Figure 2.

In depth analysis of the COPI compartment. A, Co‐localization assignments for GFP N′ tagged compartment markers of Sec13, Anp1, Sec7 or Chc1 in comparison to the Cop1‐mCherry marker. B, Isolation of intact heptameric coatomer from a GFP‐tagged Cop1/α‐COP strain, demonstrating that the GFP‐tag does not affect complex stability. All western blots were detected with an antibody raised against coatomer. C, Affinity purification of the indicated GFP‐tagged proteins. SDS/PAGE and immunoblot analysis of the eluates using antibodies specific for GFP and coatomer. D, Bar graph of the relative amounts of co‐purified COPI as quantified by the densiometric analysis of immunoblots. Quantification of 2 independent experiments. Error bars depict SEM. Avt2 was chosen as a negative control because it localizes to the vacuole/endosomes. The threshold for non‐specific binding was set at 4 times that of Avt2. All scale bars are 5 μm