| Project/Area Number |
16380192
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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 |
Applied animal science
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| Research Institution | Tokyo University of Agriculture |
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Principal Investigator |
KONO Tomohiro Tokyo University of Agriculture, Department of Bioscience, Professor, 応用生物科学部, 教授 (80153485)
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| Co-Investigator(Kenkyū-buntansha) |
IMAI Hiroshi Kyoto University, Graduate School of Agriculture, Professor, 大学院・農学研究科, 教授 (10303869)
SOTOMARU Yusuke Hiroshima University, Natural Science Center for Basic Research and Development, Assistant professor, 自然科学研究支援開発センター, 助教授 (90309352)
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| Project Period (FY) |
2004 – 2005
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| Project Status |
Completed (Fiscal Year 2005)
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| Budget Amount *help |
¥15,700,000 (Direct Cost: ¥15,700,000)
Fiscal Year 2005: ¥6,800,000 (Direct Cost: ¥6,800,000)
Fiscal Year 2004: ¥8,900,000 (Direct Cost: ¥8,900,000)
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| Keywords | Nuclear transfer / embryo cloning / gene expression analysis / reprogramming / epigenetic regulation / abnormal development / クローン / 遺伝子発現 |
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| Research Abstract |
Somatic cloning does not always result in ontogeny in mammals, and development is often associated with various abnormalities and embryo loss with a high frequency. This is considered to be due to aberrant gene expression resulting from epigenetic reprogramming errors. In the present study we conducted to explore the genes that are involved in developmental defects of somatic cloned mouse embryos. In order to address this issue, we carried out global screening of candidate genes by subtraction analysis using ES-cloned blastocysts, and differential display analysis using 2-cell embryos. From subtraction analysis 218 genes were detected as differentially expressed genes at blastocysts. Of these 198 clones were successfully sequenced and by BLAST search analysis 158 genes are detected as known genes. To confirm differentially expression, we carried out quantitative gene expression analysis by real-time PCR. The results showed that 5 of 10 genes were actually highly expressed in ES-cloned blastocysts. Further, we compared gene expression patterns using differential display RT-PCR (DDRT-PCR) between the NT and IVF embryos at the 2-cell stage to detect some abnormalities affecting later development of NT embryos. Aberrant gene expression was detected in NT embryos compared with IVF embryos, and MuERV-L and Dnaja2 genes were down-regulated and Inpp5b and Chst12 genes were up-regulated in the NT embryos. Further analysis showed that the expression of zygotically activated genes such as Interferon-?, Dub-1, Spz1, DD2106 and DD2111 were not properly activated in NT embryos, suggesting that the cellular process involved in the control of the zygotic genome activation is not appropriately regulated. These results indicate that abnormal gene expression has already occurred at the early stage of preimplantation development as a failure of nuclear reprogramming.
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