Loss of Viability in Lettuce Seeds and the Accumulation of Chromosome Damage under Different Storage Conditions

Loss of seed viability in lettuce (Lactuca sativa L.) duringstorage is associated with an increase in the frequency of cellsin the surviving seeds showing chromosome damage during firstmitoses. The relation is linear when probit of the frequencyof aberrant cells is plotted as a function of probit percentagenormal germination. The slope of the relation, however, variesaccording to moisture content so that the proportion of aberrantcells for any given loss of germination increases with decreasein moisture content over the range 13.0–5.5 per cent.At 3.3 per cent moisture content, however, the proportion ofaberrations was no greater than at 5.5 per cent moisture content;and at 18.1 per cent moisture content the proportion was noless than at 13.0 per cent moisture content. Despite these differences,the increase in chromosomal aberrations per unit time for agiven temperature was always less the lower the moisture content.Diplontic selection markedly reduced the frequency of chromosomalaberrations and eliminated the differences in these frequenciesbetween the different storage treatments. But even after fiveweeks' growth, root tips from aged seed still contained abouttwice as many aberrant cells as compared with similar root tipsderived from the original seed stock. Studies on the frequencyof recessive mutations indicated that excessive amounts of heritablemutations were not present in the progenies of aged seed, evenwhen stored at moisture contents as low as 5.5 per cent. Allthis and other evidence reinforces the view that orthodox seedsfor genetic conservation should be stored at not more than about5 per cent moisture content, and that even lower moisture contentsare worth considering. The results also emphasise the need formaintaining a high regeneration standard, i.e. the percentageto which seed viability is allowed to fall during storage beforethe seed stock is regenerated. Lactuca sativa, lettuce, seed storage, seed viability, chromosomal aberrations, phenotypic mutations     

The Influence of Pre and Post-storage Hydration Treatments on Chromosomal Aberrations, Seedling Abnormalities, and Viability of Lettuce Seeds

A 2 h soaking treatment in distilled water, or in aqueous solutionsof cysteine, potassium iodide, or sodium thiosulphate, had nosignificant effect (P > 0.25) on the subsequent longevityof lettuce seeds (Lactuca sativa L.) in two different storageenvironments. Neither did these treatments influence relationsbetween loss in germination and the frequency of chromosomalaberrations observed during first mitoses after storage. Incontrast partial hydration of lettuce seeds after storage byexposure to moist air (humidification) or to an osmoticum (priming)reversed some of the damage which resulted from ageing. Mostof the benefits occurred during the first 3 d of humidificationduring which seed moisture content rose to 34 per cent, or duringthe first 7 d of priming when seed moisture content increasedto 44 per cent. Both post-storage hydration treatments reducedthe frequency of chromosomal aberrations, increased the rateof root growth, and decreased the frequency of morphologicallyabnormal seedlings. Either treatment could be of practical use,but it is suggested that humidification is more convenient.Consideration should be given to adopting a humidification treatmentas standard practice following long-term seed storage for geneticconservation. Lactuca sativa, lettuce, seed storage, seed viability, chemical pre-treatment, seed longevity, seed humidification (conditioning), seed priming, chromosome repair, seedling abnormalities     

Seed aging: chromosome stability and extended viability of seeds stored fully imbided

Increase in moisture content of seeds of Lactuca sativa L. and Fraxinus americana L. in air-dry storage caused a rapid decline in longevity and an increase in the rate of accumulation of chromosome aberrations. Storage of seeds fully imbibed but unable to germinate allowed a high germination capacity to be maintained for long periods, together with a very low incidence of chromosome aberrations. Seedlings grown from dry-stored seeds showed an increase in morphological abnormalities with length of storage, whereas seedlings from imbibed-stored seeds appeared normal. It is suggested that in dry tissues, enzyme-controlled turnover and repair may be temporarily suspended, and that this may be an important factor in the loss of seed viability in storage. The effect of increasing seed longevity by lowering the moisture content of dry-stored seeds is discussed in relation to this hypothesis. The relevance of the proposal is also discussed in relation to ecological studies.     

