FIG. 3.

Deletion of the RAS1 gene in C. albicans. (A) The RAS1 ORF (0.9-kb), hisG-URA3-hisG (4.0-kb), and hph-URA3-hph (3.4-kb) cassettes are represented by open boxes. The arrows in the boxes indicate the direction of the transcription of URA3. 5′- and 3′-end sequences of RAS1 are represented by shaded boxes. (B) Low-stringency Southern blot analysis of RAS genes in C. albicans. Candida genomic DNAs from different strains were digested with SspI, run on 1% agarose gel, and transferred to the Hybond⁺ membrane. The membrane was hybridized at 65°C overnight and washed at 60°C in 0.5 × SSC (1× SSC is 0.15 M NaCl plus 0.015 M sodium citrate). The probe is RAS1 ORF. (C) Southern blot analysis of RAS1 deletion. Genomic DNAs were digested with SspI. The probe is the RAS1 3′ sequence. Lanes: 1, wild-type strain; 2 and 3, RAS1/ras1-1; 4 and 5, RAS1/ras1-2; 6 and 7, ras1-2/ras1-3; 8 and 9, ras1-2/ras1-4.