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1992 Fiscal Year Final Research Report Summary

Introduction of the foreign genetic information into tissue cells of living animals and its application to the medical science.

Research Project

Project/Area Number 01440087
Research Category

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

Allocation TypeSingle-year Grants
Research Field 医学一般
Research InstitutionOsaka University

Principal Investigator

NAKANISHI Mahito  Osaka University,Institute for Molecular and Cellular Biology,Research Associate., 細胞生体工学センター, 助手 (10172355)

Co-Investigator(Kenkyū-buntansha) KANEDA Yasuhumi  Osaka University,Institute for Molecular and Cellular Biology,Assistant Professo, 細胞生体工学センター, 助教授 (10177537)
YONEDA Yoshihiro  Osaka University,Medical School,Professor, 医学部, 教授 (80191667)
KOHNO Kenji  Osaka University,Institute for Molecular and Cellular Biology,Assistant Professo, 細胞生体工学センター, 助教授 (50142005)
Project Period (FY) 1989 – 1992
KeywordsGene Introduction / Liposome / Sendai virus / Virus Vector / Artificial Chromosome / Gene therapy / Membrane Fusion
Research Abstract

We obtained the following results during the term of this project. 1.Gene Introduction into Animal Tissues Using Multilamella Fusogenic Liposomes.
We prepared multilamella fusogenic liposomes encapsulated DNA and protein by incubating liposomes with Sendai virus and purifying them on sucrose gradient. These liposomes could introduce and express E.coli beta-galactosidase gene and the surface antigen gene of human hepatitis B virus into liver cells of living rats. 2.Purification and Characterization of Singlelamella Fusogenic Liposomes.
We prepared singlelamella fusogenic liposomes encapsulating DNA and protein by preparing liposomes by reverse phase method, incubating them with Sendai virus and purifying them on sucrose gradient. We found that these liposomes were essentially free from unfusogenic liposomes and virus and that they had higher efficiency of introduction of macromolecules into cells(10^<10> particles for protein and 10^7 particles for DNA per 1 OD_<540> unit). The efficiency of these purified liposomes to introduce their contents into cytoplasm was the same as that of intact Sendai virus. We also found that these liposomes had a diameter of 250nm. 3.Analysis of Mechanism of Persistent Infection of Mutant Sendai Virus.
We demonstrated the ts mutant, Cl151, Caused persistent infection because their mutant M protein was unstable at non-permissive temperature. We also showed that M protein did not affect the gene expression of the virus. This is a first step for the development of the virus vector based on Sendai virus. 4.Development of the Vectors for Human Artificial Chromosome.
We succeeded to make the vectors construct the human artificial chromosome. The vectors(pMYAC1,pMYAC2) consisted of human telomere, yeast DNA for maintaining as yeast artificial chromosome, drug resistant genes as selection markers in human cells. This is the first step for construction of human artificial chromosome, an ultimate system for human gene therapy.

  • Research Products

    (21 results)

All Other

All Publications (21 results)

  • [Publications] Kato,K.et al.: "Expression of Hepatitis B Virus Surface Antigen in Adult Rat Liver" The Jounal of Biological Chemistry. 266. 3361-3364 (1991)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Kato,K.et al.: "Direct injection of Hepatitis B virus DNA into liver induced hepatitis in adult rats" The Jounal of Biological Chemistry. 266. 22071-22074 (1991)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Kido,M.et al.: "Escherichia coli RecA protein modified with a nuclear location signal binds to chromosomes in living mammalian cells" Experimental Cell Research. 198. 107-114 (1992)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Imamoto-Sonobe,N.et al.: "A protein recognized by antibodies to Asp-Asp-Asp-Glu-Asp shows specific binding activity to heterogenous nuclear transport signals" The Jounal of Cell Biology. 265. 16504-16508 (1990)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Yoneda,Y.et al.: "A long synthetic peptide containing a nuclear localization signal and its flanking sequences of SV40 T-antigen directs the transport of IgM into the nucleus efficiently" Experimental Cell Research. 201. 313-320 (1992)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Imamoto,N.et al.: "Antibodies against 70-kD heat shock cognate protein inhibit mediated nuclear import of karyophilic proteins" The Jounal of Biological Chemistry. 119. 1047-1061 (1992)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 中西 真人: "細胞内物質導入法、リポソーム、「細胞工学(矢原一郎編)」" 丸善, 23

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 加藤 啓子・中西 真人: "細胞内への物質導入法、「ライフサイエンスにおけるリポソーム(寺田弘・吉村哲郎編)」" シュプリンガー・フェアラーク東京, 10

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Kato,K.et al: "Expression of Hepatitis B Virus Surface Antigen in Adult Rat Liver; Co-introduction of DNA and Nuclear Protein by a Simplified Liposome Method" The Journal of Biological Chemistry. 266. 3361-3364 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kato,K.et al: "Direct injection of Hepatitis B virus DNA into liver induced hepatitis in adult rats" The Journal of Biological Chemistry. 266. 22071-22074 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kido,M.et al: "Escherichia coli RecA protein modified with a nuclear location signal binds to chromosomes in living mammalian cells" Experimental Cell Research. 198. 107-114 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Imamoto-Sonobe,N.et al: "A protein recognized by antibodies to Asp-Asp-Asp-Glu-Asp shows specific binding activity to heterogeneous nuclear transport signals" The Journal of Biological Chemistry. 265. 16504-16508 (1990)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Yoneda,Y.et al: "A long synthetic peptide containing a nuclear localization signal and its flanking sequences of SV40 T-antigen directs the transport of IgM into the nucleus efficiently." Experimental Cell Research. 201. 313-320 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Imamoto,N.et al: "Antibodies against 70-kD heat shock cognate protein inhibit mediated nuclear import of karyophilic proteins." The Journal of Cell Biology. 119. 1047-1061 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Toda,K.et al: "Structure and expression of of elongation factor 2 gene during development of Dictyosterium discoideum" The Journal of Biological Chemistry. 264. 15489-15493 (1989)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Shimamoto,T.et al: "Molecular cloning of human XPAC gene homologes from chicken,Xenopus leavis and Drosophila melanogaster." Biochem.Biophys.Res.Commun. 181. 1231-1237 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kido,M.et al: "Targeted introduction of a diphtheria toxin-resistant point mutation into the chromosomal EF-2 locus by in vivo homologous recombination." Cell Struct.Funct.16. 447-453 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Tokunaga,M.et al: "Purification and characterization of BiP/Kar2 protein from Saccharomyces cerevisiae." The Journal of Biological Chemistry. 267. 17553-17559 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kohno,K.et al: "The promotor region of the yeast KAR2(BiP)gene contains a regulatory domain that responds to the presence of unfolded proteins in the endoplasmic reticulum." Mol.Cell Biol. 13. 877-890 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kimata,Y.et al: "Expression of non-ADP-ribosilatable diphtheria toxin resistant elongation factor 2 in Saccharomyces cerevisiae." Biochem.Biophys.Res.Comm.,in press.

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Nakanishi,M.et al: CRC press,UK. Liposome-mediated introduction of macromolecules into living animal cells with the aid of HVJ(Sendai virus).In Liposome Technology,2nd ed.(ed.G.Gregoriadis), 1993

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

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Published: 1994-03-24  

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