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Moonsik Yang - One of the best experts on this subject based on the ideXlab platform.
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co expression of proteinase inhibitor enhances recombinant human granulocyte macrophage colony stimulating factor production in transgenic rice Cell Suspension Culture
Protein Expression and Purification, 2008Co-Authors: Taegeum Kim, Taeho Kwon, Yongsuk Jang, Hojin Lee, Yunji Shin, Moonsik YangAbstract:Abstract The synthetic gene (sPI-II) harboring the chymotrypsin (C1) and trypsin (T1) inhibitor domains of the Nicotiana alata serine proteinase inhibitor II gene has been previously expressed, and extraCellular protease activity was shown to be reduced in the Suspension Culture medium. In this study, the sPI-II gene was introduced into transgenic rice Cells expressing rhGM-CSF (recombinant human granulocyte–macrophage colony-stimulating factor), in an effort to reduce protease activity and increase rhGM-CSF accumulation in the Suspension Culture medium. The integration and expression of the introduced sPI-II gene in the transgenic rice Cells were verified via genomic DNA PCR amplification and Northern blot analysis, respectively. Relative protease activity was found to have been reduced and rhGM-CSF production was increased 2-fold in the co-transformed Cell Suspension Culture with rhGM-CSF and the sPI-II gene, as compared with that observed in the transformed Cell Suspension Culture expressing rhGM-CSF only. These results indicate that a transformed plant Cell Suspension Culture system expressing the proteinase inhibitor can be a useful tool for increasing recombinant protein production.
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production of bioactive human granulocyte colony stimulating factor in transgenic rice Cell Suspension Cultures
Protein Expression and Purification, 2006Co-Authors: Shinyoung Hong, Taeho Kwon, Yongsuk Jang, Sooho Kim, Moonsik YangAbstract:Human granulocyte-colony stimulating factor (hG-CSF), a human cytokine, was expressed in transgenic rice Cell Suspension Culture. The hG-CSF gene was cloned into the rice expression vector containing the promoter, signal peptide, and terminator derived from a rice α-amylase gene Amy3D. Using particle bombardment-mediated transformation, hG-CSF gene was introduced into the calli of rice (Oryza sativa) cultivar Dong-jin. Expression of the hG-CSF gene was confirmed by ELISA and Northern blot analysis. The amount of recombinant hG-CSF accumulated in Culture medium from transgenic rice Cell Suspension Culture on the sugar starvation was determined by time series ELISA. Biological activity of the plant derived hG-CSF was confirmed by measuring the proliferation of the AML-193 Cells, and was similar to that of the commercial Escherichia coli-derived hG-CSF. In this paper, we discuss the attractive attributes of using rice Cell Suspension system for the expression of therapeutic recombinant hG-CSF.
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partitioning of recombinant human granulocyte macrophage colony stimulating factor hgm csf from plant Cell Suspension Culture in peg sodium phosphate aqueous two phase systems
Biotechnology and Bioprocess Engineering, 2004Co-Authors: Jaehwa Lee, Taeho Kwon, Nguyen Hoang Loc, Moonsik YangAbstract:Partitioning of human granulocyte-macrophage colony stimulating factor (hGM-CSF) was achieved in the aqueous two-phase systems (ATPSs) using a crude extract of transgenic tobacco Cell Suspension Culture. This study examined the effects of polyethylene glycol (PEG) molecular weight and concentration and the effects of sodium phosphate concentration in different PEG/sodium phosphate systems on the partition coefficient,K. The best ATPS system was 5% PEG 8,000/1.6 M sodium phosphate after 2 h of incubation at room temperature. In this system, hGM-CSF was partitioned in the PEG-rich phase with a yield of 57.99% andK hGM-CSF of 8.12. In another system, 3% PEG 10,000/1.6 M sodium phosphate, hGM-CSF was also partitioned primarily in the top phase with a yield of 45.66% andK hGM-CSF of 7.64 after 2 h of incubation at room temperature.
