MONITORING ULTRAVIOLET-B-INDUCED DNA-DAMAGE IN INDIVIDUAL DIATOM CELLS BY IMMUNOFLUORESCENT THYMINE DIMER DETECTION

We developed a method to investigate the effect of ultraviolet-B radia tion (UVBR) on the formation of thymine dimers in microalgal DIVA that can be used for both laboratory and in situ research. Antibody labeli ng of dimers was followed by a secondary antibody (fluorescein isothio cyanate) staining to allow visualization of DNA damage with flow cytom etry or fluorescence microscopy. Thymine dimer-specific fluorescence i n nuclear DNA of the marine diatom Cyclotella sp. was linearly related to the UVBR dose. Simultaneous measurements of cellular DNA content s howed that the vulnerability of G2 cells to DNA damage did not differ significantly from the vulnerability of G1 cells. The formation and re moval of thymine dimers in Cyclotella sp. cells was monitored for 3 co nsecutive days at two realistic UVBR irradiance bevels. Thymine dimers were removed within 24 h when exposed to a saturating photosynthetica lly active radiation intensity following the UVBR treatment. This new method allows the study of UVBR-induced DNA damage on a cell-to-cell b asis. It is also feasible for field studies because cells remain intac t and can be recognized readily after antibody treatment.