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. Author manuscript; available in PMC: 2015 Feb 3.
Published in final edited form as: Curr Biol. 2014 Jan 16;24(3):287–292. doi: 10.1016/j.cub.2013.11.050

Figure 1. SMC complexes are required for origin segregation.

Figure 1

(A) Heterogeneous nucleoid morphologies and cell sizes in the SMC null mutant grown under permissive conditions. Representative images of Δsmc (strain BWX2208) grown at 22°C in LB, casein hydrolysates (CH), minimal medium (S750) supplemented with glucose or sorbitol. The nucleoids (red) and the origins (green) were visualized with HbsU-GFP and TetR-mCherry bound to a tetO array inserted adjacent to the origin. (B) Spot-dilutions of indicated temperature-sensitive mutants grown on LB-agar plates at permissive (30°C) and restrictive (42°C) temperatures. Representative images of DAPI-stained nucleoids (red) and origin foci (green) in wild-type cells (BWX811) grown in CH medium at 37°C (C); smcts (BWX2090) grown in CH medium at 30°C (D) and after shift to 42°C (E–G); scpBts (BWX2092) grown in CH medium for 1.5 h at 42°C (H); smc-ssrA (BWX1497) 1.5 h after induction of SspB grown in CH medium at 37°C (I). 85% (n=1273) of the smcts cells; 81% (n=1336) of the scpBts cells; and 85% (n=1392) of smc-ssrA cells had a single origin focus or cluster of foci at hour 1.5. Membranes (false-colored blue) were stained with FM4-64. Yellow carets highlight septum formation on top of the nucleoid. (J) Origin (green), replisome foci (red, DnaX-YFP), and DAPI-stained nucleoids (blue) in the smc-ssrA strain (BWX1771) 1.5 h after induction of SspB in CH medium at 37°C. Scale bars are 4 μm. Strains harboring wild-type copies of smc, scpB, or parC with a linked antibiotic resistance gene displayed normal chromosome organization and segregation when grown at 42°C (Figure S1F). See also Figure S1.