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. 2019 Mar 8;180(1):228–239. doi: 10.1104/pp.18.01315

Figure 5.

Figure 5.

The SYP121 R20R21 motif affects binding with SEC11Δ149. A, Schematic of SEC11 domain topology. The domain break is indicated by the arrow. B, Yeast mbSUS assay is performed by using Exg2p fusions of SEC11-Cub (Zhang et al., 2018) as bait and NubG-X fusions of SNARE proteins and controls (negative, NubG; positive, NubI) as prey. Growth on CSM-LTUM was used to verify the presence of both bait and prey expression. CSM-LTUMAH was used to verify Ade- and His-independent growth of the yeast diploids. The addition of 50 μm Met to CSM-LTUMAH was used to verify interaction with SEC11-Cub expression suppressed. Yeast was dropped at 1 and 0.1 OD600 as indicated. Incubation time was 24 h for CSM-LTUM plate and 72 h for CSM-LTUMAH plates. Immunoblot analysis (5 μg of total protein per lane) of the haploid yeast used in mating (right) used the α-HA antibody for the SNARE fusions and the α-VP16 antibody for the SEC11 fusion.