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1996 Fiscal Year Final Research Report Summary

THE STUDY OF HOMOLOGOUS RECOMBINATION AND GENE REPAIR IN MOUSE SOMATIC CELLS.

Research Project

Project/Area Number 07640823
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeSingle-year Grants
Section一般
Research Field 遺伝
Research InstitutionTOKAI UNIVERSITY

Principal Investigator

GONDO Yoichi  TOKAI UNIVERSITY,THE INSTITUTE OF MEDICAL SCIENCES,ASSOCIATE PROFESSOR, 総合医学研究所, 助教授 (40225678)

Co-Investigator(Kenkyū-buntansha) SAITOH Yasushi  THE INSTITUTE OF MEDICAL SCIENCES TOKAI UNIVERSITY RESEARCH ASSOCIATE, 総合医学研究所, 助手 (70287100)
Project Period (FY) 1995 – 1996
Keywordshomologous recombination / somatic cells / gene conversion / mouse / tissue culture cells / DNA repair / molecular genetics
Research Abstract

The major objective of this study is to establish a sensitive method of detecting chromosomal recombinations and to analyze recombinational mechanisms at the molecular level. Mouse embryonic stem (ES) cell lines that carried a neo-tk cassette at the p53 locus on the chromosome 11 were established by using gene targeting (Gondo et al., Biochem.Biophys.Res.Commun., 202 : 830-837,1994). These ES cell lines were, thus, heterozygotes carrying the wild-type (W) and neo-tk (T) alleles at the p53 locus. When W/T heterozygous cells were treated with GANC,recombinational repair genotype, namely, W/W homozygotes were detected. Reciprocally, T/T homozygotes were also isolated with a high concentration of G418. In order to investigate the molecular genetic changes at and around the recombinational repair region, we also established W/T heterozygotes by using TT2 ES cells which derived from a CBAXC57BL/6 F1 mouse. Preliminary studies indicated approximately a half of chromosome 11 markers were heter … More ozygous in TT2. By using these polymorphic markers of chromosome 11 and identified recombinants of W/T to W/W or T/T,it is now possible to analyze whether the recombinational repair was manifested by 1) gene conversion with a narrow sense, 2) mitotic crossing-over between two homologous, or 3) uniparental disomy-type irregular chromosomal segregation.
We also developed a new marker gene, Eco-gpt, in addition to HSV-tk to enhance the sensitivity as well as to reduce the labor-intensive tasks of the selection system. We have established ES lines carrying neo-gpt cassette (G) allele and confirmed the genotypes were W/G heterozygotes (Ito, Gondo et al., in preparation). W/W homozygotes were also isolated from W/G ES cells by 6-tg selection. Having two distinctive negatively selectable marker genes like HSV-tk and Eco-gpt, it is plausible to construct T/G heterozygotes. For instance, neo-tk integration to one allele and puromycin^R-gpt integration to the other should provide such heterozygotes. It will provide a new tool of the selection for recombinational repair between two heterozygous homologues to both directions ; from G/T to T/T and to G/G.The tagging of HSV-tk and Eco-gpt should be applicable to any part of mammalian genome including human when a tissue culture system is available. Thus by using the established system homologous recombination is now feasible to be investigated at the molecular level to elucidate the enzymatic process at any genes and species. Less

  • Research Products

    (14 results)

All Other

All Publications (14 results)

  • [Publications] T.Kitamoto,Y.Gondo,et al.: "Humanized prion protein knock-in by Creinduced site-specific recombination in the mouse." Biochem.Biophys.Res.Commun.222. 742-747 (1996)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] H.Shibata,Y.Gondo,et al.: "Inactive allele-specific methylation and chromatin structure of the imprinted gene U2af1-rs1 on mouse chromosome 11." Genomics. 35. 248-252 (1996)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] T.Norimura,Y.Gondo,et al.: "p53-dependent apoptosis suppresses radiation-induced teratogenesis." Nature Medicine. 2. 577-580 (1996)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Y.Gondo,Y.Shioyama,et al.: "A novel positive detection system of in vivo mutations in rpsL(strA)transgenic mice." Mutation Res.360. 1-14 (1996)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 権藤洋一、勝木元也: "遺伝毒性検定のためのトランスジェニックマウス." 組織培養. 22(13). 522-525 (1996)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 権藤洋一: "発がんリスク検定のためのトランスジェニックマウス." Pharma Sandoz. 3(21). 22-23 (1996)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 権藤洋一、勝木元也: "11.4 DNA変異を検出する方法." 講談社サイエンティフィク, 11 (1995)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Tetsukuki Kitamoto, Kenji Nakamura, Kazuki Nakao, Satoshi Shibuya, Ryong-Woon Shin, Yoichi Gondo, Motoya Katsuki and Jun Tateishi: "Humanized prion protein knock-in by Cre-induced site-specific recombination in the mouse." Biochem.Biophys.Res.Commun. 222. 742-747 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Hideo Shibata, Kiyoshi Yoshino, Shoichi Sunahara, Yoichi Gondo, Motoya Katsuki, Takayuki Ueda, Mamoru Kamiya, Masami Muramatsu, Yasufumi Murakami, Iveta Kalcheva, Christoph Plass, Verne M.Chapman and Yoshihida Hayashizaki: "Inactive allele-specific methylation and chromatin structure of the imprinted gene U2af1-rs1 on mouse chromosome 11." Genomics. 35. 248-252 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Yoichi Gondo, Yoshiyuki Shioyama, Kazuki Nakao, Motoya Katsuki: "A novel positive detection system of in vivo mutations in rpsL (strA) transgenic mice." Mutation Res.360. 1-14 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Toshiyuki Norimura, Satoshi Nomoto, Motoya Katsuki, Yoichi Gondo and Sohei Kondo: "p53-dependent apoptosis suppresses radiation-induced teratogenesis." Nature Medicine. 2. 577-580 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Yoichi Gondo and Motoya Katsuki: "Transgenic system for the genotoxicity test. (in Japanese)" The Tissue Culture. 22 (13). 522-525 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Yoichi Gondo: "Transgenic mouse for the carcinogenicity test. (in Japanese)" Pharma Sandoz. No.3 (21). 22-23 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Yoichi Gondo and Motoya Katsuki: Detection methods of DNA mutations (in Japanese) in Antimutagen, Anticarcinogenand their screening methods (ed.by Y.Kuroda). Kodansha Scientific Pless, Tokyo, Japan, 387-397 (1995)

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

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Published: 1999-03-09  

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