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Methods. 2014 Sep;69(2):128-36. doi: 10.1016/j.ymeth.2014.02.019. Epub 2014 Feb 24.

CRISPR/Cas9 mediated genome engineering in Drosophila.

Author information

1
Medical Research Council Functional Genomics Unit, Department of Physiology, Anatomy and Genetics, University of Oxford, South Parks Road, Oxford OX1 3PT, United Kingdom. Electronic address: andrew.bassett@dpag.ox.ac.uk.
2
Medical Research Council Functional Genomics Unit, Department of Physiology, Anatomy and Genetics, University of Oxford, South Parks Road, Oxford OX1 3PT, United Kingdom. Electronic address: jilong.liu@dpag.ox.ac.uk.

Abstract

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.

KEYWORDS:

CRISPR; Cas9; Drosophila melanogaster; Genome engineering; RNA injection; Targeted mutagenesis

PMID:
24576617
DOI:
10.1016/j.ymeth.2014.02.019
[Indexed for MEDLINE]

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