0000000000368076
AUTHOR
Maja Fafanđel
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…
Molecular response to TBT stress in marine sponge Suberites domuncula: proteolytical cleavage and phosphorylation of KRS_SD protein kinase
Abstract Marine sponges as sessile filter feeders are inevitably under a constant influence of changes in their environment. Mediation of extracellular signals and regulation of cellular response to environmental stress is a key function of cellular protein kinases. Expression, proteolytical cleavage and phosphorylation of stress-responsive KRS_SD protein kinase, in control and tributyl-tin (TBT) treated sponges were investigated. In control sponge, two KRS_SD proteins were expressed: KRS_SD1 (54 kDa) corresponding to KRS_SD calculated molecular weight, and KRS_SD2 (50 kDa). Exposure of sponges to TBT resulted in alteration of KRS_SD1 and KRS_SD2 expression levels and their phosphorylation …
A Microplate Assay for DNA Damage Determination (Fast Micromethod)in Cell Suspensions and Solid Tissues
Abstract A rapid and convenient procedure for DNA damage determination in cell suspensions and solid tissues on single microplates was developed. The procedure is based on the ability of commercially available fluorochromes to interact preferentially with dsDNA in the presence of ssDNA, RNA, and proteins at high pH (>12.0), thus allowing direct measurements of DNA denaturation without sample handling or stepwise DNA separations. The method includes a simple and rapid 40-min sample lysis in the presence of EDTA, SDS, and high urea concentration at pH 10, followed by time-dependent DNA denaturation at pH 12.4 after NaOH addition. The time course and the extent of DNA denaturation is followed …