Rolling-circle replication of a high-copy BPV-1 plasmid

We investigated the replicating form of a bovine papillomavirus type 1 (BPV-1) deletion mutant by direct electron-microscopic analysis of low molecular weight cellular DNA fractions. The detection of viral plasmid DNA replication intermediates was facilitated by the isolation of a spontaneously tran...

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Veröffentlicht in:Journal of molecular biology 1992-11, Vol.228 (1), p.1-6
Hauptverfasser: Dasgupta, Santanu, Zabielski, Jan, Simonsson, Magnus, Burnett, Stanley
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Sprache:eng
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Zusammenfassung:We investigated the replicating form of a bovine papillomavirus type 1 (BPV-1) deletion mutant by direct electron-microscopic analysis of low molecular weight cellular DNA fractions. The detection of viral plasmid DNA replication intermediates was facilitated by the isolation of a spontaneously transformed mouse cell subclone containing an unusually high viral genome copy number (approx. 1000 per cell), and by employing a slight modification of the Hirt fractionation procedure to reduce the level of contaminating linear chromosomal DNA fragments. We observed exclusively rolling-circle-type viral DNA replication intermediates, at a frequency of detection of approximately one replication intermediate per 200 monomeric circular viral DNA molecules. The demonstration of rolling-circles with longer-than-genome-length tails indicated that this high-copy viral plasmid was not subject to a strict once-per-cell-cycle mode of DNA replication. Our observations provide further evidence in favour of an alternative replication mode of the BPV-1 genome, and may help to explain earlier conflicting findings concerning the mechanism of stable BPV-1 plasmid copy-number-control.
ISSN:0022-2836
1089-8638
DOI:10.1016/0022-2836(92)90485-3