RNA synthesis and the germination of light-sensitive lettuce seeds

Summary The germination of lettuce seeds is inhibited by the nucleotide base analogue 6-methylpurine. RNA synthesis has been measured during imbibition and germination as ³²P-phosphate incorporation into RNA species as fractionated by polyacrylamide gel electrophoresis. Seeds were surface sterilized and imbibed in the presence of various antibiotics. RNA preparations from lettuce seeds were coelectrophoresed with ³H-RNA prepared from bacteria to check for bacterial contamination of the seeds. There is a much higher rate of RNA synthesis in illuminated, germinating seeds as compared to dark, non-germinating seeds. This difference does not develop until after 12 hours of imbibition at 27°, which is the time of onset of germination and radicle growth.This investigation was supported by a contract from the United States Department of Agriculture (No. 616-15-3). Journal paper of the Purdue Agriculture Experiment Station.     

Action spectra for light-induced germination in dormant lettuce seeds

Fluence response curves for red light-induced germination of thermodormant (TD) seeds of Lactuca sativa L. show two regions that differ in their light sensitivity. In the region of high sensitivity, the germination responses differ between seed batches and can be altered by dark storage or far red irradiation. Induction of germination in far red dormant (FRD) seeds requires far higher fluences. Action spectra for induction to 60% germination were determined for these various response types. Spectra for the regions of low sensitivity response are similar for TD and FRD seeds. In comparison, the action spectrum for the highly sensitive response in TD seeds is significantly shifted to longer wavelengths. Analogous differences exist in the action spectra for far red reversal of the red induced germination responses. Germination induction in the low sensitivity region shows repeated red-far red reversibility. Far red reversal of red induction in the high sensitivity region does not saturate even at the highest far red fluences available and requires increased red fluences for subsequent reinduction. A model quantitatively accounting for these observations is presented. It is pointed out that action spectra of processes involving photoreversible pigments with partly overlapping absorption spectra in general are not identical with the absorption spectra of the partners. They should depend upon the degree of phototransformation required to elicit a given physiological response. In the case of induction of lettuce seed germination the observed action spectra can be interpreted as reflecting different requirements for P _fr of the various response types. Our results do not necessitate the assumption of spectroscopically different forms of phytochrome in these seeds.Abbreviations TD thermodormant - FRD far red dormant - P phytochrome - P _r red absorbing form of P - P _fr far red absorbing form of P     

The effects of grading and 'priming' seeds of crisp lettuce cv. Saladin, on germination at high temperature, seed 'vigour' and crop uniformity

Three seedlots of crisp lettuce cv. Saladin were ‘primed’ in 1% tri-potassium orthophosphate (K₃PO₄) or in water for 6 to 12 h and then dried at room temperature. Two of the seedlots were separated into fractions of different mean weight before ‘priming’. The effects of ‘priming’ and grading on crop uniformity, germination at 35°C and root length in a slant test were evaluated. In general, ‘priming’ increased root length in a slant test but there were no consistent beneficial effects of ‘priming’ on seedling size, head weight or uniformity of head weight. Larger seed produced longer roots in a slant test than small seed and gave larger seedlings but seed size had no effect on head weight or cv. of head weight. ‘Priming’ had no effect on germination at 20°C but significantly increased germination at 35°C. ‘Priming’ in K₃PO₄ was more effective than in water. It is suggested that the response of seeds to ‘priming’ varies with seedlot and individual seed weight.     

The growth physics and water relations of red-light-induced germination in lettuce seeds

Using lettuce (Lactuca sativa L., cv. Grand Rapids) embryos in osmotica, we have demonstrated that when the growth rates of the embryonic axes of seeds treated with red (R) or far-red (FR) light are equalized, the axes of R-treated seeds develop a 3.4-bar decrease in water potential (paper No. III).As axial growth begins, reserve protein and phytin decrease rapidly, concomitant with increases in reducing sugars, -amino nitrogen, and inorganic and esterified soluble phosphates. However, no differences between the axes of R-and FR-treated seeds are found with respect to the changes in these compounds, indicating that these changes arise as a result of growth and are not under immediate phytochrome control. Little change in the total lipid content is found in either treatment. The axes of FR-treated seeds hydrolyze endogenous sucrose at a greater rate thant those of R-treated seeds. Axes of R-treated seeds accumulate K⁺ and Na⁺ to a greater extent than those of FR-treated seeds. When potassium salts are added to the incubation medium, R induces increased K⁺ uptake by the axis and greater medium acidification by the axis. Malate and other organic acids and acidic amino acids increase at equal rates in both treatments, indicating that inorganic anions may also be taken up to balance the ionic charges. The results are compatible with the assumption that changes in the osmotic and pressure potentials of the embryonic axes of R-treated seeds are the result of a phytochrome-stimulated proton pump which, in whole dormant seeds, would initiate water-potential changes allowing the embryos to overcome the mechanical restraint of the surrounding seed layers, resulting in germination.Abbreviations FR far-red light - PEG polyethylene glyeol 4000 - Pfr far-red-absorbing form of phytochrome - R red light III=Carpita et al. 1979     

