Changes in water status and proline and abscisic acid concentrations in developing somatic embryos of pedunculate oak (Quercus robur) during maturation and germination

Somatic embryos of oak (Quercus robur L.) were matured on P24 media differing in gel strength (0.8, 0.9 and 1.0% (w/v) agar). Viscosity and osmotic potential (Psipi,medium) of the media were determined. Developing cotyledonary embryos were analyzed at maturity Stages I-III for water content, osmotic potential (Psipi,embryo) and concentrations of abscisic acid (ABA) and proline. Proliferation of embryogenic tissue, germination rates and the number of embryos formed were also determined in order to relate embryo quality to physiological parameters. Viscosity increased with agar concentration, a phenomenon apparently related to water availability. Many Stage III embryos with high germination potentials were obtained on P24 medium containing 1.0% agar. Embryo water content decreased progressively from 94 to 80% during embryo maturation. Stage I and II embryos that matured on media containing 0.8 or 0.9% agar had similar values of Psipi,embryo, whereas Psipi,embryo of Stage III embryos that matured on medium containing 1.0% agar was significantly lower, although Psipi,medium was unaffected by gel strength. Stage III embryos showed a nearly 16-fold increase in proline concentration and a 50% decrease in ABA concentration compared with Stage I embryos. We conclude that tissue water status and a complex relationship between ABA and proline concentrations, modulated by medium gel strength, are important factors in the maturation process and the quality of oak somatic embryos.     

Slash pine (Pinus elliottii Engelm.) somatic embryogenesis II. Maturation of somatic embryos and plant regeneration

Maturation of slash pine (Pinus elliottii Engelm.) somatic embryos was achieved using two protocols, each starting with a different agar incubation step to deplete plant growth regulators (PGRs) used in previous cultural steps. Strength of maturation medium (single vs. double) was found important in the first protocol to develop normal, mature embryos. In the second protocol, abscisic acid (ABA) concentrations (0, 15 and 30 M) and carbohydrate sources were tested for embryo maturation. Thirty M ABA and 6% maltose were deemed the best combination. Embryo germination was accomplished in a continuously lighted environment and embryos receiving a cold pretreatment (4 °C in darkness for 16 days) germinated better than embryos which did not receive cold pretreatment. With a survival rate of 33% after acclimation in a mist system, more than 25 plants from somatic embryos have been established in a greenhouse. Incompletely germinated embryos (lacking roots) were rooted via adventitious rooting techniques and subsequently established in the greenhouse. All established plants obtained from somatic embryogenesis appear normal in morphology.     

Screening and verification of the factors influencing somatic embryo maturation of Larix olgensis

With embryogenic callus of Larix olgensisis, we investigated the effects of inositol, glutamine, casein hydrolysate, carbohydrate, abscisic acid and silver nitrate concentration on the maturation of the somatic embryo.Three dominant factors emerged, and we developed a response surface model based on the Box-Behnken design.We defined the optimal conditions for the maturation of somatic embryos. The contents of abscisic acid, silver nitrate, sucrose and casein hydrolysis significantly affected the amount of maturing embryos, but inositol, maltose and glutamine had no effect. By establishing a response surface model with multiple factors, we predicted that the optimal number of L. olgensis somatic embryos was 204 ± 4 g~(-1) on basal medium, containing 18.28 mg L~(-1) abscisic acid,5.46 mg L~(-1) silver nitrate and 82.67 g L~(-1) sucrose. In the verification experiments, the addition of 20 mg L~(-1) abscisic acid, 5 mg L~(-1) silver nitrate and 80 g L~(-1) sucrose to BM yielded an average of 202.06 somatic embryos per gram. These results should guide large-scale breeding of L. olgensis.     

