Fig. 2.

Construction and Southern blot verification of a C. albicans als2Δ/PMAL2-ALS2 strain. The locations of the primers from Table 2 are shown by numbers in parentheses. Numbers placed above the line drawings denote forward primers; reverse primers are shown below the line drawings. (a) Both ALS2 alleles from strain SC5314 are approximately 6·2 kb and are represented in a single diagram. (b) The hisG-URA3-hisG cassette constructed by cloning PCR-amplified ALS2 upstream and downstream sequences on either side of the parent cassette. (c) The ALS2 locus with the disruption cassette integrated to replace one ALS2 allele to create strain 1442. Growth of this strain on 5-FOA created strain 1443, which was transformed with the PMAL2 construct in (d) to create strain 2342. The PMAL2-regulated ALS2 locus is shown in (e). (f) Southern blot of Bg/II-digested genomic DNA from the indicated strains hybridized with an ALS2 upstream probe to demonstrate correct strain construction. Molecular sizes (in kb) are shown on the left. Cross-hybridization between sequences upstream of ALS2 and ALS4 resulted in the appearance of the bands at approximately 9 and 11 kb, which correspond to the alleles of ALS4.