• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2005년도 생물공학의 동향(XVII)
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• pp.187-188
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• 2005

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2003년도 생물공학의 동향(XIII)
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• pp.329-333
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• 2003

본 연구에서는 형질전환된 N. tabacum 배양에 있어서 배양 중반 저온으로의 변환이 세포에 미치는 영향과 hGM-CSF의 생산성 변화를 관찰하였다. 배양 중반 저온으로의 변환은 DCW의 증가와 세포크기의 감소를 보였다. Ascorbic acid의 첨가는 배양초기 세포 생존율의 감소를 완화시켰으며, 배양 중반 저온으로의 변환은 약간의 세포생존율 감소를 보였다. 저온으로의 변환, 저온 배양에서의 betaine 첨가, ascorbic acid 첨가 모두 배양 후반 세포 lysis 억제에 효과가 있었다. 배양 중반 저온으로의 변환시 배지내 단백질 분해 효소의 활성을 측정한 결과, 대조구 세포에 비해 낮은 단백질 분해 효소 활성을 나타내었다. 그로인해 배양 중반 이후 단백질 분해 효소에 의한 급격한 hGM-CSF 분해를 감소시킴으로써 상대적으로 대조구 세포에 비해 높은 hGM-CSF 생산성을 유지시켰다. Ascorbic acid를 첨가한 후 배양 도중 betaine(1 mM)을 첨가하여 저온으로 온도를 변환시, hGM-CSF의 생산성 대조구 세포에 비하여 최대 2.1배 까지 높게 유지시켰다.

• 농업과학연구
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• 제49권4호
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• pp.795-807
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• 2022

The development and commercialization of industrial genetically modified (GM) organisms is actively progressing worldwide, highlighting an increased need for improved safety management protocols. We sought to establish an environmental monitoring method, using real-time polymerase chain reaction (PCR) and propidium monoazide (PMA) treatment to develop a quantitative detection protocol for living GM microorganisms. We developed a duplex TaqMan quantitative PCR (qPCR) assay to simultaneously detect the selectable antibiotic gene, ampicillin (AmpR), and the single-copy Escherichia coli taxon-specific gene, D-1-deoxyxylulose 5-phosphate synthase (dxs), using a direct cell suspension culture. We identified viable engineered E. coli cells by performing qPCR on PMA-treated cells. The theoretical cell density (true copy numbers) calculated from mean quantification cycle (Cq) values of PMA-qPCR showed a bias of 7.71% from the colony-forming unit (CFU), which was within ±25% of the acceptance criteria of the European Network of GMO Laboratories (ENGL). PMA-qPCR to detect AmpR and dxs was highly sensitive and was able to detect target genes from a 10,000-fold (10^-4) diluted cell suspension, with a limit of detection at 95% confidence (LOD_95%) of 134 viable E. coli cells. Compared to DNA-based qPCR methods, the cell suspension direct PMA-qPCR analysis provides reliable results and is a quick and accurate method to monitor living GM E. coli cells that can potentially be released into the environment.

• Journal of Plant Biology
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• 제20권2호
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• pp.63-69
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• 1977

Experiments were conducted to investigate the effects of several hormones on the growth of suspension culture of carrot (Daucus carota L.) cells, where changes in pH and the amount of $NH_4-Nin$ the medium were observed with regard to growth. A treatment with 2,4-D at $10^{-5}M$ resulted in a highest rate of growth; the hormone at this concentration caused an increase in dry weight by about 40 to 50% over the control, measured at a stationary phase. It was thus indicated that 2,4-D at $10^{-5}M$ provided the optimal condition for the suspension culture. Changes in pH of the medium were found to be affected by hormonal treatments during the first 2-3 days following the inoculation, after which the pattern of pH changes in hormone enriched media paralleled that of the control. Subsequent changes of $NH_4-N and NO_3-N$ from the medium by the cells, and also by growth of the cells. The uptake of $NH_4-N$ by the cells did not appear to be influenced by hormonal treatments. At a stationary phase, a considerable amout of $NO_3-N$ played a more important role than $NH_4-N$ in the growth of the carrot cell suspension culture.

• KSBB Journal
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• 제14권5호
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• pp.534-538
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• 1999

식물세포배양을 위한 bioreactor의 운전조건 최적화를 위해 Nicotiana tabacum 현탁세포를 model system으로 bioreator의 종류, 교반기의 형태, 그리고 통기량에 따른 세포생장을 관찰하였다. Bioreactor로는 stirred tank bioreactor과 airlift bioreactor를 사용하였으며 두가지 배양기 모두 flask에서의 생장보다 낮은 생장을 보였으며 stirred tank bioreactor보다는 airlift bioreator에서 높은 세포농도를 얻을 수 있었다. 교반기의 종류에 따른 세포의 생장은 큰 차이가 없었으나 hollowed paddle impeller를 사용하였을 경우에는 배양기간 동안 세포의 크기가 작게 유지되었다. 통기량을 0.30 vvm으로 유지하는 경우에 가장 좋은 세포생장을 관찰할 수 있었으며 1.0 vvm이상의 통기량에서는 과도한 foam의 형성과 세포의 갈색화 현상을 보이며 세포의 생장도 저해되었다. 또한 통기량이 증가할수록 세포크기지수가 감소하는 결과를 보였다.

