Azolla-Anabaena azollae Relationship: V. N(2) Fixation, Acetylene Reduction, and H(2) Production

In order to characterize the reactions catalyzed by nitrogenase in the Azolla-Anabaena association, ¹⁵N₂ fixation, C₂H₂ reduction, and ATP-dependent H₂ production were measured in both the Azolla-Anabaena complex and in the alga isolated from the complex.     

Improving Soybean (Glycine max L.) N2 Fixation under Stress

Soybean is a major leguminous plant that has the ability to establish a symbiotic association with the N-fixing bacteria, Bradyrhizobium japonicum. Soils are usually subjected to stress including salinity, drought, acidity, and suboptimal root zone temperature, adversely affecting the symbiotic process between soybean and the bacteria. One of the important processes affecting the performance of soybean under stress is the inhibited exchange of symbiosis-related signaling molecules, specifically genistein, between the host legume and B. japonicum during the initiation of symbiosis. Interestingly, inoculation of B. japonicum with the signal molecule genistein can partially or completely alleviate the stress. Understanding the techniques and the precise molecular pathways, which may be influenced by the signaling molecules during the stress, can be useful to determine parameters that enhance the plant’s ability to cope with stress. For example, the use of proteomic techniques to identify proteins expressed under stress can help characterize those proteins and their involvement in stress. Biotechnological-genetic techniques, either breeding or transformation, are also among the effective methods of improving soybean’s ability to fix N₂ under stress. This can be achieved by identifying the genes, which may be expressed under stress in tolerant bacterial and plant species, and inserting them into the non-tolerant species. This article highlights some important advances in soybean N₂ fixation under different stress conditions, and reviews some of the techniques used to improve the ability of plants and bacteria to efficiently fix N₂ under stress.     

野生大豆（Glycine soja）酰脲含量与根瘤固氮活力的关系

野生大豆（Ｇｌｙｃｉｎｅｓｏｊａ）酰脲含量与根瘤固氮活力的关系朱长甫,苗以农,刘学军,许守民（东北师范大学生命科学学院,长春１３００２４）郑惠玉,徐豹（吉林省农业科学院大豆研究所,公主岭１３６１００）关键词：野生大豆,固氮活力,酰脲,蛋白质根据固氮豆...     

Interrelationship between Leaf Gas-Exchange Characteristics,Area Leaf Mass,and Yield in Soybean (Glycine max L. Merr) Genotypes

Variability in leaf gas-exchange traits in thirteen soybean (Glycine max L. Merr) genotypes was assessed in a field experiment conducted at high altitude (1 950 m). Leaf net photosynthetic rate (P ^N) exhibited a high degree of variability at all the growth stages studied. P _N and other gas-exchange parameters exhibited a seasonal pattern that was similar for all the genotypes. P _N rate was highest at seed filling stage. P _N was positively and significantly associated with aboveground dry matter and seed yield. The area leaf mass (ALM) exhibited a strong positive association with leaf P _N, aboveground dry matter, and seed yield. The positive association between ALM, P _N, and seed yield suggests that this simple and easy to measure character can be used in breeding programmes as a surrogate for higher photosynthetic efficiency and eventually higher yield.     

Translocation of Sulfate in Soybean (Glycine max L. Merr)

Sulfate translocation in soybean (Glycine max L. Merr) was investigated. More than 90% of the sulfate entering the shoot system was recoverable in one or two developing trifoliate leaves. In young plants, the first trifoliate leaf contained between 10 to 20 times as much sulfate as the primary leaves, even though both types of leaf had similar rates of transpiration and photosynthesis. We conclude that most of the sulfate entering mature leaves is rapidly loaded into the phloem and translocated to sinks elsewhere in the plant. This loading was inhibited by carbonylcyanide m-chlorophenylhydrazone and selenate. At sulfate concentrations below 0.1 millimolar, more than 95% of the sulfate entering primary leaves was exported. At higher concentrations the rate of export increased but so did the amount of sulfate remaining in the leaves. Removal of the first trifoliate leaf increased two-fold the transport of sulfate to the apex, indicating that these are competing sinks for sulfate translocated from the primary leaves. The small amount of sulfate transported into the mesophyll cells of primary leaves is a result of feedback regulation by the intracellular sulfate pool, not a consequence of their metabolic inactivity. For example, treatment of plants with 2 millimolar aminotriazole caused a 700 nanomoles per gram fresh weight increase in the glutathione content of primary leaves, but had no effect on sulfate aquisition.     

