Cell cycle analysis

Cell cycle analysis of Escherichia coli cells

C period = the time for a round of chromosome replication

D period = the time between the end of a round of chromosome replication and cell division

1.Determination of initiation age (a_i) and C+D:

From flow cytometry analysis of cells treated with rifampicin and cephalexin (run-out histogram) the proportions of cells that had not initiated replication at the time of drug action (4-origin-cells, streaked) and cells that had initiated (8-origin-cells) can be estimated.The initiation age (a_i) can be found from the theoretical age distribution described by this formula,

F = 2 - 2^{((τ-a_i)/τ)}

where F is the fraction of cells that had not initiated and τ is the generation time, or from the estimated graph of the theoretical age distribution (streaked portion).

The C+D period is estimated from the initiation age (a_i), the generation time (τ) and the number of generations spanned per cell cycle.

Example:

4-origin-cells: 23 %

Generation time (τ): 27 min

Initiation age (a_i): 5 min

2. Determination of the C and D periods:

The C period is found from the oriC/terC ratio obtained by Southern blot analysis and the generation time (τ):

oriC/terC=2^C/τ

The D period is found from the C+D and C period:

D = (C+D) - C

Example (continues):

C period calculated from the oriC/terC ratio: 49 min

D period = (C+D) – C D period = 76 min – 49 min = 27 min