• 한국미생물·생명공학회지
• /
• 제29권4호
• /
• pp.206-211
• /
• 2001

Klyyveromyces marxianus exoinulinase를 Saccharomyces cerevisiae에서 구성적으로 과발현 생산하기 위해, 구성적 promoter인 GAPDH, ADH1, PGK 및 ENOI promoters 하류에 exoinulinase 유전자 (INUI)의 ORF를 in frame으로 연결한 각각의 plasmi에 YIGP, pADHI,-INU, pPGK-INU 및 pENO-INU 를 구축하였다. 이들 각 plasmid를함유한 형질전환주 4종을 포도당 농도 5% 배지에서 회분배양한 결과 균체증식은 promoter에 따라 큰 차이를 보이지 않았지만 exoinulinase 발현수준과 plasmid 안정성은 사용한 promoter 에 크게 좌우되었다. 즉 exoinulinase 발현수준은 GAPDH PGK ADH1 ENO1 promoter 각각 1.70, 1.67 1.29, 0.80 unit/ml 였으며 plasmid 안정성은 GAPDH promoter 계의 55%를 제외하고 모두 80%이상으로 높게 나타났다. 이상의 plasmid 안정성과 exoinulinae 발현수준을 고려하여 ADH1 및 PGK 발현계를 선정하여 유가배양하였다 Yeast extract와 포도당을 간헐적으로 공급한 유가배양 결과, 두 발현계에서 약 30 g-DCW/1의 균체농도를 얻었지만, ADHI promoter 계에서는 3.70 unit/ml 의 최대 exoinulinase 활성과 96%의 plasmid 안정성을 보여TRh 반면에 PGK promoter 계는 각각 2.70 unit/ml/와 80%를 나타내었다. 따라서 plasmid 안정성과 긴 배양시간을 고려할 때 비선택적 영양배지를 사용하는 고농도세포 유가배양에서 ADH1 promoter가 exoinulinase 의 구성적 과발현, 생산에 더 적합할 것으로 사료된다.

• 한국물환경학회지
• /
• 제23권1호
• /
• pp.19-26
• /
• 2007

The lab-scale BNR processes fed with Municipal Wastewater Before or After Primary Clarifier (MWBPC or MWAPC) were operated to observe the behavior of particle organic matter in terms of nitrification and denitrification efficiency. As a result of the fractionation of the COD from MWBPC or MWAPC using an aerobic respirometric serum bottle reactor, the total mass of biodegradable organic matter from MWBPC is about 52% greater than the mass from MWAPC. Batch reactors were operated to observe the effect of the Particulate Organic Matter (POM) on substrate utilization for denitrification. Although the consumption of POM for denitrification was observed, the increment of the Specific Denitrification Rate (SDNR) was not great. In terms of the effect of POM on nitrification at different HRTs, activate sludge reactors were operated to determine the optimal HRT when MWBPC and MWAPC were fed relatively. All reactors showed a great organic matter removal efficiency. Reactors fed with MWAPC had obtained the nitrification efficiency above 90% when the HRT of 4 hr, at least, was maintained, while reactors fed with MWBPC had same efficiency when the HRT longer than 5 hr was kept. Three parallel $A^2/O$ systems fed with MWBPC or MWAPC relatively were operated to investigate the effects of POM on BNR processes with varying the HRT of an anoxic reactor. For all systems, the efficiency of organic matter removal and denitrification, respectively, was great and about the same. In case of denitrification efficiency, system with MWAPC had 1.5% lower than system with MWBPC at the same HRT of anoxic reactor of 2 hr, and the increasing the HRT of the anoxic reactor by 1 hr in systems fed with MWBPC resulted in a 3.5% increment. The denitrification rate was similar while the consumption of organic matter in systems fed with MWBPC was higher than system fed with MWBPC. It suggests that POM in MWBPC was not be used significantly as a substrate for denitrification in system with the HRT of 3 hr of an anoxic reactor.

