| dc.description.abstract | There are many forms of DNA damage. One of these, DNA-metal cross-links are
particularly problematic as they not only affect the tautomeric structure of DNA, but can
also inhibit enzymatic activity and block amplification with DNA polymerases. Since the
inability to amplify DNA can affect many fields of genetic research, it is necessary to
find ways to identify this damage and repair the DNA. Copper is one metal that can form
metal-DNA cross-links and can seriously affect the recovery and analysis of degraded
DNA from forensic or archaeological material. In this research DNA-copper cross-links
were generated for the development of a repair method and applied to an archaeological
sample. The DNA-copper adducts were identified through the use of gas
chromatography-mass spectrometry on several templates of different complexity: single
deoxyribonucleotides, a synthetic 22 base pair double stranded DNA fragment, modern
amplified DNA, and an ancient extract with naturally occurring copper cross-links. A
number of chemicals were considered for direct reversal repair of copper-DNA cross-
links of which ethylenediamine was successful. Treatment of all the templates with
ethylenediamine resulted in the repair of the nucleobase, specifically guanine which is the
most susceptible to copper cross-link formation. The success of the direct reversal repair
was verified using GC-MS based on expected retention time and the identification of ion
fingerprints. The amount of copper-DNA adducts measured in each template varied
greatly as did the success of the direct reversal repair although repair was evident in all
samples. | en_US |
