• 한국동물생명공학회지
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• 제39권1호
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• pp.19-30
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• 2024

Background: Although an understanding of the proliferation and differentiation of fish female germline stem cells (GSCs) is very important, an appropriate threedimensional (3D) research model to study them is not well established. As a part of the development of stable 3D culture system for fish female GSCs, we conducted this study to establish a 3D aggregate culture system of ovarian cells in marine medaka, Oryzias dancena. Methods: Ovarian cells were separated by Percoll density gradient centrifugation and two different cell populations were cultured in suspension to form ovarian cell aggregates to find suitable cell populations for its formation. Ovarian cell aggregates formed from different cell populations were evaluated by histology and gene expression analyses. To evaluate the media supplements, ovarian cell aggregate culture was performed under different media conditions, and the morphology, viability, size, gene expression, histology, and E2 secretion of ovarian cell aggregates were analyzed. Results: Ovarian cell aggregates were able to be formed well under specific culture conditions that used ultra-low attachment 96 well plate, complete mESM2, and the cell populations from top to 50% layers after separation of ovarian cells. Moreover, they were able to maintain minimal ovarian function such as germ cell maintenance and E2 synthesis for a short period. Conclusions: We established basic conditions for the culture of O. dancena ovarian cell aggregates. Additional efforts will be required to further optimize the culture conditions so that the ovarian cell aggregates can retain the improved ovarian functions for a longer period of time.

• BMB Reports
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• 제33권4호
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• pp.349-352
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• 2000

Proteins that are involved in cellular signal cascade experience phosphorylation and dephosphorylation cycles in their tyrosine residue(s) during cell adhesion. In order to identify the protein(s), which tyrosine desidues are specifically phosphorylated when the cells attached to the substrate, we compared the tyrosine phosphorylation level of proteins between suspension and adhered culture condition in rat fibroblast 3Yl cells. We found that a cluster of 70 kDa protein was specifically phosphorylated when the cells adhered to the substrate, but did not effect the cells held in suspension. The phosphorylated protein is identified as paxillin, a focal adhesion protein in immunoprecipitation and immunobloting analysis. These results suggest that the tyrosine phosphorylation of paxillin may play a role in cell-substrate adhesion.

• 한국미생물·생명공학회지
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• 제18권4호
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• pp.327-330
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• 1990

Thalictrum rugosum 세포배양에서 여러 가지 식물생장 조절물질이 세포증식 및 berberine 생산에 미치는 영향을 조사하였다. 조사된 식물생장 조절물질들 중에서 indole-3-acetic acid(IAA)가 berberine 생산에 가장 적합함을 알 수 있었고 그 최적농도는 1$\mu \textrm m$ 이었다. 비교 기준인 2,4-D의 사용결과에 비할 때 60이상의 생산량 상승효과가 있었으며, cytokinin인 6-benzylaminopurine(BA)를 동시에 사용하는 것보다 IAA만을 단독으로 사용하는 것이 더 효과적이다.

• Journal of Microbiology and Biotechnology
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• 제9권5호
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• pp.589-592
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• 1999

Suspension cultures of Digitalis lanata were successfully performed in an aqueous two-phase system (ATPS) of 4.5％ polyethylene glycol (PEG) 20,000 and 2.8％ crude dextran. Cell growth in the medium containing an individual ATPS-forming polymer was inhibited due to the toxicity of PEG and a high viscosity of dextran. Formation of ATPS supported cell growth by showing a considerably decrease in viscosity and partitioning of cells into a PEG-lean dextran phase. It was found that an aqueous two-phase cultivation of plant cells in a stirred tank bioreactor could be successfully applied.

• 한국미생물·생명공학회지
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• 제26권2호
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• pp.130-136
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• 1998

참당귀 세포(Angelica gigas H4)를 현탁배양하여 면역 증강성 다당을 생산하기 위한 최적배지 설계 및 생산조건 최적화 연구를 수행하였다. 우선 세포의 성장에 가장 좋은 배지로서 Schenk and Hildebrandt(SH) 배지를 선택하여 배양한 결과 $25^{\circ}C$, 암배양조건에서 각각 15.8g DCW/l와 0.85 g/l의 세포 및 다당을 얻을 수 있었다. 한편 다당 생산용 배지의 개발을 위하여 여러 가지 식물세포배양용 배지 중에서 다당 생산능이 가장 높은 배지로서 Gamborg B5 배지를 선정하였으며 B5 배지를 변형하여 면역증강성 다당생산을 위한 최적배지(PPM)를 개발하였다. 그리고 최적배지를 이용한 당귀 세포증식과 다당 생산을 위한 최적 pH는 6.0-6.5, 배양온도는 $20^{\circ}C$, 그리 고 광조건보다는 암조건이 좋은 것으로 밝혀졌다. 상기 최적조건에서의 최대 세포증식과 다당생산은 각각 14.8 g/l, 1.5 g/l 이었다.

• KSBB Journal
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• 제30권3호
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• pp.103-108
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• 2015

Production of foreign proteins by transgenic plant cell cultures has several advantages such as post-translational modification, low risk of product contamination and low-cost production and purification. However, target proteins are degraded by extracellular proteases existing in the media. A solution to this problem is the use of perfusion culture and ion exchange chromatography for the application of integrated bioprocess using in situ recovery. With this method, production of human granulocyte-macrophage colony-stimulating factor (hGM-CSF) was investigated in this study. First, optimization of cell concentration during the induction phase for the production of hGM-CSF was examined. As cell concentration increased, the level of hGM-CSF was decreased due to the presence of extracellular proteases. Induction using sugarfree media produced 33% more hGM-CSF. The effects of pH on the binding of hGM-CSF to cationic and anionic exchange resins were also investigated. In terms of stability, optimal pH was found to be 5~7. In the case of using buffer exchange when CM-Sepharose was used as a cationic exchange resin, optimal pH for binding was 4.8 and adsorption yield was 77%. When DEAE-Sepharose was used as an anionic exchange resin, it was 5.5 (74%). Without buffer exchange, optimal pH was 4.6 (84%). From these results, an integrated bioprocess using in situ recovery with simultaneous production and separation of foreign protein in transgenic plant cell suspension cultures was found to be feasible.

