Specific detection of cyclobutane pyrimidine dimers in phytoplankton DNA by a non-radioactive assay based on T-4-endonuclease V digestion

The effect of artificial and natural UV irradiation on DNA in marine phytop lankton Isochrysis galbana monoculture was investigated. The presence of cy clobutane pyrimidine dimers (CPDs) in unlabelled I. galbana DNA was detecte d by a non-radiometric alkaline filter elution assay after T-4-endonuclease V digestion. The quantity of CPDs was estimated by alkaline agarose gel el ectrophoresis. Precise determination of the amount of DNA in the presence o f I. galbana pigments was achieved by oxazole yellow homodimer (YOYO) dye. T-4-endonuclease V-sensitive sites frequency (ESS/kb), measured after expos ure to 2-40 kJ m(-2) of artificial UV light, increased in a dose-dependent manner. Twelve hours after irradiation cell culture growth was disrupted, a nd 50% of initial DNA damage in the cells was observed. After 1 h of sunlig ht exposure, the incidence of CPDs increase significantly. Prolonged exposi tion to sunlight decrease CPDs incidence due to efficiency of I. galbana DN A repair mechanisms, The presence of water-soluble crude oil fraction (WSOF ) affected DNA repair efficiency resulting in accumulation of CPDs in I. ga lbana DNA. (C) 2001 Elsevier Science B.V. All rights reserved.