Prehydration and Priming Treatments that Advance Germination also Increase the Rate of Deterioration of Lettuce Seeds

The influence of prehydration in water or priming in –1.5 MPa polyethylene glycol 8000 solution for various periods,followed by redrying, on germination rate and longevity of lettuce(Lactuca sativa L.) seeds (achenes) was determined during controlleddeterioration at 10% moisture content (fresh weight basis) and40°C. Short prehydration treatments (up to 1 h) had littleeffect on either germination rate or longevity, but significantlyimproved root growth rates. Increasing durations of prehydrationor priming reduced the mean time to germination by up to 61%relative to untreated seeds, but also reduced mean seed longevityby as much as 84% Prehydration and priming altered the relationshipsbetween germination rate and viability and between normal andabnormal seedlings during ageing. Prehydration in abscisic acidor at a temperature inhibitory to germination did not preventthe reduction in longevity under controlled deterioration conditions.While prehydration or priming treatments effectively acceleratesubsequent germination rates of lettuce seeds, the redried seedsare nonetheless highly susceptible to deterioration in storage. Key words: Lettuce, Lactuca sativa L., seed priming, seed deterioration, germination rate     

Temperature and Seed Storage Longevity

Seed survival data for eight diverse species, namely the cerealbarley (Hordeum vulgare L.), the grain legumes chickpea (Cicerarietinum L.), cowpea [Vigna unguiculata (L.) Walp.] and soyabean [Glycine max (L.) Merr.], the timber trees elm (Ulmus carpinifoliaGleditsch.), mahogany (Swietenia humilis Zucc.), and terb (Terminaliabrassii Exell.), and the leaf vegetable lettuce (Lactuca sativaL.) were compared over a wide range of storage environments(temperatures from –13 °C to 90 °C, seed moisturecontents from 1.8 to 25% f. wt) using a viability equation developedpreviously. In accordance with that equation, the effect oftemperature on seed longevity was dependent upon the temperaturerange. The temperature coefficients of the viability equationdid not differ significantly (P > 0.05) among the eight speciesdespite their contrasting taxonomy. Thus the quantitative relationbetween seed longevity and temperature does not vary among diversespecies. The same conclusion was obtained for the coefficientsof a proposed alternative model of the relation between seedlongevity and temperature. The implications of the two temperaturemodels in the viability equation for extrapolations to low andvery low temperatures are discussed. Seed storage, seed longevity, seed moisture, temperature, viability equation, genetic resources conservation, Cicer arietinum L., Glycine max (L.) Merr., Hordeum vulgare L., Lactuca sativa L., Swietenia humilis Zucc., Terminalia brassii Exell., Ulmus carpinifolia Gleditsch., Vigna unguiculata (L.) Walp     

Seed Viability Constants for Lettuce

Seeds of two lettuce cultivars (Lactuca sativa L., cv. Meikoninginand cv. Grand Rapids) were hermetically stored with constantmoisture contents ranging between 3.6 and 17.9 per cent (freshweight basis) at constant temperatures ranging between 5 and75 °C. The decline with time in percentage germination andpercentage normal seedlings was determined for each storagetreatment. The data were fitted to an equation which containsthe constants: K₁, the probit of the initial percentage germinationor normal seedlings; K_E, a species constant; C_W, the constantof a logarithmic moisture term; C_H, the constant of a lineartemperature term and C_Q, the constant of a quadratic temperatureterm. Regression analysis of data from storage periods up to5.5 years at temperatures of 5–75 °C and seed moisturecontents of 3.6–13.6 per cent yielded the following values:K_E= 8.218, C_W=4.797±0.163, C_H=0.0489±0.0050 andC_Q=0.000365±0.000056. Although this equation consistentlyprovided a better fit, simplified equations, assuming eithera log-linear relationship between seed longevity and temperature,or a log-linear relationship between seed longevity and bothmoisture content and temperature, accounted for more than 94per cent of the variation at the restricted temperature rangeof 5–40 °C. Longevity of the same seed lots at sub-zero temperatures (–5,–10 and –20 °C) was studied in separate tests.Freezing damage, resulting in abnormal seedlings in the germinationtest, occurred at –20 °C when the moisture contentof the seeds exceeded 12 per cent. No decline in percentagenormal seedlings was observed after a storage period of 18 monthsor longer at –20 °C, provided the seed moisture contentdid not exceed 9.5 per cent. For seeds stored at –5 and–10 °C with 9.6–12.5 per cent moisture content,the observed rate of decline of percentage normal seedlingswas adequately predicted by the viability equation, using theabove values for the constants. This suggests that for low moisturecontents the viability equation can be applied to estimate longevityat sub-zero temperatures. Lettuce, Lactuca sativa (L.), seed longevity, seed storage, viability constants, storage conditions     

