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.     

Leakage during Seed Imbibition

When pea embryos are placed in water solutes leak from themat a rate which declines rapidly at first and then more slowly.Embryos can be dried down over calcium chloride and will thenleak as before when returned to water. Similar results wereobtained with Ricinus seeds after removal of the testa. Pea embryos that have first been allowed to imbibe some waterthrough a small part of their surface (by placing them on dampfilter-paper) leak relatively slowly when subsequently immersedin water; the greater the initial imbibition the slower thesubsequent leakage. Likewise, embryos taken from peas that wereharvested when succulent and tender show only slow leakage It is proposed that as seeds dry out in the course of developmentcell membranes lose their integrity. When such dry seeds areallowed to imbibe water there is a short period, before membraneintegrity is re-established, during which solutes can leak outof the cells.     

Nucleotide Levels and Loss of Vigour and Viability in Germinating Wheat Embryos

At an optimal germination temperature loss of viability in wheatseed is reflected in reduced levels of nucleoside triphosphatesand nucleotide sugars in the embryo compared to the levels foundin high viability embryos during the early hours of imbibition.These differences are magnified on germination at 10° Cand are accompanied by a greatly reduced rate of protein synthesisin embryos of low viability compared with the correspondingrate in embryos of high viability. Loss of vigour precedes lossof viability in seed but differences in biochemical parametersbetween high and medium vigour seed do not become apparent unlessgermination under stress conditions, e. g. low temperature,occurs. Under these stress conditions, wheat seed of reducedvigour can be distinguished from high vigour seed lots of similarviability by the decreased levels of nucleotides and nucleotidesugars found in embryos from reduced vigour seed lots duringthe early hours of imbibition and by reduced rates of proteinsynthesis in these embryos during this same period. Nucleotideanalysis is a potentially useful method for rapid assessmentof the viability or vigour of a seed lot. Key words: Nucleotides, Vigour, Viability, Wheat     

Effects of cold‐girdling on flows in the transport phloem in Ricinus communis: is mass flow inhibited?

The effects of cold girdling of the transport phloem at the hypocotyl of Ricinus communis on solute and water transport were investigated. Effects on the chemical composition of saps of phloem and xylem as well as of stem tissue were studied by conventional techniques and the water flow in the phloem was investigated by NMR imaging. Cold girdling reduced the concentration of sucrose but not that of inorganic solutes or amino acids in phloem saps. The possibility that cold treatment inhibited the retrieval of sucrose into the phloem, following leaching from the sieve tubes along a chemical gradient is discussed. Leaching of other solutes did not occur, as a result of missing promoting gradients in stem tissue. Following 3 d of cold girdling, sugar concentration increased and starch was synthesized and accumulated in stem tissue above the cold girdling region and along the cold-treated phloem pathway due to leaching of sugars from the phloem. Only in the very first period of cold girdling (<15-30 min) was mass flow inhibited, but recovered in the rest of cold treatment period to values similar to the control period before and the recovery period after the cold treatment. It is concluded that cold treatment affected phloem transport through two independent and reversible processes: (1) a permanent leaching of sucrose from the phloem stem without normal retrieval during cold treatment, and (2) a short-term inhibition of mass flow at the beginning of cold treatment, possibly involving P proteins. Possible further mechanisms for reversible inhibition of water flow are discussed.     

The effect of time of harvest on the viability and pre-emergence mortality in soil of pea (Pisutn sativum L.) seeds

A series of harvests from field plots of peas showed that the younger seeds were at the time of harvest the lower was their percentage viability and the greater their percentage mortality in soil. Young seed also showed a high degree of solute leaching into steep water and had a high moisture content at harvest compared with older seeds. Heavy rainfall just prior to harvesting reduced the percentage viability of the dried seed and increased the moisture content of the seed at harvest as well as the leaching of solutes from the dried seed into steep water. Comparisons of moisture content at harvest and seed condition after drying suggested that cotyledons and embryo axes were only able to withstand enhanced drying after removal from the plant if the seed moisture content had already begun to decline whilst the seed was still on the plant.     

The osmotic sensitivity of netropsin analogue binding to DNA

The binding of a netropsin analogue to random sequence DNA, monitored by CD, is seen dependent on the concentration of neutral solutes. The binding free energy decreases linearly with solute osmolal concentration and the magnitude of the effect is insensitive to the chemical identity of the solute fur betaine, sorbitol, and triethylene glycol. These solutes appear to modulate binding through their effect on water activity and changes in the hydration of the drug and DNA in the complex reaction, not through a direct interaction with the reactants or the product. The dependence of binding constant on solute concentration can be interpreted as an additional binding of some 50–60 extra solute excluding water molecules by the complex. A water sensitivity of drug binding is further seen from the dependence of binding constants on the type of anion in solution. Anions in the Hofmeister series strongly affect bulk water free energies and entropies. The differences in netropsin analogue binding to DNA with Cl⁻, F⁻, and CIO are consistent with the effect observed with neutral solutes. The ability to measure changes in water binding associated with a specific DNA interaction is a first step toward correlating changes in hydration with the strength and specificity of binding. © 1995 John Wiley & Sons, Inc.     

