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. 2004 Mar 29;164(7):1009–1020. doi: 10.1083/jcb.200310061

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Copyright © 2004, The Rockefeller University Press

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Figure 2. — Identification of the Golgi protein GM130 as a binding partner for YSK1. (A) Bait constructs comprising the full-length human GM130 splice variant rescued from the library screen, full-length rat GM130 or the indicated deletion constructs of rat GM130 were tested against full-length human YSK1 for the ability to grow on selective medium (QDO), compared with nonselective medium (−LW) in the yeast two-hybrid system. Lighter colony color on QDO is an indicator of a strong signal. Boxed regions indicate the p115 binding site (red) and predicted coiled-coil domains (green). (B) His-tagged YSK1 was incubated with GST-tagged fragments of rat GM130, golgin45, or GST and recovered on glutathione-agarose. Recovered complexes were analyzed by Western blotting. (C) HeLa cells depleted for lamin A or GM130 for 67 h using siRNA were stained and Western blotted for GM130, YSK1, and α-tubulin. Bar, 10 μm.