantlets consisted of leaves only. Addition of ABA during the regeneration in vitro had the opposite effect and resulted in plantlets with scales only. When applied simultaneously with fluridone, ABA nullified the effect of fluridone. This demonstrates that bulb formation in lily is under the control of ABA. Lily plantlets regenerated in vitro on scale explants at 20 or 25°C were harvested after 11 weeks, and the leaves were removed from the bulblets. The bulblets were dormant and required a cold treatment to achieve rapid emergence after planting in soil. Fluridone added during the culture in vitro prevented the development of dormancy, and the bulblets did not require a cold treatment. The effect of fluridone was nullified by simultaneous addition of ABA. Bulblets harvested after 6 weeks of culture at 20°C had not yet developed dormancy. Bulblets regenerated at 15°C were only slightly dormant. In both types of bulblets, it is unlikely that the lack of dormancy was due to low ABA-levels since addition of ABA did not affect the dormancy status. These data indicate that the level of endogenous ABA and an unknown additional factor play major roles in the development of dormancy.     

Effect of low temperature on dormancy breaking and growth after planting in lily bulblets regenerated in vitro

Lilies regenerating on scale segments may develop dormancy in vitro depending on the culture conditions. The dormancy is broken by storage for several weeks at a low temperature (5 °C). The effect of the low temperature on sprouting, time of leaf emergence and further bulb growth was studied. Dormant and non-dormant bulblets were regenerated in vitro on bulb scale segments cultured at 20 °C or 15 °C, respectively. The low temperature not only affected the number of sprouted bulblets but also the time of emergence. The longer the cold storage, the faster and more uniform leaf emergence occurred. Both dormant and non-dormant bulblets grew faster after a low temperature treatment of six weeks. Thus, during dormancy breaking the tissue is prepared not only for sprouting but also for subsequent bulb growth. These processes are rather independent as low temperature stimulates growth in non-dormant bulblets whereas these bulblets sprout also without treatment at low temperature. Moreover, the hormone gibberellin induces rapid sprouting but has no influence on further bulb growth. Good growth in bulblets exposed to the low temperature coincided with production of an increased leaf weight. However, the relationship is not absolute as bulblets that were cold-treated for six weeks grew larger than bulblets cold-treated for four weeks but the formation of leaf biomass was similar. During storage at low temperature starch was hydrolyzed in the bulb scales and sugars accumulated. This indicates that during this period, preparation for later bulb growth involves mobilization of carbohydrate reserves which play a role in leaf growth and development of the photosynthetic apparatus. Starch hydrolysis proceeded in the outer scales after planting. Approximately six weeks later, the switch from source to sink took place in the bulblet, which became visible as a deposition of starch in the middle scales.     

Phase change in lily bulblets regenerated in vitro

During the development of the lily ( Lilium ), three phases can be distinguished: the juvenile, the vegetative adult and the flowering phase. Juvenile bulblets sprout with one or a few leaves whereas vegetative adult bulblets sprout with a stem with elongated internodes. The transition to the vegetative adult phase was studied in lily ( Lilium  × cv. Star Gazer) bulblets regenerating on bulb scale segments in vitro. The phase change was marked by the development of a tunica-corpus structure in the apical meristem which leads to the formation of an actively growing stem primordium. This structure is absent in juvenile bulblets. Juvenile bulblets first developed competence for phase change during a culture period of at least 6 weeks at 25°C. Subsequent induction of the phase change occurred during a period of 2 weeks at lower temperature (15°C). A major factor influencing phase transition was bulblet weight. Small bulblets never formed a stem whereas large bulblets always formed a stem under inducing conditions. Large bulblets more often formed a stem than small ones but the relation between bulb growth and phase transition was not absolute. A high sucrose concentration, a large explant and a prolonged period for competence development stimulated bulb growth but also phase transition independently of growth. Lowering the concentration of MS-minerals reduced bulb growth but did not affect phase transition. Under these conditions, phase change was correlated with a low phosphorus content.     

