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We performed a successful transformation of the E2F1 plasmid into S. cerevisiae shown in Figure 1. Yeast with the E2F1 construct survived on uracil selective media, but not untransformed cells or cells transformed with the p14ARF sensor, which suggests no contamination.

Figure 1: Successful transformation of E2F1 plasmid into S. cerevisiae cells on a uracil dropout plate.

However, our Western Blot, as shown in Figure 2, showed no evidence of E2F1. We believe this is due to lack of E2F1 expression in the cells before our assay. Since we operated under the assumption that our construct was a constitutive promoter, we grew our stocks in glucose-based DO media instead of fructose-based media with added galactose. As a result, our actuator was likely repressed and little to no E2F1 was generated. Due to our error, we cannot extrapolate whether or not our actuator expressed E2F1 as intended.

Figure 2: Result of Western Blotting Procedure. Shown is the prestained protein ladder. No bands of protein are visible other than the ladder, indicating no present E2F1 protein.

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