Interaction of Combined Nitrogen with the Expression of Root-Associated Nitrogenase Activity in Grasses and with the Development of N(2) Fixation in Soybean (Glycine max L. Merr.)

Soluble root N concentrations of corn, sorghum, pearl millet, rice, wild rice, and soybeans were determined and related to measurements of nitrogenase activity and changes in availability of combined N to plants. In corn, sorghum, and pearl millet, applications of fertilizer N increased soluble root N concentrations, but root-associated nitrogenase activity was negligible in control and treated plants. Applications of NH₄NO₃ to rice increased the water soluble root N concentrations and inhibited root-associated nitrogenase activity. In wild rice, root-associated nitrogenase activity was absent during vegetative growth and developed at the reproductive growth stage. The soluble root N concentration decreased progressively as wild rice grew indicating that the availability of combined N in the root environment declined. Therefore, development of nitrogenase activity in wild rice is associated with the change in availability of combined N in the root environment. The development of nitrogenase activity in wild rice was probably not due to colonization of roots by N₂-fixing bacteria because most probable numbers of recovery did not significantly vary throughout the plants' growth cycle. In field-grown soybeans with or without fertilizer N application, we also observed a relationship between a decrease in soluble root N concentration and the development of nitrogenase activity.     

Early Endosperm Development in Ovules of Soybean, Glycine max (L) Merr. (Fabaceae)

Endosperm development was studied in normally setting flowersand pods of soybean from anthesis to a pod length of 10–20mm. The free-nuclear stage following double fertilization istypified by loss of starch and increasing vacuolation. The cytoplasmprovides evidence of extensive metabolic activity. Wall ingrowths,already present at the micropylar end of the embryo sac wallprior to fertilization, develop along the lateral wall of thecentral cell as well as at the chalazal endosperm haustorium.Endosperm cellularization begins when the embryo has developeda distinct globular embryo proper and suspensor. Cellularizationstarts at the micropylar end of the embryo sac as a series ofantidinal walls projecting into the endosperm cytoplasm fromthe wall of the central cell. The free, growing ends of thesewalls are associated with vesicles, microtubules, and endoplasrnicreticulum. Pendinal walls that complete the compartmentalizalionof portions of the endosperm cytoplasm are initiated as cellplates formed during continued mitosis of the endosperm nuclei.Endosperm cell walls are traversed by plasmodesmata. This studywill provide a basis for comparison with endosperin from soybeanflowers programmed to abscise. Glycine max, soybean, endosperm, ovules     

Sterol Composition and Accumulation in Glycine max (L.) Merr Leaves under Different Filtered Sunlight Conditions

Soybean plants (Merr) were grown in the field in three plots. Sixteen days after sowing, two plots were covered with blue and red polyvinylchloride filters (0.45 millimeter thick) and one remained uncovered as control. Leaves of all plots were analyzed for total, free, esterified, and glycosidic sterols at two successive stages of plant growth (flowering and podripening).     

Efficiency of Nitrogen Assimilation by N(2)-Fixing and Nitrate-Grown Soybean Plants (Glycine max [L.] Merr.)

Nodulated and non-nodulated (not inoculated) soybeans (Glycine max [L.] Merr. cv Wells) were grown in controlled environments with N₂ or nonlimiting levels of NO₃⁻, respectively, serving as sole source of nitrogen. The efficiency of the N₂-fixing plants was compared with that of the nitrate-supplied plants on the basis of both plant age and plant size. Efficiency evaluations of the plants were expressed as the ratio of moles of carbon respired by the whole plant to the moles of nitrogen incorporated into plant material.

