FIGURE 6.

Experimental strategy to determine whether a reversion phenotype is caused by a second mutation in pdr5_S558Y. Structure of JG2011 (R-1 + pS558Y) after isolation of a clo^r suppressor. In this illustration, it is assumed that the new mutation (^*) lies in S558Y rather than in some other gene. Loss of the plasmid sequences containing URA3 and 1 of 2 PDR5 cassettes, either S558Y or pdr5::KANMX4 (GEN^r), occurs by recombination in one of the two homologous regions. These events result in Ura^- colonies that are selected on 5-FOA medium (14) and are tested on clotrimazole and geneticin medium. If the reversion is attributable to a second mutation in the S558Y copy, clo^r, gen^s, and clo^s, gen^r colonies will be recovered. A mutation in another gene would yield only clo^r colonies.