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Selection of ura5 and ura3 mutants from the two varieties of Cryptococcus neoformans on 5-fluoroorotic acid medium. 1992

K J Kwon-Chung, and A Varma, and J C Edman, and J E Bennett
Laboratory of Clinical Investigation, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892.

Spontaneous mutants requiring uracil were isolated from both varieties of Cryptococcus neoformans by plating on 5-fluoroorotic acid (5-FOA) medium. Of the 36 strains tested (18 var. neoformans and 18 var. gattii), 24 (12 of each variety) generated 5-FOA-resistant cells requiring uracil for growth. Six of the 12 C. neoformans var. gattii strains produced ura3 cells while the remaining six strains produced ura5 cells. None of the 12 strains produced both ura3 cells and ura5 cells. All 12 isolates of var. neoformans, however, produced ura5 cells and one of them produced ura3 as well as ura5 cells. A genetic lesion in the URA5 gene of an isolate of C. neoformans var. gattii was confirmed by complement with the cognate URA5 gene of C. neoformans var. neoformans. The ura3 isolates were tentatively identified by their ability to grow on a medium containing uridine but not on a medium with orotic acid or orotidine. Enzymatic assays for orotidine-5'-phosphate decarboxylase activity confirmed the isolates to be ura3 mutants. Hybridization analysis of total DNA, digested with EcoRI or StuI and probed with pURA5g2, revealed the presence of only one copy of URA5 in the strains of either variety, regardless of the prevalence of ura5 mutants. Extensive polymorphism was observed in the restriction patterns of the fragments containing the URA5 locus. The prevalence of spontaneously arising ura3 mutants among the isolates of C. neoformans var. gattii, but not among the isolates of C. neoformans var. neoformans, is one more biological difference that distinguishes the two varieties.

UI MeSH Term Description Entries
D009154 Mutation Any detectable and heritable change in the genetic material that causes a change in the GENOTYPE and which is transmitted to daughter cells and to succeeding generations. Mutations
D009693 Nucleic Acid Hybridization Widely used technique which exploits the ability of complementary sequences in single-stranded DNAs or RNAs to pair with each other to form a double helix. Hybridization can take place between two complimentary DNA sequences, between a single-stranded DNA and a complementary RNA, or between two RNA sequences. The technique is used to detect and isolate specific sequences, measure homology, or define other characteristics of one or both strands. (Kendrew, Encyclopedia of Molecular Biology, 1994, p503) Genomic Hybridization,Acid Hybridization, Nucleic,Acid Hybridizations, Nucleic,Genomic Hybridizations,Hybridization, Genomic,Hybridization, Nucleic Acid,Hybridizations, Genomic,Hybridizations, Nucleic Acid,Nucleic Acid Hybridizations
D009963 Orotic Acid An intermediate product in PYRIMIDINE synthesis which plays a role in chemical conversions between DIHYDROFOLATE and TETRAHYDROFOLATE. Potassium Orotate,Sodium Orotate,Zinc Orotate,Acid, Orotic,Orotate, Potassium,Orotate, Sodium,Orotate, Zinc
D009964 Orotidine-5'-Phosphate Decarboxylase Orotidine-5'-phosphate carboxy-lyase. Catalyzes the decarboxylation of orotidylic acid to yield uridylic acid in the final step of the pyrimidine nucleotide biosynthesis pathway. EC 4.1.1.23. Orotidine Phosphate Carboxy-Lyase,Orotidylate Decarboxylase,OMP Decarboxylase,Orotidine 5 Phosphate Decarboxylase,Orotidine 5' Phosphate Decarboxylase,Orotidine-5-Phosphate Decarboxylase,Carboxy-Lyase, Orotidine Phosphate,Decarboxylase, OMP,Decarboxylase, Orotidine-5'-Phosphate,Decarboxylase, Orotidine-5-Phosphate,Decarboxylase, Orotidylate,Orotidine Phosphate Carboxy Lyase,Phosphate Carboxy-Lyase, Orotidine
D003455 Cryptococcus neoformans A species of the fungus CRYPTOCOCCUS. Its teleomorph is Filobasidiella neoformans. Blastomyces neoformans,Debaryomyces neoformans,Filobasidiella neoformans,Lipomyces neoformans,Saccharomyces neoformans,Torula neoformans,Torulopsis neoformans,Cryptococcus neoformans var. grubii
D003470 Culture Media Any liquid or solid preparation made specifically for the growth, storage, or transport of microorganisms or other types of cells. The variety of media that exist allow for the culturing of specific microorganisms and cell types, such as differential media, selective media, test media, and defined media. Solid media consist of liquid media that have been solidified with an agent such as AGAR or GELATIN. Media, Culture
D004271 DNA, Fungal Deoxyribonucleic acid that makes up the genetic material of fungi. Fungal DNA
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia
D014170 Transformation, Genetic Change brought about to an organisms genetic composition by unidirectional transfer (TRANSFECTION; TRANSDUCTION, GENETIC; CONJUGATION, GENETIC, etc.) and incorporation of foreign DNA into prokaryotic or eukaryotic cells by recombination of part or all of that DNA into the cell's genome. Genetic Transformation,Genetic Transformations,Transformations, Genetic

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