Specific detection of cyclobutane pyrimidine dimers in phytoplankton DNA by a non-radioactive assay based on T4-endonuclease V digestion.
The effect of artificial and natural UV irradiation on DNA in marine phytoplankton Isochrysis galbana monoculture was investigated. The presence of cyclobutane pyrimidine dimers (CPDs) in unlabelled I. galbana DNA was detected by a non-radiometric alkaline filter elution assay after T4-endonuclease V digestion. The quantity of CPDs was estimated by alkaline agarose gel electrophoresis. Precise determination of the amount of DNA in the presence of I. galbana pigments was achieved by oxazole yellow homodimer (YOYO) dye. T4-endonuclease V-sensitive sites frequency (ESS/kb), measured after exposure 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, and 50% of initial DNA damage in the cells was observed. After 1 h of sunlight exposure, the incidence of CPDs increase significantly. Prolonged exposition to sunlight decrease CPDs incidence due to efficiency of I. galbana DNA repair mechanisms. The presence of water-soluble crude oil fraction (WSOF) affected DNA repair efficiency resulting in accumulation of CPDs in I. galbana DNA.
Fafandel
M
M
Rud er Bosković Institute, Center for Marine Research, Rovinj, Croatia. maja@cim.irb.hr
Bihari
N
N
Krajcar
V
V
Müller
W E
WE
Zahn
R K
RK
Batel
R
R
D001681:Biological Assay; D004249:DNA Damage; D004260:DNA Repair; D043224:Deoxyribonuclease (Pyrimidine Dimer); D004706:Endodeoxyribonucleases / Q000378:metabolism*; D010578:Petroleum / Q000009:adverse effects*; D010839:Phytoplankton / Q000737:chemistry / Q000235:genetics*; D011740:Pyrimidine Dimers / Q000032:analysis*; D013472:Sunlight; D014466:Ultraviolet Rays / Q000009:adverse effects; D014764:Viral Proteins
D010578:Petroleum; D011740:Pyrimidine Dimers; D014764:Viral Proteins; D004706:Endodeoxyribonucleases; C470698:endonuclease V, phage T4; D043224:Deoxyribonuclease (Pyrimidine Dimer)