Restoration of Seed Germination at Supraoptimal Temperatures by Fluridone, an Inhibitor of Abscisic Acid Biosynthesis

Fluridone, an inhibitor of ABA biosynthesis, restored the seedgermination of lettuce (Lactuca sativa L. cv. Grand Rapids)and many other plant species at supra-optimal temperatures.ABA content in lettuce seeds after imbibition quickly decreasedat 23°C, but not at 33°C (a supraoptimal temperature).Fluridone caused a decrease in ABA content at 33°C, whichsuggests that the maintenance of high ABA content could be responsiblefor high-temperature inhibition of germination of lettuce seeds.This probably results from an increase in the rate of ABA biosynthesisat the higher temperature. The present study indicates thatABA plays a decisive role in the regulation of seed germinationat supraoptimal temperatures. ¹ Corresponding author: fax 81-22-717-8834; e-mail yoshi@bios.tohoku.ac.jp     

Interaction of Moisture Stress and Three Phenolic Compounds on Lettuce Seed Germination

No interactions between water stress and three phenolic acids(p-coumaric, caffeic and ferulic acids) on lettuce (Lactucasativa L. var. Grand Rapids) seed germination were found. Probitanalysis indicated that mechanisms of action of water stressand the phenolic inhibitors were similar. The relative effectivenessof the compounds was p-coumaric > ferulic > caffeic. Nointeraction was found between p-coumaric and ferulic acid, whereasantagonism was found between caffeic acid and each of the othertwo phenolic acids. Lactuca sativa L., lettuce, germination, phenolic compounds, moisture stress, allelopathy, seed     

Time to Seed Death as a Function of the Gibbs Free Energy of Water Vapour: Practical and Theoretical Implications

Time (t) to loss of seed viability (e.g. log t or log ) is modelledin the literature as the sum of a moisture term and a quadratictemperature term, f(T,T²). The coefficients inf(T,T²) have beenshown to be ‘identical’ in orthodox seeds. I postulatethat this identity is due to a parameter common to all seedsand, consistent with that hypothesis, report a close correlation(R²=0.9998) between f(T,T²) and the Gibbs free energy of watervapour, G, over the temperature range 0–90 °C. Thehypothesis has the statistical advantage of reducing the numberof independent variables from two to one, without changing thefit. This is demonstrated from analysis of data for lettuceand barley. An explanation for this correlation of time to seeddeath with G is that water vapour is the proximate source ofenergy leading to decomposition of individual molecules, withthe ultimate result of seed death. Copyright 1999 Annals ofBotany Company Seed longevity, temperature effect, Gibbs free energy of water vapour, moisture-temperature interactions, Hordeum vulgare L., barley, Lactuca sativa L., lettuce.     

