Gene Targeting in Drosophila

DNA double-strand breaks provide a powerful means to modify the genome. This chapter describes how to generate and use these breaks to target specific sequences, or other modifications to the Drosophila genome. Both P element dependent gene conversion, in which the chromosomal DNA is broken, and the Rong and Golic gene-targeting technique, in which the targeting vector contains the DNA break are explained. The strengths and limitations of both methods are presented so that the user can choose the appropriate method for their particular situation. The efficiency of both methods depends upon the genomic location being modified, although few, if any, genomic locations are refractory to either method. It cannot be emphasized strongly enough that the investigator should be prepared to invest sufficient time into setting up and running these experiments properly.