CRISPR pp 335-348 | Cite as

Precise Genome Editing of Drosophila with CRISPR RNA-Guided Cas9

  • Scott J. Gratz
  • Melissa M. Harrison
  • Jill Wildonger
  • Kate M. O’Connor-Giles
Part of the Methods in Molecular Biology book series (MIMB, volume 1311)


The readily programmable CRISPR-Cas9 system is transforming genome engineering. We and others have adapted the S. pyogenes CRISPR-Cas9 system to precisely engineer the Drosophila genome and demonstrated that these modifications are efficiently transmitted through the germline. Here we provide a detailed protocol for engineering small indels, defined deletions, and targeted insertion of exogenous DNA sequences within one month using a rapid DNA injection-based approach.

Key words

CRISPR Cas9 Genome engineering Nonhomologous end joining Homology-directed repair Drosophila 



Clustered regularly interspaced short palindromic repeats




Double-strand break


Double-stranded DNA


Guide RNA


Homology-directed repair




Nonhomologous end joining


Protospacer adjacent motif


Single-stranded DNA


Trans-activating CRISPR RNA



We are grateful to members of the Harrison, O’Connor-Giles, and Wildonger labs for their help in establishing CRISPR-Cas9 protocols in Drosophila, and to Dustin Rubinstein for comments on this chapter. Our work has been funded by start-up funds from the University of Wisconsin to M.M.H., J.W., and K.O.C.G. and grants from the National Institutes of Health to J.W. (R00 NS072252) and K.O.C.G. (R00 NS060985 and R01 NS078179). Plasmids and transgenic fly lines described here are available through the nonprofit distributor Addgene and the Bloomington Drosophila Stock Center, respectively. Detailed reagent information is available at


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Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Scott J. Gratz
    • 1
  • Melissa M. Harrison
    • 2
  • Jill Wildonger
    • 3
  • Kate M. O’Connor-Giles
    • 1
    • 4
    • 5
  1. 1.Genetics Training ProgramUniversity of Wisconsin-MadisonMadisonUSA
  2. 2.Department of Biomolecular ChemistryUniversity of Wisconsin School of Medicine and Public HealthMadisonUSA
  3. 3.Department of BiochemistryUniversity of Wisconsin-MadisonMadisonUSA
  4. 4.Laboratory of GeneticsUniversity of Wisconsin-MadisonMadisonUSA
  5. 5.Laboratory of Cell and Molecular BiologyUniversity of Wisconsin-MadisonMadisonUSA

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