FIG. 1.

Disruption of the C. albicans RBP1 genes encoding FKBP12. (A) Disruption alleles of the RBP1 gene. The RBP1 gene is depicted on a 5-kb genomic fragment including the 375-bp RBP1 ORF. One RBP1 wild-type allele was replaced with the CaURA3MX3 forward cassette, followed by treatment with 5-FOA to counterselect URA3. A second transformation replaced the second RBP1 allele with the CaURA3MX3 reverse cassette. Dashed lines represent PCR products used to verify homologous replacement of RBP1. Gray boxes indicate MX3 direct repeats. (B) PCR verification of homologous integration of insertion. Primers PR80 and PR92 amplified an ∼1.2-kb product containing the RBP1 ORF. Primers PR80 and JM37 amplified the rbp1::CaURA3MX3R allele. Primers PR92 and JM37 amplified the rbp1::CaURA3MX3F allele. Lanes: 1, 6, and 11, 1-kb markers; 2, 7, and 12, RBP1/RBP1 strain CAI4; 3, 8, and 13, rbp1/RBP1 strain YAG116; 4, 9, and 14, rbp1/rbp1 strain YAG171; 5, 10, and 15, no-DNA controls. (C) Southern analysis of RBP1 and rbp1 alleles. Genomic DNA from the wild-type RBP1/RBP1 strain CAI4, heterozygous rbp1/RBP1 strains YAG116 and YAG134, and homozygous rbp1/rbp1 strain YAG171 was cleaved with AflIII (A), electrophoresed in a 0.8% gel, transferred to a nylon membrane, and hybridized to a random-primed ³²P-labeled 375-bp gel-purified fragment spanning the RBP1 gene. The probe hybridizes to the RBP1 wild-type alleles in strain CAI4 (4.0 and 3.9 kb), to rbp1::URA3MX3F and RBP1 alleles in strain YAG116 (3.6 and 4.0 kb), to rbp1::MX3 and RBP1 alleles in strain YAG134 (4.2 and 4.0 kb), and to rbp1::MX3 and rbp1::URA3MX3R alleles in strain YAG171 (4.2 and 2.7 kb). The positions of DNA markers are shown on the left in kilobases.