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Journal of Virology, October 2006, p. 9905-9909, Vol. 80, No. 19
0022-538X/06/$08.00+0 doi:10.1128/JVI.01004-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.
Alicia Rodriguez-Huete, and
Gregory S. Pari*
University of Nevada, Reno, School of Medicine, Department of Microbiology and Immunology and the Cell and Molecular Biology Graduate Program, Reno, Nevada 89557
Received 16 May 2006/ Accepted 18 July 2006
The lytic origins of DNA replication for human herpesvirus 8 (HHV8), oriLyt-L and oriLyt-R, are located between open reading frames K4.2 and K5 and ORF69 and vFLIP, respectively. These lytic origins were elucidated using a transient replication assay. Although this assay is a powerful tool for identifying many herpesvirus lytic origins, it is limited in its ability to evaluate the activity of replication origins in the context of the viral genome. To this end, we investigated the ability of a recombinant HHV8 bacterial artificial chromosome (BAC) to replicate in the absence of oriLyt-R, oriLyt-L, or both oriLyt regions. We generated the HHV8 BAC recombinants (BAC36-
Ori-R, BAC36-
Ori-L, and BAC36-
Ori-RL), which removed one or all of the identified lytic origins. An evaluation of these recombinant BACs revealed that oriLyt-L was sufficient to propagate the viral genome, whereas oriLyt-R alone failed to direct the amplification of viral DNA.
Present address: G. W. Hooper Foundation, Box 0552, University of California, San Francisco, San Francisco, CA 94143-0552.
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