Evolved sequence features within the intrinsically disordered tail influence FtsZ assembly and bacterial cell division

Megan C. Cohan; Ammon E. Posey; Steven J. Grigsby; Anuradha Mittal; Alex S. Holehouse; Paul J. Buske; Petra A. Levin; Rohit V. Pappu

The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. . https://doi.org/10.1101/301622 doi: bioRxiv preprint Immunoblotting: Immunoblotting was performed as described previously (Weart and Levin, 2003) . Cells were grown overnight in Luria-Bertani (LB) medium at 37°C with 100 µg/mL ampicillin, 100 µg/mL spectinomycin, 5 µg/mL chloroamphenicol and 0.5% xylose. They were then back-diluted 1:100 and grown in 0.5% xylose until the cells reached mid-log phase. The cells were then washed twice with LB, diluted 1:100, and grown to mid-log phase in 0.1 mM IPTG. The cells were lysed with lysozyme and detergent. Loading was normalized to the OD 600 at sampling. The blot was probed using affinity-purified polyclonal rabbit anti-FtsZ antibodies and goat anti-rabbit antibodies conjugated to horseradish peroxidase (Jackson ImmunoResearch Laboratories). Immunoblots were developed using the ECL Western Blotting detection reagents (GE Healthcare) and visualized with the luminescent image analyzer ImageQuant LAS 4000 mini (GE Healthcare).