Fig. 1.

Ofm phenotype and complementation test. A schematic drawing of the 5ORIΔ-ΔR fragment of chromosome III is shown at the top. The positions of ARS elements are indicated by numbers above the line, with three digit numbers (e.g., 301) indicating dormant origins still present on the fragment and three digit numbers followed by a “triangle” symbol (e.g., 305Δ) indicating origin deletions. The arrows on the line represent the three selectable markers LEU2, ADE2 and TRP1; the latter marks the position of the chromosome fragmentation that removed the right arm of the chromosome distal to the ARS310 deletion. This fragment was introduced into both the wild-type (YKN15) and the ofm6-1 mutant (YJT417) by chromoduction. After selection, chromoductants were plated for single colonies on medium containing limiting adenine, and incubated for 5 days at 30 °C. Panels a, b and c show colonies from three different hst3 strains: the original ofm6-1 isolate (YJT417), the reconstructed point mutant (YIC257) and the hst3Δ::kanMX mutant (YIC247), respectively. Panel d shows colony-sectoring phenotypes of the wild-type strain. Loss events are visualized as red sectors in white colonies. A complementation test was done by introducing the HST3 gene into each of these mutants. A plasmid carrying the HST3 ORF under the control of its own promoter was integrated into the non-essential YGL119 W ORF by two-step gene replacement. Note that the HST3 gene complements the colony-sectoring phenotype of all mutants: e ofm6-1 (YIC275) f reconstructed point mutant (YIC271) and g hst3Δ (YIC273). Panel d shows the colony-sectoring phenotypes of a strain carrying the HST4 ORF integrated in the HST3 locus so that HST4 expression is regulated by the HST3 promoter plus the 3′ UTR (hst3::HST3pr-HST4)