Continuous 24-hour CO₂ exchange measurements on shoot and root systems made at the beginning of flowering (28 days after planting) indicated that N₂-fixing plants respired 8.28 moles of carbon per mole of N, fixed from dinitrogen, while nitrate-supplied plants respired only 4.99 moles of carbon per mole of nitrate reduced. Twenty-one-day-old nitrate-supplied plants were even more efficient, respiring only 3.18 moles of carbon per mole of nitrate reduced. The decreased efficiency of the N₂-fixing plants was not due to plant size since, on a dry weight basis, the 28-day-old N₂-fixing plants were intermediate between the 28- and 21-day-old nitrate-supplied plants.

The calculated efficiencies were predominantly a reflection of root-system respiration. N₂-fixing plants lost 25% of their daily net photosynthetic input of carbon through root-system respiration, compared with 16% for 28-day-old nitrate-supplied plants and 12% for 21-day-old nitrate-supplied plants. Shoot dark respiration was similar for all three plant groups, varying between 7.9% and 9.0% of the apparent photosynthate.

The increased respiratory loss by the roots of the N₂-fixing plants was not compensated for by increased net photosynthetic effectiveness. Canopy photosynthesis expressed on a leaf area basis was similar for 28-day-old N₂-fixing plants (15.5 milligrams CO₂ square decimeter per hour) and 21-day-old nitrate-supplied plants (14.5 milligrams CO₂ square decimeter per hour). Both were similar in total canopy leaf area. The larger nitrate-supplied plants (28-day-old) had lower photosynthetic rates (12.5 milligrams CO₂ square decimeter per hour), presumably due to self-shading of the leaves.

These data indicate that, during the early stages of plant development, dependence solely on N₂-fixation is an expensive process compared to nitrate reduction in nitrate-supplied plants, since the N₂-fixing plants retained 8% to 12% less of their photosynthate as dry matter.

     

Genetic diversity of fast-growing rhizobia that nodulate soybean ( Glycine max L. Merr)

The fast-growing Rhizobium sp. strain NGR234, isolated from Papua New Guinea, and 13 strains of Sinorhizobium fredii, isolated from China and Vietnam, were fingerprinted by means of RAPD, REP, ERIC and ARDRA. ERIC, REP and RAPD markers revealed a considerable genetic diversity among fast-growing rhizobia. Chinese isolates showed higher levels of diversity than those strains isolated from Vietnam. ARDRA analysis revealed three different genotypes among fast-growing rhizobia that nodulate soybean, even though all belonged to a subcluster that included Sinorhizobium saheli and Sinorhizobium meliloti. Among S. fredii rhizobia, two strains, SMH13 and HH303, might be representatives of other species of nitrogen-fixing organisms. Although restriction analysis of the nifD–nifK intergenic DNA fragment confirmed the unique nature of Rhizobium sp. strain NGR234, several similarities between Rhizobium sp. strain NGR234 and S. fredii USDA257, the ARDRA analysis and the full sequence of the 16S rDNA confirmed that NGR234 is a S. fredii strain. In addition, ARDRA analysis and the full sequence of the 16S rDNA suggested that two strains of rhizobia might be representatives of other species of rhizobia.     

Intraspecific Variations in Rhythmic Leaf Movements in Soybean (Glycine max L. Merr)

Intraspecific variations in the rhythmic (circadian) leaf movementsin soybean (Glycine max L. Merr) were investigated. Resultsindicated (1) that whereas there was no significant differencein the free-running periods in continuous darkness among thevarious cultivars tested, there was a significant differencein the response of the free-running rhythms to continuous light;(2) that there was a difference in the phase shifts to singlelight pulses among the cultivars. (The assumption that the responseof the rhythms to continuous light is related functionally tothe phase response to single light pulses was examined usingthe concept of the velocity response curve); and (3) that therewas a significant difference in the stability (accuracy) ofthe rhythms of the cultivars which is related to their photoperiodicsensitivity. (Received March 30, 1983; Accepted November 15, 1983)     

Scion Control of Genotypic Differences in Mineral Salts Accumulation in Soyabean (Glycine max L. Merr.) Seed

Stem-root grafts of seedling plants were used to ascertain thatgenotypic differences in P, Mg, Mn, and B accumulation in soyabean(Glycine max L. Merr.) seeds are controlled by the scion ofthe plant. The effect of the graft per se on mineral accumulationwas negligible. These results are similar to those reportedfor Sr and Ca accumulation in soyabeans. Mechanisms which couldaccount for these observations are briefly discussed.     