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high level of expression of recombinant human granulocyte macrophage colony stimulating factor in transgenic rice Cell Suspension Culture
Biotechnology and Bioengineering, 2003Co-Authors: Yunji Shin, Shinyoung Hong, Taeho Kwon, Yongsuk Jang, Moonsik YangAbstract:Recombinant human granulocyte-macrophage colony stimulating factor (hGM-CSF) has been previously produced in tobacco Cell Suspension Cultures. However, the amount of hGM-CSF accumulated in the Culture medium dropped quickly from its maximum of 150 microg/L at 5 d after incubation. To overcome this problem, we sought an expression system in which heterologous gene expression could be induced at high levels. We selected a rice amylase expression system in which the promoter Ramy3D is induced to express recombinant protein by sucrose starvation. This induction system was found to give good yield of recombinant hGM-CSF in transgenic rice Cell Suspension Culture and protease activity of this Culture medium was low compared to that of tobacco Culture system.
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establishment of a transgenic tobacco Cell Suspension Culture system for producing murine granulocyte macrophage colony stimulating factor
Molecules and Cells, 1997Co-Authors: Joonggi Lee, Taeho Kwon, Sangbong Choi, H S Kang, Kangjin Cho, Dae Hyuk Kim, Youngjoo Jang, Moonsik YangAbstract:We tested if murine granulocyte-macrophage colony stimulating factor (mGM-CSF) is produced as a biologically active form through plant Cell Culture. The mGM-CSF gene was cloned into the plant expression vector, pBI121, and Ti-plasmid mediated transformation of tobacco leaves was conducted using Agrobacterium tumefaciens harboring the recombinant mGM-CSF (rmGM-CSF) gene. Cell Suspension Culture was established from the leaf-derived calli of transgenic tobacco plants. Northern blot analysis indicated the expression of the introduced mGM-CSF gene in both transgenic plant and Cell Suspension Cultures. In addition, the biological activity of rmGM-CSF from plant Cell Culture was confirmed by measuring the proliferation of GM-CSF dependent FDC-P1 Cells.
Taeho Kwon - One of the best experts on this subject based on the ideXlab platform.
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co expression of proteinase inhibitor enhances recombinant human granulocyte macrophage colony stimulating factor production in transgenic rice Cell Suspension Culture
Protein Expression and Purification, 2008Co-Authors: Taegeum Kim, Taeho Kwon, Yongsuk Jang, Hojin Lee, Yunji Shin, Moonsik YangAbstract:Abstract The synthetic gene (sPI-II) harboring the chymotrypsin (C1) and trypsin (T1) inhibitor domains of the Nicotiana alata serine proteinase inhibitor II gene has been previously expressed, and extraCellular protease activity was shown to be reduced in the Suspension Culture medium. In this study, the sPI-II gene was introduced into transgenic rice Cells expressing rhGM-CSF (recombinant human granulocyte–macrophage colony-stimulating factor), in an effort to reduce protease activity and increase rhGM-CSF accumulation in the Suspension Culture medium. The integration and expression of the introduced sPI-II gene in the transgenic rice Cells were verified via genomic DNA PCR amplification and Northern blot analysis, respectively. Relative protease activity was found to have been reduced and rhGM-CSF production was increased 2-fold in the co-transformed Cell Suspension Culture with rhGM-CSF and the sPI-II gene, as compared with that observed in the transformed Cell Suspension Culture expressing rhGM-CSF only. These results indicate that a transformed plant Cell Suspension Culture system expressing the proteinase inhibitor can be a useful tool for increasing recombinant protein production.
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production of bioactive human granulocyte colony stimulating factor in transgenic rice Cell Suspension Cultures
Protein Expression and Purification, 2006Co-Authors: Shinyoung Hong, Taeho Kwon, Yongsuk Jang, Sooho Kim, Moonsik YangAbstract:Human granulocyte-colony stimulating factor (hG-CSF), a human cytokine, was expressed in transgenic rice Cell Suspension Culture. The hG-CSF gene was cloned into the rice expression vector containing the promoter, signal peptide, and terminator derived from a rice α-amylase gene Amy3D. Using particle bombardment-mediated transformation, hG-CSF gene was introduced into the calli of rice (Oryza sativa) cultivar Dong-jin. Expression of the hG-CSF gene was confirmed by ELISA and Northern blot analysis. The amount of recombinant hG-CSF accumulated in Culture medium from transgenic rice Cell Suspension Culture on the sugar starvation was determined by time series ELISA. Biological activity of the plant derived hG-CSF was confirmed by measuring the proliferation of the AML-193 Cells, and was similar to that of the commercial Escherichia coli-derived hG-CSF. In this paper, we discuss the attractive attributes of using rice Cell Suspension system for the expression of therapeutic recombinant hG-CSF.
