A rapid and general assay for monitoring endogenous gene modification

Methods Mol Biol. 2010;649:247-56. doi: 10.1007/978-1-60761-753-2_15.

Abstract

The development of zinc finger nucleases for targeted gene modification can benefit from rapid functional assays that directly quantify activity at the endogenous target. Here we describe a simple procedure for quantifying mutations that result from DNA double-strand break repair via non-homologous end joining. The assay is based on the ability of the Surveyor nuclease to selectively cleave distorted duplex DNA formed via cross-annealing of mutated and wild-type sequence.

MeSH terms

  • Animals
  • Biological Assay / methods*
  • DNA Breaks, Double-Stranded
  • DNA Repair
  • Electrophoresis, Polyacrylamide Gel
  • Endonucleases / genetics
  • Endonucleases / metabolism
  • Humans
  • Models, Biological
  • Polymerase Chain Reaction
  • Zinc Fingers / genetics

Substances

  • Endonucleases