Cell cycle analysis

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A theoretical exponential DNA histogram can be drawn to check whether the obtained values fit with the experimental data. From the C+D period the DNA content of the cells at different time points in the cell cycle can be calculated.
A theoretical exponential DNA histogram can be drawn to check whether the obtained values fit with the experimental data. From the C+D period the DNA content of the cells at different time points in the cell cycle can be calculated.
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Example:

Revision as of 04:28, 18 August 2008

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 (ai) 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 (ai) can be found from the theoretical age distribution described by this formula,

F = 2 - 2((τ-ai)/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 (ai), 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 (ai): 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=2C/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










3. The theoretical exponential DNA histogram:

A theoretical exponential DNA histogram can be drawn to check whether the obtained values fit with the experimental data. From the C+D period the DNA content of the cells at different time points in the cell cycle can be calculated.

Example:

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