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partitioning of recombinant human granulocyte macrophage colony stimulating factor hgm csf from plant Cell Suspension Culture in peg sodium phosphate aqueous two phase systems
Biotechnology and Bioprocess Engineering, 2004Co-Authors: Jaehwa Lee, Taeho Kwon, Nguyen Hoang Loc, Moonsik YangAbstract:Partitioning of human granulocyte-macrophage colony stimulating factor (hGM-CSF) was achieved in the aqueous two-phase systems (ATPSs) using a crude extract of transgenic tobacco Cell Suspension Culture. This study examined the effects of polyethylene glycol (PEG) molecular weight and concentration and the effects of sodium phosphate concentration in different PEG/sodium phosphate systems on the partition coefficient,K. The best ATPS system was 5% PEG 8,000/1.6 M sodium phosphate after 2 h of incubation at room temperature. In this system, hGM-CSF was partitioned in the PEG-rich phase with a yield of 57.99% andK hGM-CSF of 8.12. In another system, 3% PEG 10,000/1.6 M sodium phosphate, hGM-CSF was also partitioned primarily in the top phase with a yield of 45.66% andK hGM-CSF of 7.64 after 2 h of incubation at room temperature.
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high level of expression of recombinant human granulocyte macrophage colony stimulating factor in transgenic rice Cell Suspension Culture
Biotechnology and Bioengineering, 2003Co-Authors: Yunji Shin, Shinyoung Hong, Taeho Kwon, Yongsuk Jang, Moonsik YangAbstract:Recombinant human granulocyte-macrophage colony stimulating factor (hGM-CSF) has been previously produced in tobacco Cell Suspension Cultures. However, the amount of hGM-CSF accumulated in the Culture medium dropped quickly from its maximum of 150 microg/L at 5 d after incubation. To overcome this problem, we sought an expression system in which heterologous gene expression could be induced at high levels. We selected a rice amylase expression system in which the promoter Ramy3D is induced to express recombinant protein by sucrose starvation. This induction system was found to give good yield of recombinant hGM-CSF in transgenic rice Cell Suspension Culture and protease activity of this Culture medium was low compared to that of tobacco Culture system.
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expression and secretion of the heterodimeric protein interleukin 12 in plant Cell Suspension Culture
Biotechnology and Bioengineering, 2003Co-Authors: Taeho Kwon, Yongsuk Jang, J E Seo, Jae Bum Kim, Jinhee Lee, Myeonsik YangAbstract:It has been suggested that plant Cell Culture is the most suitable system for producing small-to-medium quantities of specialized, expensive, and high-purity proteins. Here, we report that a heterodimeric protein, human interleukin-12 (hIL-12), was expressed and secreted into Culture medium in a biologically active form. A transgenic plant expressing hIL-12 was constructed by sexual crossing of plants that expressed each subunit of the protein. From a piece of transgenic plant, callus was induced and Cell Suspension Culture was established. The biological activity and amount of hIL-12 secreted into Culture medium were analyzed using bioassays and ELISA. Analysis of Cellular localization demonstrated that the protein was secreted into the Culture medium together with its intrinsic signal peptide.
Yongsuk Jang - One of the best experts on this subject based on the ideXlab platform.
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co expression of proteinase inhibitor enhances recombinant human granulocyte macrophage colony stimulating factor production in transgenic rice Cell Suspension Culture
Protein Expression and Purification, 2008Co-Authors: Taegeum Kim, Taeho Kwon, Yongsuk Jang, Hojin Lee, Yunji Shin, Moonsik YangAbstract:Abstract The synthetic gene (sPI-II) harboring the chymotrypsin (C1) and trypsin (T1) inhibitor domains of the Nicotiana alata serine proteinase inhibitor II gene has been previously expressed, and extraCellular protease activity was shown to be reduced in the Suspension Culture medium. In this study, the sPI-II gene was introduced into transgenic rice Cells expressing rhGM-CSF (recombinant human granulocyte–macrophage colony-stimulating factor), in an effort to reduce protease activity and increase rhGM-CSF accumulation in the Suspension Culture medium. The integration and expression of the introduced sPI-II gene in the transgenic rice Cells were verified via genomic DNA PCR amplification and Northern blot analysis, respectively. Relative protease activity was found to have been reduced and rhGM-CSF production was increased 2-fold in the co-transformed Cell Suspension Culture with rhGM-CSF and the sPI-II gene, as compared with that observed in the transformed Cell Suspension Culture expressing rhGM-CSF only. These results indicate that a transformed plant Cell Suspension Culture system expressing the proteinase inhibitor can be a useful tool for increasing recombinant protein production.
