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. 1997 Jan 27;136(2):287–297. doi: 10.1083/jcb.136.2.287

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Induction of synthesis of Vps27p in vps27 cells. NBY67 (vps27) cells harboring either pSN55 (A-ALP), pSN100 ((F/A)AALP) or pSN97 (RS-ALP), and pHY5 (VPS27 under GAL1 control) (+) or empty vector (−) were grown in synthetic media containing 2% raffinose. At time t = 0, galactose was added to a final concentration of 2%, and cells were allowed to grow for an additional 90 min. At times t = 0 (− galactose) and t = 90 min (+ galactose), samples were removed and treated in the following ways (also see Fig. 5): (A) Whole-cell extracts were prepared from 10 OD units of NBY67 cells harboring pSN97 and pHY5 or empty vector to detect the presence of Vps27p using immunoblot analysis (similar results were obtained using the same cells harboring either pSN55 or pSN100 in conjunction with pHY5; data not shown); and (B) 0.5 OD units of cells were radiolabeled to determine whether the cells were secreting CPY.