• 한국생물공학회:학술대회논문집
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• 2003.04a
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• pp.245-248
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• 2003

As a result of broth substitutions when each culture-mediums were difference, whole culture-medium was found to be best feeding solution for production of PS-7 by B. indica. Maximal production of PS-7 was 1$10.0\;g/{\ell}$ and its conversion rate from 2% (w/v) glucose to PS-7 was 50%. After 48 hr, 50%(v/v) medium of working volume began to substitute in 7L jar fermenter. Production of PS-7 increased after 48hr, recovered productivity of PS-7. Following this preliminary culture, the resultant culture was subjected to continuous flow conditions controlled that the dilution rate were $0.01\;{\sim}\;0.04\;h^{-1}$. Production of PS-7 increased at dilution rate $0.0100\;h^{-1}$ whereas productivity of PS-7 decreased gradually in dilution rate $0.0200\;{\sim}\;0.0400\;h^{-1}$. Maximal production of PS-7 was $10.0\;g/{\ell}$ in continuous culture.

• Journal of Microbiology and Biotechnology
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• v.13 no.3
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• pp.451-456
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• 2003

An expression vector system was developed for the secretory production of recombinant Bacillus stearothermophilus L1 lipase in Saccharomyces cerevisiae. The mature L1 lipase gene was fused to ${\alpha}-amylase$ signal sequence from Aspergillus oryzae for the effective secretion into the culture broth and the expression was controlled under GAL10 (the gene coding UDP-galactose epimerase of S. cerevisiae) promoter. S. cerevisiae harboring the resulting plasmid successfully secreted L1 lipase into the culture broth. To examine an optimum condition for L1 lipase expression in the fed-batch culture, L1 lipase expression was induced at three different growth phases (early, mid, and late-exponential growth phases). Maximum product on of L1 lipase (1,254,000 U/l, corresponding to 0.65/1) was found when the culture was induced at an early growth phase. Secreted recombinant L1 lipase was purified only through CM-Sepharose chromatography, and the purified enzyme showed 1,963 U/mg of specific activity and thermoalkalophilic properties similar to those reported for the enzyme expressed in Escherichia coli.

• KSBB Journal
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• v.15 no.2
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• pp.134-138
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• 2000

Addition of carbon and nitrogen source to an alcohol distillery wastewater was tried to increase the cell concentration of a b baker's yeast cultivated in that wastewater. Carbon was found to be primary limiting nutrient and nitrogen secondary limiting o one. Glucose addition increased the cell concentration 1.3 times higher than no addition, and both glucose and $(NH_4)_2S0_4$ a addition did 5.8 times. A fed-batch cultivation by the automatic addition of glucose and ammonium was executed. Added g glu$\infty$se was automatically controlled to low concentration by a method using DO as control parameter. Ammonium was a automatically added as NH40H used as pH $\infty$ntrol agent after initiating glucose addition. By this simple cultivation method t the cell concentration $\infty$내d be efficiently increased from 2.6g/L to 12.0g/L, and maximum specific growth rate and biomass y yield to glu$\infty$se were $0.18hr^{-1}$ and about 0.54g/g respectively. By increasing cell concentration, COD of the wastewater m media could be additionally reduced by about 22%.

• 한국생물공학회:학술대회논문집
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• 2002.04a
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• pp.257-260
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• 2002

Dissolved oxygen level of cell culture media has a critical effect on cellular metabolism, which governs specific productivity of recombinant proteins and mammalian cell growth However, in the cores of cell aggregates or cell-immobilized beads, oxygen level frequently goes below a critical level. Mammalian cells have a number of genes induced in the lower level of oxygen, and the genes contain a common cis-acting element (-RCGTG-), hypoxia response element (HRE). By binding of hypoxia inducible factor-l (HIF-I) to the HRE, promoters of hypoxia inducible genes are activated, which is a survival mechanism. In this work, to develop a CHO cell capable of producing recombinant proteins in immobilization and high density cell culture efficiently, mammalian expression vectors containing human tissue-type plasminogen activator (t-PA) gene controlled by HRE were constructed and stably transfected into the CHO cells. In $Ba^{2+}$ -alginate immobilization culture, CHO/pCl/dhfr/2HRE-t-PA cells produced 2 folds higher recombinant t-PA activity than CHO/pCl/dhfrlt-PA cells without $CoCl_2$ treatment. Furthermore, in repeated fed batch culture, productivity of t-PA in immobilized CHO/pCI/dhfr/2HRE-t-PA cells was 121 ng/ml/day, total production of 0.968 mg/day at 11 days culture while CHO/pCIIdhfrlt-PA cells was 22.8 ng/ml/day. All these results indicate that HRE is very useful for the enhancement of protein productivity in mammalian cell cultures.