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production of bioactive human granulocyte colony stimulating factor in transgenic rice Cell Suspension Cultures
Protein Expression and Purification, 2006Co-Authors: Shinyoung Hong, Taeho Kwon, Yongsuk Jang, Sooho Kim, Moonsik YangAbstract:Human granulocyte-colony stimulating factor (hG-CSF), a human cytokine, was expressed in transgenic rice Cell Suspension Culture. The hG-CSF gene was cloned into the rice expression vector containing the promoter, signal peptide, and terminator derived from a rice α-amylase gene Amy3D. Using particle bombardment-mediated transformation, hG-CSF gene was introduced into the calli of rice (Oryza sativa) cultivar Dong-jin. Expression of the hG-CSF gene was confirmed by ELISA and Northern blot analysis. The amount of recombinant hG-CSF accumulated in Culture medium from transgenic rice Cell Suspension Culture on the sugar starvation was determined by time series ELISA. Biological activity of the plant derived hG-CSF was confirmed by measuring the proliferation of the AML-193 Cells, and was similar to that of the commercial Escherichia coli-derived hG-CSF. In this paper, we discuss the attractive attributes of using rice Cell Suspension system for the expression of therapeutic recombinant hG-CSF.
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high level of expression of recombinant human granulocyte macrophage colony stimulating factor in transgenic rice Cell Suspension Culture
Biotechnology and Bioengineering, 2003Co-Authors: Yunji Shin, Shinyoung Hong, Taeho Kwon, Yongsuk Jang, Moonsik YangAbstract:Recombinant human granulocyte-macrophage colony stimulating factor (hGM-CSF) has been previously produced in tobacco Cell Suspension Cultures. However, the amount of hGM-CSF accumulated in the Culture medium dropped quickly from its maximum of 150 microg/L at 5 d after incubation. To overcome this problem, we sought an expression system in which heterologous gene expression could be induced at high levels. We selected a rice amylase expression system in which the promoter Ramy3D is induced to express recombinant protein by sucrose starvation. This induction system was found to give good yield of recombinant hGM-CSF in transgenic rice Cell Suspension Culture and protease activity of this Culture medium was low compared to that of tobacco Culture system.
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expression and secretion of the heterodimeric protein interleukin 12 in plant Cell Suspension Culture
Biotechnology and Bioengineering, 2003Co-Authors: Taeho Kwon, Yongsuk Jang, J E Seo, Jae Bum Kim, Jinhee Lee, Myeonsik YangAbstract:It has been suggested that plant Cell Culture is the most suitable system for producing small-to-medium quantities of specialized, expensive, and high-purity proteins. Here, we report that a heterodimeric protein, human interleukin-12 (hIL-12), was expressed and secreted into Culture medium in a biologically active form. A transgenic plant expressing hIL-12 was constructed by sexual crossing of plants that expressed each subunit of the protein. From a piece of transgenic plant, callus was induced and Cell Suspension Culture was established. The biological activity and amount of hIL-12 secreted into Culture medium were analyzed using bioassays and ELISA. Analysis of Cellular localization demonstrated that the protein was secreted into the Culture medium together with its intrinsic signal peptide.
A M Nosov - One of the best experts on this subject based on the ideXlab platform.
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rare triterpene glycoside of ginseng ginsenoside malonyl rg 1 detected in plant Cell Suspension Culture of panax japonicus var repens
Russian Journal of Plant Physiology, 2017Co-Authors: D V Kochkin, B A Galishev, E S Glagoleva, M V Titova, A M NosovAbstract:This paper reports for the first time about the detection and identification of ginsenoside malonyl-Rg1 (the rare 20(S)-protopanaxatriol-type ginsenoside) in the biomass of plant Cell Suspension Culture of Japanese ginseng (Panax japonicus C.A. Mey. var. repens). Ginsenosides were analyzed by means of high-performance liquid chromatography/electrospray ionization mass spectrometry (HPLC-ESI-MS) in positive-ion mode. Malonyl-Rg1 was identified as a result of interpretation of MS spectra obtained upon fragmentation of protonated molecular ion ([M + H]+) of this compound in an ionization source. Chromatographic analysis and MS spectra showed that the Cells of P. japonicus var. repens cultivated in vitro contain several isomers of malonyl-Rg1. Thus, we ascertained for the first time that, in addition to malonyl ginsenosides of 20(S)-protopanaxadiol group, the plant Cell Culture of ginseng P. japonicus var. repens can accumulate glycosides of 20(S)-protopanaxatriol group acylated with a malonic acid residue. The obtained results showed that, in the Cells of ginseng cultivated in vitro for a long time (for 10 years and more), the assortment of secondary metabolites (ginsenosides) may be as wide as in intact plants.
