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Development of transfection technique of foreign gene using bovine ES-like cells as a vector

Research Project

Project/Area Number 09660305
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeSingle-year Grants
Section一般
Research Field Applied animal science
Research InstitutionTokyo University of Agriculture

Principal Investigator

IWASAKI Setsuo  Faculty of Applied Bioscience, Tokyo Univ. of Agric., Professor, 応用生物科学部, 教授 (50184867)

Co-Investigator(Kenkyū-buntansha) TAKADA Tatsuyuki  Faculty of Medicine, Shiga Univ. of Medical Science, Associate Professor, 医学部, 助教授
NAGASHIMA Takayuki  Faculty of Agriculture, Tokyo Univ. of Agric., Assistant Professor, 農学部, 講師 (20231483)
高田 達行  国立小児病院, 小児医療センター, 研究員
Project Period (FY) 1997 – 1998
Project Status Completed (Fiscal Year 1998)
Budget Amount *help
¥3,300,000 (Direct Cost: ¥3,300,000)
Fiscal Year 1998: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 1997: ¥1,800,000 (Direct Cost: ¥1,800,000)
KeywordsEmbryonic stem cell / Aggregation chimera / Gene transfer / Electroporation / Vector
Research Abstract

The present was carried out to develop a new system for foreign gene transfer into bovine embryos using bovine embryonic stem (ES) cells or somatic cells as a vector. 1. In order to construct a reporter gene, PGK promoter region was inserted into a cloning site of green fluorescent protein (GFP) gene which show a self fluorescent. 2. Electroporation or lipofection methods were used to transfect GDP gene into bovine somatic cells as mammary cells, ear cells or fibroblast. It was shown that electroporation was effective for mammary cells and lipofection method was effective for bovine adult ear cells and fetal fibroblasts in transfection of GDP gene. 3. Six calves were born from the transfer of the chimeric embryos, two calves died at birth or a day after birth. The microsatellite DNA analysis showed that the cells of liver, roots of hair from calves and placenta proved to be chimeric and derived from ES-like cells and tetraploid embryos. 4. We attempted aggregation of diploid or tetraploid embryos with GFP positive ES cells which were selected for only 8 days with G418. At 15 days, 7 normal embryos were obtained, 1 out of 7 embryos showed GFP expression on main five organs, and 4 embryos were detected strongly GFP expression on more one organ under a stereotypic microscope. These results indicate that transfected ES cells or somatic cells selected in a short term with G418 were able to use enough for the selection of preimplantation transgenic embryos and for aggregation chimera.

Report

(3 results)
  • 1998 Annual Research Report   Final Research Report Summary
  • 1997 Annual Research Report
  • Research Products

    (3 results)

All Other

All Publications (3 results)

  • [Publications] S.IWASAKI,Y.ITO,S.IWASAKI: "In-vitro development of aggregates of bovine inner cell moss cells or bovine mammary cells and putative tetraploid embryos produced by electrofusion"Journal of Reproduction and Development. 45. 65-71 (1999)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] IWASAKI, Y. ITO, S. IWASAKI: "In-vitro development of aggregates of bovine inner cell amass cells or bovine mammary cells and putative tetraploid embryos produced by electrofusion"Journal of Reproduction and Development. 45(1). 65-71 (1999)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] S. IWASAKI, Y ITO, S. IWASAKI: "In-vitro development of aggregates of bovine, inner cell mass cells or bovine mammary cells and putative tetraploid embryos produced by electrofusion." Jourual of Reproduction and Development. 45・1印刷中. (1999)

    • Related Report
      1998 Annual Research Report

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Published: 1997-04-01   Modified: 2016-04-21  

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