I-SceI endonuclease, a new tool for studying DNA double-strand break repair mechanisms in Drosophila

As a step toward the development of a homologous recombination system in Dr osophila, we have developed a methodology to target double-strand breaks (D SBs) to a specific position in the Drosophila genome. This method uses the mitochondrial endonuclease I-Scd that recognizes and cuts an 18-bp restrict ion site. We find that >6% of the progeny derived from males that carry a m arker gene bordered by two I-SceI sites and that express I-Scd in their ger m line lose the marker gene. Southern blot analysis and sequencing of the r egions surrounding the I-Scd sites revealed that in the majority of the cas es, the introduction of DSBs at the I-Scd sites resulted in the complete de letion of the marker gene; the other events were associated with partial de letion of the marker gene. We discuss a number of applications for this nov el technique, in particular its use to study DSB repair mechanisms.