Survival and Vigour of Lettuce (Lactuca sativa L.) and Sunflower (Helianthus annuus L.) Seeds Stored at Low and Very-low Moisture Contents

Seeds of lettuce (Lactuca sativa L.) and sunflower (Helianthusannuus L.) were stored hermetically at 35 °C with 11 differentmoisture contents between 1·3 and 6·9%, and between1·3 and 7·1% of fresh mass, respectively. Germinationand vigour (mean germination time, root length, seedling dryweight) were determined after storage for 0, 8, or 16 weeks(sunflower) or 0, 8, 16, or 48 weeks (lettuce) in these environmentsfollowed by various humidification treatments (to avoid imbibitioninjury). The range of seed storage moisture contents over whichdeterioration was minimized depended upon the criterion of deteriorationused, and varied somewhat between species. Comparison of theseranges for seeds stored for the longest durations showed thatfor some criteria seed performance was poorer (P < 0·05)at both the lowest and highest moisture contents investigatedthan at certain of the intermediate storage moisture contents(e.g, most rapid germination occurred in sunflower followingstorage at 2·2-4·7% moisture content), whereasfor other criteria all the drier storage moisture contents weresuperior to the more moist (e,g. greatest seedling growth occurredin sunflower following storage at 1·3-5·1% moisturecontent). But none of these results suggested that lettuce andsunflower seeds stored hermetically at 2·5-3·0%or 2·2-2·5% moisture content, respectively, wereless vigorous than at any other moisture content tested. Inboth species, these storage moisture contents are in equilibriumwith about 8-10% relative humidity (r.h.) at 20 °C, whichis similar to and indeed marginally less than the 10-13% r.h.recommended following earlier studies on the longevity of seedsin hermetic storage at much warmer temperatures. Thus, theseresults show no evidence that the optimum seed moisture contentfor storage increases with decrease in temperature, at leastover the range 35-65 °C, as has been suggested elsewhere.We conclude that the international recommendation for the long-termseed storage for genetic conservation at 5 ± 1% moisturecontent should not be revised upwardly, and that in situationswhere refrigeration cannot be provided storage at even lowermoisture contents is worthy of further investigation for thoseseeds in which desiccation at 20 °C to equilibrium at 10%r.h. results in moisture contents well below 5%.Copyright 1995,1999 Academic Press Helianthus annuus L., sunflower, Lactuca sativa L., lettuce, desiccation, seed storage, seed vigour     

Freezing Tolerance in Hydrated Lactuca sativa (L) Seed: A Model to Explain Observed Variation Between Seed Lots

Low temperature tolerance was investigated in the imbibed seedof 15 seed lots compnsmg seven cultivars of Lactuca sativa L.During rapid cooling (20 °C h^–1) some seeds of allseed lots survived to –16 °C but none to –20°C. The majority of seed lots retained over 50 per centviability above –14 °C due to isolation of the embryofrom external ice by the endosperm, and subsequent embryo super-cooling.Certain seed lots, including all three seed lots of cv. TomThumb, showed high mortality at temperatures above –10°C. Correlation of mortality with the formation of externalice suggested that the endosperm is not an effective nucleationbarrier in these seed lots. Survival to –20 °C was increased at slower coolingrates (6 to 1 °C h^–1) due to freeze desiccation ofthe embryo, but seed lots varied considerably in their toleranceof specific cooling rates. A model to explain this variationwas developed incorporatmg (1) seed lot super-cooling limittemperature, (2) the rate at which freeze dehydration of thesupercooled embryo took place, (3) the moisture content at whichnucleation (at –20 °C) was no longer certain and (4)the.initial equilibrium moisture content of the fully imbibedseed. Factors (1), (2) and (3) were found to be relatively constant,but low (or artificially reduced) seed moisture content wasclosely correlated with high survival at natural cooling rates.Seed size fractions of similar moisture content from a singlecultivar showed that more small seeds survive cooling at 3 °Ch^–1 to –20 °C than larger seed. Seed with pierced endosperms or ineffective nucleation barrierswere capable of surviving to at least –10 °C if cooledslowly (1 °C h^–1) but were killed by rapid (20 °Ch^–1) cooling. Lactuca sativa L, lettuce, seed germination freezing tolerance, super-cooling     

