| Project/Area Number |
12440212
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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 |
遺伝
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| Research Institution | Natiomal Institute of Genetics |
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Principal Investigator |
IKEMURA Toshimichi Natiomal Inst.of Genetics, Dept.of Popul.Genet, Professor, 集団遺伝研究系, 教授 (50025475)
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| Co-Investigator(Kenkyū-buntansha) |
FUKAGAWA Tatsuo Natiomal Inst.of Genetics,Dept.of Popul.Genet,Assis.Prof, 集団遺伝研究系, 助手 (60321600)
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| Project Period (FY) |
2000 – 2001
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| Project Status |
Completed (Fiscal Year 2001)
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| Budget Amount *help |
¥11,500,000 (Direct Cost: ¥11,500,000)
Fiscal Year 2001: ¥4,500,000 (Direct Cost: ¥4,500,000)
Fiscal Year 2000: ¥7,000,000 (Direct Cost: ¥7,000,000)
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| Keywords | Triplex / Polypurine / CENP / FISH / Nuclear organization / chromosome band boundary / Replication timing / DNA複製タイミング / ポリピリミジン |
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| Research Abstract |
The single-strand formation and transmolecular triplex formation are thought to enable sequences spaced distantly along the genome to associate with each other and organize nuclear DNA into ordered configurations. Triplex-forming DNAs in the human interphase nucleus were analyzed by combining immunodetection by antitriplex antibodies and fluorescence in situ "nondenaturing " hybridization employing polypurine/polypyrimidine (PuPy) -tract probes. The "nondenaturing" hybridization technique, which has been used to detect RNA, could detect single-stranded DNAs in nondenatured nuclei. Triplexes visualized differentially with distinct PuPy-tract probes were associated spatially with centromere sequences in the interphase nucleus in a sequence-specific manner. Therefore, we were interested in centromore formation and function. To determine the precise functions of centromere proteins, we examined their roles by generating a conditional loss-of-function mutant in the chicken DT40 cell line. We concluded that both CENP-I and CENP-H are necessary for localization of CENP-C but not CENP-A to the centromere. We also performed living cell analysis of cells expressing GFP-or YFP- tagged centromere proteins to observe the dynamics of chromosomes.
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