Functional origin of replication of pT181 plasmid DNA is contained within a 168-base-pair segment.

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Abstract
A recently developed in vitro replication system from Staphylococcus aureus was used to determine the origin and direction of replication of plasmid pT181 DNA. The origin was located to within 168 base pairs by sequential labeling of restriction endonuclease fragments after synchronous initiation in vitro in the presence of various amounts of dideoxy-TTP and by constructing in vitro deletions of pT181 DNA close to the origin of replication and testing for their ability to replicate in vitro. Plasmid pT181 replicated unidirectionally and anticlockwise, as the map is conventionally drawn. The nucleotide sequence of the region containing the origin of replication was determined; it was partially or entirely contained within the coding sequence for the repC protein, which is uniquely required for plasmid pT181 replication. Preliminary evidence suggesting that pT181 replicates by a rolling circle mechanism is discussed.