ABA content and sensitivity during the development of dormancy in lily bulblets regenerated in vitro

We measured ABA content and sensitivity in bulblels of Lilium speciosum Thunb , regenerating from scale explants in vitro at temperatures (15, 20 or 25°C) that allowed the development of various levels of dormancy (very low, intermediate or high, respectively). The one-step purification and the accuracy of the immunoassay were confirmed by HPLC and by liquid chromatography/mass spectrometry. ABA content was not correlated with dormancy development. Sensitivity to ABA was determined as the difference in sprouting performance of excised bulblets on medium with and without ABA. In bulblets regenerating at 20 or 25°C. ABA sensitivity was high during the period of dormancy establishment and decreased thereafter. Dormant hulblets were almost completely insensitive to ABA. The changes in sensitivity to ABA were confirmed by measuring the level of ABA in bulblets at the time of sprouting. This level was, as expected, highest in bulhlels with low ABA-sensitivity. Briefly cold-treated bulblets, in which dormancy may he re-established by culture at 20°C, again became sensitive to ABA. ABA sensitivity decreased with increasing temperature bulblets that regenerated at I5°C and hardly developed any dormancy, were very sensitive to ABA. It was concluded that in addition to ABA sensitivity another, still unknown, factor played a key role in dormancy development.     

Secondary dormancy in Avena fatua: Induction and characteristics in genetically pure dormant lines

The induction of secondary dormancy in caryopses of genetically pure dormant lines of Avena fatua L. is described. Seeds harvested from mature plants were after-ripened under controlled conditions (26°C, 25% relative humidity) until fully non-dormant. Secondary dormancy was then induced into these caryopses by incubation on moist filter papers in an aspirated nitrogen atmosphere at 20°C over periods from 3 h to 14 days. These caryopses failed to germinate when returned to an aerobic environment. The dose-response curves for gibberellic acid, sodium azide, sodium nitrite, sodium nitrate and ethanol show that all of these treatments can overcome the induced secondary dormancy. Drying increased the sensitivity of secondary dormant caryopses to these treatments. These treatments overcame secondary dormancy at all times, indicating the presence of only one of the two known blocks to germination that exist during primary dormancy. Similarities between primary and secondary dormancy in A. fatua are discussed.     

Role of reactive oxygen species in the regulation of Arabidopsis seed dormancy

Freshly harvested seeds of Arabidopsis thaliana, Columbia (Col) accession were dormant when imbibed at 25°C in the dark. Their dormancy was alleviated by continuous light during imbibition or by 5 weeks of storage at 20°C (after-ripening). We investigated the possible role of reactive oxygen species (ROS) in the regulation of Col seed dormancy. After 24 h of imbibition at 25°C, non-dormant seeds produced more ROS than dormant seeds, and their catalase activity was lower. In situ ROS localization revealed that germination was associated with an accumulation of superoxide and hydrogen peroxide in the radicle. ROS production was temporally and spatially regulated: ROS were first localized within the cytoplasm upon imbibition of non-dormant seeds, then in the nucleus and finally in the cell wall, which suggests that ROS play different roles during germination. Imbibition of dormant and non-dormant seeds in the presence of ROS scavengers or donors, which inhibited or stimulated germination, respectively, confirmed the role of ROS in germination. Freshly harvested seeds of the mutants defective in catalase (cat2-1) and vitamin E (vte1-1) did not display dormancy; however, seeds of the NADPH oxidase mutants (rbohD) were deeply dormant. Expression of a set of genes related to dormancy upon imbibition in the cat2-1 and vet1-1 seeds revealed that their non-dormant phenotype was probably not related to ABA or gibberellin metabolism, but suggested that ROS could trigger germination through gibberellin signaling activation.     

