• Korean Journal of Microbiology
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• v.22 no.4
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• pp.265-269
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• 1984

For the utilization as donor cells of conjugation, DAP-Auxotrophs were isolated from C. coli cells, carrying plasmid $p^{RD1}$ with(a) drug resistance makers from Pseudononas $(Km^r, \;Carb^r, \;Tc^r)$ and (b) the nif-gene group from Klebsiela. E. coli $p^{RD1}$ cells were treated with nitrosoguanidine for the mutagenesis and cephalexin for the isolation of DAP-Auxotrophs. The nature of auxotrophs was verified by suitable biochemical test and checking with 6-cyanopurine as a color indicator for the presence of nif-gene.

• Microbiology and Biotechnology Letters
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• v.14 no.5
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• pp.427-432
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• 1986

The regulation of three ammonia assimilatory enzymes, GDH (glutamate dehydrogenase), GS (glutamine synthetase) and GOGAT (glutamate synthase), has been examined in C. glutamicum. Three kinds of arginine auxotrophs blocked in each step of arginine biosynthetic pathway from glutamate were selected as arg 5, arg 6, arg 8. Histidine and tryptophan auxotrophs were also selected because histidine and tryptophan repressed GS biosynthesis in E. coli. These strains were cultured on the media containing nitrogen-excess and limited conditions, to compare the specific activities of ${\alpha}$-ketoglutarate dehydrogenase(${\alpha}-KGDH$), GDH, GS, GOGAT from the cell-free extracts. These results showed that enzyme levels of ${\alpha}-KGDH$ and GDH from 3 kinds of arginine auxotrophs, histidine and tryptophan auxotrophs in nitrogen-excess condition and those of GS and GOGAT in nitrogen limited condition were increased compared with opposite condition. The tryptophan and histidine auxotrophs showed higher level of glutamate and glutamine than parental strains and other mutants. it is assumed that the higher levels of ${\alpha-KGDH}$ and GDH from mutants in nitrogen-excess condition promoted the accumulation of glutamate and glutamine in fermentation broth. The inhibition of GS activities by ADP suggested that GS is regulated by energy charge in C. glutamicum. The results with histidine, tryptophan, glycine, alanine, serine and GMP implied that a system of feedback inhibition were effective. The GDH, GS and GOGAT biosynthesis in culture broth was markedly repressed by the nature and kinds of available nitrogen sources such as tryptophan, proline, glycine, alanine, serine and tyrosine.

• The Korean Journal of Mycology
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• v.19 no.4
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• pp.276-281
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• 1991

Auxotrophs induced by UV irradiation were Iysine, arginine, methionine, and cysteine requirement. The optimal protoplast-producing conditions of Pachysolen tannophilus were obtained in 1 hr incubation treated zymolyase 0.5 mg/ml at $37-42^{\circ}C$(pH 6.5-8.5 with 0.6 M KCI as osmotic stabilizer. Cell volumn of fusants were counted in about 4 times of parents and their morphological characteristics were similar to parents which were global or egg-shaped, though in fusant, pseudohypha were observed frequently. Ethanol production of fusants were similar to parents, but arginine and methionine auxotrophs were showed increased ethanol production respectively 1.6 times and 1.25 times more than parents.

• The Korean Journal of Mycology
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• v.24 no.2 s.77
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• pp.135-141
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• 1996

Auxotrophs and drug resistant mutants from the mycelia of Lentinus edodes were obtained by UV irradiation at survival rates of $0.024{\sim}2.45%$ and ethidium bromide (EtBr) enrichment after UV irradiation. The mutation rate was 0.40%, and back mutation rate was $4.81{\times}10^{-4}{\sim}8.46{\times}10^{-4}$. Various amino acid-, nucleic acid-, and vitamin-requiring auxotrophs were isolated. The concentrations of several fungicides, antibiotics and amino acid analogues inhibiting the growth of L. edodes were determined. The MIC values for cycloheximide, benomyl, and p-fluorophenylalanine were 2, 2000, and 1000 ug/ml respectively. Five p-fluorophenylalanine-resistant mutants and eight benomyl-resistant mutants were selected by UV irradiation.

• Applied Biological Chemistry
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• v.28 no.1
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• pp.13-18
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• 1985

To investigate the fusion of bacterial protoplasts, two auxotrophic mutants of Bacillus subtilis were isolated after treatment with nitrosoguanidine. Two auxotrophs were uracil-auxotroph and tryptophan-auxotroph. Back mutation frequencies of $ura^-$ and $trp^-$ -auxotrophs were $2.4{\times}10^{-8}$ and less than $1.0{\times}10^{-8}$, respectively. The optimal pH and temperature of protoplast formation with lysozyme were 6.5 and $30^{\circ}C$. The optimal lysozyme concentration was $200{\mu}g/ml$. The regeneration frequency of protoplast was 3.3%.

