An alternative novel tool for DNA editing without target sequence limitation: the structure-guided nuclease

Genome Biol. 2016 Sep 15;17(1):186. doi: 10.1186/s13059-016-1038-5.

Abstract

Engineered endonucleases are a powerful tool for editing DNA. However, sequence preferences may limit their application. We engineer a structure-guided endonuclease (SGN) composed of flap endonuclease-1 (FEN-1), which recognizes the 3' flap structure, and the cleavage domain of Fok I (Fn1), which cleaves DNA strands. The SGN recognizes the target DNA on the basis of the 3' flap structure formed between the target and the guide DNA (gDNA) and cut the target through its Fn1 dimerization. Our results show that the SGN, guided by a pair of gDNAs, cleaves transgenic reporter gene and endogenous genes in zebrafish embryonic genome.

Keywords: DNA editing; Fok I; Guide DNA; Structure-guided endonuclease; flap endonuclease-1 (FEN-1).

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Base Sequence
  • DNA / chemistry
  • DNA / genetics*
  • DNA / metabolism
  • DNA, Single-Stranded / genetics
  • DNA, Single-Stranded / metabolism
  • Deoxyribonucleases, Type II Site-Specific / chemistry
  • Deoxyribonucleases, Type II Site-Specific / genetics
  • Deoxyribonucleases, Type II Site-Specific / metabolism
  • Endodeoxyribonucleases / chemistry
  • Endodeoxyribonucleases / genetics
  • Endodeoxyribonucleases / metabolism*
  • Flap Endonucleases / chemistry
  • Flap Endonucleases / genetics
  • Flap Endonucleases / metabolism
  • Gene Editing*
  • Gene Targeting / methods
  • Models, Biological
  • Mutation
  • Sequence Analysis, DNA
  • Substrate Specificity
  • Zebrafish

Substances

  • DNA, Single-Stranded
  • DNA
  • Endodeoxyribonucleases
  • Flap Endonucleases
  • endodeoxyribonuclease FokI
  • Deoxyribonucleases, Type II Site-Specific