Shuttle vectors for Candida albicans: control of plasmid copy number and elevated expression of cloned genes.
Plasmids containing the inosine monophosphate dehydrogenase gene CaIMH3 from Candida albicans strain ATCC 32354 transform their host to resistance against mycophenolic acid (MPA). The transformants maintain the plasmids at a high copy number (20-40 per cell) and express the CaIMH3 gene at very high levels relative to untransformed controls. The plasmid copy number can be controlled by the concentration of MPA in the media. The transformation procedure is reproducible and the efficiency of transformation is high, up to 15,000 per microgram. Unrearranged plasmids are readily recovered by transforming total DNA from transformants back into Escherichia coli. C. albicans genes cloned into the plasmid are expressed at elevated levels relative to untransformed controls. A derivative vector containing the CaMAL2 promoter and termination sequences expresses the CaERG11 ORF at high levels and confers moderate resistance to fluconazole. These shuttle vectors should facilitate global genomics approaches in C. albicans that have been hampered by its diploid genome.
Du
Wenjin
W
Department of Biochemistry and Molecular biology, Wayne State University School of Medicine, Detroit, MI 48201, USA.
Coaker
Melisa
M
Sobel
Jack D
JD
Akins
Robert A
RA
D002176:Candida albicans / Q000235:genetics* / Q000502:physiology; D003001:Cloning, Molecular; D004261:DNA Replication / Q000235:genetics*; D004171:Diploidy; D025141:Drug Resistance, Fungal / Q000235:genetics; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015966:Gene Expression Regulation, Fungal / Q000187:drug effects; D005800:Genes, Fungal; D007168:IMP Dehydrogenase / Q000235:genetics*; D009173:Mycophenolic Acid / Q000494:pharmacology; D010957:Plasmids / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins; D014170:Transformation, Genetic
D004791:Enzyme Inhibitors; D011994:Recombinant Proteins; D007168:IMP Dehydrogenase; D009173:Mycophenolic Acid