| Project/Area Number |
10680776
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Laboratory animal science
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| Research Institution | KYOTO UNIVERSITY |
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Principal Investigator |
KITADA Kazuhiro Kyoto University, Graduate School of Medicine., Assistant, 医学研究科, 助手 (70263093)
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| Co-Investigator(Kenkyū-buntansha) |
KANEDA Yasufumi Osaka University, Graduate School of Medicine., Professor, 医学研究科, 教授 (10177537)
SERIKAWA Tadao Kyoto University, Graduate School of Medicine., Professor, 医学研究科, 教授 (30025655)
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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 |
¥3,300,000 (Direct Cost: ¥3,300,000)
Fiscal Year 1999: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 1998: ¥1,800,000 (Direct Cost: ¥1,800,000)
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| Keywords | HVJ liposome / transfection into live animals / animal models for human diseases / gene therapy / mutamt / positinal cloning / ALY mouse / tremor rat / トランスジェニック動物 / LacZ |
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| Research Abstract |
1. Establishment of transfection methods into live animals by HVJ liposome
We constructed two expression vectors, pEBActNII-LacZ and pEBActNII-GFP, and generated HVJ liposome from each of them. 50 microL of HVJ liposomes were injected into cerebral cortex of neonates(1 or 3 days of age), and the animals were examined by LacZ staining or under fluorescence microscope. As a result, we detected expression of these genes in the applied portions of the central nervous system.
2. Trial of the transgenesis by injection of HVJ liposome into testes
We injected HVJ liposome containing pEBActNII-LacZ into testes, and obtained offsprings from the manipulated animals for examining the rate of transgenesis. However, no germ-line transmission was observed in 84 animals in our research. This finding suggested that the rate of transgenesis by injection of HVJ liposome into testes is too low to generate transgenic animals efficiently.
3. Positional cloning of the causative genes in animal models for human diseases
The tremor rat is a mutant that exhibits absence-like seizure and spongiform degeneration in the central nervous system (CNS). By positional cloning, a genomic deletion was found within the critical region , in which the aspartoacylase gene is located. Accordingly, no aspartoacylase expression was detected in any of the tissues examined, and abnormal accumulation of N-acetyl-L-aspartate (NAA) was shown in the mutant brain, in correlation with the severity of the vacuole formation. Therefore, the tremor rat may be regarded as a suitable animal model of human Canavan disease, characterized by spongy leukodystrophy that is caused by aspartoacylase deficiency.
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