5-Fluoroorotic Acid

2011 ◽  
Keyword(s):  
Fluoroorotic Acid

Marker recycling via 5-fluoroorotic acid and 5-fluorocytosine counter-selection in the white-rot agaricomycete Pleurotus ostreatus

2016 ◽  
Vol 120 (9) ◽  
pp. 1146-1155 ◽  
Author(s):  
Takehito Nakazawa ◽  
Masami Tsuzuki ◽  
Toshikazu Irie ◽  
Masahiro Sakamoto ◽  
Yoichi Honda
Keyword(s):  
Pleurotus Ostreatus ◽  
White Rot ◽  
Marker Recycling ◽  
Counter Selection ◽  
Fluoroorotic Acid

The interaction of 5-fluoroorotic acid with transition metals: synthesis and characterisation of Ni(II), Cu(II) and Zn(II) complexes

2002 ◽  
Vol 89 (3-4) ◽  
pp. 227-236 ◽  
Author(s):  
Alain G. Schneider ◽  
Helmut W. Schmalle ◽  
Frédéric Arod ◽  
Erich Dubler
Keyword(s):  
Transition Metals ◽  
Fluoroorotic Acid

scholarly journals Anti-tumor activity of liposome encapsulated fluoroorotic acid as a single agent and in combination with liposome irinotecan

2011 ◽  
Vol 153 (3) ◽  
pp. 288-296 ◽  
Author(s):  
Kareen Riviere ◽  
Heidi M. Kieler-Ferguson ◽  
Katherine Jerger ◽  
Francis C. Szoka
Keyword(s):  
Single Agent ◽  
Fluoroorotic Acid ◽  
Anti Tumor Activity

scholarly journals Rare Homologous Gene Targeting in Histoplasma capsulatum: Disruption of the URA5Hc Gene by Allelic Replacement

1998 ◽  
Vol 180 (19) ◽  
pp. 5135-5143 ◽  
Author(s):  
Jon P. Woods ◽  
Diane M. Retallack ◽  
Elizabeth L. Heinecke ◽  
William E. Goldman
Keyword(s):  
Gene Targeting ◽  
Fungal Pathogen ◽  
Histoplasma Capsulatum ◽  
Selectable Marker ◽  
Pathogenic Fungi ◽  
Homologous Gene ◽  
Hygromycin B ◽  
Allelic Replacement ◽  
Fluoroorotic Acid ◽  
Potential Challenge

ABSTRACT URA5 genes encode orotidine-5′-monophosphate pyrophosphorylase (OMPpase), an enzyme involved in pyrimidine biosynthesis. We cloned the Histoplasma capsulatum URA5gene (URA5 Hc) by using a probe generated by PCR with inosine-rich primers based on relatively conserved sequences in OMPpases from other organisms. Transformation with this gene restored uracil prototrophy and OMPpase activity to UV-mutagenizedura5 strains of H. capsulatum. We attempted to target the genomic URA5 locus in this haploid organism to demonstrate homologous allelic replacement with transforming DNA, which has not been previously done in H. capsulatum and has been challenging in some other pathogenic fungi. Several strategies commonly used in Saccharomyces cerevisiae and other eukaryotes were unsuccessful, due to the frequent occurrence of ectopic integration, linear plasmid formation, and spontaneous resistance to 5-fluoroorotic acid, which is a selective agent for URA5 gene inactivation. Recent development of an efficient electrotransformation system and of a second selectable marker (hph, conferring hygromycin B resistance) for this fungus enabled us to achieve allelic replacement by using transformation with an insertionally inactivated Δura5 Hc::hph plasmid, followed by dual selection with hygromycin B and 5-fluoroorotic acid, or by screening hygromycin B-resistant transformants for uracil auxotrophy. The relative frequency of homologous gene targeting was approximately one allelic replacement event per thousand transformants. This work demonstrates the feasibility but also the potential challenge of gene disruption in this organism. To our knowledge, it represents the first example of experimentally directed allelic replacement inH. capsulatum, or in any dimorphic systemic fungal pathogen of humans.

scholarly journals sacB–5-Fluoroorotic Acid–pyrE-Based Bidirectional Selection for Integration of Unmarked Alleles into the Chromosome of Rhodobacter capsulatus

2005 ◽  
Vol 71 (6) ◽  
pp. 3014-3024 ◽  
Author(s):  
Takahiro Yano ◽  
Carsten Sanders ◽  
John Catalano ◽  
Fevzi Daldal
Keyword(s):  
Rhodobacter Capsulatus ◽  
De Novo ◽  
Model Organism ◽  
Photosynthetic Bacterium ◽  
Gene Encoding ◽  
Pyrimidine Nucleotide ◽  
Knockout Mutants ◽  
Selection For ◽  
Fluoroorotic Acid ◽  
Bidirectional Selection

ABSTRACT The gram-negative, purple nonsulfur, facultative photosynthetic bacterium Rhodobacter capsulatus is a widely used model organism and has well-developed molecular genetics. In particular, interposon mutagenesis using selectable gene cartridges is frequently employed for construction of a variety of chromosomal knockout mutants. However, as the gene cartridges are often derived from antibiotic resistance-conferring genes, their numbers are limited, which restricts the construction of multiple knockout mutants. In this report, sacB—5-fluoroorotic acid (5FOA)—pyrE-based bidirectional selection that facilitates construction of unmarked chromosomal knockout mutations is described. The R. capsulatus pyrE gene encoding orotate phosphoribosyl transferase, a key enzyme of the de novo pyrimidine nucleotide biosynthesis pathway, was used as an interposon in a genetic background that is auxotrophic for uracil (Ura−) and hence resistant to 5FOA (5FOAr). Although Ura+ selection readily yielded chromosomal allele replacements via homologous recombination, selection for 5FOAr to replace pyrE with unmarked alleles was inefficient. To improve the latter step, 5FOAr selection was combined with sucrose tolerance selection using a suicide plasmid carrying the Bacillus subtilis sacB gene encoding levansucrase that induces lethality upon exposure to 5% (wt/vol) sucrose in the growth medium. Sucrose-tolerant, 5FOAr colonies that were obtained carried chromosomal unmarked mutant alleles of the target gene via double crossovers between the resident pyrE-marked and incoming unmarked alleles. The effectiveness of this double selection was proven by seeking insertion and deletion alleles of helC involved in R. capsulatus cytochrome c biogenesis, which illustrated the usefulness of this system as a genetic means for facile construction of R. capsulatus unmarked chromosomal mutants.

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