Study of the inheritance of seed primary dormancy and the ability to enter secondary dormancy in Petunia: influence of temperature, light and gibberellic acid on dormancy

Abstract. The germination behaviour of two Petunia hybrida lines. M₃₀ and Th₇, and their reciprocal hybrids was studied. Two sets of experimental conditions appeared helped to distinguish between dormant and non-dormant parental lines: (1) 25 and 35 °C in the dark, in the latter case after 2 months of dry storage at 20 °C; (2) 35 and 40 °C in the light. Photosensitivity was tested in the first case and sensitivity to GA₃ in the second case. The predominance of paternal control over dormancy was evident. A maternal or tegumentary control of photosensitivity and of sensitivity to GA₃ was also shown. Transferring the seeds, originally imbibed in conditions expressing primary dormancy, to conditions which previously supported their germination, allowed us to show that secondary dormancy could be easily induced when a deeper primary dormancy had already developed in the seeds.     

Effect of Methyl Jasmonate on Morphology and Dormancy Development in Lily Bulblets Regenerated In Vitro

Scales of lily bulbs are swollen petioles. Lily scale fragments cultured in vitro regenerate bulblets consisting of scales that may or may not carry a leaf blade. The bulblets are dormant and require a cold treatment to sprout. We added the gaseous plant growth regulator methyl jasmonic acid (MeJA) in the headspace of the tissue-culture container and studied the effect on plantlet morphology (scale/leaf-blade formation) and dormancy development in three lilies, Lilium speciosum “Rubrum No. 10,” L. longiflorum “Snow Queen,” and the Asiatic hybrid “Connecticut King.” Methyl jasmonic acid strongly reduced leaf-blade formation in Lilium longiflorum and Connecticut King. This was a specific effect as scale formation was affected much less. The specific inhibition of leaf-blade formation was not observed in Lilium speciosum. In this lily, high concentrations of methyl jasmonic acid (MeJA) inhibited leaf-blade and scale formation to similar extents. Methyl jasmonic acid reduced dormancy development in all three lilies, with the largest effect observed in Connecticut King. In this Asiatic hybrid, almost all bulblets that had regenerated at 300 or 1000 μl l⁻¹ MeJA in the headspace, did not require a dormancy-breaking treatment to achieve sprouting after planting in soil. Previously, it has been found in lily that treatments that reduce leaf-blade formation promote dormancy development. The present findings with MeJA do not agree with this. In the three lilies, the various parameters that were studied—regeneration, scale weight, leaf-blade weight, and dormancy development—were very differently affected by MeJA.     

Factors affecting the induction and release of secondary dormancy in Kalanchoë seeds

Abstract. Several short daily R irradiations are required from the first day of incubation on water to induce germination of Kalanchoë seeds. When the same light treatment is given after a prolonged dark incubation period at 20°C, secondary dormancy prevents germination. Factors controlling the induction and breaking of secondary dormancy have been investigated. The induction of secondary dormancy is very temperature dependent. Locally puncturing the seed coat strongly delays it. Secondary dormancy is not induced in the presence of GA₃ during the first 10 d of dark incubation, although this growth substance cannot induce dark germination. Prolonged or cyclic daily R irradiations can relieve secondary dormancy of seeds kept on water, even after a dark period of 20 d. A 24 h treatment at 4°C restores responsiveness to short R exposures of slightly secondarily dormant seeds. The synergism between GA₃ and Pfr in non-dormant Kalanchoë seeds, leading to high effectiveness of even one short FR irradiation, still occurs in seeds made secondarily dormant before transfer to GA₃, but more R or FR irradiations, in combination with GA₃, are required for the release of secondary dormancy. A combination of red light and 6-benzyl-aminopurine is ineffective in removing dormancy.     