Water and Seed Survival

Between about –350 and –14 MPa the rate of lossof viability in orthodox seeds is a positive function of waterpotential. The relative effect of water potential has been analysedin an oily seed (lettuce) and a non-oily seed (barley) and foundto be more or less identical. The lower limit for the relationin various species coincides with a seed moisture content (wetbasis) between about 2 and 6%. Below this level there is littleor no improvement in longevity with reduction in moisture content.The upper limit coincides with moisture contents of between15 and 28%, depending on whether the seeds are oily or non-oily.A water potential of about –14 MPa is the threshold forrespiration which increases more-or-less linearly with increasein water potential above this level. Above this threshold, andproviding oxygen is available to sustain respiration, seed longevityincreases with increase in water potential except that, unlessthe seeds are dormant, germination may be initiated at a waterpotential of about –1·5 to –0·5 MPa.In the absence of oxygen there may be a slight further declinein longevity with increase in water potential above –14MPa before longevity reaches a minimum value Since they cannot be dried very much without immediate lossof viability, recalcitrant seeds survive longest in the presenceof oxygen at maximum water potential commensurate with preventinggermination. The threshold water potential for immediate lossof viability has not been determined for most species but itis probable that it is close to the water potential typicalof the permanent wilting point in these plants, say –2MPa Lactuca saliva L., lettuce, Hordeum oulgare L., barley, seed storage, moisture content, relative humidity, water potential, temperature, oxygen     

The Potential for Newly-Germinated Cabbage Seed Survival and Storage at Sub-Zero Temperatures

The moisture content of newly germinated cabbage seed (radicles1 05 mm long) was reduced to 14% of f.wt without loss of viability.As the moisture content was reduced below 45%, the temperatureat which the germinated seeds froze, and therefore died, decreasedprogressively to a minimum of –34 C at 19% moisture content.No freezing exotherms were recorded in seeds with moisture contentsbelow 19%. Seeds with a moisture content between 14 and 16%maintained viability for at least 1 week when cooled at 26C.min^–1to –20 C and held at this temperature, indicating thepotential for prolonged storage of these low-moisture-contentgerminated (LMCG) seeds. Brassica oleracea, cabbage, germinated seed, seed storage, fluid drilling, freezing exotherm, thermal analysis     

The Effect of Sodium Hypochlorite on Germination of Lettuce Seed at High Temperature

Investigations into the effects of treatment with sodium hypochloritesolution on high-temperature germination of lettuce seed aredescribed. A 2 h treatment in a solution with 10% availablechlorine enabled up to c. 70% of seeds to germinate at 35 °C,without impairing germination and normal seedling developmentat 20 °C. A 10 min rinse in 0.01 N HCI following such treatmentenhanced its effect and treated seeds retained their abilityto germinate at high temperature for at least 18 months. Seedlingdevelopment at 20 °C following treatment was influencedby treatment temperature and batch of hypochlorite solution.Emergence from compost in a cycling (30/15 °C) temperatureregime was improved in three out of five cultivars tested, andthere were indications that hypochlorite treatment enabled oneof the other cultivars to escape induced thermodormancy. Key words: Lactuca sativa L., Sodium hypochlorite, Thermo-inhibition     

Estimation of Provisional Seed Viability Constants for Lupinus polyphyllus Lindley

A limited amount of data on the hermetic storage of Lupinuspolyphyllus Lindley seeds in several different environmentshas been used to provide estimates of seed viability constants.These allow reasonably accurate predictions of seed storagelife under known conditions (orthodox storage), especially inthe short or medium term. Lupinus polyphyllus Lindley, seed storage, viability constants, influence of seed moisture content and temperature     

Effects of Moisture Content on Germination of Seeds of Hancornia speciosa Gom. (Apocynaceae)

