FIG. 1.

General mutation strategy. In step 1, the mTnYl1 flanking regions were sequenced to design the amplification primers Hup and Hdw. In step 2, the homologous integration of mTnYl1 was checked with the two primers. WT, wild type; fil-, Fil⁻. In step 3, the presence of a single copy of mTnYl1 in the mutants was checked by Southern blot analysis with mTnYl1 as a probe. In step 4, the wild-type locus was disrupted with a cassette derived from the mutated locus and containing the YlURA3 gene as a marker, and the phenotype was verified.