Growth,sugar accumulation,and dark respiration of suspension cell cultures derived from contrasting alfalfa cultivars

Fall dormancy results in decumbent, slow shoot growth of alfalfa (Medicago sativa L.) in autumn and reduced shoot regrowth rates after herbage removal in summer. Although fall dormancy is used to predict alfalfa adaptation, we possess a poor understanding of the biological mechanisms underlying fall dormancy. Our objective was to examine growth and carbohydrate metabolism of suspension cell cultures derived from contrasting alfalfa cultivars that genetically differed in fall dormancy. Suspension cells were grown in B5h media containing 2% sucrose. Cells derived from fall non-dormant plants accumulated sugars more rapidly after transfer to fresh media and to higher concentrations than did cells derived from fall dormant alfalfa cultivars. Dark respiration rates of cells derived from non-dormant plants were similar to those derived from fall dormant plants when growth was limited at low cell sugar concentrations. However, both cell growth and dark respiration rates increased in cells derived from non-dormant cultivars in response to greater cell sugar concentrations. High growth rates of cells derived from rapid growing, fall non-dormant alfalfa cultivars were associated with rapid sugar uptake and higher cell respiration rates when compared to cells derived from dormant alfalfa cultivars.     

Breakage of Pseudotsuga menziesii seed dormancy by cold treatment as related to changes in seed ABA sensitivity and ABA levels

The main aims of the present work were to investigate whether a chilling treatment which breaks dormancy of Douglas fir ( Pseudotsuga menziesii (Mirb.) Franco) seeds induces changes in the sensitivity of seeds to exogenous ABA or in ABA levels in the embryo and the megagametophyte, and whether these changes are related to the breaking of dormancy. Dormant seeds germinated very slowly within a narrow range of temperatures (20–30°C), the thermal optimum being approximately 25°C. The seeds were also very sensitive to oxygen deprivation. Treatment of dormant seeds at 5°C improved further germination, and resulted in a widening of the temperature range within which germination occurred and in better germination in low oxygen concentrations. In dry dormant seeds the embryo contained about one-third of the ABA in the megagametophyte. ABA content of both organs increased during the first 4 weeks of chilling. It then decreased sharply in the megagametophyte to the level in the embryo after 7–15 weeks of chilling. At 15°C, a temperature at which dormancy was expressed, the ABA level increased in the embryo and the megagametophyte of dormant unchilled seeds whereas it decreased in the organs of chilled seeds. The longer the chilling treatment, the faster the decrease in ABA after the transfer of seeds from 5°C to higher temperatures, and the decrease was faster at 25 than at 15°C. These results suggest that the breaking of dormancy by cold was associated with a lower capacity of ABA biosynthesis and/or a higher ABA catabolism in the seeds subsequently placed at 15 or 25°C. Moreover, the chilling treatment resulted in a progressive decrease in the sensitivity of seeds to exogenous ABA. However, seeds remained more sensitive to ABA at 15 than at 25°C. The possible involvement of ABA synthesis and of responsiveness of seeds to ABA in the breaking of dormancy by cold treatment is discussed.     

The control of bud dormancy in potato tubers

Potato (Solanum tuberosum L.) tuber buds normally remain dormant through the growing season until several weeks after harvest. In the cultivar Majestic, this innate dormancy persisted for 9 to 12 weeks in storage at 10° C, but only 3 to 4 weeks when the tubers were stored at 2° C. At certain stages, supplying cytokinins to tubers with innately dormant buds induced sprout growth within 2 d. The growth rate was comparable to that of buds whose innate dormancy had been lost naturally. Cytokinin-treatment did not accelerate the rates of cell division and cell expansion in buds whose innate dormancy had already broken naturally. Gibberellic acid did not induce sprout growth in buds with innate dormancy. We conclude that cytokinins may well be the primary factor in the switch from innate dormancy to the non-dormant state in potato tuber buds, but probably do not control the subsequent sprout growth.Abbreviations tio ⁶ade 6-(4-hydroxy-3-methylbut-trans-2-enyl amino)purine, zeatin - tio⁶ado 6-(4-hydroxy-3-methylbut-trans-2-enyl amino)-9--D-ribofuranosyl purine, zeatin riboside     

Changes in cis-zeatin and cis-zeatin riboside levels and biological activity during potato tuber dormancy

