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Highly Efficient Targeted Mutagenesis of Drosophila with the CRISPR/Cas9 System

2013; Cell Press; Volume: 4; Issue: 1 Linguagem: Inglês

10.1016/j.celrep.2013.06.020

2639-1856

Andrew Bassett, Charlotte Tibbit, Chris P. Ponting, Ji‐Long Liu,

Insect symbiosis and bacterial influences

Here, we present a simple and highly efficient method for generating and detecting mutations of any gene in Drosophila melanogaster through the use of the CRISPR/Cas9 system (clustered regularly interspaced palindromic repeats/CRISPR-associated). We show that injection of RNA into the Drosophila embryo can induce highly efficient mutagenesis of desired target genes in up to 88% of injected flies. These mutations can be transmitted through the germline to make stable lines. Our system provides at least a 10-fold improvement in efficiency over previously published reports, enabling wider application of this technique. We also describe a simple and highly sensitive method of detecting mutations in the target gene by high-resolution melt analysis and discuss how the new technology enables the study of gene function.