Abstract
We report a simple and efficient colorimetric method to screen large numbers of bacterial strains for UV- and X-radiation sensitivity. We used reference radiation-sensitive and control strains of Escherichia coli K-12 to compare our colorimetric method to a standard clonogenic plating method. Our colorimetric method was as accurate as the standard method and was superior in terms of savings in supplies and man-hours.
Highlights
Studies on radiation-sensitive mutants of bacteria, e.g., Escherichia coli, have been invaluable in elucidating mechanisms of DNA repair (Augusto-Pinto et al 2003; Friedberg et al 2006)
We developed a resazurin-based assay using 96-well microtiter plates to reduce the very significant time and expense normally associated with traditional clonogenic assays for radiation sensitivity
We report on the reliability of our colorimetric assay by testing (i) the radiation dosimetry among the 96 wells of a microtiter plate, (ii) the resazurin color change for reference radiation-sensitive and -resistant strains of E. coli after both UV- and X-irradiation, and (iii) the sensitivity of our colorimetric assay in comparison with a clonogenic assay for differentiating a set of reference E. coli strains based on their radiation sensitivities
Summary
Studies on radiation-sensitive mutants of bacteria, e.g., Escherichia coli, have been invaluable in elucidating mechanisms of DNA repair (Augusto-Pinto et al 2003; Friedberg et al 2006). It is common that one needs to screen, in time-consuming and expensive fashion, large numbers of strains to find or quantitate a desired phenotype. With this goal, we developed a resazurin-based assay using 96-well microtiter plates to reduce the very significant time and expense normally associated with traditional clonogenic (plating) assays for radiation sensitivity. We report on the reliability of our colorimetric assay by testing (i) the radiation dosimetry among the 96 wells of a microtiter plate, (ii) the resazurin color change for reference radiation-sensitive and -resistant strains of E. coli after both UV- and X-irradiation, and (iii) the sensitivity of our colorimetric assay (an indirect measure of cell survival) in comparison with a clonogenic assay (a more traditional and direct measure of cell survival) for differentiating a set of reference E. coli strains based on their radiation sensitivities. We report an estimate of the cost savings in using the colorimetric assay vs. the clonogenic assay
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