2005 Fiscal Year Final Research Report Summary
Reverse genetics for rice in the Post-Genome
| Project/Area Number |
15380008
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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 |
Breeding science
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| Research Institution | National Institute for Basic Biology (2004-2005)
Okazaki National Research Institutes (2003) |
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Principal Investigator |
TERADA Rie National Institute for Basic Biology, Division of Molecular Genetics, Research Associate, 分子遺伝学研究部門, 助手 (30137799)
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| Co-Investigator(Kenkyū-buntansha) |
TSUGANE Kazuo National Institute for Basic Biology, Division of Molecular Genetics, Research Associate, 分子遺伝学研究部門, 助手 (50343744)
IIDA Shigeru National Institute for Basic Biology, Division of Molecular Genetics, Professor, 分子遺伝学研究部門, 教授 (30012777)
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| Project Period (FY) |
2003 – 2005
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| Keywords | Post-Genome / Waxy / rice / gene targeting / Alcohol dehydrogenase / Adh / Cre / loxP / Gene tagging |
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| Research Abstract |
Gene targeting (GT) and gene tagging are quite powerful tools for studying gene function, after the completion of the entire rice genomic sequence. We have effectively progressed both procedures in this project. (1)Gene_targeting : Towards the wider application of our rice GT procedure, which we firstly developed for endogenous gene modification, new GT experiments have been performed for the Alcohol dehydrogenase genes, c1 and Adh2, independently. We have succeeded both GT experiments and are obtaining detailed molecular evidence for the precise recombination. The results ensure the applicability of our GT procedure to a new rice gene. Through GT experiments, we have improved each step of our GT procedure including the vector construction and the targeting transformation consisting of a strong positive-negative selection depending on a large-scale Agrobacterium-mediated transformation. In addition, we have examined the processes of homologous recombination (HR) after the T-DNA transformation. It would be another challenge in this project that the targeted excision of a 30-kb segment between the Adh1 and Adh2 on chromosome 11. We have obtained the putative recombination plant lines. To establish a conditional GT using the Cre/loxP site-specific recombination system, we attempted to remove a positive marker flanked by two loxP sequences within the targeted waxy locus by introducing the Cre gene. Our preliminary PCR analysis indicated that the positive marker can be removed from the waxy locus, and further analysis is in progress. (2)Gene tagging : We have identified non-autonomous DNA-based active rice transposon one (nDart1), belonging to the hAT superfamily, and its transposition can be induced by crossing with a line containing an active autonomous element, aDart, and be stabilized by segregating out of aDart.
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