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Rachy, Abraham and E., Sreekumar (2018) Preparation of Recombinant Alphaviruses for Functional Studies of ADP-Ribosylation. Methods in molecular biology. pp. 297-316. ISSN 1940-6029

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Recently we characterized the mono(ADP-ribosyl) hydrolase (MAR hydrolase) activity of the macrodomain of nonstructural protein 3 (nsP3MD) of chikungunya virus. Using recombinant viruses with targeted mutations in the macrodomain, we demonstrated that hydrolase function is important for viral replication in cultured neuronal cells and for neurovirulence in mice. Here, we describe the general cell culture and animal model infection protocols for alphaviruses and the technical details for biochemical characterization of the MAR hydrolase activity of nsP3MD mutants and the preparation of recombinant viruses incorporating those mutations through site-directed mutagenesis of an infectious cDNA virus clone.

Item Type: Article
Uncontrolled Keywords: Alphavirus Chikungunya virus Sindbis virus MAR hydrolase assay Infectious cDNA clone Reverse genetics Site-directed mutagenesis In vitro transcription
Subjects: Viral Disease Biology
Depositing User: Rgcb Library
Date Deposited: 25 Oct 2018 06:07
Last Modified: 25 Oct 2018 06:07
URI: http://rgcb.sciencecentral.in/id/eprint/703

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