The acetylene reduction assay has been used extensively to detect and measure nitrogenase activity of free‐living microorganisms, excised legume and non‐legume nodules, and root systems, soil cores, and entire nodulated legume plants removed from their soil medium. This investigation was undertaken to determine whether or not the acetylene reduction assay can be used to measure nitrogenase activity of cultures of nodulated soybean (Glycine max.) plants in situ and to compare the nitrogenase activity of these cultures with the nitrogenase activity of detached nodules and excised root systems from comparable cultures. Entire cultures grown in Perlite or in soil were placed in polyethylene containers, covered with Plexiglas lids, exposed to acetylene, and assayed for ethylene production. Time course experiments showed that rates of acetylene reduction of plants growing in Perlite and in soil were linear for at least 90 min following an initial lag period of 15 min. Saturation of nitrogenase in intact cultures was obtained at a pC2H2 of 0.1 atm. The apparent Km for nitrogenase was 0.05 atm. For convenience, assays in the large containers were conducted at a pC2H2 of 0.025 atm. The Michaelis‐Menten equation was used to calculate rates of acetylene reduction at saturating levels of acetylene. The correlation coefficient between rates of acetylene reduction and nodule fresh weight was 0.99 for plants of the same age and 0.79 for plants of different ages. Acetylene reduction rates of either intact nodulated plants in Perlite or nodulated root systems removed from Perlite were significantly greater than acetylene reduction rates of detached nodules from comparable cultures. Relatively little diurnal variation was observed in nitrogenase activity of potted plants grown under controlled environmental conditions. The method is useful for the assessment of nitrogenase activity of legume cultures in a porous medium under standardized conditions, but its application to legumes in soil proved to be complicated by the water content of the soil which influences rates of gas diffusion within the culture medium.
Few studies clearly compare N source effects on herbage yield of alfalfa (Medicago sativa L.) grown with Rhizobium over several harvest cycles. Therefore, the effects of combined N on nodulation, growth, N accumulation, and development of alfalfa were studied over four harvest/regrowth cycles with a genotype cloned from the nondormant cv. ‘Dohfari.’ Ramets inoculated with Rhizobium meliloti Dang. strain 102F28 and irrigated with 0,2,8, or 16 mM N provided as NH4NO3, NO3‐, or NH4+ were grown in vermiculite in a naturally illuminated greenhouse in two replicate experiments from March through June and July through October. All combined N treatments enhanced herbage dry weight and total crude protein above the nil N control during the first growth period. The promotive effect of combined N decreased with each harvest until during the fourth regrowth, plants totally dependent on Rhizobium for reduced N produced herbage and crude protein yields equivalent to plants supplied 8 or 16 mM N as NH4NO3. Plants grown with 2mM N, a more reasonable N concentration for the soil solution, were more productive than nil N plants during the first two growth cycles but had no advantage during the third and fourth cycles regardless of the form of combined N used. Nitrate suppressed nodule development more than either NH4NO3 or NH4+. Greater availability of combined N increased the soluble sugar:starch ratio in the root and crown tissues. No significant differences were observed between the two experiments except that plants grown later in the year (July to October) took longer to flower during each regrowth cycle. The data indicate that the Rhizobium symbiosis cannot produce sufficient reduced N for optimum alfalfa growth during the first few harvest/regrowth cycles. During subsequent regrowth cycles, however, plants totally dependent on Rhizobium for N compounds had overcome the period of N‐limited growth and produced as much herbage and crude protein as plants grown with levels of combined N that might be expected under normal field conditions.
In contrast with studies which have based essential element requirements of algae on nutrient solution concentrations, in this investigation the requirements of each of 6 species of green and blue-green algae for calcium, magnesium, and potossium were quantitatively evaluated in terms of critical cell concentrations of the 3 cations. A critical cell concentration was considered the minimum cell content of an element which permitted maximum or near maximum total growth of an organism. In comparison with the needs of angiosperm crop plants, the requirements of all 6 species for calcium were extremely low (critical cell contents of 0.06% or less, oven-dry basis); requirements for magnesium were equal to or only slightly less than in higher plants (0.15-0.30%) with the exception of Scenedesmus quadricauda (0.05%); and the requirements for potassium varied greatly, from critical levels less than the average values established for higher plants (0.25-0.50%) to values equal to or in excess of higher plant averages (0.80-2.40%). The nutrition of S. quadricauda was of particular interest because of extremely low requirements for all 3 cations. The results provide a more precise and meaningful expression of the essential cation requirements of algae than have previous data. The results also suggest differences in the physiology and functions of cations in the algae studied and in angiosperms which seem worthy of further investigation.
The distribution of genetic potentials for symbiotic N fixation and carbon exchange rate (CER) was examined in ‘Vernal’ alfalfa (Medicago sativa L.). Symbiotic fixation capability was correlated with whole‐plant CER in plants grown without combined N but was not correlated with innate photosynthetic efficiency measured as CER dm−2 leaf area in the presence of combined N. Values for individual leaf CER of 35 seedlings grown on combined N ranged from 24 to 82 mg CO2•dm−2•hour−1 at 300 ppm CO2 with 1,400 μE•m−2•sec ‐1 photosynthetically active radiation. Carboxylation efficiencies based on either internal or external CO2 concentrations, light and dark respiration of leaves, and leaf resistance also varied over a wide range, but there was little variation in the apparent quantum efficiencles and CO2 compensation points. Twenty‐five seedlings representative of the variation of CER observed in this initial screening were cloned, inoculated with Rhizobium meliloti strain 102F28, and grown without combined N. Leaf area, acetylene reduction, dry weight, and total N were measured for plants from each of the 25 clones. Whole‐plant CER was determined for eight selected clones that had shown high, medium, or low individual leaf CER when grown with combined N. Whole‐plant CER was correlated positively with leaf area, acetylene reduction, and total N of the eight selected clones. Leaf area was correlated positively with both acetylene reduction and total N of all 25 clones. Rankings of acetylene reduction, leaf area, total dry weight, or total N content for plants grown on N2, however, were not correlated with individual leaf CER ranking of the same genotypes grown on combined N. Six genotypes representing a wide range of photosynthetic efficiency and N fixation potential showed few corresponding differences in top growth through three harvests in the absence of combined N.
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