• Journal of Plant Biology
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• 제32권4호
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• pp.227-233
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• 1989

Single cells obtained from suspension culture of mature embryo-derived callus in wheat(Triticum aestivum L. cv Jang Kwang) were cultured to regenrated into the plantlet. Cell clusters and embryogenic calluses were efficiently developed from when the single cells clutured on the MS medium supplemented with 10${\mu}{\textrm}{m}$ 2,4-D. Upon transfer to hormone-free MS medium containing 10 mg/I AgNO3, embryogenic calluses gave rise to shoots, probably through somatic embryogenesis.

• Journal of Microbiology and Biotechnology
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• 제1권1호
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• pp.79-83
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• 1991

The effects of phosphate concentration in the medium, feeding of biosynthetic precursor, and the addition of exogenous berberine on cell growth and berberine production were studied in cell suspension cultures of Thalictrum rugosum. The depletion of phosphate in the medium enhanced the specific productivity up to twofold with significant release of berberine into the medium. Extracellular berberine was 19% of the total in the culture without phosphate while it was 2-5% of total berberine in the culture with even low amounts of phosphate. Precursor feeding was not effective in enhancing alkaloid formation. Initial presence of exogenous berberine did not have much effect on cell growth and alkaloid production. It was found that the cells have the capacity to take up large quantities of berberine. When $500{\;}mg{\cdot}l^{-1}$ of berberine was added exogenously at the beginning, 81% of total berberine was found in the cells.

• KSBB Journal
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• 제6권1호
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• pp.1-7
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• 1991

To produce economically important indole alkaloids by cell cultures, we selected protoplastsderived clones (protoclones) of vinca (Catharanthus roseus) for high yields of catharanthine and ajmalicine. Protoplasts were enzymatically isolated from suspension-cultured cells. The highest plating efficiency (1%) was obtained when protoplasts were plated at a density of 1$\times$105 protoplasts/ml in a culture medium solidified with 0.4% Seaplaque agarose. The growth rates of 40 protoclones subcultured on a solid medium varied over a wide range. Protoclone VPC-6, which had the highest growth rate, was observed to produce relatively high yields of catharanthine and ajmalicine when cultured in a liquid medium. Although the original cell line did not produce catharanthine at a detectable level by HPLC, protoclone VPC-10 produced it at a level of 5.9$\mu\textrm{g}$/g fresh weight of cells for 10 days of culture. Under the same conditions, protoclone VPC-15 produced ajmalicine at a level of 133.6$\mu\textrm{g}$/g, of which productivity was improved about ,3 times than that of the original cell line. The results indicate that differences in the growth rate and indole alkaloid yield among the protoclones reflect the somaclonal variation in suspnsion-cultured cells.

• 식물조직배양학회지
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• 제26권4호
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• pp.289-291
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• 1999

발아 7일된 오이 (Cucumis sativus L.) F$_1$ 5품종과 순계 4품종의 하배축 절편을 1mg/L 2,4-D가 첨가된 MS고체배지에 치상하여 3주간 배양하였다. F$_1$ 1품종과 순계 2품종에서 발달한 캘러스의 8%이하에서 체세포배가 발생하였다. 체세포 배 절편을 동일한 배지에 4주간 배양하여 배발생캘러스를 유도하였다. 배발생캘러스는 동일조성의 MS액체배지에 현탁되어 증식하였다. 액체배지에서 배발생캘러스는 수많은 체세포 배로 발달하였으며, 생장조절제가 첨가되지 않은 MS기본배지에서 95%이상의 체세포배가 식물체로 발아하였다. 재분화된 식물체는 화분에서 생육되어 개화 후 착과하였다.

• Applied Biological Chemistry
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• 제36권5호
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• pp.358-363
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• 1993

박하(Mentha piperita) 세포현탁배양에(-)-menthol 생합성 중간체를 투여하여 배양된 세포의 대사경로를 연구하였다. (-)-Limonene을 투여 하였을 때 이는 다른 대사물로 변환되지 않는 것으로 관찰 되었다. (+)-Pulegone은 (+)-isomenthone 및 (-)-menthone으로 변환되었으며, (-)-menthone은 (-)-menthol로 변환되었다. 이 실험은 현탁배양세포가 대부분의 생합성 활성을 유지하고 있으며 (-)-limonene hydroxylase의 활성이 제한적임을 보여 주었다. (-)-Isopiperitenone을 투여하였을 때는 (+)-pulegone, piperitenone, (-)-7-hydroxyisopiperitenone, (R)- 및 (S)-6-hydroxyisopiperitenone이 생성되었다.