The effects of postharvest storage duration and temperature on endogenous cis -zeatin ( cis -Z) and cis -zeatin riboside ( cis -ZR) levels in potato ( Solanum tuberosum L.) tubers were determined in relation to tuber bud dormancy. The tubers used in these studies were completely dormant for at least 81 days of storage. Thereafter, tuber bud dormancy diminished gradually and after 165 days of postharvest storage, the tubers were completely non-dormant. Immediately after harvest, endogenous levels of cis- Z and cis -ZR were approximately 25 pmol (g fresh weight)⁻¹ and 8 pmol (g fresh weight)⁻¹, respectively. In tubers exiting dormancy but stored at a growth-inhibiting temperature (3°C), endogenous levels of cis -Z rose over threefold after 25 days of storage and remained elevated for the duration of the study. Levels of cis -ZR remained essentially constant during this same period. In tubers transferred to a growth permissive temperature (20°C) prior to use, the rise in endogenous cis -Z was less dramatic and more protracted; increasing twofold after 53 days of storage. No change in cis -Z riboside content was observed in these tubers during this period. Dose-response studies using either cis -Z or trans -Z demonstrated a time-dependent increase in cytokinin sensitivity during postharvest storage. Immediately after harvest, dormant tubers were insensitive to both zeatin isomers. Thereafter, tubers exhibited a dose-dependent increase in premature sprouting following injection with either cytokinin isomer. After injection into dormant tubers, cis -[8-¹⁴C]-zeatin was metabolized primarily to adenine/adenosine and cis -Z riboside. Seven days after injection, less than 10% of the recovered radioactivity was associated with trans -ZR. These results are consistent with a role for endogenous cis -Z (and its derivatives) in the regulation of potato tuber dormancy.     

Abscisic acid-regulated responses of dormant and non-dormant embryos of Helianthus annuus: Role of ABA-inducible proteins

Embryos of Helianthus annuus L. became dormant 3 weeks after anthesis and their dormancy was lifted during storage in dry conditions. The objectives of this study were to investigate changes in the pattern of soluble proteins associated with the release of embryo dormancy. Sunflower dehydrins and group 3 late embryogenesis-abundant (LEA) proteins were studied in developing embryos. Three dehydrins (17, 21 and 26 kDa) and two group 3 LEA polypeptides (17 and 23 kDa) appeared during dormancy induction. Their levels remained steady until maturity. After imbibition, these polypeptides disappeared within 24 h except for the 23-kDa protein whose levels remained stable for a further 4 d, whatever the culture condition. Analysis of radiolabelled proteins by two-dimensional gel electrophoresis revealed that among dormancy-associated proteins other than dehydrin and group 3 LEA, several low molecular mass (18, 19, 20 and 21 kDa) proteins were expressed in dormant embryos but not detected in non-dormant embryos. After a treatment with fluridone, which inhibits ABA synthesis, or with GA₃, which allows germination to occur, the 19-kDa protein could not be detected. In contrast, application of ABA to non-dormant embryos arrested germination and enhanced the synthesis of the 18- and 21-kDa proteins, but not that of the 19- and 20-kDa polypeptides. These results demonstrate that steady-state levels of specific proteins change during early imbibition of dormant and non-dormant sunflower embryos and indicate that these changes may be associated with differential gene expression responsible for the maintenance of dormancy.     

Ecological genetics of seed germination regulation in Bromus tectorum L.

