| Project/Area Number |
11236208
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| Research Category |
Grant-in-Aid for Scientific Research on Priority Areas
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| Allocation Type | Single-year Grants |
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| Review Section |
Biological Sciences
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| Research Institution | KYOTO UNIVERSITY |
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Principal Investigator |
KINOSHITA Masato Kyoto University, Graduate School of Agriculture, Research Associate, 大学院・農学研究科, 助手 (60263125)
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| Project Period (FY) |
1999 – 2003
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| Project Status |
Completed (Fiscal Year 2003)
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| Budget Amount *help |
¥44,200,000 (Direct Cost: ¥44,200,000)
Fiscal Year 2003: ¥9,400,000 (Direct Cost: ¥9,400,000)
Fiscal Year 2002: ¥9,400,000 (Direct Cost: ¥9,400,000)
Fiscal Year 2001: ¥9,100,000 (Direct Cost: ¥9,100,000)
Fiscal Year 2000: ¥7,200,000 (Direct Cost: ¥7,200,000)
Fiscal Year 1999: ¥9,100,000 (Direct Cost: ¥9,100,000)
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| Keywords | MEDAKA / TRANSGENIC / CRE / loxp / INSULATOR / GREEN FLUORESCENT PROTEIN / RED FLUORESCENT PROTEIN / Cre / LoxP / GFP / 発現制御 / vasa / トランスジェニック / RFP / トランス ジェニック / パーティクルデリバリー / Vasa |
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| Research Abstract |
In this study, to elevate the usefulness of medaka as a model, I tried to adapt and improve the transgenic methods in medaka.
1)Gene induction with particle gun system: Microinjection method is a most popular one for the introduction foreign gene into fish eggs. But this method requires special skill. To develop more easy method, I adapted particle gun method in medaka transgenesis and optimized parameters. The key of this method was that bombardment should be carried out before chorion hardened. Transgenic medaka established this method revealed that transgene was transmitted stablely over 5th generation. 2)Effective selection marker of germ line transmitters : One of the most complicated works in establishing transgenic strains is the selection of the individual which harboring transgene in germ cells. We found that vasa-GFP vector was a good indicator of germ line transmitter, which told us induction of transgene in germ cell before hatch. We also found that germ cell specific cis element locates in 3'-UTR of vasa transcript. 3) Regulation of transgene expression : To prevent transgene expression from the effect of enhancer existing near inserted chromosomal position, I showed the effectiveness of sea-urchin insulator in medaka transgenesis. For the conditional gene expression, Cre-LoxP system was available in medaka. Additionally, about 200 bp sequence including loxp sequence showed internal ribosomal entry activity. 4)Transgenic medaka strain with fluorescent label: Some transgenic medaka strains with tissue specific fluorescent label were established. They had green or red fluorescent in skeletal muscle, green fluorescent in germ cells, green fluorescence in lever, green or red fluorescence in almost all tissues and green fluorescence in nerve cells
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