Abstract
Reverse genetic approach is widely used in virology as it makes possible direct identification of viral gene function and uses RNA genomes as vectors. Production of infectious cDNA clones is an essential step in developing a reverse genetic system for an RNA virus. Here, we present rapid method for generation of infectious cDNA clone for Turnip crinkle virus (TCV). The infectious cDNA clone could be used for production of in vitro transcripts with the T7 RNA polymerase which could be used for infection of plants or plant cell protoplasts. The procedure described here includes purification of TCV, viral RNA extraction, reverse transcription, PCR amplification of the full-length cDNA copy of TCV linked to a T7 RNA polymerase promoter, cloning into a plasmid vector, in vitro transcription, and selection of infectious clones.
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Acknowledgments
This work was supported by the Biotechnology and Biological Sciences Research Council.
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© 2008 Humana Press, a part of Springer Science + Business Media, LLC
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Ryabov, E.V. (2008). Construction of Infectious cDNA Clones for RNA Viruses: Turnip Crinkle Virus. In: Foster, G.D., Johansen, I.E., Hong, Y., Nagy, P.D. (eds) Plant Virology Protocols. Methods in Molecular Biology™, vol 451. Humana Press. https://doi.org/10.1007/978-1-59745-102-4_33
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DOI: https://doi.org/10.1007/978-1-59745-102-4_33
Publisher Name: Humana Press
Print ISBN: 978-1-58829-827-0
Online ISBN: 978-1-59745-102-4
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