The growth physics and water relations of red-light-induced germination in lettuce seeds

Summary A study of photodormant lettuce embryos germinating in water showed that red light induces an increased rate of water uptake. Determinations of the water potential, carried out by a modified gravimetric technique which eliminates errors introduced by solute penetration into cellular osmotic space, showed that the water potential of embryos germinating in water after dark and red light treatment was equivalent and equal to the osmotic potential of a 0.0 to 0.1 molal mannitol solution. Osmotic potentials of the embryos were determined using two new methods. One of the methods utilizes penetration of deuterated water; the other, penetration of a labeled osmoticum into the tissue. For both light- and dark-treated embryos in water, the osmotic potential was equivalent to that of a 0.34 to 0.41 molal mannitol solution. Lettuce embryos thus require that turgor pressure reach a threshold considerably above zero before growth can occur.     

Effects of long-term storage on phytochrome-mediated germination in lettuce seeds

The effects of long-term seed storage on the physiological properties of phytochrome-mediated germination including water uptake, the temperature and light flunnce dependencies of germination and dark germination were studied. The fluenceresponse relationships of the brief irradiation with monochromatic red (660 nm, 7.5 W m⁻²) and far-red (750 nm, 6.6 W m⁻²) light at various times after sowing were also studied. The samples used consisted of three lots of seeds ofLactuca sativa L. cv. MSU-16, which had been harvested in 1976, 1979 and 1985 and stored dry for 9, 6 and 0 years, respectively, in darkness at 23±2 C until the experiments were carried out in July–August, 1985. Seeds with the longer storage periods showed the higher ability to germinate in both continuous darkness and continuous white fluorescent light at 20–30 C. In the seeds stored for 6 or 9 years, red light irradiation for 20 sec given at 15 min or more after sowing at 25 C induced as high a percent germination (85–95%) as those under continuous white fluorescent light. In the freshly harvested seeds, however, germination under continuous white fluorescent light (46%) was considerably lower than the germination induced by the red pulse (97%). Germination of the seeds decreased when the intervals between sowing and a far-red irradiation for 20 sec increased up to 100 min (or 30 min in the freshly harvested seeds). The far-red pulse given later than 100 min (or 6 hr in the freshly harvested seeds) after sowing resulted in an increased germination up to the dark-germination levels with increasing intervals between sowing and the pulse irradiation. Before or at 3 min after sowing, the seeds stored for 6 or 9 years were responsive to the far-red pulse although they were not or hardly responsive to the red pulse, while the freshly harvested seeds were responsive to both the far-red and the red pulses. These data indicate that normal functions of phytochrome completely survived in the dry seeds during storage at 25 C for as long as 6 or 9 years and that these functions are restored into full operation by means of imbibition. The differences in the dependence of germination on the time and fluence of a single pulse of red or far-red light seems to be related to the smaller water content throughout the imbibition in the seeds with the longer storage periods. The greater ability to germinate in the dark indicates the greater amounts of P_FR or the greater responsivity to P_FR, in the seeds with the longer storage periods.     

