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Active Vam2p and Vam6p are required on both vacuole fusion partners. BJ3505 or DKY6281 vacuoles were mixed with buffer or antibodies against Vam2p or Vam6p. After 10 min, at 0°C, vacuoles were diluted 20-fold with PS buffer and reisolated to remove unbound antibody. Vacuole fusion partners were combined in standard fusion reactions supplemented with cytosol (30 μg/ml).
