Gene Targeting By Homologous Recombination in Drosophila

Yikang Rong
University of Utah
Department of Biology
257 S. 1400 E. Rm. 201
Salt Lake City, UT 84112 USA
telephone: (801)585-5208
fax: (801)581-4668
prestype: Platform
presenter: Yikang Rong

Yikang Rong and Kent G. Golic
Department of Biology, University of Utah, Salt Lake City, UT 84112

We have developed a method of homologous gene targeting in Drosophila. By using a pair of yeast enzymes involved in DNA metabolism, we were able to efficiently generate, in the Drosophila germ cells, a donor DNA molecule for homologous recombination which had a double strand break within the gene we wanted to target. In an attempt to repair this break, the cellular machinery pairs the extrachromosomal donor with the endogenous locus and carries out homologous recombination, leading to the integration of the donor at the target locus. Targeting was quite efficient at the yellow locus with an estimated frequency of one event in 500 female gametes. Recently, we have used a modified targeting scheme to recover the first directed knock-out in Drosophila, making it clear that this technique will be generally useful. It enables us to take full advantage of the newly sequenced Drosophila genome by allowing us to generate mutations in essentially any gene starting with only the DNA sequence of that gene and without relying on prior knowledge of the mutant phenotype.

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