Method for assaying clustered DNA damages
AssigneeBrookhaven Science Associates, LLC
InventorBetsy M. Sutherland
Life of this patent
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Abstract
Disclosed is a method for detecting and quantifying clustered damages in DNA. In this method, a first aliquot of the DNA to be tested for clustered damages with one or more lesion-specific cleaving reagents under conditions appropriate for cleavage of the DNA to produce single-strand nicks in the DNA at sites of damage lesions. The number average molecular length (Ln) of double stranded DNA is then quantitatively determined for the treated DNA. The number average molecular length (Ln) of double stranded DNA is also quantitatively determined for a second, untreated aliquot of the DNA. The frequency of clustered damages (Φc) in the DNA is then calculated.
Claims27
A method for detecting and quantifying clustered damages in DNA, the method comprising:
- a) providing a sample containing DNA to be assayed for clustered damages;
- b) contacting a first aliquot of the DNA to be tested for clustered damages with one or more lesion-specific cleaving reagents under conditions appropriate for cleavage of the DNA to produce single-strand nicks in the DNA at sites of damage lesions;
- c) quantitatively determining the number average molecular length (Ln) of double stranded DNA in the lesion-specific cleaving reagent-treated first aliquot;
- d) quantitatively determining the number average molecular length (Ln) of double stranded DNA in a second aliquot of the DNA to be tested for clustered damages, the second aliquot being untreated by lesion-specific cleaving reagents;
- e) calculating the frequency of clustered damages (Φc) in the DNA to be assayed for clustered damages using the following equation:
- Φc=1/Ln(+enzyme)−1/Ln(−enzyme)
- wherein Ln(+enzyme) is the number average molecular length of double stranded DNA determined for the lesion-specific leaving reagent-treated DNA determined in step c), and Ln(−untreated DNA in step d).
Dependencies are inferred from the claim text.