The probability that a seed will germinate depends on factors associated with genotype, maturation environment, post-maturation history, and germination environment. In this study, we examined the interaction among these sets of factors for 18 inbred lines from six populations of Bromus tectorum L., a winter annual grass that is an important weed in the semi-arid western United States. Seeds of this species are at least conditionally dormant at dispersal and become germinable through dry-afterripening under summer conditions. Populations and inbred lines of B. tectorum possess contrasting dormancy patterns. Seeds of each inbred line were produced in a greenhouse under one of three levels of maturation water stress, then subjected to immediate incubation under five incubation regimes or to dry storage at 20°C for 4 weeks, 12 weeks, or 1 year. Dry-stored seeds were subsequently placed in incubation at 20/30°C. Narrow-sense heritability estimates based on parent-offspring regressions for germination percentage of recently harvested seeds at each incubation temperature were high (0.518–0.993). Germination percentage increased with increasing water stress overall, but there were strong interactions with inbred line and incubation temperature. Inbred lines whose seeds were non-dormant over the full range of incubation temperatures when produced at low maturation water stress showed reaction norms characterized by little or no change as a function of increasing stress. For inbred lines whose dormancy status varied with incubation temperature, incubation treatments where seeds exhibited either very low or very high levels of dormancy showed the least change in response to maturation water stress. Inbred lines also varied in their pattern of dormancy loss during storage at 20°C, but maturation water stress had only a minor effect on this pattern. For fully afterripened seeds (1 year in storage at 20°C), inbred line and maturation water stress effects were no longer evident, indicating that differences in genotype and maturation environment function mainly to regulate dormancy and dormancy loss in B. tectorum, rather than to mediate response patterns of non-dormant seeds.     

Involvement of endogenous gibberellins in potato tuber dormancy and early sprout growth: a critical assessment

The role of endogenous gibberellins (GAs) in the regulation of potato (Solanum tuberosum) tuber dormancy was examined by determining: 1. changes in endogenous GA levels during natural dormancy progression, 2. the effects of GA biosynthesis inhibitors on tuber dormancy duration and 3. the dormancy status and tuber GA levels in a dwarf mutant of potato. The tubers (cv. Russet Burbank) used in these studies were still completely dormant after 98 days of storage. Between 98 and 134 days of storage, dormancy began to end and tubers exhibited limited (< 2 mm) sprout growth. Tuber dormancy weakened with further storage and tubers exhibited greater rates of sprout growth after 187 days of storage. Tubers stored for 212 days or longer were completely non-dormant and exhibited vigorous sprout growth. Immediately after harvest, the endogenous contents of GA19, GA20, and GA1 were relatively high (0.48-0.62 ng g fresh weight(-1)). The content of these GAs declined between 33 and 93 days of storage. Internal levels of GA19, GA20, and GA, rose slightly between 93 and 135 days of storage reaching levels comparable to those found in highly dormant tubers immediately after harvest. Levels of GA19, GA20, and GA1 continued to increase as sprout growth became more vigorous. Neither GA4 nor GA8 was detected in any tuber sample regardless of dormancy status. Dormant tubers exhibited a time-dependent increase in apparent GA sensitivity. Freshly harvested tubers were completely insensitive to exogenous GAs. As postharvest storage continued, exogenous GAs promoted premature dormancy release with GA1 and GA20 eliciting the greatest response. Injection of up to 5 microg tuber(-1) of kaurene, GA12, GA19 or GA8 had no effect on dormancy release. Sprout growth from non-dormant tubers was also promoted by exogenous GA in the following sequence of activity: GA1 = GA20 > GA19. Kaurene, GA12, and GA8 were inactive. Continuous exposure of developing tubers to inhibitors of GA biosynthesis (AMO-1618, ancymidol, or tetcyclasis) did not extend tuber dormancy but rather hastened dormancy release. Comparison of tuber dormancy and GA1 content in tubers of a wild-type and dwarf mutant of S. tuberosum ssp. andigena revealed a near-identical pattern of dormancy progression in spite of the absence of detectable levels of GA1 in tubers of the dwarf sibling at any time during dormancy progression. Collectively, these results do not support a role for endogenous GA in potato tuber dormancy release but are consistent with a role for GAs in the regulation of subsequent sprout growth.     

