Construction of a recombinant Anticarsia gemmatalis nucleopolyhedrovirus (AgMNPV-2D) harbouring the β-galactosidase gene

Construction of a recombinant Anticarsia gemmatalis nucleopolyhedrovirus (AgMNPV-2D) harbouring... We have constructed a transfer vector (pAgGal) containing the β-galactosidase gene under control of the Escherichia coli gpt and AgMNPV polyhedrin ( polh ) promoters. The transfer vector was cotransfected with wild type Anticarsia gemmatalis nucleopolyhedrovirus (AgMNPV) DNA into A . gemmatalis (UFL-AG-286) cells and a recombinant baculovirus (vAgGalA2) was isolated. The β-galactosidase gene insertion was checked by polymerase chain reaction (PCR) using DNA from AgMNPV and vAgGalA2 and primers specific for regions upstream and downstream of the polh gene. Insect cells (UFL-AG-286) were infected with the recombinant vAgGalA2 and wild type AgMNPV viruses and the production of the heterologous protein analyzed by SDS-PAGE and Pulse-Chase. β-galactosidase was expressed at high levels late on infection as expected for a gene under the control of the polh promoter. The highly expressed β-galactosidase protein was also shown to be biologically active by a β-galactosidase assay. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Archives of Virology Springer Journals

Construction of a recombinant Anticarsia gemmatalis nucleopolyhedrovirus (AgMNPV-2D) harbouring the β-galactosidase gene

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Publisher
Springer-Verlag
Copyright
Copyright © 2001 by Springer-Verlag/Wien
Subject
Legacy
ISSN
0304-8608
eISSN
1432-8798
D.O.I.
10.1007/s007050170096
Publisher site
See Article on Publisher Site

Abstract

We have constructed a transfer vector (pAgGal) containing the β-galactosidase gene under control of the Escherichia coli gpt and AgMNPV polyhedrin ( polh ) promoters. The transfer vector was cotransfected with wild type Anticarsia gemmatalis nucleopolyhedrovirus (AgMNPV) DNA into A . gemmatalis (UFL-AG-286) cells and a recombinant baculovirus (vAgGalA2) was isolated. The β-galactosidase gene insertion was checked by polymerase chain reaction (PCR) using DNA from AgMNPV and vAgGalA2 and primers specific for regions upstream and downstream of the polh gene. Insect cells (UFL-AG-286) were infected with the recombinant vAgGalA2 and wild type AgMNPV viruses and the production of the heterologous protein analyzed by SDS-PAGE and Pulse-Chase. β-galactosidase was expressed at high levels late on infection as expected for a gene under the control of the polh promoter. The highly expressed β-galactosidase protein was also shown to be biologically active by a β-galactosidase assay.

Journal

Archives of VirologySpringer Journals

Published: Jul 1, 2001

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