Seed-coat dormancy in Grevillea linearifolia: little change in permeability to an apoplastic tracer after treatment with smoke and heat

BACKGROUND AND AIMS: Seeds of Grevillea linearifolia germinate following fire, and have seed-coat dormancy broken by smoke and heat shock. Smoke breaks seed coat dormancy in Emmenanthe penduliflora by altering the permeability of the seed coat to an internal germination inhibitor, which subsequently escapes. This model was tested for in G. linearifolia by investigating the permeability of the seed coat to diffusion of high-molecular-weight compounds, and whether this changed after exposure to fire cues. METHODS: Germination response of the seeds to heat shock, smoke or heat + smoke was tested. Penetration of Lucifer Yellow dye into intact seeds was examined after 24 and 48 h of exposure, and penetration of the dye from the inside of the seed coat outwards was examined after 24 h. Histochemical staining with Nile Red and Acridine Orange was used to locate cuticles, suberin and lignin. KEY RESULTS: Twenty-three per cent of untreated seeds germinated; heat shock and smoke increased germination additively up to approx. 80 % for both cues combined. Lucifer Yellow did not penetrate fully through the seed coat of untreated seeds, whether diffusing inwards or outwards. Three barriers to diffusion were identified. Treatment with heat or smoke slightly increased penetration of the dye, but did not completely remove the barriers. Suberin was identified in secondary walls of exotestal and mesotestal cells, and was absent from primary cell walls. Movement of Lucifer Yellow occurred through the middle lamella and primary cell wall of suberized cells; movement of the dye was impeded where suberin was absent. CONCLUSIONS: Fire cues did not significantly decrease barriers to diffusion of high-molecular-weight compounds in the seed coat of Grevillea, and must be breaking dormancy by another mechanism.     

Fire and flood: Soil‐stored seed bank and germination ecology in the endangered Carrington Falls Grevillea (Grevillea rivularis,Proteaceae)

Abstract Seed germination is dependent on the interaction between the dormancy state of a seed and the presence of favourable environmental conditions. Thus, the spectacular pulse of seedling recruitment in many Australian vegetation communities following disturbances such as fire can be attributed to changes in microsite conditions and/or the dormancy‐breaking effect of the disturbance on accumulated seed banks. Grevillea rivularis is a threatened species endemic to the area immediately above Carrington Falls in the NSW Southern Highlands. Most of the population is confined to the riparian vegetation zone in woodland and heath, and is therefore subject to periodic disturbance from fire and flood. For this species, a pulse of seedling recruitment has been recorded after fire, flood and mechanical soil disturbance. The aims of this study were to examine the density and vertical distribution of the soil‐stored seed bank and to investigate the role of heat and scarification as cues for germination of fresh and soil‐stored seed. There was a large seed bank under the canopies of established individuals (194 ± 73 seeds m^?2) and most seeds were found in the 0–2 cm and leaf‐litter layers of the soil profile. The germination response of soil‐stored and fresh seed was examined using a hierarchical series of laboratory experiments. Seeds of G. rivularis showed marked dormancy polymorphism. Thirty‐six percent of soil‐stored seed germinated without treatment, whereas no untreated fresh seeds germinated. Scarification or heating caused significant germination of dormant soil‐stored seed, but only scarification resulted in germination of dormant fresh seeds. These results highlight important differences in the dormancy state of soil‐stored and fresh seed. Thus, being a riparian species in a fire‐prone environment, the dormancy mechanisms in seeds of G. rivularis suit this species to disturbance by both fire and flood.     