Effect of light and temperature on the germination of lettuce seeds

A short period (15–30 min) at 30° C promotes germination of seeds of Lactuca sativa L. cv. Repolhuda in darkness. Far-red light reverses this stimulation, and the escape curves for phytochrome and high-temperature action are quite similar, indicating that the two factors act at a common point in the chain of events leading to germination. It is suggested that high temperature acts by decreasing the threshold of the active, far-red absorbing, form of phytochrome (Pfr) needed to promote germination.Abbreviations FR far-red light - Pfr far-red-absorbing form of phytochrome - R red light     

The germination and the forces required to penetrate seed layers of different seedlots of three cultivars of crisp lettuce

Using 24 commercial seedlots of the lettuce cvs Ithaca, Pennlake and Saladin, the percentage germination at a range of temperatures from 20 to 35°C, and the forces required to penetrate the pericarp, the endosperm and the embryo were measured. Percentage germination declined with increasing temperature in all seedlots but there were particularly large differences between seedlots of the same cultivar at 30.0 and 32.5°C. The force required to penetrate the pericarp was on average more than twice that required to penetrate the endosperm. It is concluded that in commercial cultivars of crisp lettuce the strength of the pericarp plays a much greater role than that of the endosperm in determining germination. In all three cultivars there were significant positive correlations between seed weight and the force required to penetrate the whole seed. However only in cv. Pennlake were there significant correlations between germination at high temperature and seed penetration forces.     

Analysis of temperature effect on the germination of New York lettuce seeds

When New York lettuce seeds were imbibed below 20°C in thedark, full germination was observed, whereas none occurred above35°C even under red-light irradiation. Partial treatmentwith high temperature during imbibition revealed the involvementof a thermo-labile process in the germination. This processwas completely inhibited above 30°C in the dark, but re-activatedby the following incubation below 20°C. The longer the seedswere preincubated above 30°C, the longer the period below20°C required for re-activation. The results suggest thatthe thermo-labile process is controlled by a thermo-labile factor.The factor was inactivated slowly at 25°C and rapidly above30°C to a form which could be re-activated below 20°Cbut was irreversibly inactivated when the seeds were imbibedat temperatures above 45°C for more than 40 hr. The escapereaction of the phytochrome system proceeded even at 35°Cwhereas no germination was observed at that temperature. Thus,die germination of lettuce seeds is regulated not only by thephytochrome system but also by the thermolabile factor. (Received August 11, 1975; )     

The effects of SAN 9789 and light on phytochrome and the germination of lettuce seeds

Photoblastic seeds (akenes) of lettuce (Lactuca sativa (L.) cv. Grand Rapids) were treated with SAN 9789 [4-chloro-5-(methylamine)-2-a, a, a,-trifluoro-m-tolyl-3-(2H)-pyridasinone]. The seeds weere placed in Petri dishes on filter paper soaked with water or SAN solution. The treatment increased the germination in darkness from 17% for water to 78% for SAN treated seeds. An irradiation with 5 min red light gave a germination of 98% both in water and in SAN. In water the effect of red irradiation could be reversed with a short irradiation (8 min) of far red light (17% germination), while in SAN solution the far red reversibility was poor (92% germination). If the far red light was given repeatedly (5 min per h) it had a slightly larger effect. If given continuously for 24 hours, the germination in water was decreased to 0.3% and in SAN solution to 9%. Possible mechanisms for the SAN effect are discussed.     

Interactions between hydrostatic pressure and oxygen concentration on the germination of lettuce seeds

Lettuce seeds were germinated at different hydrostatic pressures and O₂ concentrations. Some combinations of pressure and O₂ concentration stimulated while others retarded germination. The highest germination rates which occurred with pressure application and added O₂ were equalled at atmospheric pressure if the O₂ concentration was increased sufficiently. Increasing pressure generally retarded germination, but the pressure effect was partly offset by added O₂. Sustained pressures affected subsequent germination after pressure was released, but only when O₂ was present. Seeds to which pressure was applied in the absence of O₂ germinated as well after pressure exposure as untreated seeds.     

Temperature sensitive phases during the germination of lettuce (Lactuca sativa) seeds

Lettuce seeds cvs Hilde, Feltham King and Avoncrisp were subjected, at different phases during imbibition at 22°C, to a high temperature (33°C) inhibitory for germination, for periods ranging from 4 to 144 h, before returning them to 22°C. The results showed, that the first 4h of imbibition and also the phase between the commencement of mitosis and the onset of radicle emergence were more sensitive to the effects of high temperature than other phases in the germination process. Short exposures (8–24 h) to 33°C commencing at the latter phase delayed germination by up to 4 days, and at the earlier by up to 8 days. Percentage germination was unaffectd except after prolonged exposures (> 48 h) from the beginning of imbibition, which reduced it. Seedling emergence from moist sieved soil was both delayed and reduced when imbibing seeds were exposed for a short period from the beginning of imbibition to 33°C compared with seeds imbibing continuously at 19°C. Germination was delayed and not reduced when seed was exposed to 33°C at the phase between commencement of mitosis and the onset of radicle emergence.     

