• Mycobiology
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• v.33 no.1
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• pp.51-60
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• 2005

Pot greenhouse experiments were carried out to attempt to increase the salinity tolerance of one of the most popular legume of the world; cowpea; by using dual inoculation of an Am fungus Glomus clarum and a nitrogen-fixer Azospirillum brasilense. The effect of these beneficial microbes, as single- or dual inoculation-treatments, was assessed in sterilized loamy sand soil at five NaCl levels ($0.0{\sim}7.\;2ds/m$) in irrigating water. The results of this study revealed that percentage of mycorrhizal infection, plant height, dry weight, nodule number, protein content, nitrogenase and phosphatase activities, as well as nutrient elements N, P, K, Ca, Mg were significantly decreased by increasing salinity level in non-mycorrhized plants in absence of NFB. Plants inoculated with NFB showed higher nodule numbers, protein content, nitrogen concentration and nitrogenase activities than those of non-inoculated at all salinity levels. Mycorrhized plants exhibited better improvement in all measurements than that of non-mycorrhized ones at all salinity levels, especially, in the presence of NFB. The concentration of $Na^+$ was significantly accumulated in cowpea plants by rising salinity except in shoots of mycorrhizal plants which had $K^+/Na^+$ ratios higher than other treatments. This study indicated that dual inoculation with Am fungi and N-fixer Azospirillum can support both needs for N and P, excess of NaCl and will be useful in terms of soil recovery in saline area.

• Korean Journal of Microbiology
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• v.32 no.2
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• pp.97-101
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• 1994

Fifteen strains of the nitrogen fixer Axospirillum were isolated from the rhizosphere of rice collected from Kyonggi-do and Chungcheongnam in Korea. They had strong acetylene-reducing activity of 400 of 900 nmol $C_2H_4$ per hour vial had a similar morphology in succinate-malate medium: vibrioid cells having a diameter of 1.0 ${\mu}m$ and a monopolar single flagellum in liquid media. According to their physiological and morphological characteristics, they were divided into two distinct groups, group I and group II. Group I strain were, unlike group II, distinguished by their ability to use glucose as a sole carbon source in nitrogen-free medium, requirement for biotin, and formation of wider, longer, and S-shaped cells in semisolid nitrogen-free malate medium. On the basis of their characteristics, strains belonging to group I were identified as Azospirillum lipoferum, while those belonging to group II were identified as Azospirillum brasilense.

• Journal of Plant Biology
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• v.32 no.3
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• pp.137-144
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• 1989

In order to clarify the effect of the associative nitrogen fixing bacteria on the growth of corn(Zea mays L. Suwon19), we inoculated corn seedlings with Azospirillum amazonense Y1, a micro-aerobic and acid-tolerant nitrogen fixer, and analyzed the growth and dry matter production and changes of nitrogen and phosphorus quantity of the plant during the growing period. The inoculation of associative N-fixing bacgteria increased the growth of height, leaf area, dry weight and total nitrogen quantity of the plant by 15.4%, 65.4%, 33.7% and 38.0%, respectively, on the day of 56th after sowing. Especially the inoculation of A. amazonense Y1 showed two to three-fold accumulation of phosphorus in each organ of plant. The associative bacteria accelerated the growth of the under ground parts more than those of the upper ground parts of corn plant, and caused decrease in T/R ratios.

• The Korean Journal of Food And Nutrition
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• v.5 no.2
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• pp.137-143
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• 1992

As a research for developing biofertilizers, Klebsiella oxytoca, an associative nitrogen fixer in the rhizosphere of rice plant in the soil of paddy field, was subjected to molecular breeding. The results obtained were as followings. 1). By transforming pbIC71A, Nif A overproducing plasmid, into Klebsiella oxytoca NGl3, Klebsiell6f oxytoca SH3l, and Klebsiella oxytoca SH161, nitrogenase activities in the absence of nitrogen source in the medium were increased 6.4, 17.2, and 13.5 times, respectively, in comparison with the parent strains. 2). Nitrogenase activity of Klebgiella oxytoca NGl3, Klebsiella oxytoca SH3l, and Klebsiella oxytoca SH161 was completely repressed In the presence of 15mM NH4+. But, nitrogenase activities of Klebsiella oxytoca NGl3/PMC71A, Klebsiella oxytoca SH3l /PMC71A, and Klebsiella oxytoca SH 161/pMC714 harboring PMC71A, were 13.7%, 7.7%, and 6.2% of the nitrogenase activities in the absence of nitrogen source in the medium, respectively.

• Korean Journal of Soil Science and Fertilizer
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• v.38 no.1
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• pp.8-14
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• 2005

Mass culturing of two beneficial organisms used as biofertilizers for crops would reduce the risks in production and minimize the capital involved and this demands appropriate media that supports both organism and also selection of organisms that are not antagonistic to each other. A study was initiated to culture a nitrogen fixer (Rhizobium) and phosphate solubilizer (Bacillus megaterium) in a single medium and to study their growth patterns and shelf life in carrier. The growth of Rhizobium and Bacillus megaterium was assessed in different media and a slight modification in the traditional yeast extract mannitol media promoted the growth of both the organisms. The growth of the individual organisms in the modified medium was assessed by estimating the population at regular intervals and compared to their original medium. Maximum population of Rhizobium and phosphobacteria was at 60 hr when the phosphiobacteria inoculation of later was after 48 hr of Rhizobium inoculation. The shelf life of the individual inoculants in the inoculant containing both the organism in a sterile carrier base revealed no significant differences compared to individual organisms inoculated in a sterilized carrier. The population of both organisms in carrier based mixed inoculant remained at $10^8$ cells till 90 days.

