Water and sucrose regulate canola embryo development

The effect of water and sucrose on the growth and development of zygotic, 30-day-old canola ( Brassica napus L. cv. Bounty) embryos was examined in vitro by manipulating the levels of sucrose and/or sorbitol present in the culture medium. In some experiments, the medium water potential was allowed to vary with sucrose concentration, while in other experiments, the medium water potential was held constant by adding sorbitol to varying amounts of sucrose. Our results showed that embryos cultured on sorbitol alone exhibited two developmental patterns: embryos germinated precociously on media containing up to 0.70 M sorbitol, whereas embryos became yellow and quiescent on media with higher concentrations of sorbitol. For embryos cultured on media containing sucrose alone, three distinct developmental patterns were noted: at low sucrose concentrations, embryos germinated precociously; at intermediate concentrations, embryos continued to grow in an embryonic mode; and, at high concentrations, embryos became yellow and quiescent. Continued embryonic growth was never observed in embryos cultured on media containing sorbitol alone. Embryos never germinated precociously when cultured on media maintained at a constant water potential of -1.4 MPa, rather dry weight increased in these embryos with an increase in sucrose concentration. We envision the effect of sucrose on embryo growth and development to be nested within the effect of water availability. When water availability is restricted, embryos become quiescent. When water is available, embryos have the potential to grow, but the developmental growth pattern depends on the availability of sucrose. In the absence of sucrose, embryos germinate and initiate the transition to autotrophy. If sufficient sucrose is available, embryos remain photohet-erotrophic and continue to grow in an embryonic mode.     

Oleosins prevent oil-body coalescence during seed imbibition as suggested by a low-temperature scanning electron microscope study of desiccation-tolerant and -sensitive oilseeds

In order to clarify further the physiological role of oleosins in seed development, we characterized the oil-body proteins of several oilseeds exhibiting a range of desiccation sensitivities from the recalcitrant (Theobroma cacao L., Quercus rubra L.), intermediate (Coffea arabica L., Azadirachta indica A. Juss.) and orthodox categories (Sterculia setigera Del., Brassica napus L.). The estimated ratio of putative oleosins to lipid in oil bodies of Q. rubra was less than 5% of the equivalent values for rapeseed oil bodies. No oleosin was detected in T. cacao oil bodies. In A. indica cotyledons, oil bodies contained very low amounts of putative oleosins. Oil bodies both from C. arabica and S. setigera exhibited a similar ratio of putative oleosins to lipid as found in rapeseed. In C. arabica seeds, the central domain of an oleosin was partially sequenced. Using a low temperature field-emission scanning electron microscope, the structural stability of oil bodies was investigated in seeds after drying, storage in cold conditions and rehydration. Despite the absence or relative dearth of oleosins in desiccation-sensitive, recalcitrant oilseeds, oil bodies remained relatively stable after slow or fast drying. In A. indica seeds exposed to a lethal cold storage treatment, no significant change in oil-body sizes was observed. In contrast, during imbibition of artificially dried seeds containing low amounts of putative oleosins, the oil bodies fused to form large droplets, resulting in the loss of cellular integrity. No damage to the oil bodies occurred in imbibed seeds of Q. rubra, C. arabica and S. setigera. Thus the rehydration phase appears to be detrimental to the stability of oil bodies when these are present in large amounts and are lacking oleosins. We therefore suggest that one of the functions of oleosins in oilseed development may be to stabilize oil bodies during seed imbibition prior to germination. Received: 22 April 1997 / Accepted: 5 June 1997     

Free and bound amino acids and proteins in developing grains of rice with enhanced lysine/proteins

 Free amino acids were determined in developing seed of a rice mutant with enhanced grain lysine. This phenotype frequently has enhanced protein. Some free amino acids of developing seed are inversely related to the level of total amino acids in proteins of the mature grain. Amino acids that were enhanced in protein, including aspartic acid, threonine, methionine and lysine, were notably lower in the free amino-acid pool. Our conclusion is that mutant-developing grains process aspartate amino acids more rapidly than the controls. Conversely, arginine, valine and glutamic acid/glutamine accumulate as free amino acids with mutant/control ratios of 1.39, 1.29 and 1.12, respectively. Glutamic acid/glutamine in proteins of mature seeds is lower in the mutant than the control. ³H-lysine incorporation showed enhanced isotope incorporation into at least four proteins. One mutant protein was less actively labelled than analogous controls. The ³Hlysine pattern indicates processing modifications in this useful rice mutant. Received: 14 October 1996/Accepted: 8 November 1996     

Involvement of Ca2+-calmodulin in Cd2+ toxicity during the early phases of radish (Raphanus sativus L.) seed germination

