• Journal of Microbiology and Biotechnology
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• v.12 no.5
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• pp.753-759
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• 2002

$\beta$-Cyclodextrin glucanotransferase ($\beta$-CGTase) was overproduced extracellularly using recombinant E. coli by transforming the plasmid pECGT harboring a secretive signal peptide. The $\beta$-CGTase gene of alkalophilic Bacillus firmus var alkalophilus was inserted into the high expression vector pET20b(+) containing a secretive pelB signal peptide, and then transformed into E. coli BL2l(DE3)pLysS. The optimum culture conditions fer the overproduction of $\beta$-CGTase were determined to be TB medium containing 0.5% (w/v) soluble starch at post-induction temperature of $25^{\circ}C$. A significant amount of $\beta$-CGTase, up to 5.83 U/ml, which was nine times higher than that in the parent strain B. firmus var. alkalophilus, was overproduced in the extracellular compartment. A pH-stat fed-batch cultivation of the recombinant E. coli was also performed to achieve the secretive overproduction of $\beta$-CGTase at a high cell density, resulting in production of up to 21.6 U/ml of $\beta$-CGTase.

• Biotechnology and Bioprocess Engineering:BBE
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• v.7 no.3
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• pp.163-170
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• 2002

This report describes the use of a transtubular bioreactor to study the relative effects of diffusion versus perfusion of medium on antibody production by a hybridoma cell line. The study was performed with a high-density cell culture maintained in a serum-free, low-protein medium for 77 days. It was determined that the reactor possessed a macro-mixing pattern residence time distribution similar to a continuous stirred tank reactor (CSTR), However, due to the arrangement of the medium lines in the reactor, the flow patterns for nutrient distribution consist of largely independent medium path lengths ranging from short to long. When operated with cyclic, reversing, transtubular medium flow, some regions of the reactor (with short residence times) are more accessible to medium than others (with long residence times). From this standpoint, the reactor can be divided into three regions: a captive volume, which consists of medium primarily delivered via diffusion; a lapped volume, which provides nutrients through unilateral convection; and a swept volume, which operates through bilateral convection. The relative sizes of these three volumes were modified experimentally by changing the period over which the direction of medium flow was reversed from 15 min (larger captive volume) to 9 h (larger swept volume). The results suggest that antibody concentration increases as the size of the diffusion-limited (captive) volume is increased to a maximum at around 30 min with a sharp decrease thereafter. As reflected by changes in measured consumption of glucose and production of lactate, no significant difference in cellular metabolism occurred as the reactor was moved between these different states. These results indicate that the mode of operation of the transtubular bioreactor may influence antibody productivity under serum-free, low-protein conditions with minimal effects on cellular metabolism.

• KSBB Journal
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• v.11 no.2
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• pp.132-139
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• 1996

The growth of recombinant E. coil and by-product production were investigated. D-cycloserine was added to increase the secretion of ${\alpha}$-amylase from recombinant E.coil. Even though cell growth was increased for optimal d-cycloserine concentration $0.1g/\ell$, extracellular ${\alpha}$-amylase activity remained almost the same as the case without d-cycloserine. It is important to achieve the high cell density for commercial production of methaboliles including ${\alpha}$-amylase. To achieve this goal, culture conditions should be selected carefully and optimized considering cell growth and by-products production. In cultivating retombinant E. coli, lactic acid and acetic acid turned out to be important by-product which affected cell yield and growth rate.

• Development and Reproduction
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• v.16 no.4
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• pp.339-351
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• 2012

Fetal bovine serum (FBS) is the most frequently used serum for the cultivation of mammalian cells. However, since animal-derived materials might not be appropriate due to safety issues, allogeneic human serum (HS) has been used to replace FBS, particularly for the culture of human cells. While there has been a debate about the advantages of HS, its precise effect on human adult stem cells have not been clarified. The present study aimed to investigate the effect of HS on the human eyelid adipose stem cells (HEACs) in vitro. When HEACs were cultivated in a medium containing 10% HS, many cells moved into several spots and aggregated there. The phenomenon was observed as early as 9 days following 10% HS treatment, and 12 days following 5% HS plus 5% FBS treatment. However, the aggregation was never observed when the same cells were cultivated with 10% FBS or bovine serum albumin. To examine whether cell density might affect the aggregation, cells were seeded with different densities on 12-well dish. Until the beginning of aggregation, cells seeded at low densities exhibited the longest culture period of 16 days whereas cells seeded at high densities showed the shortest period of 9 days to form aggregation. The number of cells was $15.1{\pm}0.2{\times}10^4$ as the least for the low density group, and $29.3{\pm}2.8{\times}10^4$ as the greatest for the high density group. When human cord blood serum or normal bovine serum was examined for the same effect on HEACs, interestingly, cord blood serum induced the aggregation of cells whereas bovine serum treatment has never induced. When cells were cultivated with 10% HS for 9 days, they were obtained and analyzed by RT-PCR. Compared to FBS-cultivated HEACs, HS-cultivated HEACs did not express VIM, and less expressed GATA4, PALLD. On the other hand, HS-cultivated HEACs expressed MAP2 more than FBS-cultivated HEACs. In conclusion, human adult stem cells could move and form aggregates by the treatment with human body fluids.

