| Project/Area Number |
10470464
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Chemical pharmacy
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| Research Institution | The University of Tokyo |
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Principal Investigator |
EBIZUKA Yutaka The University of Tokyo, Graduate School of Pharmaceutical Sciences, Professor, 大学院・薬学系研究科, 教授 (90107384)
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| Co-Investigator(Kenkyū-buntansha) |
ORIHARA Yutaka The University of Tokyo, Graduate School of Pharmaceutical Sciences, Associate Professor, 大学院・薬学系研究科, 助教授 (30137905)
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| Project Period (FY) |
1998 – 1999
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| Project Status |
Completed (Fiscal Year 1999)
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| Budget Amount *help |
¥9,600,000 (Direct Cost: ¥9,600,000)
Fiscal Year 1999: ¥3,000,000 (Direct Cost: ¥3,000,000)
Fiscal Year 1998: ¥6,600,000 (Direct Cost: ¥6,600,000)
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| Keywords | Triterpene / Saponin / Triterpene Synthase / Oxidosqualene / cDNA / PCR / Transgenic Plant / Medicinal Plant / オタネニンジン / カンゾウ |
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| Research Abstract |
(1) Molecular cloning of OSC cDNAs and identification of their enzyme functions. By employing homology based PCRs, more than twenty oxidosqualene cyclase (OSC) cDNAs with different product specificities have been cloned from various higher plants. They show more than 60% sequence identity even though they are derived from Plants belonging to different Plant families and giving rise to different cyclization products. Identification of their enzyme functions as synthases of cycloartenol, β-amyrin lupeol, cucurbitadienol were established by product analysis of transformants of Saccharomyces cerevisiae mutant, which lacks lanosterol synthase and accumulates oxidosqualene in the cells carrying expression plasmid with respective cDNA. Isolation of β-amyrin synthase clone, giving rise to most popular oleanane type triterpenes, would be efficiently applied for transformation of medicinally important saponin producing plants. (2) Transgenic Arabidopsis thaliana with antisense LUP1 lupeol synthase DNA and its chemotype analysis. Antisense LUP1 lupeol synthase DNA was introduced into A. thaliana via pBIX vector system driven by 35S promoter by means of in planta infiltration method. Transgenic plants were selected on plates and further grown in artificial soil. T1 progeny showed marked dwarfism. Triterpene contents in T2 progeny were analyzed by means of LC-MS method revealing the complete absence of lupeol in the transgenic plants. These results, combined with those described in (1), transgenic technology with these triterpene synthase cDNAs would lead to the development of overproducers of pharmacologically active saponins.
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