• 대한수학회지
• /
• 제40권1호
• /
• pp.87-107
• /
• 2003

In this paper, we consider a discrete time queueing system fed by a superposition of an ON and OFF source with heavy tail ON periods and geometric OFF periods and a D-BMAP (Discrete Batch Markovian Arrival Process). We study the tail behavior of the queue length distribution and both infinite and finite buffer systems are considered. In the infinite buffer case, we show that the asymptotic tail behavior of the queue length of the system is equivalent to that of the same queueing system with the D-BMAP being replaced by a batch renewal process. In the finite buffer case (of buffer size K), we derive upper and lower bounds of the asymptotic behavior of the loss probability as $K\;\longrightarrow\;\infty$.

• 한국미생물·생명공학회지
• /
• 제17권3호
• /
• pp.263-268
• /
• 1989

유전자의 재조합 균주의 생산성을 향상시키기 위한 발효시스템 개발을 목적으로 regulated promoter 인 SUC2 gene 갖는 유전자 재조합 S. cerebisiae를 모델로하여 유전자 산물인 Invertase 발현에 미치는 글루코오스 농도의 영향, 발효 중 플라스미드의 불안정성, 생육특성 및 continuous fed batch system을 연구하였다. 유전자 재조합 균주는 biphasic growth 현상을 보였으며 글루코오스 농도가 0.9g/$\ell$에서 2.2g/L로 증가함에 따라 비증식 속도는 0.224 h$^{-1}$에서 0.226 h$^{-1}$로 증가했으며 숙주효모보다 낮은 값을 나타내었다. 유전자 재조합 균주의 invertase 생산은 발효조내의 글루코오스 농도에 크게 영향을 받아 글루코오스 농도가 0.25~0.4g/L로 감소될 때 invertase 생산이 시작되었으며 회분발효중 플라스미드의 분리는 글루코오스 자화기간 동안에 빈번히 일어났으나 에탄을 자화기간에는 완만해지는 경향을 보였다. 또한 통기를 해주지 않고 배지의 용존산소만으로 배양시킨 결과 통기를 한 경우에 비하여 invertase의 비활성과 총활성이 각각 1.5 및 1.3 배 증가되었다. 균체의 증식단계와 유전자의 발현단계로 구분하여 발효시키기 위하여 영양분을 함유한 글루코오스 용액을 48m1/h(0.096g 글루코오스/48L의 유량으로 12시간 연속적으로 공급하여 fed batch 배양한 결과 invertase의 비활성과 총활성이 비통기적 회분배양 보다 각각 1.74, 2.74배 증가되었다.

• 한국미생물·생명공학회지
• /
• 제28권6호
• /
• pp.341-348
• /
• 2000

Selection of high producer strain, optimization of production medium and cultivation in bioreactor system were carried out in order to produce an antifungal substance, PAFS in large amounts which sources and 41 kinds of nitrogen sources, a synthetic medium consisting of fructose(70 g/1) and ammonium sulfate (10g/l) and a complex medium including galactose(30g/l), fructose(20g/l) and cottonseed flour(35g/l) were determined as opti-mized media for PAFS production. In bioreactor studies examining physiological characteristics of the pro- ducer microorganism with the complex medium, typical pattern of diauxic growth was observed as demonstrated by the result that fructose was not used before almost exhaustion on readily utilizable carbon source, galactose. When galactose was supplemented additionally during the fermentation period. PAFS pro-ductivity did no increases any more, indicating that large portion of the added galactose was used for cell growth instead of biosynthesis of the secondary metabolite. It was deduced that PAFS production could be enhananced by employing fed-batch operation in order to overcome the apparent phenomenon of catabolite repression and /or inhibition.

• Journal of Microbiology and Biotechnology
• /
• 제12권5호
• /
• pp.753-759
• /
• 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.