Charcoal affects early development and hormonal concentrations of somatic embryos of hybrid larch

Embryogenic tissue of hybrid larch (Larix x marschlinsii Coaz) was multiplied on Medium M (modified MSG medium supplemented with the plant growth regulators (PGRs) 2,4-dichlorophenoxyacetic acid (2,4-D; 9 microM) and N-6-benzyladenine (2.25 microM)). After 1 week, cultures were transferred to either MSG lacking PGRs (Medium C-) or MSG lacking PGRs but supplemented with 1% activated charcoal (Medium C+). Embryos were sampled after 1 week on Medium M, C- or C+. Embryos were analyzed by ELISA for abscisic acid (ABA), abscisic acid-glucose ester, 2,4-D, indole-3-acetic acid (IAA), indole-3-aspartate (IAAsp), zeatin (Z), zeatin riboside (ZR), isopentenyladenine (iP) and isopentenyladenosine (iPA). Transfer of embryos to Medium C+ reduced the embryo concentrations of 2,4-D and iPA, but resulted in elevated concentrations of IAA, IAAsp, ABA, Z, ZR and iP. Charcoal reduced 2,4-D concentrations of embryos by an order of magnitude greater than PGR-free medium alone. Charcoal affected embryo concentrations of five of the eight PGRs quantified. Use of either C+ or C- medium as part of the maturation protocols also affected germination and plantlet establishment of the embryos. A 1-week treatment on Medium C+ positively influenced plantlet establishment and generally reduced variability during both germination and plantlet establishment.     

火炬松体细胞胚胎发生和干化体细胞的氧化物酶活性(英文)

培养于附加2,4-D、BA和KT的愈伤组织诱导培养基上的火炬松成熟合子胚在培养3-9周后形成白色、半透明、有光泽的粘性愈伤组织。这类愈伤组织形成于成熟合子胚的子叶,但当用NAA或者IBA代替愈伤组织诱导培养基中的2,4-D时,它的诱导频率明显降低。这种粘性愈伤组织在分化培养基上形成体细胞胚。体细胞胚经过去50μm ABA和8.5%PEG600处理后成为耐干化胚。扫描电镜观察表明,萌发处理36小时后,耐干化胚恢复到干化处理之前的状态且大小和形态正常,而不耐干化胚不能恢复到干化处理之前的状态且表面撕裂。过氧化物酶活性的分析结果表明,耐干化胚有更高的过氧化物酶活性。耐干化胚的高过氧化物酶活性可能与催化H2O2的分解和保护体细胞胚免受氧化的伤害有关。     

Defense chemistry of cyanogenic Eucalyptus cladocalyx seedlings is affected by water supply

Cyanogenesis is a widespread and effective defense mechanism in plants. Published evidence suggests that cyanogenic capacity (i.e., cyanogenic glycoside concentration) is enhanced in response to water stress, although potentially confounding variables preclude a definite conclusion. We used highly cyanogenic Eucalyptus cladocalyx var. nana F. Muell. seedlings grown with varying amounts of water and nitrogen (N) to determine the relationship between cyanogenic capacity and water stress. We also examined whether variation in cyanogenic capacity affects phenolic biosynthesis because both pathways use phenylalanine as a substrate. Cyanogenic capacity in fully expanded leaves increased 70% in response to moderate water stress when N availability was high but only 30% when growth was N-limited. Absolute cyanogenic capacity also increased with increasing N supply. Total phenolics and condensed tannins decreased with increasing N supply, but these compounds were unaffected by water stress. We conclude that, under the influence of water stress, the enhanced demand for phenylalanine for cyanogenic glycoside biosynthesis can be sustained by enhanced shikimate pathway flux without affecting phenolic metabolism.     

Thidiazuron-induced somatic embryos,their multiplication,maturation, and conversion in <Emphasis Type="Italic">Cinnamomum pauciflorum</Emphasis> Nees (Lauraceae)

Thidiazuron (TDZ) induced somatic embryogenesis from immature zygotic embryos in Cinnamomum pauciflorum Nees while 2,4-dichlorophenoxyacetic acid (2,4-D), 6-benzylaminopurine (BA) or picloram only induced callus and/or adventitious buds. The highest induction frequency for somatic embryogenesis was achieved with MS medium (Murashige and Skoog in Physiol Plant 15:473–497 1962) supplemented with 2.5 μM TDZ using torpedo-shaped embryos (3–5 mm in length) as explants. In addition, induction medium was supplemented with 0.8 g l⁻¹ casein, 0.4 g l⁻¹ glutamine, and 10 g l⁻¹ sucrose. Somatic embryos (SEs) initiated from root tips or hypocotyls without callus formation. SEs were maintained and multiplied via secondary somatic embryogenesis. Embryo maintenance medium was similar to induction medium except that TDZ was reduced to 0.5 μM. Secondary embryogenesis was enhanced by supplementation of 5 g l⁻¹ activated charcoal in the culture. The best medium for embryo maturation was MS medium containing 30 g l⁻¹ sucrose and 5 g l⁻¹ Phytagel without plant growth regulators. A typical mature SE consisted of two large cotyledons and a short embryo proper. Approximately 82% of selected mature SEs were able to germinate and 63% could convert into plantlets on germination medium that was composed of half strength MS medium salts, 10 g l⁻¹ sucrose, 3 g l⁻¹ Phytagel, and 5 g l⁻¹ activated charcoal.     