• KSBB Journal
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• v.9 no.2
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• pp.211-223
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• 1994

In fermentations with a 4-liter stirred tank bioreactor, a better than two-fold enhancement of the gas-liquid mass transfer coefficient$(k_La)$ in the celite-immobilized fungal cultures of Tolypocladium in flatum over the parallel conventional free-cell was observed at identical biomass concentrations, despite the higher specific oxygen uptake rate of the immobilized fungi during exponential growth. As a result oxygen sufficient conditions, i. e., dissolve oxygen(D.O.) concentrations exceeding 75% air saturation, could be maintained throughout exponential growth period of the immobilized culture, in contrast to the suspended fungal culture, whose D.O. levels fell below 50% air saturation. A linear monotonic dependence of $k_La$ upon impeller agitaion rate was found for both immobilized and conventional cultivation modes over a range of 250 to 550rpm, the slope being a function of biomass concentration for the free but not for the immobilized cell system In contrasts oxygen transfer rate was a much weaker function of aeration rate up to about 2.5 vvm for both culture configurations. Above this level, aeration rate had no further effect on the mass transfer. In addition, the immobilized cultures sustained good morphological and physiological states, leading to almost two times higher cyclosporln A (CyA) productivity overt the parallel free cell system. These experiments suggest that the celite-immobilized fungal system in a stirred tank reactor has considerable promise for scaling up cyclosporin A production in terms of high-density cultivation.

• Journal of Microbiology and Biotechnology
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• v.4 no.4
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• pp.316-321
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• 1994

We investigated the possibility of industrial application and economit process of high temperature fermentation by thermotolerant alcohol producing yeasts as previously reported. From the 20% glucose media, the RA-74-2 produced 11.8% (v/v) ethanol at $32^{\circ}C$ (0.5% inoculum) and 10.6% (v/v) ethanol at $40^{\circ}C$ (3% inoculum), respectively. Also, 11.3% (v/v) ethanol was produced for 96 hours in the temperature-gradient fermentation. These results suggest that the RA-74-2 could isuccessfully be applied to save the cooling water and energy in industrial scale without re-investment or modification of established fermentation systems. When potato starch was used as the substrate for the RA-74-2, high temperature fermentation above $40^{\circ}C$ was more appropriate for industrial utilization because organic nitrogen was not necessary to economical fermentation. As the naked barley media just prior to industrial inoculation, taken from the Poongkuk alcohol industry Co., were used, 9.6% (v/v) ethanol was produced at $40^{\circ}C$ for 48 hours in jar-fermentor scale (actually, 9.5-9.8% (v/v) ethanol was produced at 30~$32^{\circ}C$ for 100 hours in industrial scale). The ethanol productivity was increased by the high glucoamylase activity as well as the high metabolic ratio at $40^{\circ}C$ Therefore, if the thermotolerant yeast RA-74-2 would be used in industrial scale, we could obtain a high productivity and saving of the cooling water and energy. Meanwhile, the RA-912 produced 6%(v/v) ethanol in 10% glucose media at $45^{\circ}C$ and showed the less ethanol-tolerance compared with industrial strains. As the produced alcohol was recovered by the vacuum evaporator at $45^{\circ}C$ in 15% glucose media, the final fermentation ratio was enhanced (76% of theoretical yields). This suggest that a hyperproductive process could be achieved by a continuous input of the substrate and continuous recovery of the product under vacuum in high cell-density culture.

