• Microbiology and Biotechnology Letters
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• v.14 no.4
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• pp.311-318
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• 1986

Glucoamylase and ethanol productivities of HSDD-170 and HSDM-119 formed by S. cerevisiae and S. diastaticus protoplast fusion were investigated. For the production of the glucoamylase, soluble starch as carbon source, yeast extract and C. S. L as nitrogen source added into the basal medium were favorable. The production of the enzyme reached at maximum after cultivation of the fusant for 4 days at 3$0^{\circ}C$, aerobically. The properties of glucoamylase produced by fusants were very similar to those produced by S. diastaticus as based on optimum temperature, pH stability. In alcohol fermentation from starch, strain HSDD-170 fermented starch faster than either of its parental strains. The maximum of alcohol yield in 15% of liquefied potato starch was 7.5% (v/v).

• Microbiology and Biotechnology Letters
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• v.49 no.1
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• pp.45-56
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• 2021

Improved amylases were developed from protoplast fusants of two amylase-producing Aspergillus species. Twenty regenerated fusants were screened for amylase production using Remazol Brilliant Blue agar. Crude enzyme extracts produced by solid state fermentation of rice bran were assayed for activity. Three variable factors (temperature, pH and enzyme type) were optimized to increase the amylase activity of the parents and selected fusants using rice bran medium and solid state fermentation. Analysis of this optimization was completed using the Central Composite Design (CCD) of the Response Surface Methodology (RSM). Amylase activity assays conducted at room temperature and 80℃ demonstrated that Aspergillus designates, T5 (920.21 U/ml, 966.67 U/ml), T13 (430 U/ml, 1011.11 U/ml) and T14 (500.63 U/ml, 1012.00 U/ml) all exhibited improved function making them the preferred fusants. Amylases produced from these fusants were observed to be active over the entire pH range evaluated in this study. Fusants T5 and T14 demonstrated optimal activity under acidic and alkaline conditions, respectively. Fusants T13 and T14 produced the most amylase at 72 h while parents TA, TC and fusant T5 produced the most amylase after 96 h of incubation. Response surface methodology examinations revealed that the enzyme from fusant T5 was the optimal enzyme demonstrating the highest activity (1055.17 U/ml) at pH 4 and a temperature of 40℃. This enzyme lost activity with further increases in temperature. Starch hydrolysis using fusant T5 gave the highest yield of glucose (1.6158 g/100 ml). The significant activities of the selected fusants at 28 ± 2℃ and 80℃ and the higher sugar yields from cassava starch hydrolysis over their parental strains indicate that it is possible to improve amylase activity using the protoplast fusion technique.

• Microbiology and Biotechnology Letters
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• v.17 no.1
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• pp.14-18
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• 1989

As the final experiment to assess the possibility of industrial application of FSC14-75, ethanol productivity from liquefied sweet potato starch was examined in a pilot scale of 300 liters. FSC14-75 produced 6.6％(v/v) of ethanol from 13.3％ of liquefied sweet potato starch in 8 days, and the residual sugar was 3.15％. The corresponding efficiency was 70％ of the theoretical maximum. Since we could isolate unicellular cell and flocculent cell from the fermentation broth, we designated them FSC14-75(S) and FSC14-75(F), respectively. We investigated ethanol productivity of FSC14-75(F) compared with that of FSC14-75(S) from liquefied potato starch in a mini·tar tormentor scale of 2.5 liters. FSC14-75(F) was found more favorable than the counterpart in terms of ethanol productivity, and produced 8.1％(v/v) of ethanol from 15％ of liquefied potato starch with an efficiency of 75％. In a pilot scale fermentation with 15％ of liquefied sweet potato starch, ethanol productivity of FSC14-75(F) reached maximum level of 7.7％(v/v) after 8 days, and the residual sugar was 1.9％. However, the ethanol productivity was not enhanced by a supplementary addition of Thermamyl to the fermentation broth after sterilization.

• The Korean Journal of Mycology
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• v.20 no.4
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• pp.324-336
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• 1992

On the five interspecific protoplast fusants of Ganoderma lucidum and G. applanatum was the antitumor test performed. The fusant F-2 was selected, to examine the cultured mycelia (protein bound polysaccharide) as antitumor components. When a dose of 20 mg/kg/day of each components purifed from F-2 fusant was, i.p., injected into ICR mice, the inhibition ratio of Fr. II against the solid form of sarcoma 180 increased to 1.5 times as compared with that of their parents. When Fr. II was examined for immunopotentiation activity, it increased the amount of the superoxide anion in activated macrophages to 1.2 times and the count of hemolytic plaque forming cells in the spleen to 4.3 times as compared with that of each control group. Its chemical analysis showed 85.2% polysaccharide which consisted of glucose, galactose, mannose, fucose and xylose, and 0.39% protein of 15 amino acids. The content of hexosamine was 0.39% and the molecular weight of Fr. V was $5.6{\times}10^4$ dalton.

