Fig. 3. Sly1 is included in the Bet1-Sed5 v-t-SNARE complex in vivo. The SLY1 gene of the wild-type strain W303 was substituted with an epitope-tagged SLY1-6myc. Cells incubated in YEPD medium at 30°C were lysed by vortexing with glass beads on ice. After adding 1% Triton X-100, the lysate was divided into three parts, and each was immunoprecipitated at 4°C by a monoclonal anti-myc antibody, by an affinity-purified polyclonal anti-Sed5 antibody or by the preimmune rabbit serum from which the anti-Sed5 antibody had arisen. Anti-myc, anti-Sed5 and anti-Bet1 antibodies were used to detect Sly1-6myc, Sed5 and Bet1 proteins in the immunoprecipitates (lanes 1-3) and one fifth of their supernatants (lanes 4-6). One fifth of starting lysate is shown in lane 7.