2003 Fiscal Year Final Research Report Summary
DEVELOPMENT OF INTRODUCTION OF TRANSGENE INTO MEDAKA EGGS AND MANUPILATION OF GENE EXPRESION
| 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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| Keywords | MEDAKA / TRANSGENIC / CRE / loxp / INSULATOR / GREEN FLUORESCENT PROTEIN / RED FLUORESCENT PROTEIN |
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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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Research Products
(13 results)
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[Publications] Kinoshita, M.Yamauchi, M., Sasanuma, M., Ishikawa, Y., Osada, T., Inokue, K., Wakamatus, Y., Ozato, K.: "A transgene and its expressiion profile are stably transmitted to offspring in transgenic medaka generated by the particle gun method."Zool. Sci.. 20. 869-875 (2002)
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「研究成果報告書概要(欧文)」より
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[Publications] Ju, B., Pristyazhnyuk, I., Ladygina, T., Kinoshita., Ozato, K., Wakamatsu, Y.: "Development and gene expression of nuclear transplants generated by transplantation of cultured cell nuclei into nonenucleated eggs in medaka, Oryzias latipes."Dev. Growth Dif.. 45. 167-174 (2003)
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「研究成果報告書概要(欧文)」より
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[Publications] Wakamatsu, Y., Ju, B., Pristyaznhyuk, I., Niwa, K., Ladygina, T., Kinoshita, M., Araki, K., Ozato, K.: "Fertile and diploid nuclear transplants derived from embryonic cells of a small laboratory fish, medaka (Oryzias latipes)."Proc Natl Acad Sci USA. 98. 1071-1076 (2001)
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「研究成果報告書概要(欧文)」より
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[Publications] Yamauchi, M., Kinoshita, M., Sasanuma, M., Tsuji, S., Terada, M., Morimyo, M., Ishikawa, Y.: "Introduction of a foreign gene into medakafish using the particle gun method."J. Exp. Zool.. 287. 285-293 (2000)
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「研究成果報告書概要(欧文)」より
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[Publications] Kinoshita, M., Tanaka, M.: "Transgenic medaka as a model for fish biology and aquaculture"Aquatic Genomics-Steps Toward a Great Future edited by Shimizu, N., Aoki, T., Hirono, I., Takashima, F.. 320-328 (2002)
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「研究成果報告書概要(欧文)」より