Flower and Pod Development in Water-Deficient Soybeans (Glycine max L. Merr.)

Water deficits during flowering decrease the number of seed-bearingpods in soybean (Glycine max L. Merr.). Failure to set podsmay indicate an inherent sensitivity to low tissue water potential(     

Synchronization of protoplasts from Glycine max (L.) Merr. and Brassica napus (L.)

Cells of Glycine max originating in a suspension culture and cells of Brassica napus prepared from hypocotyls were synchronized. Synchronization was achieved by preparing protoplasts in the usual way and subsequently letting the protoplasts regenerate into cells by removing the cell-wall-digesting enzymes. More than 70% of the cells had divided synchronously at the end of the first cycle as determined by the mitotic index. The high frequency of mitosis critically depended on the osmolality of the medium. The duration of the S-phase was estimated by measuring the activity of thymidylate kinase as well as incorporation of [³H]deoxythymidine into acid-insoluble material. The data indicate that synchronization is induced by resetting the cell cycle.Abbreviations dTMP deoxythymidine 5-monophosphate - TCA trichloroacetic acid     

Growth and Development of Soybean (Glycine max [L.] Merr.) Pods: CO(2) Exchange and Enzyme Studies

The rates of CO₂ exchange and ¹⁴CO₂ incorporation in the light and dark and the activities of several photosynthetic, photorespiratory, and respiratory enzymes of soybean (Glycine max [L.] Merr. cv. Wye) reproductive structures were determined at weekly intervals from anthesis to pod maturity. At all stages of pod development soybean reproductive structures were found to be incapable of net photosynthesis under the experimental conditions employed, but capable of gross photosynthesis and light-induced ¹⁴CO₂ uptake. Consistent with the lack of net photosynthesis throughout the development of the reproductive structure, the maximum in vitro activity of ribulose 1,5-bisphosphate carboxylase (EC 4.1.1.39) in pod tissue was only 3% of that in leaf extracts when expressed on a fresh weight basis. We concluded that the major role of the reproductive structure of the soybean with respect to photosynthetic carbon metabolism is the reassimilation of its respiratory CO₂.     

Characterization of Solute Efflux from Dehydration Injured Soybean (Glycine max L. Merr) Seeds

Soybean (Glycine max L. Merr) seeds lose their tolerance of dehydration between 6 and 36 hours of imbibition. Soybean axes and cotyledons were excised 6 hours (tolerant of dehydration) and 36 hours (susceptible) after commencing imbibition and subsequently dehydrated to 10% moisture. Kinetics of the efflux of potassium, phosphate, amino acid, sugar, protein, and total electrolytes were compared in the four treatments during rehydration. Only slight differences were observed in the kinetics of solute efflux between the two cotyledon treatments dehydrated at 6 and 36 hours suggesting that the cotyledons may retain their tolerance of dehydration at this stage of germination. Several symptoms of injury were observed in the axes dehydrated at 36 hours. An increase in the initial leakage of solutes during rehydration, as quantified by the y-intercept of the linear regression line for solute efflux between 2 and 8 hours suggests an increased incidence of cell rupture. An increase in the rate of solute efflux (slope of regression line between 2 and 8 hours) from fully rehydrated axes was observed in comparison to axes dehydrated at 6 hours. The Arrhenius activation energy for potassium, phosphate, and amino acid efflux decreased and for protein remained unchanged. Both observations indicate an increase in membrane permeability in dehydration-injured tissue. Increasing the H⁺ concentration of the external solution increased K⁺ efflux from both control and dehydrated/rehydrated samples, increased sugar efflux from axes at 6 hours imbibition but decreased sugar efflux from axes at 36 hours imbibition, indicating changes in membrane properties during germination. The dehydration treatment did not alter the pattern of the pH response of axes dehydrated at 6 or 36 hours but did increase the quantity of potassium and sugar efflux from dehydration injured axes. These results are interpreted as indicating that dehydration of soybean axes at 36 hours of imbibition increased both the incidence of cell rupture during rehydration and altered membrane permeability of the rehydrated tissue.     

