title: Going green in Cryptococcus neoformans: the recycling of a selectable drug marker.
abstract: Cryptococcus neoformans is an opportunistic fungal pathogen that primarily affects immunocompromised individuals. Reverse genetics is commonly used to identify and characterize genes involved in a variety of cellular processes. In C. neoformans there is a limited set of positive selectable markers available to make gene deletions or other genetic manipulations. This has hampered the application of reverse genetics in this organism. We have adapted the Bacteriophage P1 Cre-loxP system for use in C. neoformans and successfully excised and reused the same drug marker, G418, to make two sequential gene deletions, lac1Delta and cap59Delta, in the same strain. This tool will allow investigators to make multiple sequential gene deletions in the same strain, which should facilitate the analysis of multigene families.
authors: Department of Molecular Microbiology, Washington University School of Medicine, 660 S. Euclid Avenue, Saint Louis, MO 63110, USA.
authors: Raunak D
authors: RD
authors: Patel
authors: Jennifer K
authors: JK
authors: Lodge
authors: Lorina G
authors: LG
authors: Baker
mesh_terms: D015825:Chromosomes, Fungal; D003455:Cryptococcus neoformans / Q000187:drug effects / Q000235:genetics*; D004271:DNA, Fungal / Q000737:chemistry; D005656:Fungal Proteins / Q000235:genetics; D017353:Gene Deletion; D015966:Gene Expression Regulation, Fungal; D005819:Genetic Markers; D005822:Genetic Vectors; D016681:Genome, Fungal; D005839:Gentamicins / Q000494:pharmacology*; D019426:Integrases / Q000235:genetics; D008969:Molecular Sequence Data; D005810:Multigene Family; D016254:Mutagenesis, Insertional; D010641:Phenotype; D011993:Recombinant Fusion Proteins / Q000378:metabolism
chemical_list: D004271:DNA, Fungal; D005656:Fungal Proteins; D005819:Genetic Markers; D005839:Gentamicins; D011993:Recombinant Fusion Proteins; C010680:antibiotic G 418; D019426:Integrases