Seed anatomy and water uptake in relation to seed dormancy in Opuntia tomentosa (Cactaceae, Opuntioideae)

BACKGROUND AND AIMS: There is considerable confusion in the literature concerning impermeability of seeds with 'hard' seed coats, because the ability to take up (imbibe) water has not been tested in most of them. Seeds of Opuntia tomentosa were reported recently to have a water-impermeable seed coat sensu lato (i.e. physical dormancy), in combination with physiological dormancy. However, physical dormancy is not known to occur in Cactaceae. Therefore, the aim of this study was to determine if seeds of O. tomentosa are water-permeable or water-impermeable, i.e. if they have physical dormancy. METHODS: The micromorphology of the seed coat and associated structures were characterized by SEM and light microscopy. Permeability of the seed-covering layers was assessed by an increase in mass of seeds on a wet substrate and by dye-tracking and uptake of tritiated water by intact versus scarified seeds. KEY RESULTS: A germination valve and a water channel are formed in the hilum-micropyle region during dehydration and ageing in seeds of O. tomentosa. The funicular envelope undoubtedly plays a role in germination of Opuntia seeds via restriction of water uptake and mechanical resistance to expansion of the embryo. However, seeds do not exhibit any of three features characteristic of those with physical dormancy. Thus, they do not have a water-impermeable layer(s) of palisade cells (macrosclereids) or a water gap sensu stricto and they imbibe water without the seed coat being disrupted. CONCLUSIONS: Although dormancy in seeds of this species can be broken by scarification, they have physiological dormancy only. Further, based on information in the literature, it is concluded that it is unlikely that any species of Opuntia has physical dormancy. This is the first integrative study of the anatomy, dynamics of water uptake and dormancy in seeds of Cactaceae subfamily Opuntioideae.     

Anatomical and chemical characteristics of the seed coat of <i>Medicago sativa</i> L. (alfalfa) cv. Baralfa 85 seeds and their association with seed dormancy

Seeds of alfalfa (Medicago sativa L.) can exhibit seedcoat imposed dormancy, which produces hard seeds within a seed lot. These seeds do not germinate because they do not imbibe water due to a barrier to water entry in the seed coat. The aim of this work was to analyze the anatomical and chemical characteristics of the testa of alfalfa seeds with respect to water permeability levels. The anatomy of seeds of the cv. Baralfa 85 was studied and structural substances, polyphenols, tannins and cutin present in the testa of seeds of different water permeability levels were determined. The anatomical characteristics of the seed coat and the proportions of components were found to determine the permeability level of the seed coat, an aspect that is associated with the physical seed dormancy level. Anatomically, increased thickness of the testa was associated with a lower permeability level. The difference may be attributed to the variation in cuticle thickness, length of macrosclereids and thickness of the cell wall, and presence and development of osteosclereids. From the physiological and chemical points of view, the mechanism of physical dormancy of the testa is explained by a greater amount of components that repel water and cement the cell wall, such as polyphenols, lignins, condensed tannins, pectic substances, and a lower proportion of cellulose and hemicellulose.     

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     

The size,distribution and germination requirements of the soil-stored seed-bank of Grevillea barklyana (Proteaceae)

Abstract Fire-triggered release from seed dormancy is a characteristic of many Australian plant species. We investigated aspects of the seed-bank dynamics and dormancy characteristics in seeds of Grevillea barklyana, an understorey shrub of coastal sclerophyll vegetation in the Jervis Bay Region on the south coast of New South Wales. We used two soil core sizes to compare the number and distribution of stored seeds in soil cores taken from underneath and outside the limits of plant canopies at three study sites. Core size did not affect the estimate of seed density. No seeds were found outside the existing canopies. Even in under-canopy samples, seed numbers were small. Mean seed densities were estimated as 10. 9, 14. 1 and 4. 3 seeds per for the three sites. A hierarchical series of laboratory experiments was used to test the germination response of both fresh and soil-stored seeds. Likewise, we attempted to simulate ‘natural’ disturbance conditions in a glasshouse experiment. Both laboratory and glasshouse experiments indicated polymorphism in germination behaviour. A constant proportion of seeds exhibited enforced dormancy, when moist at room temperature, while a smaller number of seeds showed either an induced dormancy or a non-seed-coat linked innate dormancy. The majority (75%) of seeds were innately dormant due to a hard seed-coat. This dormancy was broken when the seed-coat was damaged, for instance, by heat. The level of polymorphic germination behaviour will be dependent on the length of the inter-fire periods. We conclude that the expression of polymorphism within and between species across a range of environments and fire regimes is an important consideration for any further study attempting to assess the role of the seed-bank.     

