Isolation of Mutations in the Drosophila Homologues of the Human Neurofibromatosis 2 and Yeast Cdc42 Genes Using a Simple and Efficient Reverse-Genetic Method

Reverse genetic analysis in Drosophila has been greatly aided by a growing collection of lethal P transposable element insertions that provide molecular tags for the identification of essential genetic loci. However, because the screens performed to date primarily have generated autosomal P-element insertions, this collection has not been as useful for performing reverse genetic analysis of X-linked genes. We have designed a reverse genetic screen that takes advantage of the hemizygosity of the X chromosome in males together with a cosmid-based transgene that serves as an autosomally linked duplication of a small region of the X chromosome. The efficacy and efficiency of this method is demonstrated by the isolation of mutations in Drosophila homologues of two well-studied genes, the human Neurofibromatosis 2 tumor suppressor and the yeast CDC42 gene. The method we describe should be of general utility for the isolation of mutations in other X-linked genes, and should also provide an efficient method for the isolation of new alleles of existing X-linked or autosomal mutations in Drosophila.