The original polymerase I mutant was not completely defective in that enzyme, and later experiments showed that the residual polymerase I activity in this strain plays a key role in the replication process. It is now known that, in addition to polymerase III, polymerase I is also required for replication of E. Temperature-sensitive polymerase III mutants, however, were unable to replicate their DNA at high temperature, and subsequent studies have confirmed that polymerase III is the major replicative enzyme in E. coli with mutations in polymerase II were found to grow and otherwise behave normally, so the role of this enzyme in the cell is unknown. The potential roles of these enzymes were investigated by the isolation of appropriate mutants. coli must contain other DNA polymerases, and subsequent experiments led to the identification of two such enzymes, now called DNA polymerases II and III. The conclusion that polymerase I is not required for replication implied that E. Several thousand colonies were then cultured and screened to identify (more.) coli was treated with a chemical mutagen, and individual bacterial colonies were isolated by growth on semisolid medium. Isolation of a mutant deficient in polymerase I. On the other hand, the mutant bacteria were extremely sensitive to agents that damage DNA (e.g., ultraviolet light), suggesting that polymerase I is involved primarily in the repair of DNA damage rather than in DNA replication per se. Surprisingly, the mutant bacteria grew normally, leading to the conclusion that polymerase I is not required for DNA replication. Analysis of a few thousand colonies led to the isolation of the desired mutant, which was almost totally defective in polymerase I activity. coli were treated with a chemical (a mutagen) that induces a high frequency of mutations, and individual bacterial colonies were isolated and screened to identify a mutant strain lacking polymerase I. coli that was deficient in polymerase I ( Figure 5.1). The multiplicity of DNA polymerases was first revealed by the isolation of a mutant strain of E. Instead, it is now clear that both prokaryotic and eukaryotic cells contain several different DNA polymerases that play distinct roles in the replication and repair of DNA. Ironically, however, this first DNA polymerase to be identified (now called DNA polymerase I) is not the major enzyme responsible for E. The ability of this enzyme to accurately copy a DNA template provided a biochemical basis for the mode of DNA replication that was initially proposed by Watson and Crick, so its isolation represented a landmark discovery in molecular biology. DNA polymerase was first identified in lysates of E.

Describe the process of dna replication