种壳厚度对鼠类种子扩散行为及种子命运的影响

种子扩散是植物更新和扩大分布区的一种重要途径。鼠类采取不同的种子扩散和贮藏策略,以应对食物短缺,同时也促进了植物种子扩散。为应对鼠类对植物种子的过度取食,种子进化出了一系列物理、化学等防御特征。其中种壳厚度作为一种物理防御策略,是影响鼠类贮藏行为和种子命运的关键因素。本研究拟通过去除天然栓皮栎（Quercus variabilis）种子的外壳,再在种仁外包被1、2、4、6不同层数的聚乙烯薄膜,模拟种壳厚度,准确控制种壳厚度。2020年10月—2021年1月,在四川都江堰森林生境中释放人工种壳包被的种子,研究人工种壳厚度对鼠类介导的种子扩散和命运的影响。结果表明：（1）鼠类优先扩散种壳较薄（1层薄膜包被）的人工种子;随着种壳厚度的增加,扩散速率逐渐降低,种壳最厚（6层薄膜包被）的种子扩散最慢（P < 0.001）;（2）鼠类喜好分散贮藏1层、2层薄膜包被的种子;当种壳厚度增加至包被4层、6层薄膜时,分散贮藏比例显著降低（P < 0.05）;（3）鼠类偏好集中贮藏4层薄膜包被的种子（P < 0.05）;（4）不同种壳厚度的种子扩散距离无显著差异（P > 0.05）;（5）种壳较薄（1层薄膜包被）的种子分散贮藏率在3 m范围内比例较高。采用聚乙烯薄膜包被是模拟种子外壳的可行方法,并可用于评估种壳厚度对鼠类种子贮藏行为和种子命运的影响等相关研究。     

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     

Development and structure of the seed-coat of Lannea discolor (Sonder) Engl. (Anacardiaceae)

VON TEICHMAN, I., 1988. The development and structure of the seed-coat of Lannea discolor (Sonder) Engl. (Anacardiaceae). The bitegmic, anatropous ovule contains a group of nucellar cells with slightly thickened and intensively staining cell walls. Besides this hypostase sensu stricto, the nucellus cells in the chalaza become tanniniferous. This tanniniferous chalazal-nucellar tissue is intially plate-like. It is referred to as the hypostase sensu lato. The latter and the chalaza enlarge significantly. The raphe, extensive chalaza and well-developed cup-like hypostase sensu lato play an important role in the development of the seed-coat. The inner, tanniniferous epidermis of the inner integument persists in parts of the mature seed-coat. The outer, distinctly tanniniferous epidermis of the outer integument shows in the mature seed-coat a degree of secondary wall thickening. This undifferentiated type of seed-coat of L. discolor (tribe Spondieae) is remarkably similar to that of Camnosperma minor (tribe Rhoideae), both also showing tendency towards the exotestal type. In the Rhoideae the endotestal, i.e. differentiated type, of seed-coat is also present. The exalbuminous seed of L. discolor represents a derived and advanced type.     

Phylogeny of seed dormancy in Convolvulaceae, subfamily Convolvuloideae (Solanales)