Osmotic and cryoprotective effects of glycerol-sucrose solutions on day-3 mouse embryos

The relative volume of Day-3 mouse embryos changed as a linear function of the reciprocal of osmolality [corrected] of non-permeating solutes after 10 min exposure to sucrose and glycerol-sucrose solutions at 20 degrees C. The slope of the linear regression line was less in glycerol-sucrose than in sucrose solutions because glycerol permeation caused re-expansion. Before freezing by direct transfer to -180 degrees C the embryos were placed into glycerol-sucrose in 1-step (1-step equilibration) or first into glycerol and then into glycerol-sucrose (2-step equilibration). Using 2-step equilibration the post-thaw survival rate was substantially higher at 3.0 and 4.0 M-glycerol levels and less dependent on changes in the sucrose concentration within the range of 0.125 to 1.0 M than with 1-step equilibration. Under optimal conditions 90-95% of rapidly frozen embryos developed to blastocysts in vitro and 30% into live young in vivo. It is suggested that the cryoprotective role of glycerol is due to its ability to reduce osmotic pressure differences between the extra and intracellular spaces during rapid freezing of embryos.     

Relationship between stage of development and sex of bovine IVM-IVF embryos cultured in vitro versus in the sheep oviduct

We have confirmed more rapid development of male compared with female in vitro-cultured bovine embryos during the first 7 d after in vitro fertilization. The male-to-female sex ratio of expanded blastocysts after 10 d of in vitro culture was 1.37:1.00, which was significantly different from the expected 1:1 ratio, but no deviation from a 1:1 ratio was observed for male and female expanded blastocysts derived from culture of bovine embryos in the sheep oviduct (1.11:1.00). When embryos that developed only to the morula stage were analyzed for sex, a greater number of female than male bovine embryos was observed from in vitro culture but not after culture in the sheep oviduct. Possible causes of these sex-related differences in development of cultured bovine embryos are discussed.     

Osmoregulation, growth and sucrose accumulation in germinated Trigonella foenum-graecum (fenugreek) seeds treated with polyethylene glycol

Germinated seeds of Trigonella foenum-graecum L. (fenugreek) were grown in water or in polyethylene glycol (PEG) solutions. After endosperm removal, the water relations, growth, dry weight, sucrose and reducing sugar content of the embryo were determined. Under water sstress conditions, water content and osmotic potential (π₀) at saturation, growth and dry weight were lower than in non-stressed controls. The reduction in dry weight indicated a lower uptake of solutes from the endosperm and the decrease in π₀ was not accompanied by an increase in the amount of the accumulated solutes. It is suggested that embryos of stressed fenugreek seeds control osmotic potential by reduction of water uptake and that this results in reduction of growth. Embryos isolated from germinated seeds ("naked" embryos) were grown in water or in PEG solutions, with or without galactose (as an external solute source substituting for the endosperm). The results indicate that a decrease in the external solute did not account for growth reduction under conditions of water stress, and that decreased solute transport to the embryo may be important. The sucrose contents of "naked" embryos and of embryos from whole seeds were higher after PEG treatment, while reducing sugar contents were lower compared to non-stressed controls. The increased sucrose accumulation may be due to decreased sucrose hydrolysis.     

Anaerobiosis and Release from Dormancy in Apple Embryos: Leaching of (+/-) [C]Abscisic Acid and its Metabolites under Aerobic and Anaerobic Conditions

An anaerobic treatment released Pyrus malus L. cv Golden Delicious embryos from their primary dormancy. It also suppressed the inhibitory effect induced by exogenous abscisic acid (ABA) on after-ripened embryos. For the study of ABA metabolism, a two-step culture method was developed. Embryos in primary dormancy were cultivated aerobically in the presence of [¹⁴C]ABA (first culture). Some were directly analyzed to evaluate metabolism of absorbed ABA. The remaining embryos were cultivated on moist cotton without ABA, either in aerobic or anaerobic conditions (second culture). The amounts of ABA and its metabolites were measured both in the embryos and the water-leachates. After the second culture, the embryos showed a spectacular decrease in ABA content, with no difference between anaerobic and aerobic cultures. The amount of ABA glucose ester increased slightly in aerobiosis but diminished markedly in anaerobiosis. Radioactivity of the butanol fraction, which corresponded to polar conjugates, decreased considerably in anaerobiosis, whereas it increased in aerobiosis.

