EFFECT OF BROMODEOXYURIDINE ON RADIATION-INDUCED DNA-DAMAGE AND REPAIR BASED ON DNA FRAGMENT SIZE USING PULSED-FIELD GEL-ELECTROPHORESIS

We have used biphasic linear ramping pulsed-field gel electrophoresis (PFGE) to understand the effect of incorporation of bromodeoxyuridine (BrdUrd) on radiation-induced DNA damage and repair. This technique pe rmits a determination of the fragment size distribution produced immed iately after irradiation as well as during the repair period. We found that incorporation of BrdUrd increased the induction and decreased th e repair of radiation damage, The fragment size distribution was consi stent with a random breakage model, When we found that significantly m ore damage was detected after irradiation of deproteinized DNA compare d to intact cells, we studied the effects of BrdUrd incorporation on t he radiation response of cells or DNA at various phases of preparation for electrophoresis: cells adherent to the culture dish (A), trypsini zed cells (B), agarose-embedded cells (C) and deproteinized DNA (D). A lthough there was a general tendency to detect more damage when irradi ation was performed later in the preparation process, steps B and C we re the only successive steps which were significantly different. These findings demonstrate that incorporation of BrdUrd randomly increases the induction of radiation damage and decreases its repair at the leve l of 200 kbp to 5 Mbp fragments, Furthermore, they confirm that the am ount of damage detected depends upon the conditions of the cells or DN A at the time of irradiation. (C) 1995 by Radiation Research Society