• The Korean Journal of Mycology
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• v.13 no.3
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• pp.185-189
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• 1985

This experiment was undertaken to investigate considerable economy of labor for selection of auxotrophs from strains of Lentinus edodes. Various types of auxotrophs of this strain were isolated after treatment of basidiospores with ultraviolet light, and the highest proportion of putative mutants was also obtained from isolates irradiated to give 1.30 % survival.

• Korean Journal of Microbiology
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• v.21 no.3
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• pp.156-162
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• 1983

Conditions for effcient formation and regeneration of protoplasts of Micromonospora rosaria and Micromonospora purpurea were investigated. The state of inoculm, culture stage and growth in a medium containing partially growth-inhibiting concentration of glycing have significant effects on portoplasting. A high frequency of regeneration (up to 30%) was accomplished with a hypertonic regeneration agar medium defined by Okanishi for Strptomyces. Using the optimal conditions for protroplasting and regeneration, protoplast fusion of auxotrophic M.rosaria was carried out. Polyethylene glycol 1,000 was chosen for fusogenic agent. When signgle auxotrophs were used, the recombinant frequency of auxortrophic markers varied from 1.3 to 3.2%. Using two double auxotrophs, the recombinant frequencies of 0.7-4.3% were obtained. Much lower frequencies(three or more orders of magnitude) were observed by the conventional matings.

• Applied Biological Chemistry
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• v.28 no.2
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• pp.92-97
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• 1985

UV irradiation method was applied to Agrobacterium tumefaciens A 136 to obtain arginine auxotrophic mutant which is applicable as a host of Ti-plasmid. When the bacterial growth was measured at 600 nm, it showed the exponential phase between 7 and 16 hours after 2% inoculation (v/v) in TY medium and the generation time of 4.8 hours. Survival rate of $1{\sim}0.1%$ was reserved when irradiated at the intensity of $800\;{\mu}w/cm^2$ for $30{\sim}50sec$. Fifteen mutants were selected among 5,000 colonies after UV irradiation. Two of them were identified as arginine auxotrophs, three of them as asparagine auxotrophs, ana the other not as arginine, asparagine, glycine nor cysteine auxotrophs.

• Microbiology and Biotechnology Letters
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• v.16 no.5
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• pp.407-412
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• 1988

In order to increase the tryptophan productivity of E. coli SB1007, a mutant resistant to sulfanilamide was isolated and then a tyrosine auxotroph TY-90 was derived from the sulfanilamide-resistant mutant SA3-39-16. In the test-tube culture a quantitative amount of tryptophan was accumulated in strain TY-90 but in a jar fermentor culture the productivity was lower as compared to the level obtained by the parent strain. From the double auxotrophic mutant SB2756, a revertant resistant to 2, 000$\mu\textrm{g}$/$m\ell$ of $\beta$-thienylalanine, TA 40-10, was selected and then phenylalanine auxotrophs were derived from the revertant strain TA-40-10. One of the phenylalanine auxotrophs, TP-4, accumulated 3.7g/$\ell$ of L-tryptophan after 71-hr cultivation in a jar fermentor experiment.

• Archives of Pharmacal Research
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• v.23 no.1
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• pp.79-86
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• 2000

Heterokaryotic nuclear hybrids overcoming the natural barriers of incompatibility have been studied in basidiomycetes. To produce these nuclear hybrids between incompatible mushrooms, which have several potent pharmacological effects, nuclear transfer was performed between Lentinula edodes and Coriolus versicolor. Nuclei from serine auxotrophs of Lentinula edodes, $LE207(Ser^{-})$ were transferred into the protoplasts of arginine auxotrophs of Coriolus versicolor, $CV17(Ser^{-})$using 30% polyethylene glycol 4000 in 10 mM $Cacl_{2}$-glycine solution (pH 8.0). Nulcear transfer progenies were selected by nutritional complementation on minimal media supplemented with 0.6 M sucrose. The progenies were classified based on colony morphology to L. edodes-like, C, versicolor-like and non-parental type. Most of the progenies grew slower than either parent. The number of nuclei per cell was similar but the DNA content varied between progenies. The isozyme patterns of nuclear hybrids resembled either of the parent porfiles or showed a mixed profile.