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. 2010 Nov 8;107(47):20364–20369. doi: 10.1073/pnas.1011069107

Fig. 2.

Fig. 2.

The sororin-binding site remains after DNA replication is complete. (A) Experimental scheme to test whether sororin binds chromatin only during DNA replication or after replication is complete. Aliquots of a nuclear assembly reaction in sororin-depleted extract were supplemented with in vitro-translated sororin at indicated times after the start of the reaction (t = 0, 30, 60, 90, or 120′, Upper). Following sororin addition, chromatin fractions were isolated immediately (0′) and at 7.5 and 15′ and assayed by immunoblot for cohesin (Smc3) and sororin (B). (C) DNA replication was assessed in extract depleted for sororin (as used in B) and mock-depleted extract. (D) Chromatin was collected from all reactions shown in (B) at 150′ after the start of the experiment and analyzed for cohesin (Smc3) and sororin by immunoblot. Ongoing replication is not required for sororin recruitment. (E) Nuclei were added to a sororin-depleted extract and the reaction was allowed to proceed for 60′. Aphidicolin (or DMSO control) was added. After 10 min, in vitro-translated sororin was added to both extracts and the levels of chromatin-associated Smc3 and sororin were assessed by immunoblot. (F) DNA replication levels over a 30-min period from the time of sororin addition.