2001 Fiscal Year Final Research Report Summary
Functional analysis of a role of prolylendopeptidase in brain by constructing gene targeting mice
| Project/Area Number |
12672105
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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 |
Biological pharmacy
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| Research Institution | The University of Tokyo |
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Principal Investigator |
AKIMITSU Nobuyoshi The University of Tokyo, Graduate School of Pharmaceltical Sciences, Developmental Biochemistry, Assistant professor, 大学院・薬学系研究科, 助手 (40294962)
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| Project Period (FY) |
2000 – 2001
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| Keywords | Prolylendopeptidase / knock-out mouse / ES cells |
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| Research Abstract |
Mouse prolylendopeptidase consists of the peptidase domain (N-terminal and C-terminal) and the propeller domain (intermediate region). The gene encoding this protein consists of 15 exons. Among them, the exon 6 encodes the propeller domain. We tried to construct mutant mice lacking the exon 6 of the prolylendopeptidase gene. First, we cloned the gene from the genomic library of 129 strain mice. We next constructed a targeting vector whose exon 6 was disrupted by the insertion of the neomycine-resistant gene. The targeting vector was introduced into mice ES cells by electroporation, followed by disruption of the endopeptidase gene by homologous recombination. The mutant ES cells were used for construction of chimera mice. We are now testing the germ line transmission of the mutant ES cells by analysis of Fl mice. We also tested whether ES cells lacking both alleles of the prolylendopeptidase gene. The results showed that we could construct a null mutant which expressed no prolylendopeptidase activity. The growth of the mutant was indistinguishable from that of wild type ES cells, indicating that the protein is dispensable for proliferation of ES cells.
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