• Korean Journal of Plant Tissue Culture
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• v.21 no.2
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• pp.91-97
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• 1994

To improve the productivity of peroxidase (POD) of cell line SP-47 derived from cell suspension cultures of sweet potato (Ipomoea batatas (L) Lam.cv White Star), we optimized culture conditions including the composition and concentration of plant growth regulators and carbon source, and the cell inoculum size. When one g (fr wt) of cells was inoculated into 50 mL TL medium supplemented with l mg/L 2,4-D and 30g/L sucrose in 300 mL Erlenmeyer flask at 25$^{\circ}C$ in the dark (100rpm), the POD activity per g cell dry wt was maximized to be about 6,800 units after 25 days of subculture, which was about 30 times higher than that of intact roots of horseradish plants grown in the greenhouse, but the cell growth was maximum after 15 days of subculture. The protein content per g cell dry wt maintained almost plateau and after 25 days of subculture decreased as culture Proceeded further whereas the POD specific activity (unit/mg protein) was about two times higher after subculture and continuously increased from 12 days to the end of cultures (40 days). The POD isozyme patterns showed almost the same regardless of cell growth stage, but some acidic isozymes were slightly increased after 25 days of subculture. These results indicate that POD activity in suspension cultures of sweet potato is closely associated with cell growth and stresses derived from cell culture renditions and medium depletion. Due to its high POD activity the SPL47cell line seems to be suitable for the mass production of POD.

• Journal of Plant Biotechnology
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• v.3 no.1
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• pp.1-5
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• 2001

Traditional culture technology of medicinal plants mainly depends on the field culture, which has many problems. With progress of modern culture technology, it has become possible to produce valuable secondary metabolites from medicinal plants. In this paper, we discuss about the pigment and saikosaponin production from too medicinal plants, Carthamus tinctorius and Bupleurum falcatum, through bioreactor culture system. A two-stage bioreactor culture system was established for the production of yellow and red pigments and saikosaponins by cell suspension cultures of Carthamus tinctorius and Bupleurum falcatum. In Carthamus tinctorius, balloon type airlift bioreactors and column type airlift bioreactors were employed for the tell culture and for the pigment production, respectively. The greatest pigment production was obtained on White medium supplemented with 4 mg/L kinetin, high levels of sucrose concentration and photosynthetic photon flux. In Bupleurum falcatum, adventitious roots were cultured in balloon type airlift bioreactors and the root growth was greatest on SH medium containing 5 mg/L IBA and 0.2 mg/L kinetin. HPLC analysis showed that the contents of main active saikosaponins a, c, and d in adventitious roots were almost the same as those in field cultured root.

• Journal of Microbiology and Biotechnology
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• v.1 no.1
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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.

• Journal of Plant Biology
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• v.36 no.1
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• pp.19-27
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• 1993

Effects of polyamines on DNA synthesis were studied in synchronized culture of Nicotiana tabacum L. When DFMO and DFMA, inhibitors of ornithine decarboxylase and arginine decarboxylase, respectively were initially applied to the cells, the polyamine contents were rapidly dropped and [methyl-3H] thymidine incorporation into DNA was markedly reduced during the early stage of culture period. Inhibition of DNA synthesis, however, was partially reversed when these inhibitors were applied simultaneously with putrescine. In addition, exogenous administration of putrescine also increased the DNA synthesis during the all over the culture period. In vitro activity of DNA polymerase from Nicotiana tabacum L. was promoted by increasing concentrations of polyamines in the reaction mixture. Maximal activity was shown at 5 mM putrscine, 0.5 mM spermidine and spermine, respectively. Lack of Mg2+ ion in the reaction buffer resulted in an inhibition of the enzyme activity by about 30%. The inhibition could not be completely reversed by application of polyamines at optimal concentrations. These results suggest that polyamines promote the DNA synthesis in vivo and in vitro by stabilizing the DNA-helix upon binding to negatively charged groups on DNA or increasing the activity of DNA polymerase in Nicotiana tabacum L.

