A method of alphavirus replicon particle titration based on expression of functional replicase/transcriptase

Alphavirus replicon particles are being exploited for a variety of purposes both in vitro as gene expression vectors, and in vivo as vaccines or gene therapy vectors. There is a need for a simple and universal method of titration of replicon particles that is independent of expression of the foreign...

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Bibliographic Details
Published in:Journal of virological methods 2003-05, Vol.109 (2), p.133-138
Main Authors: Gangolli, Seema S., Vasilakis, Nikolaos, Kovacs, Gerald R., Zamb, Timothy J., Kowalski, Jacek
Format: Article
Language:English
Subjects:
Alphaviruses
Animals
Biological and medical sciences
Cercopithecus aethiops
Defective Viruses - genetics
DNA-Directed RNA Polymerases - physiology
Encephalitis Virus, Venezuelan Equine - genetics
Fundamental and applied biological sciences. Psychology
Green Fluorescent Proteins
Luminescent Proteins - genetics
Microbiology
Replicon - genetics
Replicon particles
Techniques used in virology
Titration
Vaccinia virus - genetics
Venezuelan equine encephalitis virus
Vero Cells
Virology
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Summary:Alphavirus replicon particles are being exploited for a variety of purposes both in vitro as gene expression vectors, and in vivo as vaccines or gene therapy vectors. There is a need for a simple and universal method of titration of replicon particles that is independent of expression of the foreign protein. We devised a method that uses modified vaccinia virus Ankara (MVA) as an indicator virus, to deliver a Venezuelan equine encephalitis virus (VEE) defective helper RNA encoding green fluorescent protein (GFP). Co-infection of cells with the MVA-based indicator and Venezuelan equine encephalitis virus replicon particles (VRP) results in expression of the GFP gene. VRP titer is readily determined by counting fluorescent cells.
ISSN:0166-0934
1879-0984
DOI:10.1016/S0166-0934(03)00063-6