Negative control of DNA replication in composite SV40-bovine papilloma virus plasmids

Abstract

To identify DNA sequences that function in the control of DNA replication, we designed a hybrid replicon consisting of linked SV40 and BPV DNA sequences. In the composite SV40-BPV plasmid negative control encoded by BPV is dominant over the uncontrolled replication encoded by the positive factor, SV40 T antigen. Using a transient replication assay, we show that replication control requires three BPV elements. Two cis-acting sequences are closely linked to BPV replication origins. A third trans-acting element is encoded within the 5′ part of the BPV E1 open reading frame (ORF) and is separable from the positive replication factor encoded within the 3′ part of the same ORF. The controlled replication of SV-BPV composite replicons has enabled us to create permanent COS cell lines that stably maintain these plasmids as episomes.