• Proceedings of the Korean Society of Life Science Conference
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• 2002.12a
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• pp.48-60
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• 2002

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a cytokine that stimulates the production of granulocytes, macrophages, and white blood cells. The effects of osmotic pressure on secretion of human GM-CSF into the culture medium were investigated in suspension cultures of transgenic tobacco cells. An increase in osmotic pressure caused by the addition of mannitol decreased the cell size index, with the effect being more pronounced when cells were measured wet rather than dry. Increased osmotic pressure enhanced the secretion of hGM-CSF. At 90 g/L mannitol, the maximum concentration tested, hGM-CSF was present in the culture medium at 980 ug/L. As the concentration of mannitol increased, the total amount of protein secreted also increased, but was disproportionately enriched in GM-CSF NaCl, another osmoticum, had very similar effects on cell growth and hGM-CSF production, but did not cause enrichment for hGM-CSF Additionally, protein-stabilizing polymer was added to culture broth to enhance stability of secreted recombinant protein. Finally, above two method were applied together to maximize the productivity.

• Proceedings of the Botanical Society of Korea Conference
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• 1995.06a
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• pp.40-56
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• 1995

During the past two decades, China has seen her great progress in plant biotechnology. Since the Chinese market of herb medicine is huge, while the plant resources are shrinking, particular emphasis has been placed in plant tissue and cell cultures of medicinal plants, this includes fast propagation, protoplast isolation and regeneration, cell suspension cultures and large scale fermentation. To optimize culture conditions for producing secondary compounds in vitro, various media, additives and elicitors have been tested. Successful examples of large scale culture for the secondary metabolite biosynthesis are quite limited : Lithospermum ery throrhizon and Arnebia euchroma for shikonin derivatives, Panax ginseng, P. notoginseng, P. quinquefolium for saponins, and a few other medicinal plants. Recent development of genetic transformation systems of plant cells offered a new approach to in vitro production of secondary compounds. Hairy root induction and cultures, by using Ri-plasmid, have been reported from a number of medicinal plant species, such as Artemisia annua that produces little artemisinin in normal cultured cells, and from Glycyrrhiza uralensis. In the coming five years, Chinese scientists will continue their work on large scale cell cultures of a few of selected plant species, including Taxus spp. and A. annua, for the production of secondary metabolites with medicinal interests, one or two groups of scientists will be engaged in molecular cloning of the key enzymes in plant secondary metabolism.

• Journal of Plant Biotechnology
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• v.36 no.1
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• pp.7-12
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• 2009

This study was conducted to establish the optimal suspension culture system for both the propagation of embryogenic cells (ECs) and the induction of somatic embryos (SEs) of Kalopanax septemlobus. The proliferation rate of ECs was reduced as the inoculum density was increased; the highest rate was obtained when 0.1 g/100 ml of cells was initially inoculated. According to the analysis of cell growth pattern and cell growth cycle (G1, Sand G2/M), the cell growth started in 5 days culture initiation, grew rapidly until 15 days and then decreased gradually. Distinctive changes of the cell growth cycle by the culture periods was also observed; the growth cycle was doubled from initial 5.6% to 11.7% of S stage in 5 days culture and then reached in stable stages again. Therefore, the results indicated that a 15-day-cycle was the optimal culture period for the propagation of the ECs through the suspension culture. Furthermore, the cell inoculum density was also important for the induction of SE; more than 65% of SEs at the torpedo stage was induced by using the low level of cell inoculum (0.5 g/L), while the higher inoculum densities were rapidly reduced the proportion of SEs at that stage. Although the higher inoculum density delayed the development of SE, it did not affect the proportion of SEs at the globular and heart stage. In conclusion, this study showed that the suspension culture of the Kalopanax septemlobus ECs through the control of inoculum density was an efficient way for both the propagation of ECs and the induction of SEs, suggesting that the development of this system might help to reduce the culture period for the somatic embryo production.

• Korean Journal of Plant Tissue Culture
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• v.27 no.3
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• pp.155-161
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• 2000

This experiment was carried out to examine the effects of culture medium on cell growth of the viola (Viola patrinii DC.) suspension culture. The results are as follows: The greatest cell growth rates were found with MS medium suggesting that this medium could be recommendable for the viola suspension cell culture. When the nitrogen sources (NH$_4$NO$_3$ and KNO$_3$) of MS salts were diluted at half concentrations, the cell growth rates were slightly increased, but when the combined concentration rations of NH$_4$+ and NO$_3$ions were 25 to 75 the greatest cell growth rates were obtained. This result imply that the nitrogen ion sources had slight influence on the rates. Another feature was obtained. This result implys that the nitrogen ion sources had slight influence on the rates. Another feature was that as the concentration of NH$_4$+ ion lowered, the callus color changed to pale yellow with some red spots. The addition of casein hydrolysate (5 g/L) was more effective for the cell growth. On the basis of microscopic observation, the highest cell growth rates were detected during 2-4 weeks culture and after 6 weeks of the culture, some elongated and vacuolated cells were determined.

• Journal of Plant Biotechnology
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• v.36 no.4
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• pp.309-319
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• 2009

Transgenic plant cell cultures for the production of biopharmaceuticals including monoclonal antibodies, recombinant proteins have been regarded as an alternative platform in addition to traditional microbial fermentation and mammalian cell cultures. Plant-made pharmaceuticals (PMPs) have several advantages such as safety, cost-effectiveness, scalability and possibility of complex post-translational modifications. Increasing demand for the quantity and diversity of pharmaceutical proteins may accelerate the industrialization of PMP technology. Up to date, there is no plant-made recombinant protein approved by USFDA (Food and Drug Administration) for human therapeutic uses due to the technological bottlenecks of low expression level and slight differences in glycosylation. Regarding expression levels, it is possible to improve the productivity by using stronger promoter and optimizing culture processes. In terms of glycosylation, humanization has been attempted in many ways to reduce immune responses and to enhance the efficacy as well as stability. In this review article, all these respects of transgenic plant cell cultures were summarized. In addition, we also discuss the general characteristics of plant cell suspension cultures related with bioreactor design and operation to achieve high productivity in large scale which could be a key to successful commercialization of PMPs.