• Journal of Microbiology and Biotechnology
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• v.20 no.11
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• pp.1529-1533
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• 2010

The inulinase gene (INU1) from Kluyveromyces marxianus NCYC2887 was overexpressed by using the GAL10 promotor in a ${\Delta}ga180$ strain of Saccharomyces cerevisiae. The inulinase gene lacking the original signal sequence was fused in-frame to a mating factor ${\alpha}$ signal sequence for secretory expression. Use of the ${\Delta}ga180$ strain allowed for the galactose-free induction of inulinase expression using a glucose-only medium. Shake-flask cultivation in YPD medium produced 34.6 U/ml of the recombinant inulinase, which was approximately 13-fold higher than that produced by K. marxianus NCYC2887. It was found that the use of the ${\Delta}ga180$ strain improved the expression of inulinase in the recombinant S. cerevisiae in both aerobic and anaerobic conditions by about 2.9- and 1.7-fold, respectively. A 5-l fed-batch fermentation using YPD medium was performed under aerobic condition with glucose feeding, which resulted in the inulinase production of 31.7 U/ml at the $OD_{600}$ of 67. Ethanol fermentation of dried powder of Jerusalem artichoke, an inulin-rich biomass, was also performed using the recombinant S. cerevisiae expressing INU1 and K. marxianus NCYC2887. Fermentation in a 5-l scale fermentor was carried out at an aeration rate of 0.2 vvm, an agitation rate of 300 rpm, and with the pH controlled at 5.0. The temperature was maintained at $30^{\circ}C$ and $37^{\circ}C$, respectively, for the recombinant S. cerevisiae and K. marxianus. The maximum productivities of ethanol were 59.0 and 53.5 g/l, respectively.

• Applied Chemistry for Engineering
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• v.5 no.2
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• pp.305-312
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• 1994

The fed-batch process which combinded high selectivity of affinity chromatography and membrane process was developed. The mixture of trypsin and chymotrypsin, having almost the same molecular weight and the chemical structure, were used as model enzymes. The water soluble polymer having more affinity for trypsin and celluose acetate membrane gelated in 50vol.% ethanol for removing free enzymes and retentating trypsin-affinity polymer complex simutaneously were used in this system. The membrane pore size was controlled by ethanol concentration in the gellation bath, and the affinity polymer was prepared by polymerization of acrylamide with N-acryloyl-m-aminobenzamidine at $4^{\circ}C$. The trypsin could be effectively concentrated by utilizing an affinity polymer and a prepared UF-50 ultrafiltration membrane. As a result, 86% purity trypsin was recovered by the current purification process.

• Journal of Korean Society of Water and Wastewater
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• v.27 no.6
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• pp.689-701
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• 2013

Bacterial growth and corresponding consumption of carbon and phosphorus were examined in which tap water samples containing a very low concentration of free chlorine were supplemented with organic carbon and/or phosphorus. The experiments were performed in a fed-batch mode under a controlled temperature of $20^{\circ}C$. In the phosphorus alone-added water, there was no significant increase in bacterial numbers measured as heterotrophic plate count (HPC) in the bulk water. However, bacterial growth was stimulated by the addition of carbon (e.g., bulk HPC levels increased to $10^3CFU/mL$) and further stimulated by the combined addition of carbon and phosphorus (e.g., bulk HPC to $10^5CFU/mL$). The same effects were observed in biofilm HPC and biomass formed on polyethylene (PE) slide surfaces. In the water where organic carbon and phosphorus were added together, the highest biofilm HPC and biomass (measured as extracellular polymeric substance components) densities were observed which were $7.6{\times}10^5CFU/cm^2$ and $5.3{\mu}g/cm^2$, respectively. In addition to the bacterial growth, additions of organic carbon and/or phosphorus resulted in different bacterial carbon-to-phosphorus (C/P) consumption ratios. Compared to a typical bacterial C/P consumption ratio of 100:1, a higher C/P ratio (590:1) occurred in the carbon alone-added water, while a lower ratio (40:1) in phosphorus alone-added water. Comparative value (80:1) of C/P ratio was also observed in the water where organic carbon and phosphorus were added together. At the given experimental conditions, bacterial growth was deemed to be more sensitive to microbially available organic carbon than phosphorus. The effect of phosphorus addition, which resulted in a lower C/P consumption ratio, seemed to be tightly associated with the presence of microbially available organic carbon. These results suggested that the control of extrinsic carbon influx seemed to be more important to minimize bacterial regrowth in drinking water system, since even low content of phosphorus naturally occurring in drinking water was enough to allow a bacterial growth.