• Korean Journal of Plant Tissue Culture
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• v.27 no.3
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• pp.185-189
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• 2000

For the micropropagation, node explants of cassava were cultured in liquid MS medium with various concentrations of cytokinins on a rotary shaker (100 rpm) for 2 weeks. The adventitious roots and shoots from the explants were differentiated more efficiently in liquid medium than in solid. But root formation was not inhibited in medium with BAP and kinetin at low concentration (＞0.05 mg 1/sup -1/), while in medium added with BAP and zeatin at high level (＜0.25 mg 1/sup -1/), it was inhibited by callus forming on cut end of the cuttings. However, all of plantlets grown in liquid medium for more than 2 weeks showed symptoms of hyperhydricity. The plantlets grown in liquid medium were transferred into culture bottles filled with fine sand or artificial soil (pitmoss：perlite：vermiculite, 1：1：1 v/v) wetted with half strength of Knop's solution. After transplanted to culture bottles, some of vitriscent leaves were defoliated and new leaves were normally formed from shoot apex. Most of plantlets (＞95%) were hardened-off successfully only in culture bottles with fine sand, and grew into 3-5 cm seedlings possessing 4-6 nodes after 4 weeks. Thus, the mass propagation of cassava on medium containing cytokinin could be established based on the suspension culture using node explants.

• 한국생물공학회:학술대회논문집
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• 2003.10a
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• pp.329-333
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• 2003

Human granulocyte-macrophage colony-stimulating factor (hGM-CSF) is a glycoprotein that stimulates the production of granulocytes, macrophages and white blood cells. hGM-CSF secreted by transgenic Nicotiana tabacum suspension cells was unstable in the culture medium and rapidly degraded by extracellular preteases. In order to reduce extracellular pretense activity, culture temperature was lowered. Then, the production of hGM-CSF by transgenic plant suspension cell cultures could be enhanced by reduced degradation of hGM-CSF at low temperature.

• KSBB Journal
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• v.30 no.3
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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.

• Korean Journal of Plant Tissue Culture
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• v.21 no.6
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• pp.357-362
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• 1994

Chitinases and $\beta$-1,3-glucanases are believed to be important in defending plane against pathogens. Here, we investigated the expression of chitinase(s) in Arabidopsis thaliana cell suspension culture system in response to ethephon (2-chloroethyl phosphonic acid) which produces ethylene or a microbial elicitor, a bacterial pectin-degrading enzyme, $\beta$-1, 4-endopolygalactronic acid Iyase (PGA Iyase), treatment. Chitinase activity was measured either by radio chemical assay using $^3$H-labeled regenerated chitin as substrate or western blot analysis using antibody raised against tobacro chitinase(S). With 1 mg/mL of ethephon or 100 m units/mL of elicitor treatment, maximum levels of activity were reached after 48h. We also investigated distribution of chitinase activity in seedlings, leaves, and root of A. thaliana and found that root have the highest chitinase activity.

• Journal of Plant Biotechnology
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• v.34 no.3
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• pp.207-212
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• 2007

This study describes a simple and rapid method for quantitative determination of $\gamma$-aminobutyric acid (GABA) using $^1H$ NMR spectroscopy from whole cell extracts of plant suspension cultures. When 9 cell lines derived from 8 species of higher plants maintained in liquid Marashige and Skoog (MS) medium supplemented with 1 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D) were subjected to $^1H$ NMR, a cell line of Coriandrum sativum L. exhibited the highest level of GABA. The level reached up to 16.9 mg/dry wt when cells were cultured in MS medium supplemented with 0.5 mg/L 2,4-D after 3 weeks of incubation. The method for quantitative determination of GABA using $^1H$ NMR established in this study could be applied to high-throughput screening of various plant resources for GABA production and the cell suspension culture system of C. sativum could be further developed for commercial production of GABA.

• Korean Journal of Plant Tissue Culture
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• v.27 no.2
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• pp.125-131
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• 2000

Protoplasts of Arabidopsis thaliana were easily isolated from the shoot-forming (SF) suspension-cultured cell clusters with 4 hours-shaking condition (40 rpm) on CPD enzyme solution containing 1% cellulase R-10, 0.25% pectolyase Y-23 and 0.5% driselase. Protoplasts were cultured on liquid KAO medium supplemented with 1 mg/L 2,4-D, 0.5 mg/L kinetin, 200 mg/L spermidine and 68 g/L glucose. Also, protoplasts were cultured on 0.2 $\mu$M membrane filter placed onto CP solid medium containing the suspension cells as feeder cells in the dark at $25^{\circ}C$ for 4 weeks. Protoplast-derived-SF calli were cultured on MS medium containing 0.05 mg/L IAA, 7 mg/L 2 ip and 30 g/L sucrose under the continuous illumination for four weeks. The frequency of shoot formation was about 60%. The regenerants were transferred into potting soil to grow mature plants. The regenerants formed the silques with seeds after 8 weeks of cultures.