Biochemical studies were carried out to determine the effect of χ mutation on T4 DNA synthesis. The rate and final extent of DNA synthesis are almost the same with T4D- and T4χ-infected cells, although the burst size of T4χ is about one-sixth that of the wild type. The DNA synthesis of T4χ-infected cells is more readily inhibited by mitomycin C than is that of T4 wild type. When mitomycin C was added during active phage growth, DNA synthesis of T4χ halted almost immediately. T4 DNA polymerases isolated from χ + - and χ-infected cells, however, exhibit no difference with regard to their sensitivities to mitomycin C, priming activities with alkylated or ultraviolet light-irradiated templates and other enzymatic properties.