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AssigneeChiron Corporation
InventorsThomas W. Dubensky JR., John M. Polo, Silvia Perri, Barbara Belli
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Isolated nucleic acid molecules are disclosed, comprising an alphavirus nonstructural protein 2 gene which, when operably incorporated into an alphavirus replicon particle, eukaryotic layered vector initiation system, alphavirus vector construct or RNA vector replicon, provides a noncytopathic phenotype or confers the ability to establish persistent replication. Also disclosed are RNA vector replicons, alphavirus vector constructs, alphavirus replicon particles and eukaryotic layered vector initiation systems which contain the above-identified nucleic acid molecules, as well as methods of using such replicons, constructs, particles and eukaryotic layered vector initiation systems for expression of recombinant proteins.
An isolated nucleic acid molecule, comprising an alphavirus nonstructural protein gene which, when operably incorporated into an alphavirus replicon particle, has a reduced level of vector-specific RNA synthesis, as compared to the wild-type replicon particle, and the same or greater level of proteins encoded by RNA transcribed from the subgenomic junction region promoter, as compared to the wild-type alphavirus replicon particle, and wherein said alphavirus nonstructural protein gene encodes a protein with a substitution in an amino acid residue of nsP2 selected from the group consisting of residues 1, 10, 468, 469, 472, 708, 712, 713, and 721.
An isolated nucleic acid molecule, comprising an alphavirus nonstructural protein gene which, when operably incorporated into an alphavirus replicon particle, increases the time required to reach 50% inhibition of host-cell directed macromolecular synthesis following expression in mammalian cells, as compared to the wild-type alphavirus replicon particle, and wherein said alphavirus nonstructural protein gene encodes a protein with a substitution in an amino acid residue of nsP2 selected from the group consisting of residues 1, 10, 468, 469, 472, 708, 712, 713, and 721.
An isolated nucleic acid molecule, comprising an alphavirus nonstructural protein gene which, when operably incorporated into an alphavirus RNA vector replicon, alphavirus replicon particle, or eukaryotic layered vector initiation system, results in a vector capable of persistent replication following introduction into a mammalian cell, and wherein said alphavirus nonstructural protein gene encodes a protein with a substitution in an amino acid residue of nsP2 selected from the group consisting of residue 1, 10, 468, 469, 472, 708, 712, 713, and 721.
An expression cassette, comprising a promoter operably linked to a nucleic acid molecule, said nucleic acid molecule encoding a temperature sensitive R17 coat protein.
A method of making a selected recombinant protein, comprising:
Dependencies are inferred from the claim text.