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Genome engineering has revolutionised genetic analysis in many organisms. Here we describe a simple and efficient technique to generate and detect novel mutations in desired target genes in Drosophila melanogaster. We target double strand breaks to specific sites within the genome by injecting mRNA encoding the Cas9 endonuclease and in vitro transcribed synthetic guide RNA into Drosophila embryos. The small insertion and deletion mutations that result from inefficient non-homologous end joining at this site are detected by high resolution melt analysis of whole flies and individual wings, allowing stable lines to be made within 1 month.

Original publication

DOI

10.1016/j.ymeth.2014.02.019

Type

Journal article

Journal

Methods

Publication Date

09/2014

Volume

69

Pages

128 - 136

Keywords

CRISPR, Cas9, Drosophila melanogaster, Genome engineering, RNA injection, Targeted mutagenesis, Animals, Animals, Genetically Modified, Base Sequence, Binding Sites, CRISPR-Cas Systems, Clustered Regularly Interspaced Short Palindromic Repeats, DNA-Binding Proteins, Drosophila, Drosophila Proteins, Genetic Engineering, Genome, Molecular Sequence Data