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1994 Fiscal Year Final Research Report Summary

Precise Manipulation of Developemental transgenic cells using a microwichirned Technology

Research Project

Project/Area Number 05455005
Research Category

Grant-in-Aid for General Scientific Research (B)

Allocation TypeSingle-year Grants
Research Field 広領域
Research InstitutionJapan Advanced Institute of Science & Technology (JAIST)

Principal Investigator

TAMIYA Eiichi  JAIST,Materials Science Professor, 材料科学研究所, 教授 (60179893)

Co-Investigator(Kenkyū-buntansha) YOKOYAMA Kenji  JAIST,Materials Science Assoc. Prof., 材料科学研究所, 助教授 (80242121)
Project Period (FY) 1993 – 1994
KeywordsMicromachinning technique / Embryo Engineering / Medaka / Gene transfer / Electroporation / Transgluic fish / トランスジェニック生物
Research Abstract

In this study, a novel system for gene transfer has been fabricated using micro-machining techniques. Micromachining techniques have been developed for fabricating semiconductor devices. Microelectrodes and can be fabricated onto the glass plate using lithography and vapor deposition techniques.
We have used Medaka (Oryzias latipes) as an experimental fish, because it is a suitable material for embryo engineering research based on the following reasons. Compared with mice, fish fertilizes eggs extemally, and there is no need for implant eggs into the uterus of pseudo pregnant animal. Since fish has a transparent egg membrane, both developement and differentiation process are easy to observe.
A high electric field induces a hole of lipid bilayr of the cell membranes and results in destroying the egg yolk and a limited yield. Therefore, the electric field should be localized to a certain part of cytoplasm of the cell to increase the yield. Micro-electrodes were fabricated on glass plate with micro-machining techniques to introduce gene to them. The material of the bottom of the electrode is Cr, and the upper part is Au. A polymer film with a hole (1.2mm in the diameter) is attached with an adhesive.
Microelectrodes was used for application of localized high electric field at the animal pole of the fertilized egg. High electric field pulses (750 V/cm-1000V/cm) of duration fifty microseconds was applied at a repetition rate of 1Hz for five seconds. Microscopic observation showed no damage of eggs using our system.
Forty-one percent of the eggs show luminescence (19 samples in 46 samples). The present system gave a higher ratio of the expression of luciferase than a conventional electroporation method did.
The dependence on the cleavage stages about electroporation was measured.
Gene transfer was observed in the range from 1-cell stage to 64-cell stage embryo.

  • Research Products

    (11 results)

All Other

All Publications (11 results)

  • [Publications] Y.Murakami: "Micromachined electroporation system for transglnic fish" J.Biotechnology. 34. 35-42 (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 民谷栄一: "微小薄膜電極を用いる生物分子・細胞の操作" 化学. 49. 746-747 (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 民谷栄一: "マイクロバイオセンサーの開発動向と展望" 計測と制御. 134. 36-41 (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Y.Murakami: "Micromachied Electrochemical Flow Ceel for Bioseusing" Electroavalysis. 6. 735-739 (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] E.Tamiya: "Direct Monitoring of Glutamate Deleased from-" Sensors & Materials. 7(in press). (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 民谷栄一: "バイオセンサーによる細胞情報の解析" 農林水産技術「研究ジャーナル」. 18. 29-33 (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 民谷栄一: "機械分析ガイドブック" 丸善(印刷中), (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] E.Tamiya: "Olfaction and TasteXI" Springer-velag, (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Y.Murakami et al.: "Micromachined electroporation system for transgluic fish." J.Biotechnology. 34. 35-42 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Y.Murakami et al.: "Micromachined Electrochemical Flow Cell for Biosensing." Electroanalysis. 6. 735-739 (1994)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] E.Tamiya et al.: "Direct Monitoring of Glutamate Released from Cerekellum neurous" Sensors & Materials. 7(in press). (1995)

    • Description
      「研究成果報告書概要(欧文)」より

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Published: 1996-04-15  

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