• 한국환경과학회지
• /
• 제7권6호
• /
• pp.867-873
• /
• 1998

Laboratory study was conducted to provide basic data for operating anaerobic sequencing batch reactor(ASBR) process for treatment of nightsoil. The experiments were concerned with digestion characteristics, settleabiltity and dewaterability of digested sludge in ASBR system. Completely-mixed dally-fed control reactor without solid-liquid separation step was also operated to evaluate the baseline performance since the nature of nightsoil was changed with time. In all case, gas production from the ASBR shows 1.3 to 1.44 times higher than that from control, in spite of almost similar trend in organics removal. During thickening period, remarkable decrease in surface settling velocity was observed at the ASBRs compared with the control. In case of the ASBR run, flotation of mixed digested sludge was not occurred. Also, ultimate thickened volume of ASBRs increased 1.2~1.5 times compared with control. Dewaterability of digested sludge without conditioning decreased when the completely-mixed daily-fed reactor for ASBR run was converted to the ASBR. However, improvement of dewaterability of digested sludge from the ASBRs was observed as a result of addition of FeCl$_3$ to digested sludge for conditioning.

• Biotechnology and Bioprocess Engineering:BBE
• /
• 제3권2호
• /
• pp.51-54
• /
• 1998

It has been proved that co-cultivation of human neroblastoma cells and human fibroblast cells can enhance nerve cell growth and the production of BDNF in perfusion cultivation. In batch co-cultivation, maximum cell density was increased up to 1.76${\times}$106 viable cells/mL from 9${\times}$105 viable cells/mL of only neuroblastoma cell culture. The growth of neuroblastoma cells was greatly improved by culturing both nerve and fibroblast cells in a perfusion process, maintaining 1.5${\times}$106 viable cells/mL, which was much higher than that form fed-batch cultivation. The nerve cell growth was greatly enhance in both fed-batch and perfusion cultivations while the growth of fibroblast cells was not. It strongly implies that the factors secreted from human fibrobast cells and/or the environments of co-culture system can enhance both cell growth and BDNF secretion. Specific BDNF production rate was not enhanced in co-cultures; however, the production period was increased as the cell growth was lengthened in the co-culture case. Competitive growth between nerve cells and fibroblast cells was not observed in all cases, showing no changes of fibroblast cell growth and only enhancement of the neuroblastoma cell growth and overall BDNF production. It was also found that the perfusion cultivation was the most appropriate process for cultivating two cell lines simultaneously in a bioreactor.

• 한국생물공학회:학술대회논문집
• /
• 한국생물공학회 2000년도 춘계학술발표대회
• /
• pp.243-246
• /
• 2000

Clostridium.aetobutylicum Bl8에 의한 부탄올 발효에서 농도, extra nutrient, pH대한 부탄올 생성량을 알아보았다. 실험결과 extra nutrient를 주입했을 때 부탄올의 농도가 주입하지 않았던 배양에서보다 약 17%정도 증가하는 결과를 보였고, pH를 control 했을 때에는 glucose 60g/L 회분 실험을 기준으로 부탄올 생산량이 약 41%정도가 증가하는 것으로 나타났다.

• KSBB Journal
• /
• 제13권1호
• /
• pp.38-43
• /
• 1998

Application of pervaporative extraction of ethanol to simultaneous saccharification and fermentation(SSF) of cellulose was investigated. From batch experiments, optimum cellulose substrate and enzyme loadings were found to be 10% and 15 IFPU/g cellulose, respectively. The cellulose conversion was lowered in fed-batch system due to the ethanol accumulation. The activity of the yeast Saccharomyces uvarum used in this study was significantly reduced at ethanol concentrations above around 40 g/L. From pervaporation experiments using PDMS membrane, ethanol was efficiently separated at 38$^\circ C$ and 10 mmHg of a down stream pressure. The pervaporation unit with 240 cm$^2$ of surface area was combined into the SSF reactor. The continuous removal of ethanol by pervaporation during SSF resulted in an improved cellulose conversion. Within the scope of this experiment, ethanol yields in the pervaporative SSF and simple SSF were 68.3% and 56.6%, respectively. The permeate flux for SSF broth pervaporation was about one-half that for the pervaporation of aqueous ethanol solution. Accordingly, the development of a membrane with higher ethanol selectivity and flux will increase the feasibility of this technology.