Infection of duck embryo fibroblasts by Marek's disease herpesvirus (MDHV), strain GA, led to the induction of a novel DNA polymerase. This novel DNA polymerase, designated MDHV-induced DNA polymerase, could be distinguished from the DNA polymerase activities of uninfected duck embryo fibroblasts by its chromatographic behavior on phosphocellulose, by its sedimentation coefficient, and by its catalytic properties. The characteristics of MDHV-induced DNA polymerase which had been purified by phosphocellulose chromatography were investigated. The sedimentation coefficient of the enzyme, as determined by sucrose density gradient centrifugation in the presence of 0.25 M KCl, was 5.9S. From this sedimentation coefficient, the molecular weight of MDHV-induced DNA polymerase was estimated to be 100,000. MDHV-induced DNA polymerase could not effectively use either poly(dA).oligo(dT)(12-18) or poly(dC).oligo(dG)(12-18) as a template-primer. The DNA polymerases from uninfected duck embryo fibroblasts could use these synthetic template-primers. MDHV-induced DNA polymerase also could not use poly(rA).oligo(dT)(12-18) or poly(rC).oligo(dG)(12-18) as template-primers or oligo(dT)(12-18) as a primer, indicating that it was not a polymerase of the type R-DNA polymerase, reverse transcriptase, or a terminal nucleotidyl transferase. In vitro synthesis of DNA by MDHV-induced DNA polymerase was markedly inhibited by the addition of (NH(4))(2)SO(4) to the reaction mixture.