| Project/Area Number |
11670211
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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 |
Experimental pathology
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| Research Institution | Osaka University |
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Principal Investigator |
KIMURA Tohru Research Institute for Micobial Diseases, Osaka University, Research Associate, 微生物病研究所, 助手 (50280962)
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| Co-Investigator(Kenkyū-buntansha) |
NAKANO Toru Research Institute for Micobial Diseases, Osaka University, Professor, 微生物病研究所, 教授 (00172370)
伊川 正人 大阪大学, 遺伝情報実験施設, 助手 (20304066)
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| Project Period (FY) |
1999 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥4,000,000 (Direct Cost: ¥4,000,000)
Fiscal Year 2000: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 1999: ¥2,700,000 (Direct Cost: ¥2,700,000)
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| Keywords | Germ lineage / primordial germ cells / TNAP gene / Oct-3 / 4 gene / transgenic mice / EGFP / Cre / loxP system / loxP系 / 生殖質 / germ cell-less(gcl)遺伝子 |
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| Research Abstract |
Primordial germ cells (PGC) are founder cells of all gametes. Because of the absence of efficient experimental system, it remains largely unknown what kind of molecular mechanisms are operating during the commitment and maintenance of mouse germ lineage. In order to establish the experimental system to investigate the PGC development, we generated the following transgenic mice using the promoters of TNAP (tissue nonspecific alkaline phosphatase) and Oct-3/4 genes, the marker genes for PGC.
(1) We established the mice expressing fluorescent protein EGFP in PGC by knocking-in EGFP cDNA into TNAP locus. In vivo visualization of PGC was successfully achieved in this mouse. Furthermore, live PGC can be isolated at 100% purity using cell sorter. These convenient visualization and purification techniques should be useful for analyzing PGC development at cellular and molecular levels.
(2) We tried to generate a model mouse for PGC-specific gene expression system. For this purpose, we produced the transgenic mouse containing the transgene in which floxed-EGFP cDNA followed by _-galactosidase cDNA is under the control of Oct-3/4 gene promoter. As expected, EGFP but not _-galactosidase was expressed in PGC of the transgenic mice. On the other hand, it had been expected that expression of _-galactosidase would have turned on in PGC when this mouse was crossed with transgenic mouse expressing Cre recombinase in PGC.However, _-galactosidase activity was not detected in PGC.The failure to detect the transgene expression is probably due to the excesives loss of transgenes that resulted from the recombination between loxP sites of tandemly inserted transgenes.
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