Abstract
It is known that DNA lesions induced by ionizing radiation and chemicals can cause mutation and carcinogenesis. In particular, "clustered damage" site, that is a DNA region with multiple lesions within a few helical turns, is believed to hardly be repaired. This type of damage is considered to be induced around high-LET radiation tracks and at track-end of secondary electron. However, details of the clustered damage sites are not known. In order to get experimental information about the hypothetical damage, we have developed an analytical method for measuring the degree of dispersion of distances between DNA lesions using fluorescence resonance energy transfer (FRET). Complementary two 31-mer single-strand DNA with an AP (apurinic/apyrimidinic)-site at each center nucleotide were synthesized to obtain relationship between FRET efficiency and AP-site- AP-site distance. One DNA strand with an AP-site was labeled with a donor or fluorescent probe at the lesion, and another strand was labeled with an acceptor. As a result of fluorescence spectral analyses with annealing these strands, fluorescence intensity from acceptor increased with proceeding the annealing. D-A distance calculated from the FRET efficiency was 4 nm, which was reasonable with the estimated one taking the diameter of B-form DNA duplex and length of each probe into account. Now we have tried to apply the FRET method to plasmid DNA irradiated with some radiation qualities. The results and the perspective will be discussed.