• Korean Journal of Microbiology
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• v.24 no.3
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• pp.211-220
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• 1986

Nitrogen fixation (nif) genes of Rhizobium leguminosarum were transferred to nif Klebsiella pneumoniae and E. coli by conjugation after partial heat induction of $RP_4$ :: Mu cts in Rhizobium $R^+$ transconjugant, and the hybrid plasmids in the transconjugant strains were isolated and characterized. In order to transfer the nif genes from Rhizobium, the hybrid plasmid $RP_4$ :: Mu cts was transferred by conjugation from E. coil to the symbiotic nitrogen fixer, R. leguminosarum. After stabillity test, the $RP_4$ :: Mu cts in Rhixobium $R^+$ transconjugant was subjected to partial heat induction by culturing it statically at $38^{\circ}C$ for 16 hours, and then conjugated with the nif defective mutant strains of K. pneumoniae or nif mutant strains of E. coli having whole nif gene plasmid. Recombinant strains of K. pneumoniae, which could grow in a N-free medium and exhibit the nitrogenase activity were selected. However, in the case of E. coli, they could grow well in a NA medium containing antibiotices, but hardly frow in a N-free medium. The hybrid plasmids in these transconjugal strains were isolated by gel electrophoresis and compared their molecular sizes.

• Applied Biological Chemistry
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• v.48 no.3
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• pp.189-200
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• 2005

Biological nitrogen fixation is an important process for academic and industrial aspects. This review will briefly compare industrial and biological nitrogen fixation and cover the characteristics of biological nitrogen fixation studied in Azotobacter vinelandii. Various organisms can carry out biological nitrogen fixation and recently the researches on the reaction mechanism were concentrated on the free-living microorganism, A. vinelandii. Nitrogen fixation, which transforms atmospheric $N_2$ into ammonia, is chemically a reduction reaction requiring electron donation. Nitrogenase, the biological nitrgen fixer, accepts electrons from biological electron donors, and transfers them to the active site, FeMo-cofactor, through $Fe_4S_4$ cluster in Fe protein and P-cluster in MoFe protein. The electron transport and the proton transport are very important processes in the nitrogenase catalysis to understand its reaction mechanism, and the interactions between FeMo-cofactor and nitrogen molecule are at the center of biological nitrogen fixation mechanism. Spectroscopic studies including protein X-ray crystallography, EPR and $M{\ddot{o}}ssbauer$, biochemical approaches including substrate and inhibitor interactions as well as site-directed mutation study, and chemical approach to synthesize the FeMo-cofactor model compounds were used for biological nitrogen fixation study. Recent research results from these area were presented, and finally, a new nitrogenase reaction mechanism will be proposed based on the various research results.

• Korean Journal of Microbiology
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• v.23 no.2
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• pp.107-114
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• 1985

In order to use for recipient strains of RP4:Mu cts, 5 strainsof Rhizobium were selected among 32 strains, which were isolated and identified in this study. Hybrid plasmin RP4::Mu cts, which, is temperature sensitive and confers resistance to ampicillin, kanamycin and tetracycline was transfered by conjugation from E. coli to other atrains of C. coli and the symbiotic nitrogen fixer, Rhizobium leguminosarum. Transfer frequencies of RP4::Mu cts plasmid from E. coli to Rhizobium were about $10^{-8}-10^{-7}$ in LB agar and YMA media. The transconjugants were confirmed by demonstrating that the drug-resistant and temperature-sensitive clones isolated were drug-resistant and temperature-sensitive clones isolated were capable of releasing phage and forming plaques. The plaque-forming units of transconjugants were about $10^2\;to\;10^3$. Stability test of RP4::Mucts in Rhizobium represented that most of the transconjugants had drug resistance and produce phage Mu cts.

• Korean Journal of Soil Science and Fertilizer
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• v.10 no.1
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• pp.23-28
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• 1977

Acetylene-ethylene($C_2H_2-C_2H_4$) assay was carried out to find the heterotrophic nitrogen-fixing activities and distribution of nitrogen fixers in eleven different soil series which are located in Kyeonggi province. Following are the summaries of this study. 1. Acetylene-reducing activities were higher in Gwanghwal, Mangyeong and Buyong series which are derived from fluviomarine deposite than in Yeongsan, Hamchang and Pyeongtack series which are observed from continental alluvial plain. The lowest activities are observed in Yecheon, Hoegog, and Jisan series which are situated in local valley region. 2. Estimated amouts of $N_2$ per annum fixed by $N_2$-fixing heterotrophs were about 3.2kg in fuluvio marine soils, 2.6kg in continental alluvial plain and 2.0kg/10a in local valley region, respectively. 3. Azotobacter and Beijerinckia were not detected in any of the ten different soil series except in Pyeongtaeg series. However, Clostridia, anaerobic nitrogen fixer, was detected in order of ${\times}10^2$. It is assumed that these population are not enough to contribute to the nitrogen supply by the biological fixation in paddy soil. 4. For the assesment of heterotrophic nitrogen fixation in paddy soil, it must be presumed that aerobes, anaerobes and phototrophs which can grow on nitrogen free media may greatly contribute for the asymbiotic netrogen fixation.