栽培和野生大豆及其杂交后代幼苗的耐氯性与多胺积累的关系

对经逐代耐盐性筛选的栽培和野生大豆杂交组合(‘Jackson’בBB52’)F4代‘JB185’株系及其亲本幼苗以不同浓度NaCl和等渗(-0.53 MPa)PEG-6000、NaCl、钠盐(无Cl-)和氯盐(无Na )溶液处理6d。结果表明:(1)随NaCl浓度的提高,3种遗传材料幼苗叶片相对电解质渗漏率和MDA含量均呈上升趋势,叶绿素含量除‘BB52’和‘JB185’在NaCl 50mmol/L处理时显著上升外,其余处理呈下降趋势,‘JB185’变化介于两亲本之间。(2)不同离子胁迫下,它们叶片相对电解质渗漏率和MDA含量较对照多表现增加趋势,其中‘BB52’和‘JB185’在钠盐(无Cl-)处理下的变化明显大于氯盐(无Na )处理。叶片中游离态和束缚态Put、Spd和Spm含量都较对照明显提高,但‘BB52’和‘JB185’在钠盐(无Cl-)处理下游离态(Spd Spm)/Put比值和束缚态多胺总量为3种盐处理中最低。表明‘JB185’与野生大豆‘BB52’种群一样对Na 敏感而对Cl-表现较强的耐性。     

Effects of Bradyrhizobium japonicum and Soybean (Glycine max (L.) Merr.) Phosphorus Nutrition on Nodulation and Dinitrogen Fixation

Cells of Bradyrhizobium japonicum were grown in media containing either 1.0 mM or 0.5 μM phosphorus. In growth pouch experiments, infection of the primary root of soybean (Glycine max (L.) Merr.) by B. japonicum USDA 31, 110, and 142 was significantly delayed when P-limited cells were applied to the root. In a greenhouse experiment, B. japonicum USDA 31, 110, 122, and 142 grown with sufficient and limiting P were used to inoculate soybeans which were grown with either 5 μM or 1 mM P nutrient solution. P-limited cells of USDA 31 and 110 formed significantly fewer nodules than did P-sufficient cells, but P-limited cells of USDA 122 and 142 formed more nodules than P-sufficient cells. The increase in nodule number by P-limited cells of USDA 142 resulted in significant increases in both nodule mass and shoot total N. In plants grown with 1 mM P, inoculation with P-limited cells of USDA 110 resulted in lower total and specific nitrogenase activities than did inoculation with P-sufficient cells. Nodule numbers, shoot dry weights, and total N and P were all higher in plants grown with 1 mM P, and plants inoculated with USDA 31 grew poorly relative to plants receiving strains USDA 110, 122, and 142. Although the effects of soybean P nutrition were more obvious than those of B. japonicum P nutrition, we feel that it is important to develop an awareness of the behavior of the bacterial symbiont under conditions of nutrient limitation similar to those found in many soils.     

大豆磷、硫转运蛋白研究进展

磷、硫转运蛋白是大豆（Glycine max（L.）Merr.）体内磷、硫转运的重要载体,参与调节磷和硫酸盐的吸收与转运,对提高大豆的磷、硫利用效率至关重要。大豆磷转运蛋白可划分为Pht1、Pht2、Pht3、Pho1和Pho2 5大家族,目前对Pht1的研究最为深入。大豆14个Pht1家族可分为3个亚家族,他们对磷吸收和转运具有重要作用。大豆硫转运蛋白基因GmSULTR1;2b可在大豆根中特异性表达并被低硫胁迫诱导。本文基于大豆磷、硫的营养吸收、转运与利用过程中的相关性,对Pht1家族以及GmSULTR1;2b基因在大豆中的研究进展进行了综述,并对近年来大豆磷、硫转运蛋白的研究进展及未来的研究方向进行了展望。