Opposing effects of strychnine and brucine on the germination and growth of lettuce seeds

Strychnine and brucine, two related alkaloids that occur in plants belonging to theStrychnos species, were shown to have opposing effects on the elongation of the radicle of lettuce seeds. Strychnine was found to be inhibitory, whereas brucine was found to be stimulating to radicle elongation. Alkaloids, generally, are more commonly known for their inhibitory effects on plant growth rather than on their stimulating effects.Paper No. 2435 of the Journal Series of the South Dakota Agricultural Experiment Station, Brookings, South Dakota, USA.     

The effects of pH during treatment of lettuce seeds with chlorine-releasing compounds on germination and seedling development

Hypochlorite seed treatments have been reported in the literature as giving variable results. We report experiments on this topic in which lettuce seeds were used as a model biological system. Seeds were treated for 2 h with solutions of sodium hypochlorite diluted with buffers of pH 9·8, 10·8 and 11·8, or with the buffers alone. Only the seeds treated with hypochlorite diluted with pH 9·8 buffer showed increased numbers of abnormal seedlings when subsequently germinated. In an experiment in which lettuce seeds were treated with sodium dichloroisocyanurate (a chlorine-releasing triazine compound) and related non-chlorinated compounds, seedling abnormalities were related to chlorine in solution. Measurements of pH together with the known chemistry of hypochlorite suggest that the damaging agent in these experiments was the biocidal undissociated hypochlorous acid.     

Opposing effects of strychnine and brucine on the germination and growth of lettuce seeds

Strychnine and brucine, two related alkaloids that occur in plants belonging to theStrychnos species, were shown to have opposing effects on the elongation of the radicle of lettuce seeds. Strychnine was found to be inhibitory, whereas brucine was found to be stimulating to radicle elongation. Alkaloids, generally, are more commonly known for their inhibitory effects on plant growth rather than on their stimulating effects.     

Actions of gibberellic Acid and phytochrome on the germination of grand rapids lettuce seeds

Red light and gibberellic acid were about equally effective in promoting germination of Grand Rapids lettuce (Lactuca sativa L.) seeds. With initial far red light treatment more than 80% remained dormant in subsequent dark storage. After 2 days of dark storage, red light effectively promoted germination, while gibberellic acid action was weak. With between 2 and 10 days of dark storage, gibberellic acid had little effect, while promotion by red light decreased slowly and finally disappeared. After 10 days of dark storage, both gibberellic acid and red light were required for germination. The dark storage treatment interferes with phytochrome-independent germination processes and cannot be overcome by added gibberellic acid. However, storage may also decrease the effectiveness of endogenous gibberellins. Phytochrome-dependent germination seems to require only low levels of endogenous gibberellin activity or the addition of gibberellic acid. Gibberellins and red light appear to act on germination by regulation of sequential sites of a branched-looped pathway.     

Temperature and other factors affecting chloramphenicol stimulation of the germination of light-sensitive lettuce seeds

Summary D-threo-chloramphenicol at concentrations ranging from 1000 to 3000 g/ml stimulated the germination of the light-sensitive seeds of the lettuce (Lactuca sativa L.) varieties Attractie and Grand Rapids. This stimulatory effect of chloramphenicol was markedly temperature dependent, increasing with decrereasing temperature. Seeds showed little response to chloramphenicol at temperatures of 28°C and above except in the case of light treated Attractie seed. The failure of one batch of Grand Rapids seed to respond to chloramphenicol was associated with the low degree of dormancy in this batch.When the germination of half-seeds or intact excised embryos of Attractie seed was inhibited osmotically with 0.15 M NaCl a stimulatory response to chloramphenicol was obtained suggesting that the site of action was in the embryo itself.Other inhibitors of protein synthesis, cycloheximide, puromycin and p-fluorophenylalanine, did not stimulate germination. Cycloheximide at concentrations of 10 g/ml and above inhibited germination whereas puromycin and p-fluorophenylalanine were relatively ineffective as germination inhibitors.