Background and Aims: The water gap is an important morphoanatomical structure inseeds with physical dormancy (PY). It is an environmental signaldetector for dormancy break and the route of water into thenon-dormant seed. The Convolvulaceae, which consists of subfamiliesConvolvuloideae (11 tribes) and Humbertoideae (one tribe, monotypicHumberteae), is the only family in the asterid clade known toproduce seeds with PY. The primary aim of this study was tocompare the morphoanatomical characteristics of the water gapin seeds of species in the 11 tribes of the Convolvuloideaeand to use this information, and that on seed dormancy and storagebehaviour, to construct a phylogenetic tree of seed dormancyfor the subfamily. Methods: Scanning electron microscopy (SEM) was used to define morphologicalchanges in the hilum area during dormancy break; hand and vibratomesections were taken to describe the anatomy of the water gap,hilum and seed coat; and dye tracking was used to identify theinitial route of water entry into the non-dormant seed. Resultswere compared with a recent cladogram of the family. Key Results: Species in nine tribes have (a) layer(s) of palisade cells inthe seed coat, a water gap and orthodox storage behaviour. Erycibe(Erycibeae) and Maripa (Maripeae) do not have a palisade layerin the seed coat or a water gap, and are recalcitrant. The hilarfissure is the water gap in relatively basal Cuscuteae, andbulges adjacent to the micropyle serve as the water gap in theConvolvuloideae, Dicranostyloideae (except Maripeae) and theCardiochlamyeae clades. Seeds from the Convolvuloideae havemorphologically prominent bulges demarcated by cell shape inthe sclereid layer, whereas the Dicranostyloideae and Cardiochlamyeaehave non-prominent bulges demarcated by the number of sub-celllayers. The anatomy and morphology of the hilar pad follow thesame pattern. Conclusions: PY in the subfamily Convolvuloideae probably evolved in theaseasonal tropics from an ancestor with recalcitrant non-dormantseeds, and it may have arisen as Convolvulaceae radiated tooccupy the seasonal tropics. Combinational dormancy may havedeveloped in seeds of some Cuscuta spp. as this genus movedinto temperate habitats.     

Proteomic analysis of seed dormancy in Arabidopsis

The mechanisms controlling seed dormancy in Arabidopsis (Arabidopsis thaliana) have been characterized by proteomics using the dormant (D) accession Cvi originating from the Cape Verde Islands. Comparative studies carried out with freshly harvested dormant and after-ripened non-dormant (ND) seeds revealed a specific differential accumulation of 32 proteins. The data suggested that proteins associated with metabolic functions potentially involved in germination can accumulate during after-ripening in the dry state leading to dormancy release. Exogenous application of abscisic acid (ABA) to ND seeds strongly impeded their germination, which physiologically mimicked the behavior of D imbibed seeds. This application resulted in an alteration of the accumulation pattern of 71 proteins. There was a strong down-accumulation of a major part (90%) of these proteins, which were involved mainly in energetic and protein metabolisms. This feature suggested that exogenous ABA triggers proteolytic mechanisms in imbibed seeds. An analysis of de novo protein synthesis by two-dimensional gel electrophoresis in the presence of [(35)S]-methionine disclosed that exogenous ABA does not impede protein biosynthesis during imbibition. Furthermore, imbibed D seeds proved competent for de novo protein synthesis, demonstrating that impediment of protein translation was not the cause of the observed block of seed germination. However, the two-dimensional protein profiles were markedly different from those obtained with the ND seeds imbibed in ABA. Altogether, the data showed that the mechanisms blocking germination of the ND seeds by ABA application are different from those preventing germination of the D seeds imbibed in basal medium.     

Functional aspects of mature seed coat of theCannaceae

Cannaceae seeds have been analysed regarding seed coat structure, germination and macromolecular composition of the seed coats. Data of several mass spectrometric techniques were combined with those of microscopic and histochemical techniques to acquire insight into the functions of the seed coat.Cannaceae seeds have an exotestal layer of Malpighian cells with a hydrophobic and a hydrophilic part. The hydrophobic part is mainly responsible for the impermeability of the seed and contains silica, callose, lignin as water repellent substances. Water can only enter the seed after a certain temperature-induced opening of an imbibition lid. During imbibition the hydrophilic part of the Malpighian cells swells and the seed coat ruptures due to differences in pressure in the upper and lower part of the Malpighian cells.     

