The indicated Deinococci were inoculated into TGY medium at 106 CFU/ml (CFU - colony-forming unit) and grown at 32oC to the early plateau phase (108 cells/ml). E. coli was similarly pre-grown in LB medium at 37oC. Cells were then irradiated without change of broth on ice with 60Co at 10 kGy/h. At the indicated doses, viable cell counts were determined by plating appropriate cell dilutions for CFU on TGY agar at 32oC. E. coli was allowed to recover on LB medium at 37oC. For a typical culture of D. radiodurans, cell grouping will vary with growth stage.
Cells grow as clusters of two cells (diplococci) in the early stages of growth, and as four cells (tetracocci) in the late stages [Thornley et al., 1965]. The slightly greater resistance of tetracocci compared to dipplococci at doses > 10 kGy has been reported previously [Venkateswaran et al., 2000] and is believed to be the result of cell grouping. Whereas four cells of a tetracoccus need to be killed to eliminate a colony-forming unit (CFU), only two cells of a diplococcus need to be killed, as outlined in cell-grouping.