title: Production of mouse granulocyte-macrophage colony-stimulating factor by gateway technology and transgenic rice cell culture.
abstract: To establish a production platform for recombinant proteins in rice suspension cells, we first constructed a Gateway-compatible binary T-DNA destination vector. It provided a reliable and effective method for the rapid directional cloning of target genes into plant cells through Agrobacterium-mediated transformation. We used the approach to produce mouse granulocyte-macrophage colony-stimulating factor (mGM-CSF) in a rice suspension cell system. The promoter for the αAmy3 amylase gene, which is induced strongly by sugar depletion, drove the expression of mGM-CSF. The resulting recombinant protein was fused with the αAmy3 signal peptide and was secreted into the culture medium. The production of rice-derived mGM-CSF (rmGM-CSF) was scaled up successfully in a 2-L bioreactor, in which the highest yield of rmGM-CSF was 24.6 mg/L. Due to post-translational glycosylation, the molecular weight of rmGM-CSF was larger than that of recombinant mGM-CSF produced in Escherichia coli. The rmGM-CSF was bioactive and could stimulate the proliferation of a murine myeloblastic leukemia cell line, NSF-60.
authors: Graduate Institute of Biochemical and Biomedical Engineering, Chang Gung University, Kwei-Shan, Taoyuan County, Taiwan, ROC. ykliu@mail.cgu.edu.tw
authors: Yu-Kuo
authors: YK
authors: Liu
authors: Li-Fen
authors: LF
authors: Huang
authors: Shin-Lon
authors: SL
authors: Ho
authors: Chun-Yu
authors: CY
authors: Liao
authors: Hsin-Yi
authors: HY
authors: Liu
authors: Ying-Hui
authors: YH
authors: Lai
authors: Su-May
authors: SM
authors: Yu
authors: Chung-An
authors: CA
authors: Lu
mesh_terms: D060054:Agrobacterium / Q000235:genetics; D000818:Animals; D019149:Bioreactors; D001709:Biotechnology / Q000379:methods*; D018929:Cell Culture Techniques; D003470:Culture Media / Q000737:chemistry; D004269:DNA, Bacterial; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008970:Molecular Weight; D012275:Oryza / Q000235:genetics / Q000378:metabolism*; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D003470:Culture Media; D004269:DNA, Bacterial; D011994:Recombinant Proteins; C036483:T-DNA; D016178:Granulocyte-Macrophage Colony-Stimulating Factor