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Journal of Virology, June 2000, p. 5182-5189, Vol. 74, No. 11
0022-538X/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Differential Activity of Two Non-hr Origins during Replication of the Baculovirus Autographa californica Nuclear Polyhedrosis Virus Genomedagger

Saman Habib1,* and Seyed E. Hasnain2,3

Membrane Biology Division, Central Drug Research Institute, Chattar Manzil, Lucknow-226001,1 Eukaryotic Gene Expression Laboratory, National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi-110067,2 and Centre for DNA Fingerprinting and Diagnostics, Nacharam, Hyderabad-500007,3 India

Received 28 September 1999/Accepted 13 March 2000

The identification of potential baculovirus origins of replication (ori) has involved the generation and characterization of defective interfering particles that contain major genomic deletions yet retain their capability to replicate by testing the replication ability of transiently transfected plasmids carrying viral sequences in infected cells. So far, there has not been any evidence to demonstrate the actual utilization of these putative origins in Autographa californica multinucleocapsid nuclear polyhedrosis virus (AcMNPV) replication. By using the method of origin mapping by competitive PCR, we have obtained quantitative data for the ori activity of the HindIII-K region and the ie-1 promoter sequence in AcMNPV. We also provide evidence for differential activity of the two ori in the context of the viral genome through the replication phase of viral infection. Comparison of the number of molecules representing the HindIII-K and ie-1 origins vis-à-vis the non-ori polH region in a size-selected nascent DNA preparation revealed that the HindIII-K ori is utilized ~14 times more efficiently than the ie-1 region during the late phase of infection. HindIII-K also remains the more active ori through the early and middle replication phases. Our results provide in vivo evidence in support of the view that AcMNPV replication involves multiple ori that are activated with vastly different efficiencies during the viral infection cycle.

* Corresponding author. Mailing address: Membrane Biology Division, Central Drug Research Institute, Chattar Manzil, Post Box 173, Lucknow-226001, India. Phone: 91-522-212-411, ext. 4282. Fax: 91-522-223-405. E-mail: samamit{at}lw1.vsnl.net.in.

dagger This is CDRI communication no. 5989.

Journal of Virology, June 2000, p. 5182-5189, Vol. 74, No. 11
0022-538X/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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