A sensitive model system for in vivo monitoring of baculovirus gene expression in single infected insect cells.
We have developed a fast and sensitive system for the in vivo analysis of gene expression in baculovirus infected lepidopteran insect cells. A recombinant baculovirus containing a luciferase gene from the click beetle, Pyrophorus plagiophthalamus, under transcriptional regulation of the polyhedrin gene promoter of Autographa californica nuclear polyhedrosis virus (AcNPV) was used to infect a Spodoptera frugiperda cell line. Recombinant luciferase could be monitored by luminometry in real-time without disruption of the infected cells, allowing detection of synthesis as early as one hour after infection. The range of luminescence measurements was normally over four orders of magnitude, and the kinetics of luciferase synthesis and the levels of light produced in vivo closely correlated with the expression of polyhedrin in AcNPV infected cells when analyzed by SDS-PAGE. Additionally, single infected cells could be identified by CCD image analysis and flow cytometry.
Centre for Biotechnology, Turku, Finland.
M
M
Karp
K
K
Akerman
C
C
Lindqvist
A
A
Kuusisto
P
P
Saviranta
C
C
Oker-Blom
D000818:Animals; D016367:Baculoviridae / Q000235:genetics*; D002478:Cells, Cultured; D001517:Coleoptera / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D005434:Flow Cytometry; D015870:Gene Expression / Q000502:physiology*; D005814:Genes, Viral / Q000235:genetics*; D007091:Image Processing, Computer-Assisted; D007313:Insecta / Q000235:genetics*; D008027:Light; D008156:Luciferases / Q000235:genetics*; D008957:Models, Genetic; D010957:Plasmids / Q000235:genetics; D012680:Sensitivity and Specificity
D008156:Luciferases