Method for production of virus-like particles for gene delivery
AssigneeCarnegie Mellon University
InventorsYongxin ZHAO, Yu Hong WANG, Emma DiBernardo
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Abstract
Methods for production of gene-delivery vectors suitable for animal models and humans using a bacteria/mammalian dual-expression plasmid, wherein the virus-like particles may be massively produced via heterologous expression in bacteria. Also provided are methods of increasing virus-like particle transduction efficiency and methods of directed evolution.
Claims28
A method of producing virus-like particles in bacteria comprising:
- mutating a DNA sequence of a gene of interest;
- synthesizing the mutated DNA sequence with a plasmid, wherein a generated plasmid is formed;
- transforming the generated plasmid into bacteria;
- producing a protein of interest via a method of inducing gene expression from at least one transformed bacterial cell;
- lysing a volume of the at least one transformed bacterial cell;
- separating the protein of interest from cellular debris;
- collecting the virus-like particles;
- purifying the virus-like particles, wherein the virus-like particles comprise a structure in which at least two proteins are present in an aggregated form, and wherein the at least two proteins enclose a cavity;
- and
- mixing the virus-like particles with liposomes, wherein the virus-like particles are complexed with the liposomes.
A dual expression plasmid, wherein the dual expression plasmid is capable of producing at least one virus-like particle, and wherein the dual expression plasmid is selected for at least one phenotype in a mammalian cell.
A method of producing virus-like particles in bacteria comprising:
- mutating a DNA sequence of a gene of interest;
- synthesizing a mutated sequence with a plasmid, wherein a generated plasmid is formed;
- transforming the generated plasmid into a bacteria cell;
- producing a protein of interest via a method of inducing gene expression from at least one transformed bacterial cell;
- lysing a volume of the at least one transformed bacterial cell;
- separating the protein of interest from cellular debris;
- collecting the virus-like particles;
- and
- purifying the virus-like particles, wherein the virus-like particles comprise a structure in which at least two proteins are present in an aggregated form, and wherein the at least two proteins enclose a cavity.
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Dependencies are inferred from the claim text.