Abstract: The present invention provides methods of inhibiting picornavirus genome replication in a subject. In particular, methods for interfering with VPg uridylylation and elongation are provided. The methods comprise administering to a subject an effective amount of at least one of VPg, VPg analog, VPg homology or biologically active fragment thereof as well as oligonucleotides, divalent cations, ribonucleotide or deoxyribonucleotide. Also provided are methods of identifying an inhibitor of picornaviral replication which comprise adding a potential inhibitor of picornaviral replication to an in vitro assay and analyzing levels of VPg uridylylation reaction products.

Abstract: A process for the de novo synthesis of a picornavirus using a cell-free medium has been developed. The cell-free medium is prepared from a lysate of mammalian cells from which the nuclei and mitochondria has been removed, endogenous mRNA deactivated and supplemented with nucleoside triphosphates, tRNA, amino acids and precursors necessary for generating proteins. The genomic RNA of the picornavirus or rhinovirus is added to the cell-free medium and after incubation at about 30.degree. C. to 40.degree. C. of from about 4 to 24 hours infectious, mature virus particles are formed.