Analysis of Replication of oriP-Based Plasmids by Quantitative, Competitive PCR

The quantitative, competitive polymerase chain reaction (PCR) assay outlined in this chapter was designed for the detection and quantitation of replicated DNAs in both short-term and long-term assays (1 ). Quantitative, competitive PCR can be used to study both the contribution of proteins to the replication of oriP-based plasmids (1 ) as well as the requirements for specific DNA sequences to support replication of a plasmid (2 ). Advantages of this assay include an increased sensitivity and a decreased time required to analyze samples relative to DNA blots, the traditional assay used to study replication of oriP-containing plasmids in the presence of EBNA-1 (3 –8 ).