Radiolysis of Chromatin Extracted from Cultured Mammalian Cells: Production of Alkali-labile Strand Damage in DNA

Summary

Chromatin has been isolated from cultured Chinese-hamster lung fibroblasts as an expanded aqueous gel. The DNA in isolated chromatin has been examined by sedimentation on alkaline sucrose gradients. The average molecular weight of the DNA has been determined to be 50 million. γ-irradiation of isolated chromatin degrades the DNA to lower molecular weight. The yield of single-strand breaks in the DNA is 0·02 single-strand breaks per krad-10⁶ dalton, calculated from a dose-range of 1–400 krad and covering a DNA molecular weight range of 2 × 10⁷−1·4 × 10⁵. There is a considerable difference in the efficiency of the formation of single-strand breaks in DNA irradiated as isolated chromatin compared with chromatin irradiated in whole cells before isolation. For isolated chromatin, values of 6 eV per break have been calculated compared with about 80 eV per break for chromatin irradiated in whole cells, which suggest a large contribution from indirect action by aqueous radicals in isolated chromatin.