Thymidine labeling of pyrimidine isostichs from human lymphocyte DNA during repair after damage with N-acetoxy acetylaminofluorene or nitrogen mustard

Evidence for the faithfulness of DNA repair in mammalian cells is presented. Human peripheral blood lymphocytes with very low background levels of DNA synthesis were prepared by using populations that are 99.8 to 99.9% free of dividing cells and by further supressing DNA synthesis in these with hydroxyurea. These cells were damaged with nitrogen mustard or the proximate carcinogen N-acetoxy-2-acetylaminofluorene and allowed to undergo repair in the presence of tritiated thymidine. Pyrimidine isostichs were prepared from the DNA by formic acid and diphenylamine digestion and were separated on DEAE-cellulose. The distribution of counts in the pyrimidine isostichs from treated and control cells was found to be the same. The latter served as a measure of the frequency of various isostichs present in the normal DNA. Thus, it is suggested that during repair, bases are incorporated in a fashion which approximates their distribution in the original DNA.