• 생명과학회지
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• 제17권2호통권82호
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• pp.293-297
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• 2007

형광단백질 (green fluorescent protein, GFP)은 생물공정을 살피데 지표 단백질로 유용하게 사용이 된다. 본 연구에서는 쌀세포에서 외래 단백질의 발현양상을 관찰하기 위해서, 표지 단백질로 GFP를 형질전환 후 이것에서 유도된 현탁세포에서 GFP의 발현 양상을 관찰하였다. 형질전환시 GFP의 발현을 위한 프로모터로 RAmysE를 사용하였으며 이것은 배양액 중에서 당이 고갈되었을 때 강력히 작동된다. 그래서 본 연구에서는 배양액 중에 다양한 슈크로오스 농도로 쌀세포를 배양하여 세포의 성장양태 및 GFP의 발현양에 미치는 영향을 관찰한 결과 세포의 성장은 12%의 당농도에서 7.06g/L로 최적이였으며 GFP는 당을 가장 적게 사용한 3%에서 최적임을 알 수가 있었다. 이것은 세포의 성장과 GFP의 생산에 사용된 당이 반대로 영향을 미침을 알 수가 있었으며 향후 최적의 대량배양을 위해서는 세포의 성장과 산물의 생산시기를 분리한 이단계 배양법이 필요함을 암시한다.

• Biotechnology and Bioprocess Engineering:BBE
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• 제9권6호
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• pp.423-427
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• 2004

A simple purification procedure of bioactive human granulocyte macrophage colony stimulating factor (hGM-CSF) secreted in rice cell suspension culture has previously been described. In this study the protein was purified to apparent homogeneity with an overall yield of $80.1\%$ by ammonium sulfate precipitation and a single chromatographic step involving FPLCanion exchange chromatography. The purified hGM-CSF revealed at least five glycosylated forms ranging from $21.5{\~}29$ kDa, and its biological activity was independent of the glycosylation pattern. This is the first purification report of recombinant hGM-CSF to apparent homogeneity from rice cell suspension cultures.

• Biotechnology and Bioprocess Engineering:BBE
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• 제6권1호
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• pp.51-55
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• 2001

This study was conducted to establish a plant cell culture system for the production of medically important secondary metabolites from Xanthium strumarium. The effects of plant growth regulators including NAA, 2,4-D, kinetin, and ABA were examined in terms of callus induction, maintenance of callus and suspension cultures. It was shown that callus was induced upon treatment with NAA while embryo was induced after treatment with 2,4-D. Callus formation was further improved by treatment with ABA and NAA. The level of callusing increased by 17-29% for the seed case, cotyledon, leaf, and hypocotyl and by 96% in the case of the root. Suspension cell lines were established using calli produced from cotyledon, hypocotyl and root and cultured at 25$\^{C}$ under light conditions. The cells grew up to 15g/L with NAA 2ppm, BA 2ppm, and ABA 1ppm treatment. Supernatants of suspension cultures of cell lines derived from coyledon and hypocotyl produced some distinctive secondary metabolites, one of which was identified as 8-epi-tomentosin, which belongs to the xanthanolides. The amounts of 8-epi-tomentosin produced by the cotyledon- and hypocotylderived cell lines were 13.4mg/L and 11.0mg/L, respectively.

• Journal of Plant Biotechnology
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• 제5권2호
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• pp.131-135
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• 2003

Callus and cell suspension cultures of Eurycoma longifolia Jack were initiated from leaves of different trees. The leaf explant of tree Eu9 produced the most calli and also induced high cell biomass in the cell suspension culture. Optimum production of cell biomass could be initiated in proliferating culture medium with a pH of 5.75 prior to autoclaving. The effects of macronutrient inorganic salts of Murashige and Skoog (MS) liquid medium supplemented with X on production of cell biomass of Eurycoma longifolia were also investigated. The highest cell biomass was produced in MS medium containing macronutrients of $21\;mM\;NH_4NO_3,\;12.25\;mM\;KNO_3,\;3.00\;mM\;CaCl_2.2H_2O,\;0.575\;mM\;MgSO_4.7H_2O$, and $1.83\;mM\;KH_2PO_4$. A new medium labeled as TAM was formulated for the production of Eurycoma longifolia cell biomass in the cell suspension culture.

• 한국산업융합학회 논문집
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• 제2권2호
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• pp.73-81
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• 1999

Shake mixing has been widely used in cell culture. The mixing performance for shake mixing, however, has not been reported quantitatively. The critical circulating frequency and the power consumption for complete suspension of particles, based on the definition of Zwietering, were measured in a shaking vessel containing a solid-liquid system. The critical suspension frequency was correlated by the equation from Baldi's particle suspension model modified with the physical properties of the particles. Critical suspension frequency was correlated as following ; $$N_{JS}={\frac{0.58\;d{_p}^{0.06}(g{\Delta}{\rho}/{\rho}_L)^{0.004}X^{0.03}}{D^{0.35}d^{0.17}{\upsilon}^{0.04}}}$$ The power consumption at the critical suspension condition in the shaking vessel was less than that in an agitated vessel with impeller.

