| Project/Area Number |
59870010
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| Research Category |
Grant-in-Aid for Developmental Scientific Research
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| Allocation Type | Single-year Grants |
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| Research Field |
General medical chemistry
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| Research Institution | Institute of Medical Science, University of Tokyo |
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Principal Investigator |
NAGATA Shigekazu Research Associate, Institute of Medical Science, University of Tokyo, 医科学研究所, 助手 (70114428)
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| Co-Investigator(Kenkyū-buntansha) |
SOKAWA Yoshihiro Assistant Proffesor, Institute of Virus Research, University of Kyoto, ウィルス研究所, 助教授 (30012727)
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| Project Period (FY) |
1984 – 1986
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| Project Status |
Completed (Fiscal Year 1986)
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| Budget Amount *help |
¥8,400,000 (Direct Cost: ¥8,400,000)
Fiscal Year 1986: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 1985: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 1984: ¥5,600,000 (Direct Cost: ¥5,600,000)
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| Keywords | Recombinant DNA technology / Humoral factors / glycoproteins / Interferons / Granulocyte colony-stimulating factor / Bovine papilloma virus / トランスフォーメーション / ウシパヒローマウィルス / マウスC127細胞 / ヒトインターフェロンα,γ / モノクローナル抗体 |
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| Research Abstract |
In this project, we have developed the mammalian cell system to produce human interferon ( IFN ) and granulocyte colony-stimulating factor ( G-CSF ) using the recombinant DNA technology.
The cDNAs for human IFNs and G-CSF have been cloned from the Con A- stimulated human spleen cells or human squamous carcinoma cells ( CHU-2 ) producing G-CSF, respectively. By cloning of two different G-CSF cDNAs, it was revealed that two different G-CSF mRNAs are produced by the alternative splicing from the single precursor mRNA. The cDNAs for human IFNs or G-CSF were placed under the promoter of SV40 early promoter and introduced into mouse C127 I cells using bovine papilloma virus as a vector. The mouse C127 I cells were transformed by bovine papilloma virus-IFN ( G-CSF ) hybrid plasmid, and the transformed cells could secrete human IFNs or G-CSF, constitutively. Some of the transformed cells could produce IFNs or G-CSF very efficiently at the rate of 1.0 mg / l for IFNs or 10-20 mg /l for G-CSF. Human IFN- <gamma> and G-CSF have been purified to homogeneity from the medium conditioned with the transformed cells. IFN- <gamma> and G-CSF produced by mouse cells were glycosylated, and the NH-terminal amino acid sequence of those proteins were identical to that of the native proteins, indicating both proteins were processed correctly. When the recombinant human G-CSF was subcutasneously administrated into mice, a remarkable stimulation of granulopoiesis and splenomegaly was observed.
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