Efficient genome engineering by targeted homologous recombination in mouse embryos using transcription activator-like effector nucleases

Nat Commun. 2014;5:3045. doi: 10.1038/ncomms4045.


Generation of mouse models by introducing transgenes using homologous recombination is critical for understanding fundamental biology and pathology of human diseases. Here we investigate whether artificial transcription activator-like effector nucleases (TALENs)-powerful tools that induce DNA double-strand breaks at specific genomic locations-can be combined with a targeting vector to induce homologous recombination for the introduction of a transgene in embryonic stem cells and fertilized murine oocytes. We describe the generation of a conditional mouse model using TALENs, which introduce double-strand breaks at the genomic locus of the special AT-rich sequence-binding protein-1 in combination with a large 14.4 kb targeting template vector. We report successful germline transmission of this allele and demonstrate its recombination in primary cells in the presence of Cre-recombinase. These results suggest that TALEN-assisted induction of DNA double-strand breaks can facilitate homologous recombination of complex targeting constructs directly in oocytes.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Cells, Cultured
  • DNA / genetics
  • Deoxyribonucleases / genetics*
  • Deoxyribonucleases / physiology*
  • Embryo, Mammalian / cytology*
  • Embryo, Mammalian / physiology
  • Embryonic Stem Cells / cytology
  • Embryonic Stem Cells / physiology
  • Gene Targeting / methods*
  • Genetic Engineering / methods*
  • Genetic Vectors / genetics
  • Genetic Vectors / physiology
  • Integrases / physiology
  • Matrix Attachment Region Binding Proteins / genetics
  • Matrix Attachment Region Binding Proteins / physiology
  • Mice
  • Models, Animal
  • Molecular Sequence Data
  • NIH 3T3 Cells
  • Oocytes / cytology
  • Oocytes / physiology
  • Recombination, Genetic / genetics*
  • Transcriptional Activation / genetics*
  • Transcriptional Activation / physiology*


  • Matrix Attachment Region Binding Proteins
  • Satb1 protein, mouse
  • DNA
  • Cre recombinase
  • Integrases
  • Deoxyribonucleases