Morphology and ontogeny of directly differentiating shoot buds and somatic embryos in Santalum album L.

Santalum album L.is a commercially important tree that yields essential oil of high medicinal value.Regeneration research through organogenesis and embryogenesis has been documented but no report depicts comparative ontogeny of directly differentiating shoot buds(SB)and somatic embryos(SE).In the present study aseptic seedling derived hypocotyl segments(HC)and hypocotyl+root junction(HC+R)were used to induce SBs and SEs,respectively.Ontogenic differences between the structures were confirmed using scanning electron microscopy and histological analysis.MS medium containing 6-benzyladenine or BA(2.5μM)produced highest number of direct SB,while MS+BA(7.5μM)proved suitable for higher frequency of SE differentiation.The differentiating structures attained growth when transferred to MS medium containing a combination of BA and anaphthalene acetic acid or silver nitrate(AgNO3).A combination of indole-3-butyric acid and silver nitrate(AgNO3)in half-strength woody plant medium and lesser osmotic concentration(2%sucrose),induced rhizogenesis.     

<Emphasis Type="Italic">Pinus halepensis</Emphasis> somatic embryogenesis is affected by the physical and chemical conditions at the initial stages of the process

Pinus halepensis has been described as a drought-tolerant species with high plasticity to growth in different environments. Its eco-physiological characteristics could facilitate the use of this species in large afforestations in the future scenery of climate change. Somatic embryogenesis is a biotechnological tool with potential for large-scale clonal propagation. In order to establish an improved regeneration protocol for Pinus halepensis, the effects of different temperatures (18, 23, and 28 °C) and water availability conditions (2, 3, and 4 g L^?1 Gelrite^®), during initiation of embryonal masses on the rate of initiation, proliferation, maturation, and the number of embryos developed, were evaluated. It was found that environmental conditions during the initiation stage of Pinus halepensis somatic embryogenesis influence the success of initiation and proliferation. In contrast, there was no effect of these conditions on the maturation rates and the number of somatic embryos. Somatic embryos were obtained in all treatments tested, indicating that plants can be produced from extreme conditions of induction, such as high temperatures (28 °C) and low water availability conditions (4 g L^?1).     

Endogenous ascorbic acid modulates meristem reactivation in white spruce somatic embryos and affects thymidine and uridine metabolism

Previously, we demonstrated that, in some cell lines, exogenous applications of ascorbic acid (ASC) enhance the conversion frequency of white spruce (Picea glauca (Moench) Voss) somatic embryos, by stimulating mitotic activity in the apical meristems. To examine this event in more detail, we investigated the effects of ASC on de novo, salvage and degradation pathways of pyrimidine metabolism by following the metabolic fate of (14)C-labeled orotic acid, thymidine, uridine and uracil in shoot and root poles of germinating embryos, after altering the cellular ASC content of the embryos. Alterations in endogenous ASC content did not affect the utilization of either orotic acid or uracil, but affected the metabolism of thymidine and uridine. Specifically, a lowering of endogenous ASC content by applications of lycorine (L), an inhibitor of the last enzyme of the ASC de novo biosynthetic pathway, resulted in a lower embryo conversion frequency, as well as a reduced percentage of thymidine and uridine incorporated into nucleotides and nucleic acids. The reduction in thymidine and uridine anabolism was mainly ascribed to the decreased activities of thymidine kinase (TRK) and uridine kinase (URK), the respective salvage enzymes of thymidine and uridine, measured in L-treated embryos. These effects were solely a result of a decrease in endogenous ASC content because applications of ascorbic acid plus lycorine (ASC + L) increased embryo conversion frequency, thymidine and uridine salvage activities, and TRK and URK activities to near control values. Inclusion of exogenous ASC in the germination medium did not affect the percentage of embryos able to convert to viable plantlets, although it increased thymidine and uridine utilization for nucleic acid synthesis in the shoot and root poles of the embryos. Taken together, these findings confirm that cellular ASC plays a key role in the reactivation of the apical meristems of germinating white spruce somatic embryos.     

