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. 1998 Dec 28;143(7):1789–1800. doi: 10.1083/jcb.143.7.1789

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ndc1-1 strains do not exhibit defects in NPC assembly. An assay to examine NPC assembly was carried out using either wild-type (WT) or ndc1-1 cells (HC10-42a/42d and HC10-42b/42c, respectively; Table I) transformed with a plasmid containing NIC96-ProA expressed under the control of a galactose-inducible promoter (pGAL-NIC96-ProA). Asynchronous cultures were shifted to 13.5°C for 18 h (18h); at this point, the ndc1-1 culture contained 80% large budded cells. NIC96-ProA expression was induced for 12 h (30h), followed by a glucose chase for 4 h to determine whether Nic96p-ProA was properly assembled into NPCs (34h). Nic96p-ProA localization was visualized using indirect IF microscopy with an FITC-conjugated secondary antibody (see Materials and Methods).