This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Crouch, E. A. Articles by Passarelli, A. L. Search for Related Content PubMed PubMed Citation Articles by Crouch, E. A. Articles by Passarelli, A. L.

 Previous Article  |  Next Article 

Journal of Virology, September 2002, p. 9323-9334, Vol. 76, No. 18
0022-538X/02/$04.00+0     DOI: 10.1128/JVI.76.18.9323-9334.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Genetic Requirements for Homologous Recombination in Autographa californica Nucleopolyhedrovirus

Division of Biology, Molecular, Cellular, and Developmental Biology Program, Kansas State University, Manhattan, Kansas 66506

Received 9 April 2002/ Accepted 11 June 2002

It is known that baculovirus infection promotes high-frequency recombination between its genomes and plasmid DNA during the construction of recombinant viruses for foreign gene expression. However, little is known about the viral genes necessary to promote homologous recombination (HR). We developed an assay to identify viral genes that are necessary to stimulate HR. In this assay, we used two plasmids containing extensive sequence homology that yielded a visible and quantifiable phenotype if HR occurred. The plasmids contained the green fluorescent protein gene (gfp) that was mutated at either the N or the C terminus and a viral origin of DNA replication. When the plasmids containing these mutant gfp genes were transfected into insect cells alone or together, few green fluorescent protein (GFP)-positive cells were observed, confirming that the host cell machinery alone was not able to promote high levels of HR. However, if viral DNA or viral genes involved in DNA replication were cotransfected into cells along with the mutant gfp-containing plasmids, a dramatic increase in GFP-positive cells was observed. The viral genes ie-1, ie-2, lef-7, and p35 were found to be important for efficient HR in the presence of all other DNA replication genes. However, ie-1 and ie-2 were sufficient to promote HR in the absence of other viral genes. Recombination substrates lacking a viral origin of replication had similar genetic requirements for recombination but were less dependent on ie-1. Interestingly, even though HR was stimulated by the presence of a viral origin of DNA replication, virally stimulated HR could proceed in the presence of the DNA synthesis inhibitor aphidicolin.

Contribution 02-390-5 from the Kansas Agricultural Experiment Station.

This article has been cited by other articles:

• Kamita, S. G., Maeda, S., Hammock, B. D. (2003). High-Frequency Homologous Recombination between Baculoviruses Involves DNA Replication. J. Virol. 77: 13053-13061 [Abstract] [Full Text]  
• Olson, V. A., Wetter, J. A., Friesen, P. D. (2003). The Highly Conserved Basic Domain I of Baculovirus IE1 Is Required for hr Enhancer DNA Binding and hr-Dependent Transactivation. J. Virol. 77: 5668-5677 [Abstract] [Full Text]