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Transgene excision in zebrafish using the phiC31 integrase.

James A Lister1

  • 1Department of Human and Molecular Genetics, Virginia Commonwealth University School of Medicine, Richmond, Virginia 23298-0033, USA. jalister@vcu.edu

Genesis (New York, N.Y. : 2000)
|January 23, 2010
PubMed
Summary

The Streptomyces bacteriophage phiC31 integrase enables genome engineering in zebrafish. This site-specific recombinase can excise transgenes and function in the germline, offering a new tool for zebrafish research.

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Area of Science:

  • Genetics
  • Molecular Biology
  • Developmental Biology

Background:

  • Site-specific recombinases (SSRs) are crucial for studying gene function in model organisms.
  • The Streptomyces bacteriophage phiC31 integrase mediates recombination between attP and attB sites.

Purpose of the Study:

  • To evaluate the feasibility of using phiC31 integrase for intramolecular recombination in zebrafish embryos.
  • To assess the functionality and applications of phiC31 integrase in zebrafish genome engineering.

Main Methods:

  • Testing phiC31 integrase activity in zebrafish cells and germline.
  • Excision of transgene cassettes flanked by attB and attP sites.
  • Utilizing a phiC31 integrase-estrogen receptor fusion protein dependent on 4-hydroxytamoxifen.

Main Results:

  • phiC31 integrase is functional in zebrafish cells and germline.
  • phiC31 integrase successfully excised transgene cassettes.
  • A tamoxifen-inducible phiC31 integrase variant showed recombination activity in zebrafish embryos.

Conclusions:

  • phiC31 integrase is a viable tool for genome manipulation in zebrafish.
  • This system offers a new strategy for intramolecular recombination and gene function studies in zebrafish.
  • The tamoxifen-inducible variant provides temporal control over recombination events.