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DETECTION OF LOW DOSE RADIATION INDUCED DNA DAMAGE USING TEMPERATURE DIFFERENTIAL FLUORESCENCE ASSAY
Citation:
Rogers, K R., A. B. Apostol, S. J. Madsen, AND C. W. Spencer. DETECTION OF LOW DOSE RADIATION INDUCED DNA DAMAGE USING TEMPERATURE DIFFERENTIAL FLUORESCENCE ASSAY. Analytical Chemistry 71(19):4423-4426, (1999).
Impact/Purpose:
The overall objective of this task is to develop rapid, cost-effective and scientifically sound techniques for measuring chemically induced DNA damage. This method is expected to provide the Agency with rapid, sensitive, and simple techniques that can be used among a panel of methods to determine the genotoxic potential of polluted samples.
Description:
A rapid and sensitive fluorescence assay for radiation-induced DNA damage is reported. Changes in temperature-induced strand separation in both calf thymus DNA and plasmid DNA (puc 19 plasmid from Escherichia coli) were measured after exposure to low doses of radiation. Exposures of between 0.004 and 1 Gy were measured with doses as low as 0.008 Gy yielding significant responses. The double-strand, sensitive dye PicoGreen was used as an indicator of DNA denaturation. Calibration plots indicate that fluorescence changes corresponding to amounts as low as 1 ng of double stranded DNA (10(6) copies for plasmid puc 19) are detected by this method.
The U.S. Environmental Protection Agency, through its Office of Research and Development (ORD), funded this research through a competitive internal grant (to K.R.). It has been subject to the EPA's peer and administrative review and has been approved for publication. Mention of trade names or commercial products does not constitute endorsement or recommendation by EPA for use.