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Rolling-circle replication of a high-copy BPV-1 plasmid
S Dasgupta1, J Zabielski, M Simonsson
1Department of Medical Genetics, Biomedical Centre, Uppsala, Sweden.
Journal of Molecular Biology
|November 5, 1992
Summary
This study reveals bovine papillomavirus type 1 (BPV-1) DNA replicates via a rolling-circle mechanism. This finding clarifies BPV-1 plasmid replication and copy-number control.
Area of Science:
- Molecular Biology
- Virology
- Genetics
Background:
- Bovine papillomavirus type 1 (BPV-1) exists as a high-copy-number plasmid in infected cells.
- The precise replication mechanism and copy-number control of BPV-1 have been debated.
Purpose of the Study:
- To investigate the replicating form of a BPV-1 deletion mutant.
- To elucidate the DNA replication mechanism of BPV-1 plasmids.
Main Methods:
- Direct electron microscopy of low molecular weight cellular DNA fractions.
- Isolation of a mouse cell subclone with high BPV-1 genome copy number (~1000 per cell).
- Modified Hirt fractionation to enrich for viral DNA and reduce chromosomal DNA contamination.
Main Results:
- Exclusively observed rolling-circle DNA replication intermediates.
- Detected rolling-circles with longer-than-genome-length tails.
- Replication intermediates found at a frequency of approximately 1 per 200 monomeric circular viral DNA molecules.
Conclusions:
- BPV-1 DNA replication predominantly occurs via a rolling-circle mechanism.
- Replication is not strictly limited to once per cell cycle in high-copy situations.
- Findings support an alternative replication model and explain previous conflicting results on BPV-1 copy-number control.