Analysis of the water-leachates indicated that, compared to aerobic conditions, anaerobiosis increased total leaching of radioactive materials (× 4.2) as well as leaching of ABA (× 1.4). In addition, anaerobiosis induced leaching of conjugates, such as ABA glucose ester and butanol-soluble metabolites. We concluded that the anaerobic treatment affects mainly membrane permeability.

     

Analysis of apoplastic solutes in the cortex of soybean nodules

Various techniques were used to extract solutes from the free space of intact soybean [ Glycine max (L.) Merr.] nodules. A variety of solutes (carbohydrates, amino acids, organic acids, ions) was found, but the major solute obtained with all methods was allantoic acid. Most work was done with a technique involving vacuum infiltration of intact detached nodules with water. This approach provided rapid sampling of the apoplastic solutes, and the results indicated that solutes were not derived from the xylem and phloem of ruptured vascular bundles. Infiltration of intact nodules with Fast Green showed dye penetration only to the barrier in the inner cortex, indicating that infected tissues did not contribute to solute composition. Although allantoic acid was the only ureide which could be detected in solute samples, no evidence was obtained for the presence of allantoinase in the cortical apoplast. The results suggest the transport of allantoic acid by an apoplastic route in nodules or the release of allantoic acid to the cortical apoplast in response to treatments which disrupt ureide export. Calculated values for solute concentrations in the cortical apoplast were in the hundred millimolar range, suggesting that apoplastic solutes may represent a significant osmotic component in the nodule cortex.     

In vitro development of bovine embryos as affected by different lots of bovine serum albumin and citrate

The effect of bovine serum albumin (BSA) lots on the development of in vitro-derived bovine embryos in synthetic oviductal fluid was investigated. Citrate concentration was determined for each lot of BSA, and then correlated with differences noted in the ability of BSA lots to support embryo development. Development of bovine embryos to the blastocyst stage was also compared after culture in chemically-defined medium with varying levels of citrate. There were distinct differences in the ability of the different BSA lots to support embryo development to the blastocyst stage (P相似文献   

The use of plant stress biomarkers in assessing the effects of desiccation in zygotic embryos from recalcitrant seeds: challenges and considerations

Zygotic embryos from recalcitrant seeds are sensitive to desiccation. In spite of their sensitivity, rapid partial dehydration is necessary for their successful cryopreservation. However, dehydration to water contents (WCs) that preclude lethal ice crystal formation during cooling and rewarming generally leads to desiccation damage. This study investigated the effects of rapid dehydration on selected stress biomarkers (electrolyte leakage, respiratory competence, rate of protein synthesis, superoxide production, lipid peroxidation, antioxidant activity and degree of cellular vacuolation) in zygotic embryos of four recalcitrant‐seeded species. Most biomarkers indicated differences in the levels of stress/damage incurred by embryos dried to WCs < and >0.4 g·g^?1, within species; however, these changes were often unrelated to viability and percentage water loss when data for the four species were pooled for regression analyses. Dehydration‐induced electrolyte leakage was, however, positively related with percentage water loss, while biomarkers of cellular vacuolation were positively related with both percentage water loss and viability. This suggests that electrolyte leakage and degree of cellular vacuolation can be used to quantify dehydration‐induced stress/damage. Biomarkers such as superoxide production, whilst useful in establishing the nature of the dehydration stress incurred may not be able to distinguish the effects of different WCs/drying times. Irrespective of which biomarker is used, the data suggest that understanding differences in desiccation sensitivity across recalcitrant‐seeded species will remain a challenge unless these biomarkers are related to a generic desiccation stress index that integrates the effects of percentage water loss and drying time.     

Influence of gelling agents on culture medium gel strength, water availability, tissue water potential, and maturation response in embryogenic cultures of Pinus strobus L.

Summary Maturation of somatic embryos of Pinus strobus L. was evaluated on media containing various types (agars and gellan gum), brands and concentrations of gelling agents in the presence of 80 μM ABA and 0.09 M sucrose. The media were characterized with respect to gel strength, water potential and water availability. Embryogenic tissue and somatic embryos cultured on medium with various concentrations of gellan gum were used to determine their water potential (Ψ). Regardless of the type of gelling agent used, gel strength increased with gelling agent concentration and was critical to the maturation response. High gel strength was associated with reduced water availability from the medium to the cultures. The water potential of gelled maturation medium remained constant between 0.4 and 1.0% gellan gum. It is concluded that the embryogenic tissue was exposed to varying amounts of water at the onset of and during the culture period, and that the amount of water in the culture environment in turn influenced the maturation response. Cotyledonary somatic embryos derived from gellan gum medium of high gel strength had a lower Ψ than somatic embryos matured on medium of lower gel strength. Once somatic embryos developed to the cotyledonary stage on the maturation medium, they were transferred to the germination medium. The germination frequency and the number of morphologically normal germinants were higher for somatic embryos matured on medium of high gel strength. Raising the concentration of the gelling agent in the maturation medium may be an alternative to the use of solutes to restrict water available to the embryogenic cultures.     

