Poliovirus Replicon RNA Generation, UNIT 15H.4Transfection, Packaging, andQuantitation of Replication

摘要

Poliovirus is a prototype member of the Enterovirus genus of the Picornaviridaefamily of small positive strand RNA viruses, which include importanthuman and animal pathogens. Quantitative assessment of viral replication isvery important for investigation of the virus biology and the development ofanti-viral strategies. The poliovirus genome structure allows replacement ofstructural genes with a reporter protein, such as a luciferase or a fluorescentprotein, whose signals can be detected and quantified in vivo, thus permittingobservation of replication kinetics in live cells. This paper presents protocolsfor poliovirus replicon RNA production, purification, packaging and transfection,as well as techniques for monitoring Renilla luciferase replication signalin living cells.