Isolation of mutations in the Drosophila homologues of the human Neurofibromatosis 2 and yeast CDC42 genes using a simple and efficient reverse-genetic method

• Published in: Genetics (Austin) 1997-05, Vol.146 (1), p.245-252
• Main Authors: Fehon, R.G. (Duke University, Durham, NC.), Oren, T, LaJeunesse, D.R, Melby, T.E, McCartney, B.M
• Format: Article
• Language: English
• Subjects: ADN; Animals; cdc42 GTP-Binding Protein, Saccharomyces cerevisiae; Cell Cycle Proteins - genetics; CHROMOSOME; CHROMOSOMES; COMPLEMENTARY DNA; COMPLEMENTATION; COMPLEMENTATION GROUPS; COSMID TRANSGENES; Cosmids; CROMOSOMAS; DESARROLLO EMBRIONARIO; DEVELOPPEMENT EMBRYONNAIRE; DNA; Drosophila; Drosophila - genetics; DROSOPHILA MELANOGASTER; EMBRYONIC DEVELOPMENT; ENSAYO; Female; GENE; GENE LETAL; GENES; GENES LETALES; Genes, Neurofibromatosis 2; GENETIC ANALYSIS; Genetic Complementation Test; GENETICA; GENETICS; GENETIQUE; GTP-Binding Proteins - genetics; HEMIZYGOSITY; Humans; Insects; Investigations; LETHAL GENES; Male; Membrane Proteins - genetics; MERLIN GENE; Molecular Sequence Data; MUTACION; MUTAGENESIS; Mutagenesis, Insertional; MUTATION; Neurofibromin 2; NUCLEIC PROBES; POLYTENE CHROMOSOMES; REVERSE GENETIC ANALYSIS; SCREENING; SONDAS NUCLEICAS; SONDE NUCLEIQUE; TESTAGE; TESTING; Tumors; Yeast
• ISSN: 0016-6731; 1943-2631; 1943-2631
• DOI: 10.1093/genetics/146.1.245

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 allcles of existing X-linked or autosomal mutations in Drosophila.