The toxicity of Cd²⁺in vivo during the early phases of radish (Raphanus sativus L.) seed germination and the in vitro Cd²⁺ effect on radish calmodulin (CaM) were studied. Cd²⁺ was taken up in the embryo axes of radish seeds; the increase in fresh weight of embryo axes after 24 h of incubation was inhibited significantly in the presence of 10 mmol m^?3 Cd²⁺ in the external medium, when the Cd²⁺ content in the embryo axes was c. 1.1 μmol g^?1 FW. The reabsorption of K⁺, which characterizes germination, was inhibited by Cd²⁺, suggesting that Cd²⁺ affected metabolic reactivation. The slight effect of Cd²⁺ on the transmembrane electric potential of the cortical cells of the embryo axes excluded a generalized toxicity of Cd²⁺ at the plasma membrane level. After 24 h of incubation, Cd²⁺ induced no increase in total acid-soluble thiols and Cd²⁺-binding peptides able to reduce Cd²⁺ toxicity. Ca²⁺ added to the incubation medium partially reversed the Cd²⁺-induced inhibition of the increase in fresh weight of embryo axes and concomitantly reduced Cd²⁺ uptake. Equilibrium dialysis experiments indicated that Cd²⁺ bound to CaM and competed with Ca²⁺ in this binding. Cd²⁺ inhibited the activation of Ca²⁺-CaM-dependent calf-brain phosphodiesterase, inhibiting the Ca²⁺-CaM active complex. Cd²⁺ reduced the binding of CaM to the Ca²⁺-CaM binding enzymes present in the soluble fraction of the embryo axes of radish seeds. The possibility that Cd²⁺ toxicity in radish seed germination is mediated by the action of Cd²⁺ on Ca²⁺-CaM is discussed in relation to the in vivo and in vitro effects of Cd²⁺.     

Recruitment and survival of native annual Trifolium species in the highlands of Ethiopia

Germination and survival of indigenous annual Trifolium species were studied. Seedlings naturally emerging in the field and from sown seeds in pots were regularly counted and uprooted, and survival was studied by monitoring colour‐coded seedlings. Differences in recruitment of Trifolium species were strongly related to the rainfall pattern. In the fallow (crop) lands, no seedling survived the dry season between the short and main rainy periods during the year, while in the natural pasture, 8% of the seedlings survived into the main growing (rainy) season. Occasional rains occurring in the dry period (between the two rainy periods) also induced successions of germination and seedlings death, and therefore depleted the soft seed reserve in the soil by the beginning of the main growing season. As temperature fluctuations were minimal during the main rainy season, the rate of seed softening was low, affecting new germinations. This had a significant impact on the quality of the natural pastures on which livestock in the highlands are dependent.     

Plant-growth regulators derived from the sweetener stevioside

A series of isosteviol derivatives such as aromatic esters and amino acid amides were synthesized. Their effects on seed germination and root elongation of the crop plants, Capsicum annuum, Lens culinaris medicus, Lycopersicon esculentum, Trifollium spp. and Triticum vulgare, were studied. The derivatives could be arranged into four groups: (1) seed germination inhibitors (2) root elongation inhibitors (3) root elongation inducers (4) general inhibitors.     

Plantlets from encapsulated micropropagated buds of M.26 apple rootstock

In order to be considered usable as synthetic seeds, encapsulated explants sown underin vitro orex vitro conditions must be able to produce whole plantlets. Ninety percent of non-encapsulated M.26 apple rootstock single nodes produced a plantlet (i.e., a well-formed shoot with a root system) after 30 days of culturein vitro if the explants were previously given a 24-hour treatment with 24.6 μM IBA and 15 g 1⁻¹ sucrose in darkness. In contrast, when the single nodes were encapsulated in a calcium-sodium alginate bead immediately after the same treatment only 10% of the encapsulated explants formed a plantlet. Addition of growth regulators to the artificial endosperm and culture of the single nodes for root primordia initiation for 3, 6 or 9 days in darkness before encapsulation allowed production of 58%, 60% and 66% of plantlets, respectively.     

Comparison among DNA polymerases 1, 2 and 3 from maize embryo axes. A DNA primase activity copurifies with DNA polymerase 2

Three DNA polymerase activities, named 1, 2 and 3 were purified from maize embryo axes and were compared in terms of ion requirements, optimal pH, temperature and KCl for activity, response to specific inhibitors and use of templates. All three enzymes require a divalent cation for activity, but main differences were observed in sensitivity to inhibitors and template usage: while DNA polymerases 1 and 2 were inhibited by N-ethyl maleimide and aphidicolin, inhibitors of replicative-type enzymes, DNA polymerase 3 was only marginally or not affected at all. In contrast, DNA polymerase 3 was highly inhibited by very low concentrations of ddTTP, an inhibitor of repair-type enzymes, and a 100-fold higher concentration of the drug was needed to inhibit DNA polymerases 1 and 2. Additionally, DNA polymerases 1 and 2 used equally or more efficiently the synthetic template polydA-oligodT, as compared to activated DNA, while polymerase 3 used it very poorly. Whereas DNA polymerases 1 and 2 shared properties of replicative-type enzymes, DNA polymerase 3 could be a repair-type enzyme. Moreover, a DNA primase activity copurified with the 8000-fold purified DNA polymerase 2, strenghtening the suggestion that polymerase 2 is a replicative enzyme, of the -type. This DNA primase activity was also partially characterized. The results are discussed in terms of relevant data about other plant DNA polymerases and primases reported in the literature.