Viability of onion (Allium cega) seed as influenced by temperature during seed growth

The yield and performance of seeds from crops of winter-hardy, bolting-resistant onion grown at temperatures of 15–16, 18–19 and 22–23°C in 1979, 1980 and 1982 were compared. Yields of seed from crops grown at 22–23°C were lower than those from crops grown at lower temperatures but the seeds ripened between 11 and 32 days earlier. Seeds from crops grown at mean temperatures of above 18°C gave higher percentage germination when imbibed at 30°C than 20°C and they also gave higher percentage seedling emergence than those from crops grown at lower temperatures. Seedlings from seeds produced at mean temperatures above 18°C were heavier than those from seeds of a similar weight but produced at lower temperatures. None of these differences were associated with differences in seed weight, embryo weight or seed dormancy but were positively correlated with differences in seed N-concentration. The differences were also associated with the rate of imbibition of water as high germination, high N-content seeds had a slower rate of imbibition than low germination, low N-content seeds of the same weight.     

Ecophysiology of seed dormancy in the Australian endemic species Acanthocarpus preissii (Dasypogonaceae)

BACKGROUND AND AIMS: Seedlings of Acanthocarpus preissii are needed for coastal sand dune restoration in Western Australia. However, seeds of this Western Australian endemic have proven to be very difficult to germinate. The aims of this study were to define a dormancy-breaking protocol, identify time of suitable conditions for dormancy-break in the field and classify the type of seed dormancy in this species. METHODS: Viability, water-uptake (imbibition) and seed and embryo characteristics were assessed for seeds collected in 2003 and in 2004 from two locations. The effects of GA(3), smoke-water, GA(3) + smoke-water and warm stratification were tested on seed dormancy-break. In a field study, soil temperature and the moisture content of soil and buried seeds were monitored for 1 year. KEY RESULTS: Viability of fresh seeds was >90 %, and they had a fully developed, curved-linear embryo. Fresh seeds imbibed water readily, with mass increasing approx. 52 % in 4 d. Non-treated fresh seeds and those exposed to 1000 ppm GA(3), 1 : 10 (v/v) smoke-water/water or 1000 ppm GA(3) + 1 : 10 (v/v) smoke-water/water germinated <8 %. Fresh seeds germinated to >80 % when warm-stratified for at least 7 weeks at 18/33 degrees C and then moved to 7/18 degrees C, whereas seeds incubated continuously at 7/18 degrees C germinated to <20 %. CONCLUSIONS: Seeds of A. preisii have non-deep physiological dormancy that is released by a period of warm stratification. Autumn (March/April) is the most likely time for warm stratification of seeds of this species in the field. This is the first report of the requirement for warm stratification for dormancy release in seeds of an Australian species.     

Development and structure of the seed of Ozoroa paniculosa (Anacardiaceae) and taxonomic notes

The anatropous, bitegmic and crassinucellar ovule has a nuclear endosperm development. It is further characterized by a hypostase sensu lato. This hypostase being an integral part of the chalaza undergoes a secondary extension with it. At maturity the exalbuminous seed is partially pachychalazal and therefore two anatomically distinct larger parts can be distinguished in the mature seed coat. An endotegmen typifies the integumentary seed coat, while a saddle-shaped hypostase characterizes the chalazal seed coat. This seed coat shows several characteristics of the typical anacardiaceous pachychalazal seed. The cotyledons store lipids and protein as nutrient reserveS. A well-developed cuticle, cuticular layer, cutin and callose in the hypostase cell walls, as well as tannin-like deposits in the seed coat, protect the physiologically ripe seed against dehydration.     

Effect of tannin content of faba bean (Vicia faba) seed on seed vigour, germination and field emergence

Differences in seed vigour of zero- and high-tannin faba beans were investigated using 25 seed lots of 12 cultivars following earlier reports of poor emergence in the zero-tannin types. Field emergence ranged from 54–96% indicating differences in seed vigour between cultivars all having high laboratory germination (>91%). Seed from zero-tannin accessions with poor emergence had a higher incidence of testa and cotyledon cracking, a smaller percentage of hard seeds, more rapid water uptake, a lower percentage of vital staining of cotyledons and a greater leaching of solutes than high-tannin types. Nevertheless, variation in these characteristics existed between cultivars and lines of both types. Seeds with more cracks in the seed coat and fewer hard seeds imbibed water more rapidly and consequently showed lower levels of vital staining and more cracks in the abaxial surface of the cotyledons. Slower imbibition in polyethylene glycol lessened the incidence of these deleterious characteristics and may provide a practical resolution to the problem of poor field emergence in zero-tannin lines of faba bean with low seed vigour.     