Incubation media affect the survival,pathway and time of embryo development in Neotropical annual fish Austrolebias nigrofasciatus (Rivulidae)

To analyse the survival, pathway and time of embryo development in the annual fish Austrolebias nigrofasciatus eggs were monitored in four liquid media and two damp media under experimental conditions for 130 days until their development was complete. Eggs kept in the same breeding water from oviposition remained in diapause I (DI) during all experiments. In constrast, up to the stage prior to entering diapause II (DII), the other media had no influence on development. Embryos at this stage (DII), however, show longer development time when treated in medium with water and powdered coconut shell so that about 80% of embryos remained in DII at 100 days. In contrast, all other treatments had a significantly lower proportion of embryos remaining in DII. When treated with Yamamoto's solution in humid media, embryos showed the fastest development. The first fully developed embryos (DIII) were seen at 27 days after oviposition. It took an average of 46–58 days for 50% of eggs in each treatment to reach DIII. Compared with other studies, survival in all incubation media was high at between 70 and 98%. Taken together, it can be concluded that all incubation media were found to be viable for maintaining embryos. Altering developmental trajectories through the manipulation of diapauses in different media makes this species a potential model organism for laboratory studies.     

Water relations parameters and tissue development in somatic and zygotic embryos of three pinaceous conifers

There is increasing interest in using somatic embryogenesis to meet the demand for high quality seedlings. However, in vitro production of propagules on a large scale depends on the optimization of the maturation and germination steps promoted by desiccation and subsequent imbibition of the embryo, respectively. It is therefore important to characterize zygotic and somatic embryos in terms of their water relations. Bound water, elastic modulus, osmotic potential at full turgor, and relative water content at turgor loss point were determined for somatic and zygotic embryos of western larch and loblolly pine and somatic embryos of white spruce at two developmental stages. These water relations parameters were derived from water-release curves obtained by suspending tissue samples in sealed vials over unsaturated NaCl solutions of known water potential. There was little difference in water relations parameters among species but marked stage dependency for bound water and elastic modulus. The amount of bound water was lowest in western larch somatic embryos (0.02-0.07) and highest in zygotic loblolly pine embryos (0.10-0.18). Elastic modulus ranged from 2.5 to 6.2 MPa in somatic embryos but varied between 1.4 and 1.8 MPa in zygotic embryos. The osmotic potential was lower in somatic embryos than in their zygotic counterparts. Our results show that water relations parameters are remarkably conservative across species but that, within a given species, these parameters are stage specific. It would seem, therefore, that desiccation protocols might be developed for a given developmental stage and applied across a range of species without the confounding effects of differences in water relations parameters.     

Hydrophobic tendency of polar group hydration as a major force in type I antifreeze protein recognition

The random network model of water quantitatively describes the different hydration heat capacities of polar and apolar solutes in terms of distortions of the water-water hydrogen bonding angle in the first hydration shell (Gallagher and Sharp, JACS 2003;125:9853). The distribution of this angle in pure water is bimodal, with a low-angle population and high-angle population. Polar solutes increase the high-angle population while apolar solutes increase the low-angle population. The ratio of the two populations quantifies the hydrophobicity of the solute and provides a sensitive measure of water structural distortions. This method of analysis is applied to study hydration of type I thermal hysteresis protein (THP) from winter flounder and three quadruple mutants of four threonine residues at positions 2, 13, 24, and 35. Wild-type and two mutants (VVVV and AAAA) have antifreeze (thermal hysteresis) activity, while the other mutant (SSSS) has no activity. The analysis reveals significant differences in the hydration structure of the ice-binding site. For the SSSS mutant, polar groups have a typical polar-like hydration, that is, more high-angle H-bonds than bulk water. For the wild-type and active mutants, polar groups have unusual, very apolar-like hydration, that is, more low-angle H-bonds than bulk water. This pattern of hydration was seen previously in the structurally distinct type III THPs (Yang & Sharp Biophys Chem 2004;109:137), suggesting for the first time a general mechanism for different THP classes. The specific shape, residue size, and clustering of both polar and apoler groups are essential for an active ice binding surface.