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. 2014 Aug 18;289(40):27418–27431. doi: 10.1074/jbc.M114.569608

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© 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

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FIGURE 7. — Mutations in Smc5-3KE do not affect Smc5-Smc6 complex formation and protein abundance. A, Smc5 (or Smc5-3KE) and Smc6 proteins were immunoprecipitated (IP) from whole cell extract using anti-StreptagII (STII) and anti-Myc antibodies to evaluate the association between Smc5-STH (or Smc5-3KE) and Smc6-13xMYC by Western blotting with the reciprocal antibodies, as indicated. Cell lysate (Lysate) and beads alone (Beads) were used as controls. The strains used in this experiment are as follows. No-tag, SMC5/SMC5; SMC6/SMC6; WT, SMC5-STH/SMC5; SMC6-13MYC/SMC6; 3KE, smc5-3KE-STH/SMC5; SMC6-13MYC/SMC6. B, Western blotting analysis of Smc5-3KE protein abundance in yeast upon α-factor arrest and release into a synchronous cell cycle at 18 °C, using a StreptagII antibody. Pgk1 protein was used as a loading control.