• The Plant Pathology Journal
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• v.30 no.3
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• pp.288-298
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• 2014

We examined the efficacy of a bacterium for biocontrol of the root-knot nematode (RKN) Meloidogyne hapla in carrot (Daucus carota subsp. sativus) and tomato (Solanum lycopersicum). Among 542 bacterial isolates from various soils and plants, the highest nematode mortality was observed for treatments with isolate C1-7, which was identified as Bacillus cereus based on cultural and morphological characteristics, the Biolog program, and 16S rRNA sequencing analyses. The population density and the nematicidal activity of B. cereus C1-7 remained high until the end of culture in brain heart infusion broth, suggesting that it may have sustainable biocontrol potential. In pot experiments, the biocontrol efficacy of B. cereus C1-7 was high, showing complete inhibition of root gall or egg mass formation by RKN in carrot and tomato plants, and subsequently reducing RKN damage and suppressing nematode population growth, respectively. Light microscopy of RKN-infected carrot root tissues treated with C1-7 showed reduced formation of gall cells and fully developed giant cells, while extensive gall cells and fully mature giant cells with prominent cell wall ingrowths formed in the untreated control plants infected with RKNs. These histopathological characteristics may be the result of residual or systemic biocontrol activity of the bacterium, which may coincide with the biocontrol efficacies of nematodes in pots. These results suggest that B. cereus C1-7 can be used as a biocontrol agent for M. hapla.

• Journal of Microbiology and Biotechnology
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• v.28 no.10
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• pp.1691-1699
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• 2018

A metabolically-engineered Deinococcus radiodurans R1 strain capable of producing phytoene, a colorless $C_{40}$ carotenoid and a promising antioxidant, has been developed. To make this base strain, first, the crtI gene encoding phytoene desaturase was deleted to block the conversion of phytoene to other carotenoids such as lycopene and ${\gamma}$-carotene. This engineered strain produced $0.413{\pm}0.023mg/l$ of phytoene from 10 g/l of fructose. Further enhanced production of phytoene up to $4.46{\pm}0.19mg/l$ was achieved by overexpressing the crtB gene encoding phytoene synthase and the dxs genes encoding 1-deoxy-$\text\tiny{D}$-xylulose-5-phosphate synthase gene, and by deleting the crtD gene. High cell-density culture of our final engineered strain allowed production of $10.3{\pm}0.85mg/l$ of phytoene with the yield and productivity of $1.04{\pm}0.05mg/g$ and $0.143{\pm}0.012mg/l/h$, respectively, from 10 g/l of fructose. Furthermore, the antioxidant potential of phytoene produced by the final engineered strain was confirmed by in vitro DPPH radical-scavenging assay.

• Korean Journal of Ecology and Environment
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• v.41 no.1
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• pp.26-33
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• 2008

To understand the effect of water temperature on growth pattern of Stephanodiscus sp., we weekly or biweekly investigated in the lower part of the Nakdong River from 1994 to 2006 and performed a laboratory experiment. Stephanodiscus was the most dominant species among phytoplankton in winter when low flow persisted and the high abundances of the species were maintained from December to February. Three strains of Stephanodiscus sp. were isolated for the in vitro experiment from the Nakdong River in January 2005. Over the water temperature range of $4^{\circ}C$ to $20^{\circ}C$, the growth patterns of Stephanodiscus sp. were different in the short-term batch culture. The maximum cell density of Stephanodiscus sp. was observed at approximately $5^{\circ}C$ in the river systems, but the optimum water temperature of Stephanodiscus sp. was $10^{\circ}C$ for the growth in the laboratory experiment. However, the proliferation of Stephanodiscus sp. was related to low water temperature in the Nakdong River.

• KSBB Journal
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• v.20 no.6
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• pp.458-463
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• 2005

[ $H_2$ ] from CO and water was continuously produced in a trickle bed reactor(TBR) using Citrobacter amalonaticus Y19. When the strain C. was cultivated in a stirred-tank reactor under a chemoheterotrophic and aerobic condition, the high final cell concentration of 13 g/L was obtained at 10 hr. When the culture was switched to an anaerobic condition with the continuous supply of gaseous CO, CO-dependent hydrogenase was fully induced and its hydrogen production activity approached 16 mmol/g cell/hr in 60 hr. The fully induced C. amalonaticus Y19 cells were circulated through a TBR packed with polyurethane foam, and the TBR was operated for more than 20 days for $H_2$ production. As gas retention time decreased or inlet CO partial pressure increased, $H_2$ production rate increased but the conversion from CO to $H_2$ decreased. The maximum $H_2$ production rate obtained was 16 mmol/L/hr at the gas retention time of 25 min and the CO inlet partial pressure of 0.4 atm. The high $H_2$ production rate was attributed to the high cell density in the liquid phase circulating the TBR as well as the high surface area of polyurethane foam used as packing material of the TBR.