CRISPR/Cas9-mediated genome engineering and the promise of designer flies on demand

Fly (Austin). Oct-Dec 2013;7(4):249-55. doi: 10.4161/fly.26566. Epub 2013 Oct 2.

Abstract

The CRISPR/Cas9 system has attracted significant attention for its potential to transform genome engineering. We and others have recently shown that the RNA-guided Cas9 nuclease can be employed to engineer the Drosophila genome, and that these modifications are efficiently transmitted through the germline. A single targeting RNA can guide Cas9 to a specific genomic sequence where it induces double-strand breaks that, when imperfectly repaired, yield mutations. We have also demonstrated that 2 targeting RNAs can be used to generate large defined deletions and that Cas9 can catalyze gene replacement by homologous recombination. Zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) have shown similar promise in Drosophila. However, the ease of producing targeting RNAs over the generation of unique sequence-directed nucleases to guide site-specific modifications makes the CRISPR/Cas9 system an appealingly accessible method for genome editing. From the initial planning stages, engineered flies can be obtained within a month. Here we highlight the variety of genome modifications facilitated by the CRISPR/Cas9 system along with key considerations for starting your own CRISPR genome engineering project.

Keywords: CRISPR; Cas9; genome engineering; homologous recombination; site-directed mutagenesis.

Publication types

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

MeSH terms

  • Animals
  • CRISPR-Cas Systems*
  • Drosophila / genetics*
  • Drosophila / metabolism
  • Genetic Engineering / methods*
  • Genome, Insect*
  • Models, Genetic
  • Mutagenesis, Site-Directed