Induction of vivipary in Avena fatua

An investigation was conducted under controlled conditions to determine whether treatments designed to maximize the availability of water during seed development could induce viviparous germination in wild oats ( Avena fatua L.). Panicles of three genetic lines, which differed in their degree of dormancy, were kept in darkness at ca 100% RH and 20±1°C and were either supplied with water through the cut end of the rachis or left attached to the plant which was exposed to light. In the non-dormant line, germination of both primary and secondary caryopses on excised panicles increased with their stage of development when treated, i.e., 0, 5 and 10 days after anthesis. Germination of primary caryopses varied between 70 and 80% and was similar on both isolated and attached panicles treated at 10 and 5 days after anthesis, respectively. The percentage germination was considerably lower in all treatments of the two dormant lines and was inversely related to the genetically determined difference in their degree of dormancy. In these dormant lines germination was significantly lower on the intact plant than on the detached panicles. Water potential measurements suggested that this difference may be due partly to the transpiration-induced negative ψ_xyin the stem which may contribute to the inhibition of embryo growth and thus to the prevention of viparous germination.     

Seed Germination Biology of the Narrowly Endemic Species Lesquerella stonensis (Brassicaceae)

Abstract Lesquerella stonensis (Brassicaceae) is an obligate winter annual endemic to a small portion of Rutherford County in the Central Basin of Tennessee, where it grows in disturbed habitats. This species forms a persistent seed bank, and seeds remain viable in the soil for at least 6 years. Seeds are dormant at maturity in May and are dispersed as soon as they ripen. Some of the seeds produced in the current year, as well as some of those in the persistent seed bank, afterripen during late spring and summer; others do not afterripen and thus remain dormant. Seeds require actual or simulated spring/summer temperatures to come out of dormancy. Germination occurs in September and October. Fully afterripened seeds germinate over a wide range of thermoperiods (15/6–35/20°C) and to a much higher percentage in light (14 h photoperiod) than in darkness. The optimum daily thermoperiod for germination was 30/15°C. Nondormant seeds that do not germinate in autumn are induced back into dormancy (secondary dormancy) by low temperatures (e.g., 5°C) during winter, and those that are dormant do not afterripen; thus seeds cannot germinate in spring. These seed dormancy/ germination characteristics of L. stonensis do not differ from those reported for some geographically widespread, weedy species of winter annuals and thus do not help account for the narrow endemism of this species.     

Hypoxia interferes with ABA metabolism and increases ABA sensitivity in embryos of dormant barley grains

Two mechanisms have been suggested as being responsible for dormancy in barley grain: (i) ABA in the embryo, and (ii) limitation of oxygen supply to the embryo by oxygen fixation as a result of the oxidation of phenolic compounds in the glumellae. The aim of the present work was to investigate whether hypoxia imposed by the glumellae interferes with ABA metabolism in the embryo, thus resulting in dormancy. In dormant and non-dormant grains incubated at 20 degrees C and in non-dormant grains incubated at 30 degrees C (i.e. when dormancy is not expressed), ABA content in the embryo decreased dramatically during the first 5 h of incubation before germination was detected. By contrast, germination of dormant grains was less than 2% within 48 h at 30 degrees C and embryo ABA content increased during the first hours of incubation and then remained 2-4 times higher than in embryos from grains in which dormancy was not expressed. Removal of the glumellae allowed germination of dormant grains at 30 degrees C and the embryos did not display the initial increase in ABA content. Incubation of de-hulled grains under 5% oxygen to mimic the effect of glumellae, restored the initial increase ABA in content and completely inhibited germination. Incubation of embryos isolated from dormant grains, in the presence of a wide range of ABA concentrations and under various oxygen tensions, revealed that hypoxia increased embryo sensitivity to ABA by 2-fold. This effect was more pronounced at 30 degrees C than at 20 degrees C. Furthermore, when embryos from dormant grains were incubated at 30 degrees C in the presence of 10 microM ABA, their endogenous ABA content remained constant after 48 h of incubation under air, while it increased dramatically in embryos incubated under hypoxia, indicating that the apparent increase in embryo ABA responsiveness induced by hypoxia was, in part, mediated by an inability of the embryo to inactivate ABA. Taken together these results suggest that hypoxia, either imposed artificially or by the glumellae, increases embryo sensitivity to ABA and interferes with ABA metabolism.