• Journal of Plant Biotechnology
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• v.6 no.4
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• pp.265-268
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• 2004

Cell growth and ginseng saponin production by large-scale suspension (bioreactor) cultures of Panax ginseng were investigated under various inoculum sizes. Cell growth was low at an inoculum size of 40 g FW/L, and the maximum cell growth was obtained with increasing inoculum size up to 100 g FW/L. The cell density of 333 g FW/L and 12.7 g DW/L was obtained at inoculum size of 100 g FW/L after 30 days of cultivation. Maximum saponin production of $4.40\;\cal{mg/g}$ DW was achieved at 60 g FW/L of inoculum size. Thus, inoculum size 60 g FW/L was suitable for optimum biomass accumulation as well as saponin production during bioreactor cultivation of ginseng suspension cells.

• Proceedings of the Korean Society of Crop Science Conference
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• 2007.04a
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• pp.65-73
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• 2007

The objective of this study is to understand how regulatory mechanisms respond to sugar status for more efficient carbon utilization and source-sink regulation in plants. So, we need to identify and characterize many components of sugar-response pathways for a better understanding of sugar responses. For this end, genes responding change of sugar status were screened using Arabidpsis cDNA arrays, and confirmed thirty-six genes to be regulated by sucrose supply in detached leaves by RNA blot analysis. Eleven of them encoding proteins for amino acid metabolism and carbohydrate metabolism were repressed by sugars. The remaining genes induced by sugar supply were for protein synthesis including ribosomal proteins and elongation factors. Among them, I focused on three hydrolase genes encoding putative $\beta$-galactosidase, $\beta$-xylosidase, and $\beta$-glucosidase that were transcriptionally induced in sugar starvation. Homology search indicated that these enzymes were involved in hydrolysis of cell wall polysaccharides. In addition to my results, recent transcriptome analysis suggested multiple genes for cell wall degradation were induced by sugar starvation. Thus, I hypothesized that enzyme for cell wall degradation were synthesized and secreted to hydrolyze cell wall polysaccharides producing carbon source under sugar-starved conditions. In fact, the enzymatic activities of these three enzymes increased in culture medium of Arabidopsis suspension cells under sugar starvation. The $\beta$-galactosidase encoded by At5g56870 was identified as a secretory protein in culture medium of suspension cells by mass spectrometry analysis. This protein was specifically detected under sugar-starved condition with a specific antibody. Induction of these genes was repressed in suspension cells grown with galactose, xylose and glucose as well as with sucrose. In planta, expression of the genes and protein accumulation were detected when photosynthesis was inhibited. Glycosyl hydrolase activity against galactan also increased during sugar starvation. Further, contents of cell wall polysaccharides especially pectin and hemicellulose were markedly decreased associating with sugar starvation in detached leaves. The amount of monosaccharide in pectin and hemicellulose in detached leaves decreased in response to sugar starvation. These results supported my idea that cell wall has one of function to supply carbon source in addition to determination of cell shape and physical support of plant bodies.

• Proceedings of the Korean Society of Plant Biotechnology Conference
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• 2003.04a
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• pp.162-162
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• 2003

• Journal of Plant Biotechnology
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• v.47 no.3
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• pp.227-234
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• 2020

To produce the miraculin protein in suspension cultures, rice (Oryza sativa L.) was transformed with Agrobacterium tumefacience EHA105 containing the miraculin AB512278 gene. The cell suspension cultures were established using cell lines selected from transgenic rice callus. The integration of the miraculin gene into the rice chromosome was confirmed using genomic PCR analysis. In addition, RT-PCR analysis indicated that the miraculin gene is expressed in the selected suspension cell lines. Thus, the recombinant miraculin was expressed in the transgenic suspension cell line, HK-2. Therefore, we have successfully developed a HK-2 line that produces miraculin. These results demonstrate that transformed cell suspension cultures can be used to produce a taste-modifying protein such as miraculin.

• Korean Journal of Plant Tissue Culture
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• v.25 no.2
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• pp.81-88
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• 1998

The Southeast Asian javanica rice variety Tinawen was investigated for efficient protoplast culture and plant regeneration from cell suspension-derived protoplasts using a feeder cell culture method. Feeder cells of both Lolium multiflorum and Oryza ridleyi, either alone, or in combination, were employed and plants were regenerated from protoplast-derived colonies on several plant regeneration media. Dehydration of protoplast-derived colonies was also investigated as a means of enhancing plant regeneration. In the presence of L. multiflorum or O. ridleyi feeder cells, the protoplast plating efficiency ranged from 0.09% to 1.48%, depending on the feeder cell type and the age of the cell suspension. L. multiflorum feeder cells induced approximately 6-fold higher plating efficiency compared with those of O. ridleyi. The plant regeneration frequencies were 19.3-31.7% with L. multiflorum, 13.0-18.0% with O. ridleyi and 18.0-22.0% with a mixture of both in various plant regeneration media when protoplast-derived colonies were dehydrated, while for the non-dehydrated colonies, the values were 2.0-7.0%, 3.0-5.0% and 0-4.0%, respectively. Flow cytometric analysis of 34 protoplast-derived plants showed that the majority of plants were diploids and only 2 plants were tetraploids. The plants which were transferred to glasshouse were fertile.