Marine Biotechnology

, Volume 5, Issue 2, pp 174–184

RecA-Mediated, Targeted Mutagenesis in Zebrafish

  • Zongbin Cui
  • Ying Yang
  • Christopher D. Kaufman
  • Dritan Agalliu
  • Perry B. Hackett
Article

DOI: 10.1007/s10126-002-0059-0

Cite this article as:
Cui, Z., Yang, Y., Kaufman, C.D. et al. Mar. Biotechnol. (2003) 5: 174. doi:10.1007/s10126-002-0059-0

Abstract

We have evaluated the efficacy of RecA, a prokaryotic protein involved with homologous recombination, to direct site-specific mutagenesis in zebrafish embryos. For this we coinjected a vector containing a mutated enhanced green fluorescent protein (EGFP) gene plus 236-nucleotide corrective single-stranded DNAs coated with RecA into 1-cell zebrafish embryos. Twenty-hours after fertilization, about 5% to 20% of injected embryos showed EGFP expression in 1 or more cells when RecA-coated corrective DNAs were used, but not when RecA was omitted. Mutated EGFP genes with 1-bp insertions or deletions were inefficiently activated, whereas those with 7-bp insertions were activated about 4-fold more efficiently. RecA-coated template strand had a higher efficiency than its complementary strand in activation of EGFP expression. Prior irradiation of the embryos with UV light enhanced RecA-mediated restoration of gene activity, suggesting that the effects we observed were augmented by one or more factors of zebrafish DNA repair systems.

Keywords

RecA protein gene targeting site-specific mutagenesis zebrafish 

Copyright information

© Springer-Verlag New York Inc. 2003

Authors and Affiliations

  • Zongbin Cui
    • 1
    • 2
  • Ying Yang
    • 1
  • Christopher D. Kaufman
    • 1
  • Dritan Agalliu
    • 1
  • Perry B. Hackett
    • 1
  1. 1.Department of Genetics, Cell Biology and Development and The Arnold and Mabel Beckman Center for Transposon ResearchUniversity of Minnesota, St. Paul, MN 55108-1095USA
  2. 2.Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072People's Republic of China

Personalised recommendations