Figure 2. Performance of P_F139-URA3 and P_F19-URA3 plasmids as [SWI⁺] reporters for 74D-694 derivatives.

(A) DNA sequencing data demonstrate that the 74D-694 strain contains the same nonsense mutation in the FLO8 ORF as S288C-derivated strains. (B) Plasmid p413FLO8 was introduced into the four indicated strains to ectopically express Flo8 under its own promoter. Transformants were then tested for adhesion on SC selective plates using a wash assay (see Experimental Procedures). The remaining cells were imaged prior to and after washing (a representative result of at least three repeated assays). (C) The same strains used in panel B were assayed for flocculation (cell-cell adhesion, or cell aggregation, see details in Experimental Procedures). 0 min, immediately after vortexing the cell cultures; 15 min, 15 min after keeping cultures still on bench post-vortex. Shown is a typical result of at least three repeated tests. (D) Indicated isogenic 74D-694 strains were examined for raffinose phenotype (right) and aggregation patterns of Swi1-NQ-YFP (NQ), Sup35-NM-GFP (NM) and Rnq1-GFP (RNQ1) after transforming them with p416TEF-NQYFP, pCUP1-NMGFP, or pCUP1-RNQ1GFP. (E) Indicated strains were co-transformed with p415F139-URA3 and p413FLO8 or p413TEF (vector), and assayed for their growth on glucose-containing SC medium (glucose) without uracil (−uracil), or with 5-FOA (+5FOA). Shown are representative results (3 days post-spotting) of at least three repeated assays. (F) Experiment was carried out similar to that in panel E except that p415F19-URA3 was used instead of p415F139-URA3.