Probing DNA Replication Mechanisms with the T4 Bacteriophage In Vitro System

Abstract

We have previously reported the development of a multienzyme in vitro DNA replication system which replicates double-stranded (DS) DNA templates efficiently. This system consists of six highly purified bacteriophage T4-encoded proteins, the products of genes 32, 41, 43, 44, 45, and 62 (Morris et al. 1975; Alberts et al. 1975, 1977). Each of these proteins has been shown to be essential for DNA replication in vivo (Epstein et al. 1964). An amber mutation in any of these genes results in a phenotype showing very little or no DNA synthesis (Warner and Hobbs 1967), even though adequate deoxyribo- and ribonucleoside triphosphate levels are maintained (Mathews 1972). Additionally, temperature-sensitive mutants have been isolated in genes 32, 41, 43, and 45 which cease DNA replication immediately upon transfer to the restrictive temperature (Riva et al. 1970; Curtis and Alberts 1976).