Continuous directed evolution of DNA-binding proteins to improve TALEN specificity

Nat Methods. 2015 Oct;12(10):939-42. doi: 10.1038/nmeth.3515. Epub 2015 Aug 10.


Nucleases containing programmable DNA-binding domains can alter the genomes of model organisms and have the potential to become human therapeutics. Here we present DNA-binding phage-assisted continuous evolution (DB-PACE) as a general approach for the laboratory evolution of DNA-binding activity and specificity. We used this system to generate transcription activator-like effectors nucleases (TALENs) with broadly improved DNA cleavage specificity, establishing DB-PACE as a versatile approach for improving the accuracy of genome-editing agents.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • DNA-Binding Proteins / chemistry
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism*
  • Deoxyribonucleases / metabolism*
  • Directed Molecular Evolution / methods*
  • Gene Targeting / methods
  • High-Throughput Screening Assays / methods
  • Humans
  • Polycomb Repressive Complex 1 / genetics
  • Polycomb Repressive Complex 1 / metabolism
  • Protein Engineering / methods


  • CBX8 protein, human
  • DNA-Binding Proteins
  • Polycomb Repressive Complex 1
  • Deoxyribonucleases