Seeds of Hancornia speciosa germinated best at a temperatureof 20–30 °C. The viability of the seeds during storagewas short and the best storage conditions for viability entailedkeeping the seeds in polyethylene bags. Seed viability was maintainedonly when the seeds were stored at a moisture content above30%; storage conditions which allowed dehydration resulted ina rapid loss of viability (the seeds showed recalcitrant behaviour). Low temperature during storage did not improve longevity. Arelationship between germination and moisture content was established,but when the moisture content fell below 25% there was a drasticreduction of germination. After 9 weeks of storage, even athigh moisture content, seeds lost viability. Loss of seed viability during seed dehydration was associatedwith increased leakage of electrolytes and organic solutes,and reduced tetrazolium staining during subsequent imbibition. Hancornia speciosa, germination, recalcitrant seeds, storage, moisture     

Germination and Dormancy in Immature and Fresh-mature Lettuce Seeds

The effects of red light, far-red light, Gibberellin A₃, andethephon were studied on the germination of lettuce seeds cv.Grand Rapids harvested at different stages of development. Seeds did not become capable of germination until 8 days afteranthesis. Red light promoted seed germination from the age of8–9 days following anthesis up to the newly mature stage.Ten or 11 days following anthesis, a large percentage of seedsbecame capable of germination in the dark and therefore couldbe considered not dormant. They were affected by far-red light,but less so than the mature seeds. The effect of light on the germination of developing seeds appearedto be similar to the known light effect on mature lettuce seedgermination. Gibberellin A₃ and ethephon had no effect on immatureand fresh seed germination. Lactuca sativa L., Lettuce, germination, dormancy, red light, far-red light, gibberellin A₃, ethephon     

Changes in Endogenous Cytokinins of Lettuce Seed during Germination

Using the soybean callus bioassay it has been shown that dormant lettuce seeds (Lactuca sativa L. cv. Grand Rapids) contain large amounts of water soluble cytokinins and small amounts of butanol soluble ones. When the seeds are irradiated with red light, or imbibed with 5 mg/1 gibberellic acid in the dark, the total cytokinin content of the seeds decreases, the level of water soluble cytokinins decreases, and the level of the butanol soluble cytokinins increases. Far-red light does not reverse this effect completely although cytokinin activity in the butanol extracts decreases following such irradiation. It is proposed that the interconversion of cytokinins initiated by red light, or gibberellic acid in the dark, is one of the primary events leading to radicle elongation in light-sensitive lettuce seed.     

Flow Cytometric Determination of Nuclear Replication Stage in Seed Tissues

Flow cytometric determination of DNA levels in embryos of fullymatured seeds of various plant species revealed large amountsof 2C DNA signals, indicating that most cells had arrested thecell cycle at the presynthetic G₁ phase of nuclear division.The accumulation of cells at G₁ was found both in orthodox andin recalcitrant (i.e. Castanea sativa) seed species. As recalcitrantseeds are characterized by the absence of maturation drying,the arrest of the cell cycle in the presynthetic phase may notbe linked to the seed water status. Apart from the 2C signal, 4C values were found in the embryoof some seed species (e.g. Raphanus sativus) indicating thatcells were arrested in G₂ Cells arrested in G₂ were primarilylocated in the root-tip region of the embryo. In addition, combinationsof higher C values (i.e. 8C, 12C, 16C and 64C) were observedin the endosperm of Solanum melongena and Lycopersicon esculentum,and in the root-tip cells of Phaseolus vulgaris and Spinaciaoleracea. These mixtures of polyploid nuclei (also called 'polysomaty')may arise from a developmentally controlled cellular endoreduplicationand indicates that in each cell type of the seed the amountof DNA is regulated both spatially and temporally.Copyright1993, 1999 Academic Press Endive, Cichorium endiva, lettuce, Lactuca sativa, egg-plant, Solanum melongena, pepper, Capsicum annuum, tomato, Lycopersicon esculentum, radish, Raphanus sativus, bean Phaseolus vulgaris, spinach, Spinacia oleracea, chestnut, Castanea sativa, beech, Fagus sylvatica, pine, Pinus nigra, DNA content, flow cytometry, seed, nuclear replication stage, C levels, storage