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. 2005 Apr 11;169(1):61–71. doi: 10.1083/jcb.200411056

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Copyright © 2005, The Rockefeller University Press

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Figure 6. — Xnf7 associates with the core APC. (A) Sucrose gradient cosedimentation of Xnf7 and APC subunits. CSF egg extract was fractionated on a 10–40% sucrose gradient. Fractions were collected and analyzed by SDS-PAGE and immunoblotting for the indicated proteins. Asterisks indicate the fractions used in the immunoprecipitations in B. (B) Xnf7 coimmunoprecipitates with Cdc27 from sucrose gradient fractions. Purified anti-Xnf7 antibodies or purified preimmune IgG were coupled to protein A–Sepharose and incubated with sucrose gradient fractions 4, 10, 11, and 18 (1 h at 4°C). Beads were washed and bound proteins were analyzed by SDS-PAGE and immunoblotting with the indicated antibodies. (C) Xnf7 coimmunoprecipitates with Cdc27 during CSF arrest and during exit from arrest. Purified anti-Xnf7 antibodies were coupled to protein A–Sepharose and incubated with aliquots of CSF extracts removed at the indicated times after CaCl₂ addition (1 h at 4°C). Beads were washed and bound proteins were analyzed by SDS-PAGE and immunoblotting with anti-Cdc27 antibodies.