• Biomolecules & Therapeutics
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• v.8 no.2
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• pp.199-205
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• 2000

On the inter-order protoplast fusants of Lentinus edodes and Ganoderma lucidum was the antitumor activity test performed and the fusant P22 was selected. The hot water extract of the cultured mycelia of P22 were purified by DEAE-cellulose chromatographya and the resulting purified fraction was designated as P22A. It was found to be a proteoglycan whose molecular weight was 47 kDa. When examined for immunopotentiation activity, P22A increased the number of colony forming unit in the bone marrow stem cells to 3-folds. It also potentiated the secretion of nitric oxide in activated macrophages to 2-folds. In humoral immune response, it increased the activities of the alkaline phosphatase in differentiated B cells to 1.6-folds and the number of plaque forming cells to 1.8-folds. In cellular immune response, it restored the depressed response of delayed type hypersensitivity in tumor bearing mice to normal level. These results suggest that P22A have potential to restore the decreased immune activity of the tumor bearing mice to normal level.

• Korean Journal of Pharmacognosy
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• v.27 no.3
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• pp.224-230
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• 1996

To find pharmacologically active hybrids among the inter-order protoplast fusants of Lentinula edodes and Ginoderma lucidam the antitumor test was performed and the fusant P22 was selected among them. The hot water and alkaline extracts from the cultured mycelia of P22 were purified and separated into four fractions by DEAE-cellulose anion exchange chromatography. When a dose of 20 mg/kg/day of each fractions was injected into ICR mice by i.p., the tumor inhibition ratio of Fr. IV against solid sarcoma 180 was the higher than any other fraction. Fr. IV was a protein-bound polysaccharide which was composed of 69. 12% polysacchafide and 9.76% protein and the molecular weight of Fr. IV was $6.7{\times}10^4$ dalton.

• KSBB Journal
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• v.25 no.6
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• pp.565-571
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• 2010

To develop thermostable ethanol fermentative yeast strain for lignocellulosic simultaneous saccharification and fermentation, high ethanol producing yeast, Saccharomyces cerevisiae CHY1012 and thermostable yeast, Kluyveromyces marxianus CHY1703 were fused by protoplast fusion. The thermostable fusant, CHY1612 was identified as a Kluyveromyces marxianus by phenotypic and physiological characteristics, as well as molecular analysis based on the D1/D2 domains of the large subunit (26S) rDNA gene and the internally transcribed spacer (ITS) 1 + 2 regions. For lignocellulosic ethanol production, AFEX pretreated barley straw at $150^{\circ}C$ for 90 min was used in a simultaneous saccharification and fermentation (SSF) process using thermotolerant CHY1612. The SSF from 16% pretreated barley straw at $43^{\circ}C$ gave a saccharification ratio of 90.5%, a final ethanol concentration of 38.5 g/L, and a theoretical yield of 91.2%. These results show that K. marxianus CHY1612 has potential for lignocellulosic ethanol production through simultaneous saccharification and fermentation with further development of process.

• The Korean Journal of Mycology
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• v.17 no.4
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• pp.197-201
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• 1989

The fusion between protoplasts of Ganoderma applanatum and oidia of Lyophyllum ulmarium (Hypsizigus marmoreus) was induced with polyethylene glycol and $CaCl_2$. When transferred to Ganoderma complete medium plates, fusants showed mixed morphologies both parents. During three times subcultivation the fusants were changed similar to those of L. ulmarium type. All fusants produced oidia, clamp connections and basidiocarps similar to those of L. ulmarium. Isozyme pattern of esterase of interorder fusants showed both parental and non-parental bands. Each individual fusant did not showed both parental and non-parental bands. Each individual fusant did not show any differences in mycelial growth rate, colony morphology, esterase band pattern and basidiocarp.

• Journal of Microbiology and Biotechnology
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• v.7 no.1
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• pp.25-31
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• 1997

Protoplast fusion between L. casei KCTC 1121 and L. acidophilus 88 was attempted to obtain improved strains. The fusants produced a bacteriocin against indicator strains, making a smaller inhibition zone compared to that of L. acidophilus 88. After culturing for 2 months on selective medium, the selected fusants were still stable without segregation. Fusants showed higher lipase activity compared to those of the two parent strains. Fusant No.4, 11, and 15 exhibited excellent lactic acid productivity. Fusant No.4 and 15 exhibited improved proteolysis ability compared to the two parent strains. Whereas L. casei possessed both ${\beta}-galactosidase$ and $phospho-{\beta}-galactosidase$ activities, and L. acidophilus 88 had only ${\beta}-galactosidase$ activity, the fusants had both the intermediate enzyme activities. Cell size of the fusants was greater than that of the parents.

• Microbiology and Biotechnology Letters
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• v.36 no.2
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• pp.158-164
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• 2008

The fusants which contain starch utilizing ability and alcohol fermenting ability were developed by protoplast fusion of Saccharomyces cerevisiae KOY-1 and Saccharomyces diastaticus KCTC 1804. Sacharomyces cerevisiae KH-12 was obtained by haploid induction from Saccharomyces cerevisiae KOY-1. The auxotropic mutants of yeast were obtained by using an ethylmethane sulfonate (EMS). The frequency of protoplast formation in Saccharomyces cerevisiae KOY-1 $(Met^-)$ and Saccharomyces diastaticus KCTC 1804 $(Trp^-)$ were 90.5% and 97.7%, respectively. The frequency of fusant formation was $1.79{\times}10^{-4 }$ for the regenerated protoplast and the 1,000 fusants were obtained. Fusant FA 776 was selected as a potential yeast which contain an alcohol fermenting ability in the starch medium. The genetic stability was 4.64% for 10 passages of generation. Fusant FA 776 produced 13mg/ml of alcohol in 24% starch medium and showed 1.86-fold higher alcohol fermenting ability than Saccharomyces diastaticus KCTC 1804.