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A bacmid approach to the genetic manipulation of granuloviruses
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Hilton, Sally, Kemp, E. (Elizabeth), Keane, G. (Gary) and Winstanley, Doreen (2008) A bacmid approach to the genetic manipulation of granuloviruses. Journal of Virological Methods, Vol.152 (No.1-2). pp. 56-62. doi:10.1016/j.jviromet.2008.05.015
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Official URL: http://dx.doi.org/10.1016/j.jviromet.2008.05.015
Abstract
A Cydia pomonella granulovirus (CpGV) bacmid has been constructed, which allows rapid and efficient production of recombinant baculoviruses in Escherichia coli. An 8.6 kbp bacterial DNA cassette derived from the AcMNPV Bac-to-Bac (R) system was ligated into a unique Pacl restriction site within an intergenic region flanking the DNA ligase gene of the CpGV genome. The CpGV bacmids produced in E. coli were transfected into a CpGV-permissive C. pomonella cell line and the transfected cells fed to larvae to amplify the virus. The enhanced green fluorescent protein (EGFP) gene under the constitutive Drosophila heat-shock promoter was transposed into the mini-attTn7 transposition site, using a modified pFASTBAC (TM) donor plasmid, to generate a recombinant CpGV bacmid which caused infected larvae to glow under UV light. Targeted homologous recombination was also achieved in a recombinant proficient E. coli strain (BJ5183). A chloramphenicol acetyl transferase (CAT) gene replaced the cathepsin (v-cath) gene in the bacmid to produce a v-cath-deletion mutant. This is the first published report of a granulovirus bacmid, which will allow easy manipulation of the CpGV genome, enabling future studies on granulovirus genes and biology. (C) 2008 Elsevier B.V. All rights reserved.
| Item Type: | Journal Article | ||||
|---|---|---|---|---|---|
| Subjects: | Q Science > QH Natural history > QH426 Genetics Q Science > QR Microbiology > QR355 Virology |
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| Divisions: | Faculty of Science > Life Sciences (2010- ) > Warwick HRI (2004-2010) | ||||
| Library of Congress Subject Headings (LCSH): | Baculoviruses -- Genetics, Codling moth -- Pathogens, Microbial genetic engineering | ||||
| Journal or Publication Title: | Journal of Virological Methods | ||||
| Publisher: | Elsevier BV | ||||
| ISSN: | 0166-0934 | ||||
| Official Date: | September 2008 | ||||
| Dates: |
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| Volume: | Vol.152 | ||||
| Number: | No.1-2 | ||||
| Number of Pages: | 7 | ||||
| Page Range: | pp. 56-62 | ||||
| DOI: | 10.1016/j.jviromet.2008.05.015 | ||||
| Status: | Peer Reviewed | ||||
| Publication Status: | Published | ||||
| Access rights to Published version: | Restricted or Subscription Access | ||||
| Funder: | Biotechnology and Biological Sciences Research Council (Great Britain) (BBSRC) |
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