Production of baculovirus defective interfering particles during serial passage is delayed by removing transposon target sites in fp25k

Lopamudra Giri; Michael G Feiss; Bryony C Bonning; David W Murhammer

doi:10.1099/vir.0.036566-0

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Production of baculovirus defective interfering particles during serial passage is delayed by removing transposon target sites in fp25k

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Production of baculovirus defective interfering particles during serial passage is delayed by removing transposon target sites in fp25k

Lopamudra Giri, Michael G Feiss, Bryony C Bonning and David W Murhammer

Journal of general virology, Vol.93(Pt 2), pp.389-399

02/2012

DOI: 10.1099/vir.0.036566-0

PMCID: PMC3352349

PMID: 21994323

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Abstract

Accumulation of baculovirus defective interfering particle (DIP) and few polyhedra (FP) mutants is a major limitation to continuous large-scale baculovirus production in insect-cell culture. Although overcoming these mutations would result in a cheaper platform for producing baculovirus biopesticides, little is known regarding the mechanism of FP and DIP formation. This issue was addressed by comparing DIP production of wild-type (WT) Autographa californica multiple nucleopolyhedrovirus (AcMNPV) with that of a recombinant AcMNPV (denoted Ac-FPm) containing a modified fp25k gene with altered transposon insertion sites that prevented transposon-mediated production of the FP phenotype. In addition to a reduction in the incidence of the FP phenotype, DIP formation was delayed on passaging of Ac-FPm compared with WT AcMNPV. Specifically, the yield of DIP DNA in Ac-FPm was significantly lower than in WT AcMNPV up to passage 16, thereby demonstrating that modifying the transposon insertion sites increases the genomic stability of AcMNPV. A critical component of this investigation was the optimization of a systematic method based on the use of pulsed-field gel electrophoresis (PFGE) to characterize extracellular virus DNA. Specifically, PFGE was used to detect defective genomes, determine defective genome sizes and quantify the amount of defective genome within a heterogeneous genome population of passaged virus.

Sequence Deletion

Animals

Nucleopolyhedroviruses - genetics

Lepidoptera - virology

Virulence

Defective Viruses - genetics

Nucleopolyhedroviruses - ultrastructure

Nucleopolyhedroviruses - growth & development

Serial Passage

DNA Transposable Elements

Defective Viruses - isolation & purification

Details

Title: Subtitle: Production of baculovirus defective interfering particles during serial passage is delayed by removing transposon target sites in fp25k
Creators: Lopamudra Giri - Department of Chemical and Biochemical Engineering, University of Iowa, Iowa City, IA, USA
Michael G Feiss - Department of Microbiology, University of Iowa, Iowa City, IA, USA
Bryony C Bonning - Department of Entomology, Iowa State University, Ames, IA, USA
David W Murhammer - Department of Chemical and Biochemical Engineering, University of Iowa, Iowa City, IA, USA
Resource Type: Journal article
Publication Details: Journal of general virology, Vol.93(Pt 2), pp.389-399
DOI: 10.1099/vir.0.036566-0
PMID: 21994323
PMCID: PMC3352349
NLM abbreviation: J Gen Virol
ISSN: 0022-1317
eISSN: 1465-2099
Publisher: England
Grant note: GM-51611 / NIGMS NIH HHS R01 GM051611 / NIGMS NIH HHS
Language: English
Date published: 02/2012
Academic Unit: Microbiology and Immunology; Chemical and Biochemical Engineering
Record Identifier: 9984003955102771

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