• Journal of Microbiology and Biotechnology
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• 제1권3호
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• pp.220-226
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• 1991

Large and rapid increases in benzophenanthridine alkaloid production occured in suspension cultures of Eschscholtzia californica cells treated with elicitors. Response to different biotic elicitors showed that elicitors prepared from yeast extract, Collectotrichum lindemuthianum and Verticillium dahliae induced alkaloid formation. Highest alkaloid accumulation was obtained with $60\;\mu\textrm{g}$ of yeast extract elicitor per gram of fresh cell weight. In time course performance after elicitor addition, more than 40 hours were required to obtain saturated alkaloid accumulation. Compounded silicone fluid, an ideal accumulation phase for two-phase culture of E. californica, accumulated a large amount of alkaloids produced in a specific manner. Elicitation in two-phase culture clearly increased net alkaloid production as well as their concentrations in the accumulation phase.

• Plant Biotechnology Reports
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• 제2권1호
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• pp.87-92
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• 2008

An improved protocol for high frequency plant regeneration via somatic embryogenesis from zygotic embryo-derived cell suspension cultures of watershield (Brasenia schreberi) was developed. Zygotic embryos formed pale-yellow globular structures and white friable callus at a frequency of 80% when cultured on halfstrength MS medium supplemented with $0.3mg\;l^{-1}$ 2,4-D. However, the frequency of formation of pale-yellow globular structures and white friable callus decreased slightly with increasing concentrations of 2,4-D up to $3mg\;l^{-1}$, where the frequency reached ~50% of the control. Cell suspension cultures from zygotic embryoderived white friable callus were established using half-strength MS medium supplemented with $0.3mg\;l^{-1}$ 2,4-D. Upon plating of cell aggregates on half-strength MS basal medium, approximately 8.3% gave rise to somatic embryos and developed into plantlets. However, the frequency of plantlet development from cell aggregates was sharply increased (by up to 55%) when activated charcoal and zeatin were applied. Regenerated plantlets were successfully transplanted to potting soil and grown to normal plants in a growth chamber. The distinctive feature of this study is the establishment of a high frequency plant regeneration system via somatic embryogenesis from zygotic embryo-derived cell suspension cultures of water-shield, which has not been previously reported. The protocol for plant regeneration of watershield through somatic embryogenesis could be useful for the mass propagation and transformation of selected elite lines.

• 한국식생활문화학회지
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• 제27권6호
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• pp.743-750
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• 2012

A tissue cultured wild ginseng (TCWG) suspension was inoculated with lactic acid bacteria and fermented to improve the functionality of TCWG. The utilization of TCWG was increased directly using the freeze-dried powder. The optimal ratio of TCWG powder and water for fermentation was 1:19 (5%), which was selected by measuring the fluidity and viable cell count according to concentration. The effects on ADH activation and immune cell activation by each ferments with 10 kinds of Lactobacillus sp. strains were examined. The ferments with the Lactobacillus casei KFRI 692 strain showed 5.4 times higher ADH activity and 1.3 times higher ALDH activity than the non-fermented TCWG powder (control). The level of NO production and cytotoxicity was also measured by Raw 264.7 cells. The ferment with the Lac. casei KFRI 692 strain showed the highest level of NO production and lower cytotoxicity than the others. Therefore, the Lac. casei KFRI 692 strain was selected as a strain for fermentation of a TCWG suspension to maximize its functionality. To identify the optimal fermentation time of the selected Lac. casei KFRI 692 strain on the 5% TCWG suspension, the viable cell count of lactic acid bacterial and the changes in pH were observed for 72 hours. 24-hrs was found to be the optimal fermentation time. In this way, fermented TCWG with lactic acid bacteria showed higher ADH activation efficacy and immune cell activation than non-fermented TCWG.

• Plant Biotechnology Reports
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• 제4권2호
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• pp.125-128
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• 2010

Culture conditions were established for high frequency plant regeneration via somatic embryogenesis from cell suspension cultures of Nymphoides coreana. Zygotic embryos formed pale-yellow globular structures and calluses at a frequency of 85.6% when cultured on half-strength Murashige and Skoog (MS) medium supplemented with 0.3 $mg\;l^{-1}$ of 2,4-D. However, the frequency of pale-yellow globular structures and white callus formation decreased slightly with an increasing concentration of 2,4-D up to 10 $mg\;l^{-1}$ with the frequency rate falling to 16.7%. Cell suspension cultures were established from zygotic embryo-derived calluses using half-strength MS medium supplemented with 0.3 $mg\;l^{-1}$ of 2,4-D. Upon plating onto half-strength MS basal medium, over 92.3% of cell aggregates gave rise to numerous somatic embryos and developed into plantlets. Regenerated plantlets were successfully transplanted into potting soil and achieved full growth to an adult plant in a growth chamber. The high frequency plant regeneration system for Nymphoides coreana established in this study will be useful for genetic manipulation and cryopreservation of this species.