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 allcles of existing X-linked or autosomal mutations in Drosophila.
Fehon
R G
RG
Department of Zoology, Duke University, Durham, North Carolina 27708-1000, USA. rfehon@acpub.duke.edu
Oren
T
T
LaJeunesse
D R
DR
Melby
T E
TE
McCartney
B M
BM
D000818:Animals; D018797:Cell Cycle Proteins / Q000235:genetics*; D003360:Cosmids; D004330:Drosophila / Q000235:genetics*; D005260:Female; D019204:GTP-Binding Proteins / Q000235:genetics*; D016515:Genes, Neurofibromatosis 2; D005816:Genetic Complementation Test; D006801:Humans; D008297:Male; D008565:Membrane Proteins / Q000235:genetics; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D025581:Neurofibromin 2; D020846:cdc42 GTP-Binding Protein, Saccharomyces cerevisiae
D018797:Cell Cycle Proteins; D008565:Membrane Proteins; D025581:Neurofibromin 2; D019204:GTP-Binding Proteins; D020846:cdc42 GTP-Binding Protein, Saccharomyces cerevisiae