MET15 as a visual selection marker for Candida albicans.
To develop better molecular genetic tools for the diploid yeast Candida albicans, the suitability of the MET15 gene as a visual selection marker was studied. Both MET15 alleles of C. albicans CAI-4 were isolated by functional complementation of a Saccharomyces cerevisiae strain lacking the MET15 gene. Growth of this complemented strain on Pb(2+)-containing medium was associated with a colour shift of brown into white colonies. The MET15 alleles of C. albicans were located on chromosome 4 by pulsed-field gel electrophoresis and Southern blotting. A met15-deficient strain of C. albicans CAI-4 was generated using the ura blaster technique. This strain showed a brown colony colour on Pb(2+)-containing medium, which corresponded with the colony colour of a S. cerevisiae strain lacking the MET15 gene. Unexpectedly, the met15-deficient strain of C. albicans still grew on methionine-depleted medium. However, this growth was severely delayed. In addition, complementation of this strain with an integrative or replicative plasmid containing either of the MET15 alleles resulted in the formation of white transformants on Pb(2+)-containing medium. These transformants grew very well on methionine-depleted medium. Colony sectoring was obtained with the replicative plasmid and not with the integrative one. This study demonstrates that the MET15 gene of C. albicans is suitable as a visual marker and therefore can be used to identify transformants and study plasmid stability. GenBank Accession Nos for MET15 nucleotide sequences are AF188273, AF188274 and AF188275.
Viaene
J
J
Department of Molecular Biology, Unit of Fundamental and Applied Molecular Biology, University of Ghent and Flanders Interuniversity Institute for Biotechnology, K. L. Ledeganckstraat 35, B-9000 Ghent, Belgium.
Tiels
P
P
Logghe
M
M
Dewaele
S
S
Martinet
W
W
Contreras
R
R
D000595:Amino Acid Sequence; D001483:Base Sequence; D015415:Biomarkers; D002176:Candida albicans / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development; D003547:Cysteine Synthase; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D005800:Genes, Fungal / Q000235:genetics; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes; D009154:Mutation; D029701:Saccharomyces cerevisiae Proteins; D017422:Sequence Analysis, DNA
D015415:Biomarkers; D018076:DNA, Complementary; D004271:DNA, Fungal; D005656:Fungal Proteins; D009097:Multienzyme Complexes; D029701:Saccharomyces cerevisiae Proteins; D003547:Cysteine Synthase; C103381:MET17 protein, S cerevisiae