Effect of Rhizosphere pO 2 on Nitrogen Fixation by Excised and Intact Nodulated Soybean Roots 1

117

113

ABSTRACIAn open ps exchange system was used to monitor the nonsteady state and steady state changes in nitrogenase activity (H2 evolution in N2:02 and Ar:02) and respiration (CO2 evolution) in attached, excised, and sliced nodules of soybean (Glycine max L. Menf.) exposed to extenal P02 of 5 to 100%. In attached nodules, increases in external P02 in steps of 10 or 20% resulted in sharp declines in the rates of H2 and CO2 evolution. Recovery been proposed (18). Evidence for a distinct barrier to gas diffusion in the nodule cortex has been obtained experimentally by direct measurements of P02 in the outer and central tissues of soybean nodules (2 1).The exact nature of the diffusion barrier and the manner in which it is regulated are unknown, but it has been suggested that the nodules of many symbiotic associations increase their diffusion resistance to O2 entry when they are exposed to an atmosphere containing 10% C2H2, or one in which N2 is replaced by Ar (25). This increase in diffusion resistance is apparent as a decline in nodule respiration rate, with a concomitant decline in C2H2 reduction or H2 production rate, during the first 30 min of exposure to C2H2 or Ar. The presence or absence of an C2H2-or Ar-induced decline has been correlated with the optimum P02 for N2 fixation within a specific legume-Rhizobium association and, by inference, with the speed with which the diffusion barrier is regulated in that association (25). It has also been suggested that the regulation of the diffusion resistance of the nodule requires physical changes in the diffusion barrier (4), and this may limit the speed with which the nodule responds to changes in its gaseous environment.The aims of this study were (a) to determine whether the gas exchange characteristics of soybean nodules following changes in external P02 are consistent with the presence of a variable diffusion barrier in the nodule cortex, and (b) if so, to identify experimental conditions which could be used to vary this diffusion barrier. An open circuit gas exchange system was used to monitor continuously changes in respiration (CO2 evolution) and nitrogenase activity (H2 evolution in N2:02 and Ar:02) as the P02 surrounding the nodules was varied. Nonsteady state measurements of CO2 production and H2 evolution were used to determine the speed with which the nodules adjust to changes in rhizosphere O2 concentration. Steady

Section: Assessment Of Methods In the Study Of 02 Effects On In Vivosupporting

confidence: 61%

Section: Assessment Of Methods In the Study Of 02 Effects On In Vivomentioning

confidence: 99%

Steady and Nonsteady State Gas Exchange Characteristics of Soybean Nodules in Relation to the Oxygen Diffusion Barrier

Hunt

King²,

Canvin³

et al. 1987

117

113

“…Thereafter a decline is observed in the dinitrogen fixation rates (2,12). The trend in nitrogenase activity, established during the reproductive growth period, is likely a result of diminished photosynthate supply to the root and nodule tissues.…”

mentioning

confidence: 99%

Studies on Genetic Male-Sterile Soybeans

Wilson

Burton²,

Buck³

et al. 1978

Soybeans (Glycine max IL.l Menf. cv. NC 69-2774) were used to study the nonstructural carbohydrate and nitrogen content of plant tissues, and nitrogenase activity tbrougbout the development of male-sterile and malefertile plants. Male (10,15,17), and the influence of source-sink manipulation on the carbohydrate content of soybean leaves, stems, pods, and seed is shown in several studies (4,5,13,15,19,20). Only a few reports mention the effect of altered source-sink relations on soybean nitrogenase activity (7,12) Sampling was initiated at 80 DAE and continued at 10-day intervals until 140 DAE. The final sampling (140 DAE) was taken when the fertile siblings were physiologically mature. At each sampling date five sterile and five fertile plants were harvested from one plot in each of the eight replications. The plants were cut off at ground level, and separated into stems plus petioles, trifoliolate leaves, and pods plus seed. The numbers of pods and leaves from each plant were counted and all samples were maintained separately. The samples were dried, weighed, and prepared for Kjeldahl nitrogen analysis.Plants selected for tissue extraction and acetylene reduction assay were taken from one of the harvested plots in two randomly selected replications. In each of the two replications, the excised root systems of the five sterile and five fertile harvested plants were removed from the ground, and plant identity was recorded. Four root samples of each plant type in each replication were used in the acetylene reduction assay. Carbohydrate and lipid were extracted from the entire root of the fifth plant of each type in each replication, and from 10-g fresh wt portions of leaf, stem plus petioles, and pods plus seed from these same plants. All

“…Prolonged exposures of soybean root to 1.0 atm 02 never completely destroyed nitrogenase activity. Criswell et aL (6,7) similarily detected trace nitrogenase activity even after 2 d exposure of roots to 0.89 atm 02.…”

Section: Resultsmentioning

confidence: 82%

Effect of Supra-Ambient Oxygen on Nitrogenase Activity (C₂H₂) and Root Respiration of Soybeans and Isolated Soybean Bacteroids

Patterson

Peterson²,

LaRue³

1983

The legume nodule requires 02 to support the respiration associated with fixation. Because the availability of 02 might be a factor limiting symbiotic fixation in the field, Criswell and coworkers (6, 7) made a detailed study of the effect of PO2 on nitrogenase (C2H2) activity of excised soybean roots and intact soybean plants. Most (15). Bacteroids were isolated from 10 g nodules and assayed for ARA3 as described (15) except that freshly prepared bacteroids were used instead of being stored frozen. The assays were performed in serum-stoppered vials with a gas phase of 0.1 atm C2H2 and low (0.005-0.03 atm) levels of 02. The balance was Ar. The assay vials, containing buffer, were prepared by first changing the gas phase to Ar by alternate evacuations and additions of Ar using the manifold described earlier (14). 02 and acetylene were then added with syringes.In one type of experiment, the 02 concentration was rapidly changed from 0.03 atm to 0.01 atm during the assay. After the initial incubation, one series of vials was attached to the manifold and the atmospheres changed to Ar by alternately evacuating to 0.3 atm and then adding Ar to 1.0 atm. This was repeated eight times. Acetylene and 02 were then added to give 0.1 and 0.01 atm, respectively. The entire procedure took less than 2 min. The incubation and assay was then continued.Bacterial protein was measured by the method of Lowry et al. (12)