Analysis of UV-induced DNA photoproducts by 32P-postlabelling.
UV-induced cyclobutane dimers and 6-4 photoproducts, containing an unmodified nucleotide at the 5'-position were released from DNA by means of digestion with DNase I, snake venom phosphodiesterase and prostatic acid phosphatase. The enzymes were deactivated by proteinase K followed by ethanol precipitation. The products were phosphorylated by polynucleotide kinase and [gamma-32P]ATP. The TLC system used for the analysis enables separation of the different photoproducts and detection at a fmol level. T4 endonuclease treatment was applied to confirm the positions of cyclobutane dimers.
Center for Nutrition and Toxicology, Karolinska Institute, Novum, Huddinge, Sweden.
V J
VJ
Bykov
R
R
Kumar
A
A
Försti
K
K
Hemminki
D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D004247:DNA / Q000032:analysis / Q000378:metabolism / Q000528:radiation effects*; D018736:DNA Adducts / Q000032:analysis* / Q000302:isolation & purification / Q000528:radiation effects; D006801:Humans; D007553:Isotope Labeling; D010761:Phosphorus Radioisotopes; D010777:Photochemistry; D011740:Pyrimidine Dimers / Q000032:analysis; D014466:Ultraviolet Rays
D018736:DNA Adducts; D010761:Phosphorus Radioisotopes; D011740:Pyrimidine Dimers; D004247:DNA