Fig. 3.
Fig. 3. Changes of [3H]ara-C incorporation into DNA by dFdC. Exponentially growing CCRF-CEM cells were treated without (A), or with 8 nmol/L (B) and 80 nmol/L (C) nonradioactive dFdC, simultaneously adding 0.6 nmol/L [3H]ara-C (1.5 × 10−2 μCi/mL) in the presence of 1.0 μmol/L BrdUrd and 10 nmol/L FdUrd for 13 hours. DNA isolation and CsCl ultracentrifugation were performed as described in Materials and Methods. Fractions were collected for the measurement of radioactivity (○) and UV absorption at 260 nm (×). DNA-specific activity (dpm/UV absorbance unit) from [3H]ara-C labeling of cells were calculated. The high-density peaks between fractions 45 and 49 (A, B), or fractions 40 and 44 (C) were designated as representing replication. To calculate the specific activities of replicating DNA, the total dpm in these fractions was divided by the sum of the UV absorption at 260 nm in each set of fractions. The DNA specific activities in fractions located at the replication peaks were 20,432 dpm/absorbance unit for ara-C alone (A); 32,452 dpm/absorbance unit for ara-C in the presence of 8 nmol/L dFdC (B); and 48,969 dpm/absorbance unit for ara-C in the presence of 80 nmol/L dFdC (C).

Changes of [3H]ara-C incorporation into DNA by dFdC. Exponentially growing CCRF-CEM cells were treated without (A), or with 8 nmol/L (B) and 80 nmol/L (C) nonradioactive dFdC, simultaneously adding 0.6 nmol/L [3H]ara-C (1.5 × 10−2 μCi/mL) in the presence of 1.0 μmol/L BrdUrd and 10 nmol/L FdUrd for 13 hours. DNA isolation and CsCl ultracentrifugation were performed as described in Materials and Methods. Fractions were collected for the measurement of radioactivity (○) and UV absorption at 260 nm (×). DNA-specific activity (dpm/UV absorbance unit) from [3H]ara-C labeling of cells were calculated. The high-density peaks between fractions 45 and 49 (A, B), or fractions 40 and 44 (C) were designated as representing replication. To calculate the specific activities of replicating DNA, the total dpm in these fractions was divided by the sum of the UV absorption at 260 nm in each set of fractions. The DNA specific activities in fractions located at the replication peaks were 20,432 dpm/absorbance unit for ara-C alone (A); 32,452 dpm/absorbance unit for ara-C in the presence of 8 nmol/L dFdC (B); and 48,969 dpm/absorbance unit for ara-C in the presence of 80 nmol/L dFdC (C).

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