Recombinant circovirus capsid-virus-like particle (vlp): compositions, methods and uses
AssigneesTechnoVax, Inc., Huvepharma
InventorsJose M. GALARZA, Ke WEN, Boris GAVRILOV
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Abstract
Provided herein is a mammalian expression system for producing recombinant porcine circovirus type 2 (PCV2) virus-like particles (VLPs). The expression system includes a mammalian cell and a plasmid that comprises a PCV2 gene encoding a capsid protein. The PCV2 gene is codon optimized, and the mammalian cell is transfected with the plasmid. The expression system produces recombinant PCV2 VLPs, such as PCV2d VLPs. Also provided herein are a method for producing porcine circovirus type 2 (PCV2) virus-like particles (VLPs), as well as a PCV2 VLP generated by the method.
Claims24
A mammalian expression system for producing recombinant porcine circovirus type 2 (PCV2) virus-like particles (VLPs), the expression system comprising:
- a mammalian cell;
- a plasmid comprising a PCV2 gene encoding a capsid protein, wherein the PCV2 gene is codon optimized;
- wherein the mammalian cell is transfected with the plasmid;
- and
- wherein the expression system produces recombinant PCV2 VLPs.
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A method for producing porcine circovirus type 2 (PCV2) virus-like particles (VLPs), comprising:
- providing a suspension of cultured mammalian cells;
- transfecting the mammalian cells with a plasmid comprising a PCV2 gene encoding a capsid protein;
- adding valproic acid (VPA) sodium salt to the transfected mammalian cells, wherein the addition of the VPA sodium salt inhibits cell proliferation;
- centrifuging and washing the transfected mammalian cells;
- suspending the centrifuged mammalian cells in a phosphate buffered saline (PBS) solution;
- performing multiple freeze and thaw cycles on the mammalian cells;
- sonicating the mammalian cells in multiple cycles;
- and
- performing two successive centrifugation cycles of the mammalians cells to produce the PCV2 VLPs,
- wherein a majority of the produced PCV2 VLPs are present in the nucleus of the mammalian cells.
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Dependencies are inferred from the claim text.