Dormancy of<Emphasis Type="Italic"> Arabidopsis</Emphasis> seeds and barley grains can be broken by nitric oxide

Seeds of Arabidopsis thaliana (L.) Heynh. and grains of barley (Hordeum vulgare L.) were used to characterize the affects of nitric oxide (NO) on seed dormancy. Seeds of the C24 and Col-1 ecotypes of Arabidopsis are almost completely dormant when freshly harvested, but dormancy was broken by stratification for 3 days at 4°C or by imbibition of seeds with the NO donor sodium nitroprusside (SNP). This effect of SNP on dormancy of Arabidopsis seeds was concentration dependent. SNP concentrations as low as 25 M reduced dormancy and stimulated germination, but SNP at 250 M or more impaired seedling development, including root growth, and inhibited germination. Dormancy was also reduced when Arabidopsis seeds were exposed to gasses that are generated by solutions of SNP. Nitrate and nitrite, two other oxides of nitrogen, reduced the dormancy of Arabidopsis seeds, but much higher concentrations of these were required compared to SNP. Furthermore, the kinetics of germination were slower for seeds imbibed with either nitrate or nitrite than for seeds imbibed with SNP. Although seeds imbibed with SNP had reduced dormancy, seeds imbibed with SNP and abscisic acid (ABA) remained strongly dormant. This may indicate that the effects of ABA action on germination are downstream of NO action. The NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3 oxide (cPTIO) strengthened dormancy of unstratified and briefly stratified Arabidopsis seeds. Dormancy of three cultivars of barley was also reduced by SNP. Furthermore, dormancy in barley grain was strengthened by imbibition of grain with cPTIO. The data presented here support the conclusion that NO is a potent dormancy breaking agent for seeds and grains. Experiments with the NO scavenger suggest that NO is an endogenous regulator of seed dormancy.Abbreviations ABA Abscisic acid - cPTIO 2-(4-Carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3 oxide - GA Gibberellin - SNP Sodium nitroprusside - NO_x Gaseous oxides of nitrogen     

Freezing and desiccation tolerance of imbibed canola seed

Depending on the environmental conditions, imbibed seeds survive subzero temperatures either by supercooling or by tolerating freezing-induced desiccation. We investigated what the predominant survival mechanism is in freezing canola ( Brassica napus cv. Quest) and concluded that it depends on the cooling rate. Seeds cooled at 3°C h⁻¹ or faster supercooled, whereas seeds cooled over a 4-day period to −12°C and then cooled at 3°C h⁻¹ to−40°C did not display low temperature exotherms. Both differential thermal analysis and nuclear magnetic resonance (NMR) spectroscopy confirmed that imbibed canola seeds undergo freezing-induced desiccation at slow cooling rates. The freezing tolerance of imbibed canola seed (LT₅₀) was determined by slowly cooling to −12°C for 48 h, followed with cooling at 3°C h⁻¹ to −40°C, or by holding at a constant −6°C (LD₅₀). For both tests, the loss in freezing tolerance of imbibed seeds was a function of time and temperature of imbibition. Freezing tolerance was rapidly lost after radicle emergence. Seeds imbibed in 100 μ M abscisic acid (ABA), particularly at 2°C, lost freezing tolerance at a slower rate compared with water-imbibed seeds. Seeds imbibed in water either at 23°C for 16 h, or 8°C for 6 days, or 2°C for 6 days were not germinable after storage at −6°C for 10 days. Seeds imbibed in ABA at 23°C for 24 h, or 8°C for 8 days, or 2°C for 15 days were highly germinable after 40 days at a constant −6°C. Desiccation injury induced at a high temperature (60°C), as with injury induced by freezing, was found to be a function of imbibition temperature and time.