Storage lipid dynamics in somatic embryos of Norway spruce (Picea abies): histochemical and quantitative analyses

Adequate storage compounds are a prerequisite for successful development during the later stages of somatic embryogenesis; however, the critical amount of reserves below which somatic embryos fail to mature and germinate has not been determined. We analyzed storage lipids during Norway spruce (Picea abies (L.) Karst.) somatic embryogenesis. As maturation progressed, lipids, which were stored as lipid bodies in the cytoplasm, were localized first in suspensor cells of the early embryos, and later in the embryonic root pole, superficial layers of the hypocotyl and in cotyledons. The concentration of total lipids exhibited marked variation, with values peaking during cotyledon development and then decreasing during maturation. Linoleic (18:2), oleic (18:1), palmitic (16:0) and 5,9-octadecenic (5,9-18:2) acids were the most abundant fatty acids in embryos. As embryos developed, linoleic acid concentration increased slightly, whereas oleic acid concentration decreased. Oleic acid was the most prominent component of the fatty acid spectrum in isolated dormant zygotic embryos and megagametophytes. Addition of 5% polyethylene glycol to the medium during somatic embryo maturation caused a shift in the fatty acid spectrum toward that of zygotic embryos. During maturation, changes in the exogenous carbohydrate supply had no significant effect on total lipid concentration in mature embryos. A marked decrease in lipid concentration was detected during desiccation, indicating the importance of adequate lipid reserves during this developmental stage. The lipid content of zygotic embryos differed considerably with harvest year and location, suggesting that zygotic embryo data cannot be an indicator of somatic embryo quality.     

Production of phenols and charcoal from bagasse by a rapid continuous pyrolysis process

Summary Tar and charcoal could be produced in high yields from bagasse by applying a rapid continuous pyrolysis process at a relatively low temperature. The ether extractives of the pyrolytic tar and oil amounted to 9.4% based on bagasse. Phenols represented 79% of these extractives. Gas chromatographic separation showed that guaiacol and its derivatives constituted 38% of the identified simple phenols. There were much smaller amounts of syringol and none at high pyrolysis temperatures. Depithing did not reduce the ash content of the charcoal, but it yielded an environmentally clean charcoal containing practically no sulfur or nitrogen. It was necessary to remove the fine particle size fraction of the bagasse after grinding in order to reduce the ash content of the charcoal. The carbon content of the charcoal increased rapidly with increasing temperature, and reached 96% at 720°C. The charcoal had a remarkably high adsorption capacity despite the fact that it had not been subjected to any activation treatment.The first two authors wish to express their thanks to the Institut für Holzchemie, Hamburg for providing facilities during this work and to the Alexander von Humboldt-Stiftung, and the Deutsche Forschung Gemeinschaft for their financial support     

Changes in soil chemistry and element uptake by Oak seedlings after application of soil amendment

ABSTRACT

The purpose of this study is to investigate the effect of cellulosic wastes and nanoscale zero-valent iron (nZVI) on the soil properties and the nutrient uptake by Quercus castaneifolia C.A.Mey seedlings. The seedlings were planted in pots (unpolluted soil) and cadmium (Cd) was added to the pots in three concentrations (10, 20, 30?mg.kg^?1). Cellulosic wastes were mixed with the soil (at the same time as the planting) in four levels (W: 0, 10, 20, 30?g.100g^?1 soil). The reduction method was used to prepare nZVI, then the nanoparticles were injected into the polluted pots in four levels (N: 0, 1, 2, and 3?mg.kg^?1). The lowest concentration of Cd in leaves, stems and roots and for all the contamination levels was observed in N3. All the cellulosic wastes and N3 significantly increased the soil pH at all the levels of contamination. An increasing trend of the soil carbon (C) content was observed by adding the cellulosic wastes. The highest pH and C content were observed in W3. The lowest bioavailable Cd contents for all the contamination levels was observed in N3. According to the results, nZVI and cellulosic wastes could help the plants establishing in heavy metal-polluted lands.     

