Deletion mutagenesis is one of the most efficient approaches to studying gene function. However, conventional methods of inducing targeted mutations in the drosophila genome are timeand labor-consuming. This work proposes a new, simple, and effective method of producing drosophila mutants with gene deletions. The method involves the insertion of I-SceI and I-CreI recognition sites and a fragment homologous to the target sequence into the chromosome region of interest by means of an attB-containing construct, the induction of double-strand DNA breaks by the appropriate meganuclease, and their repair by homologous recombination. The procedure results in a deletion extending from the attP-site to the target locus. A cassette was designed to enable single-step construct production for the deletion of any given genomic region. A set of markers facilitates the selection of recombination events. The efficacy of the proposed technique was confirmed by the induction of a 47-kb deletion containing the qtc gene.
Russian Journal of Genetics – Springer Journals
Published: Nov 13, 2015
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