Figure 6.

Condensin binding to chromatin at the rDNA locus. (A) A significant portion of Smc2p is chromatin-bound throughout the cell cycle. Chromatin binding of Smc2p and Hmo1p (a reference chromatin protein) throughout the cell cycle determined by chromatin solubilization assay (Liang and Stillman 1997). Strain YPH499bp2 was grown asynchronously or arrested in G1, S, or G2/M phase. Lysates were centrifuged to yield a chromatin-unbound fraction (supernatant 2 according to Liang and Stillman 1997) and a chromatin-bound fraction (pellet 3). Unfractionated lysate (T), supernatant (S), and pellet (P) were probed with antibodies to Smc2p and Hmo1p (Lu et al. 1996). (B) Agarose gel electrophoresis of ChIP analysis of the rDNA repeat in the asynchronously growing YPH499bp2 strain with PCR probes shown in C. (D) Quantification of the Smc2p-HA ChIP in strain YPH499bp2 arrested with α-factor and nocadazole. The PCR signals were quantified using ImageQuant (Molecular Dynamics). All volume scans were normalized to the signal from probe 12 that was only marginally above background (Relative ChIP) and to the corresponding PCR on the input DNA. (E) Quantification of the Ycs5p-HA ChIP in strain YPH499bp5 arrested with α-factor and nocadazole. Analysis was as in Fig. 5 D.