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malonyl ginsenoside content of a Cell Suspension Culture of panax japonicus var repens
Phytochemistry, 2013Co-Authors: D V Kochkin, Vadim V Kachala, Alexander S Shashkov, Alexander O Chizhov, Vasily Y Chirva, A M NosovAbstract:The presence of large amounts of ginsenosides malonyl-Rb1, -Rc, -Rb2, and -Rd in a Suspension Culture of Panax japonicus var. repens Cells was demonstrated for the first time. Identification of ginsenoside malonyl-Rb1 was based on chromatographic, chemical, and spectroscopic evidence. Ginsenosides malonyl-Rc, -Rb2, and -Rd were identified on the basis of chromatographic and chemical data. Content and composition of the individual ginsenosides (Rg1, R0, malonyl-Rb1, Rb1, Rc, Rb2, and Rd) were monitored in the Suspension Culture over 4 years. The RP-HPLC-UV analysis showed that Rg1, R0, and malonyl-Rb1 accounted for more than 75% of the total pool of ginsenosides. In accordance with this result, and data analysis reported in the literature, we propose that ginsenoside formation in the Cells of P. japonicus var. repens in vitro is closely related to the Cellular compartmentation of these substances. In particular, the accumulation of the 20(S)-protopanaxadiol ginsenosides (especially Rb1) is strongly dependent on their pattern of malonylation, which likely targets them for transport into the vacuole.
Yunji Shin - One of the best experts on this subject based on the ideXlab platform.
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co expression of proteinase inhibitor enhances recombinant human granulocyte macrophage colony stimulating factor production in transgenic rice Cell Suspension Culture
Protein Expression and Purification, 2008Co-Authors: Taegeum Kim, Taeho Kwon, Yongsuk Jang, Hojin Lee, Yunji Shin, Moonsik YangAbstract:Abstract The synthetic gene (sPI-II) harboring the chymotrypsin (C1) and trypsin (T1) inhibitor domains of the Nicotiana alata serine proteinase inhibitor II gene has been previously expressed, and extraCellular protease activity was shown to be reduced in the Suspension Culture medium. In this study, the sPI-II gene was introduced into transgenic rice Cells expressing rhGM-CSF (recombinant human granulocyte–macrophage colony-stimulating factor), in an effort to reduce protease activity and increase rhGM-CSF accumulation in the Suspension Culture medium. The integration and expression of the introduced sPI-II gene in the transgenic rice Cells were verified via genomic DNA PCR amplification and Northern blot analysis, respectively. Relative protease activity was found to have been reduced and rhGM-CSF production was increased 2-fold in the co-transformed Cell Suspension Culture with rhGM-CSF and the sPI-II gene, as compared with that observed in the transformed Cell Suspension Culture expressing rhGM-CSF only. These results indicate that a transformed plant Cell Suspension Culture system expressing the proteinase inhibitor can be a useful tool for increasing recombinant protein production.
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high level of expression of recombinant human granulocyte macrophage colony stimulating factor in transgenic rice Cell Suspension Culture
Biotechnology and Bioengineering, 2003Co-Authors: Yunji Shin, Shinyoung Hong, Taeho Kwon, Yongsuk Jang, Moonsik YangAbstract:Recombinant human granulocyte-macrophage colony stimulating factor (hGM-CSF) has been previously produced in tobacco Cell Suspension Cultures. However, the amount of hGM-CSF accumulated in the Culture medium dropped quickly from its maximum of 150 microg/L at 5 d after incubation. To overcome this problem, we sought an expression system in which heterologous gene expression could be induced at high levels. We selected a rice amylase expression system in which the promoter Ramy3D is induced to express recombinant protein by sucrose starvation. This induction system was found to give good yield of recombinant hGM-CSF in transgenic rice Cell Suspension Culture and protease activity of this Culture medium was low compared to that of tobacco Culture system.