Figure 1.

The cell cycle and experimental approach. (A) At the slowest growth rates, the chromosome rests in B/G1 before the replication period C/S starts. This is as in eukaryotes. At faster growth rates, however, replication is continuous, and the B/G1 period disappears. The growth condition in our study is such that a new round of replication is initiated just before the previous one is terminated, as illustrated in the right panel. (B) We constructed a set of strains with two independent fluorescent chromosome markers. One type of parS sequence (pMT1 parS) was inserted at either 22′ or 98′ to serve as a reference marker, and another type of parS sequence (P1 parS) was inserted at one of the 13 positions on the map. Each strain has a pair of parS sequences and a matching pair of ParB proteins fused to CFP and YFP (CFP-P1DOParB and yGFP-pMT1D23ParB). (C) Cell size distributions of a population of cells growing under steady-state growth conditions in this study (the color of the lines matches the chromosome marker in B). Typically, we analyzed 10⁴–10⁵ cells per strain. Approximately 10⁶ cells were studied in this work. This was sufficient to obtain reproducible actual cell size distributions from each strain. Both the tails of the distributions and the Gaussian-looking dividing cell size distributions indicate significant intrinsic stochasticity of growth and cell cycle regulation. The black dashed line is the theoretical distribution in the absence of cell-to-cell variation. The inset graph also illustrates the stochasticity of the cell cycle. There is a considerable variation in the length of dividing cells, defined in the measurement program as cells with a central constriction with <90% of the average cell diameter of the rest of the cell. (D) A typical field of view of one of the strains (22′ and 54.2′ markers) grown under overlapping cell cycle condition is shown. Due to multifork replication, many cells contain multiple fluorescent foci per locus.