Article Figures & Data
Figures
- FIG. 1.
Properties of Cdc11-GFP fusion proteins. Cells were grown to log phase overnight at the indicated temperature, and then Cdc11-GFP fluorescence was analyzed. (A) Signal intensity for the different versions of Cdc11-GFP was compared in three independent assays in which 50 septin rings per assay were quantified for each different Cdc11-GFP. The average fluorescence intensity was normalized to 100 for Cdc11-YeGPF3. The Cdc11-CaGFPγ variant gave a significantly stronger signal than the other variants (P < 0.001). (B) Western blot analysis comparing the levels of Cdc11-GFP produced in the indicated strains. The lane labeled “neg” refers to the negative-control strain (BWP17) that lacks GFP. Blots were probed with anti-GFP to detect Cdc11-GFP, anti-glucose-6-phosphate dehydrogenase (αG6PD) as a control, and anti-Cdc11 to detect the untagged version of Cdc11.
- FIG. 2.
Photostability of GFP variants. (A and B) Relative fluorescence intensity of the GFP variants at 4-s intervals over a time course of 1 min of continuous exposure to the fluorescence excitation lamp after growth at 30°C (A) and at 37°C (B). CaGFPγ showed the best photostability (t1/2 of ∼2 min). The relative fluorescence was normalized to 100 for each Cdc11-GFP variant at the start of the time course. The results represent the average of three independent assays in which three septin rings were analyzed for each mutant. Error bars indicate standard deviations. (C) Cells carrying Cdc11 fused to YeGFP3 or CaGFPγ were continuously exposed to the fluorescence excitation lamp, and then images of septin rings were captured at the indicated times.
