http://orcid.org/0000-0003-3463-8072
Figures & data
Figure 1. Growth of the wild-type and spontaneous resistant mutant strains on azole-containing medium. Both strains were inoculated on Czapek–Dox agar medium supplemented with 10 μg/mL miconazole and grown for 10 days at 30°C.
Figure 2. Phylogenetic trees of the PDR-type (A) and MDR-type (B) ABC transporters from A. oryzae, A. fumigatus, and A. nidulans. The trees were generated using a multiple sequence alignment program ClustalW (http://align.genome.jp/) with default settings. Amino acid sequences of the proteins were obtained from the Aspergillus Genome Database (AspGD; http://www.aspgd.org/).
Figure 3. Northern blot analysis of the ABC transporter genes in the wild-type and spontaneous mutant strains with or without 10 μg/mL miconazole.
Figure 4. Growth phenotypes of atrA, atrG, and atrH overexpression transformants. Approximately 104 conidiospores were inoculated and grown on Czapek–Dox agar medium (containing 0.5% maltose as a carbon source) supplemented with 2 μg/mL clotrimazole or 20 μg/mL miconazole for 14 days at 30°C.
Figure 5. Construction of the deletion mutants for the three ABC transporter genes in A. oryzae. (A) Strategies for single gene deletion by homologous recombination and Southern blot analysis of the deletion mutants. Genomic DNAs of the wild-type and deletion mutant strains were digested with the restriction enzyme indicated. Digested DNAs were electrophoresed on a 0.8% agarose gel and transferred onto a nylon membrane. The predicted sizes of the signals resulted by digestion with restriction enzymes are shown in the panel. (B) Strategies for construction of double deletion mutants using the ΔatrG mutant as a host strain. Southern blot analysis was performed as described in (A).
Figure 6. Susceptibility of the deletion mutant strains against the azole fungicides. (A) Growth of the single deletion mutants on the azole-containing medium. Approximately 104–10 conidiospores were inoculated on Czapek–Dox agar medium containing 0.15 µg/mL clotrimazole or 0.375 μg/mL miconazole supplemented with methionine and cultivated for 3 days at 30°C. (B) Growth of the double deletion mutants on the azole-containing medium. The strains were grown under the same culture conditions as described in (A).
