Targeted mutagenesis by homologous recombination in D. melanogaster
Abstract
We used a recently developed method to produce mutant alleles of five endogenous Drosophila genes, including the homolog of thep53 tumor suppressor. Transgenic expression of the FLP site-specific recombinase and the I-SceI endonuclease generates extrachromosomal linear DNA molecules in vivo. These molecules undergo homologous recombination with the corresponding chromosomal locus to generate targeted alterations of the host genome. The results address several questions about the general utility of this technique. We show that genes not near telomeres can be efficiently targeted; that no knowledge of the mutant phenotype is needed for targeting; and that insertional mutations and allelic substitutions can be easily produced.
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Footnotes
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Present addresses: 4Laboratory of Molecular Cell Biology, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA; 5Program in Gene Function and Expression, University of Massachusetts Medical School, Worcester, MA 01655, USA.
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↵6 Corresponding author.
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E-MAIL kgg{at}stowers-institute.org; FAX (816) 926-2065.
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Article and publication are at http://www.genesdev.org/cgi/doi/10.1101/gad.986602.
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- Received February 21, 2002.
- Accepted May 3, 2002.
- Cold Spring Harbor Laboratory Press