Integrated management of the charcoal disease by silviculture,chemical and biological methods in forest parks

Biscogniauxia mediterranea (De Not.) Kuntze, is a widespread fungus, causes outbreaks of Charcoal disease on cork oak and dieback thousands oak trees in recent years in Iran. Yet no efficient and effective management methods have been found to control the disease. The present study aimed to provide a suitable method for charcoal disease management through silviculture operations, systemic pesticides and biological compounds. The results showed that, simultaneous implementation of silvicultural practices (scarification+sanitation), use of systemic fungicides and Trichoderma compounds, reduce the number and depth of cankers, increase callus thickness, and ultimately improve health of the diseased trees. But disease severity and fungi activity were increased when scarification operation implements alone. The greatest effectiveness of the treatments and health condition improvement of the diseased trees was observed 12 months after applying the treatments. Then, preventive and therapeutic effectiveness of the treatments reduced over time. Sanitation and scarification practices can increase soil permeability and also destroy the larvae, pupa or adult insects and oak borer beetles that play an important role in disease outbreaks. Therefore, simultaneous implementation of sanitation and scarification practices with application of biological and systemic compounds can reduce primary source of pathogen inoculum and help trees to recover their health.     

Biology and efficiency of phytoseiid mites as affected by cucumber leaves

The biology of the phytoseiid mitesPhytoseiulus persimilis Athias-Henriot,Phytoseius finitimus Ribaga andAmblyseius gossipi Elbadry was studied using the cucumber leaves as a rearing substrate. The consumption of the two-spotted spider miteTetranychus urticae Koch by the different stages ofP. persimilis was higher than that of their counterparts of the other two predators. The present study revealed that cucumber leaves were adequate fot the development and reproduction ofP. persimilis andP. finitimus, while this rearing substrate was inadequate for the biology ofA. gossipi.     

Stand-level allometry in Pinus taeda as affected by irrigation and fertilization

Changing environmental conditions have the potential to alter allometric relationships between plant parts, possibly leading to ecosystem-level feedbacks. We quantified allometric shifts in field-grown loblolly pine (Pinus taeda L.) in response to altered resource availability based on data from multiple harvests to correct for size-related changes in biomass partitioning. A replicated factorial arrangement of irrigation and fertilization treatments was applied for 4 years to an 8-year-old loblolly pine plantation on a well-drained, low fertility site in North Carolina. Destructive and nondestructive growth measurements were used to develop treatment-specific regressions to estimate stand-level biomass for ephemeral and perennial plant parts, both above- and belowground. Stand-level allometric analysis indicated that irrigation increased biomass partitioning to fine roots and decreased partitioning to foliage, relative to other plant parts. Fertilization increased partitioning to perennial tissues (coarse roots, taproots, and branches) and decreased partitioning to ephemeral tissues (foliage and fine roots). Changes in allometry were small (< 6 %) but statistically significant, indicating that biomass partitioning in loblolly pine changes with altered resource availability, but is probably under strong ontogenetic control.     

Gas exchange and water relations of Fraxinus americana affected by flurprimidol

Effects of flurprimidol on plant water relations and leaf gas exchange were investigated in one-year-old white ash (Fraxinus americana L.) seedlings subjected to soil water deficits. Flurprimidol (20 mg kg(-1) of soil equivalent) was applied to the soil surface of pot-grown seedlings after shoot growth was completed. Two months after flurprimidol application, water was withheld from one-half of the seedlings. Leaf water relations and gas exchange parameters were measured 5, 7, 10, 14, 18 and 22 days after withholding water. Under both irrigated and nonirrigated conditions, flurprimidol treatment resulted in reduced net CO(2) assimilation rate and transpirational water loss of seedlings as a result of decreased stomatal conductance. Consequently, flurprimidol-treated seedlings had higher leaf water potential and relative water content than untreated seedlings. Nonirrigated flurprimidol-treated seedlings also had greater turgor and sap osmolality and lower osmotic potential at full turgor than seedlings in the other treatments, indicating that flurprimidol increased osmotic adjustment. Under water-stress conditions, water use efficiency was lower and gas exchange efficiency was higher in flurprimidol-treated seedlings than in untreated seedlings, suggesting that flurprimidol treatment enhances survival of plants subjected to soil water deficits.