• 식물조직배양학회지
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• 제21권1호
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• pp.47-53
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• 1994

본 실험은 포도와 미국자리공의 세포현탁배양계에서 세포생장과 athocyaninn 및 betacyanin 색소생성에 미치는 광의 영향을 조사하였다. 그 결과, 포도의 세포현탁배양계에서의 세포증식 패턴은 광의 효과가 미약하였으나, 미국자리공의 경우는 암배양에 비하여 광조사구에서 뚜렷한 세포증식 패턴을 나타내었다. 포도세포는 12일간의 배양기간중 한번의 생장 피크를 보여주는데 비하여 미국자리공의 배양세포는 두 번의 증식 피크를 보여 광이 세포의 생장 및 분열 주기를 촉진시킬 뿐만 아니라 색소의 축적도 촉진시키는 것으로 나타났으며, 미국자리공 배양세포는 배양 후 4일째에 액포화에 따른 색소의 축적이 확인되었다. 한편, 색소생성에 미치는 광의 영향은 포도 현탁배양계에서는 배양후 6일째부터 생성되기 시작하여 배양 후 10일 전후에 최고치를 보이는데 비하여, 미국자리공의 현탁배양계에서는 세포증식 패턴에 따라 배양 후 4일째와 8일째에 두 번의 피크를 보였다. 그러나 이들 두 가지 모두 암배양하에서는 거의 색소 생합성이 이루어지지 않고 광상태하에서만 색소의 축적을 볼 수 있었다. 한편, 포도 세포배양계에서는 배양 후 4일째에 처리한 광은 약간의 효과가 있었으나, 배양 후 7일째 12시간 이상의 광에 의하여 색소생성이 크게 촉진되는 것을 알 수 있었다. 미국자리공의 경우에는 배양 후 7일 째보다는 오히려 배양 후 4일째에 광이 더욱 효과적이었다. 이상의 사실은 이들 두 가지 색소의 생합성과정에는 광이 절대적으로 필수적이지만 광을 필요로 하는 시기 및 요구하는 광량 등이 서로 다르다는 것을 시사해 준다.

• Plant Biotechnology Reports
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• 제5권2호
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• pp.127-134
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• 2011

The biotransformation potential of cell suspension cultures generated from Withania somnifera leaf was investigated, using withanolides, i.e. withanolide A, withaferin A, and withanone as precursor substrates. Interestingly, the cell suspension cultures showed inter-conversion of withanolides, as well converted to some unknown compounds, released to the culture media. The bio-catalyzed withanolide was detected and quantified by TLC and HPLC, respectively. There is noticeable conversion of withanolide A to withanone, and vice versa though at a lower level. The type of reaction of this biotransformation appears to be substitution of 20-OH group to 17-OH in withanolide A. In this paper, we present for the first time the possibility of biotransformation by inter-conversion of withanolides of pharmacological importance through cell suspension culture of W. somnifera. The possible role of putative cytochrome $P_{450}$ hydroxylases is implicated in the conversion.

• Journal of Plant Biology
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• 제33권3호
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• pp.189-196
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• 1990

Suspension cell lines have been newly established from the calli derived from the immuature embryo culture of hexapolid (Triticum aestivum var. sicco), tetrapolid (T. durum) and diploid (T. tauchii or Aegilops squarrosa) wheat species. The chromosomal variation in suspension cultured cell lines was examined and old cell line, C82d, established from T. aestivum var. copain was also used. New method using 1-bromonaphthalene for metaphase rapping of suspension cells was developed. Variation in chromosome number was observed among all the suspension lines. Cells with doubled chromosome number and deleted chromosome were also observed. Extensive structural changes in chromosome were found in C82d line. Chromosome aberrations showed loss of chromosome arms and chromosome segment. The mean chromosome number in suspension cells of T. aestivum var. sicco was 40, in C82d line 33, in T. durum 28 and in T. tauchii 14. The stability of chromosome in suspension cells of diploid and tetrapolid wheats was higher than that of hexaploid wheat.

• 생명과학회지
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• 제12권5호
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• pp.577-583
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• 2002

$\beta$-glucuronidase 유전자를 형질전환한 담배에서 여러 조직중에서 가장 activity가 높은 줄기에서 현탁세포를 유도하여 생장 및 발현 양상을 조사하였다. Flask culture와 2.5 L bubble column culture시 세포의 성장에는 큰 차이가 없었지만 $\beta$-glucuronidase의 발현은 bubble column culture시 약 2850 U/mg으로 95배 향상되는 결과를 나타내었다. 하지만 두 경우 모두 $\beta$-glucuronidase의 생산량이 증가와 감소를 반복하는 양상을 나타내었다. 이것은 외래 단백질의 생산과 파괴가 반복적으로 일어나는 것으로 추정이 된다. 또한 bubble column culture의 생산성을 향상시키기 위해 중요한 영양분인 sucrose을 30 g/L로 두 번 첨가한 경우, 다른 영양분의 고갈로 인하여 세포의 최종 생산량은 향상되지가 않았고 $\beta$-glucuronidase는 안정적으로 생산되었지만 최종 생산량은 향상되지가 않았다. 하지만 세포의 크기가 감소하여 배양기 운전이 쉬워지는 장점이 있었고 이것은 세포의 고농도 배양을 통한 재조합 단백질 생산성을 향상시키는 유리하게 작용할 것이다.

• 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.