DNA repair synthesis induced by methyl methanesulfonate in preconditioned HeLa cells in which DNA replicative synthesis had been highly suppressed was inhibited by aphidicolin (an inhibitor of DNA polymerases alpha and delta) and dideoxythymidine (ddThR, an inhibitor of DNA polymerase beta). Incomplete repair patches sensitive to exonuclease III were accumulated in the presence of aphidicolin while not in the presence of ddThR. These patches were comopleted by the combined action of Klenow fragment and T4 DNA ligase, indicating that the single-stranded gaps were formed during the repair synthesis. Moreover, ddThR had little effect on the repair synthesis in the presence of aphidicolin. Thus, the results suggest that the single-stranded gaps may be sealed first by aphidicolin-sensitive polymerase followed by ddThR-sensitive DNA polymerase on the same site of the repair patch.