Cell Harvest and Immunoblot Analysis

Cells were harvested from 2 mL of the synchronous culture at the indicated time points by centrifugation. Cells were then resuspended in the SDS-PAGE sample buffer (50 mM Tris, pH 6.8, 6% 2-mercaptoethanol, 2% sodium dodecyl sulfate, 10% glycerol). Protein content was quantified with XL-Bradford (APROSCIENCE), and equal amounts of the total protein were subjected to immunoblot analysis. Immunoblotting was performed as described in Miyagishima et al.^{34 (link)} using 12% SDS-polyacrylamide gels. The anti-FtsZ, anti-nucleomorph H2A or anti-H3S10Ph^{35 (link), 36 (link)} antibodies was diluted to 1:1000. The primary antibodies were detected by HRP-conjugated goat anti-rabbit antibody (Thermo Scientific) diluted to 1:20,000.