FIGURE 3: Pif1 overexpression rescues lagging-CEB1 instability in the rfa1-D228Y mutant. (A) Colonies containing the leading- or lagging-CEB1, and overexpressing Pif1 (Pif1 overexpression) or not (Empty vector) were plated on galactose medium and subsequently incubated at 30°C. (B) Left: cDNA was prepared from the indicated strains. The ration of PIF1 transcript to that of ACT1 was determined using qPCR. (-): Empty vector, (+): Pif1 overexpression. Right, genomic DNAs from wild-type (WT) or rfa1-D228Y cells overexpressing Pif1 (Pif1 overexpression) and containing the leading-CEB1 were treated as in Figure 2. M: ladder DNA. (C) Left: cDNA was prepared from the indicated strains. The ration of PIF1 transcript to that of ACT1 was determined using qPCR. (-): Empty vector, (+): Pif1 overexpression. Right, genomic DNAs from wild-type (WT) or rfa1-D228Y cells overexpressing Pif1 (Pif1 overexpression) and containing the lagging-CEB1 were treated as in Figure 2. M: ladder DNA. Yeast strains were grown in medium containing galactose (2%). The number of colonies analysed per well, the percentage of rearrangement frequencies, and the total numbers of colonies are indicated in Table 1.