Self-deleting plasmid
AssigneeCobra Biologics Limited
InventorsRocky Marc Cranenburgh, Matthew William Leckenby
Life of this patent
Estimating the term…
Abstract
A method of producing a selectable marker gene-free plasmid by culturing a plasmid containing a selectable marker gene flanked by site specific recombinase target sites in a host cell environment incapable of effecting recombination between the site specific recombinase target sites and subsequently culturing the plasmid in another host cell environment which is capable of effecting recombination between the site specific recombinase target sites, so that the selectable marker gene is excised. Uses of plasmids produced by the method for the production of recombinant protein for therapeutic and vaccine purposes, production of therapeutic DNA and DNA vaccines and delivery of recombinant protein and DNA to a patient using live bacterial vectors.
Claims11
A method of producing a selectable marker gene-free plasmid comprising the steps of:
- a) culturing a plasmid containing a selectable marker gene flanked by site specific recombinase target sites selected from Ecdif, cer, psi, pif and mwr in a first host cell environment which is incapable of effecting recombination between the site specific recombinase target sites, wherein the first host cell environment comprises an inactivating mutation in one or more of the genes encoding PepA, ArgR and ArcA;
- and
- b) subsequently culturing the plasmid in a second host cell environment which is capable of effecting recombination between the site specific recombinase target sites, such that the selectable marker gene is excised, wherein the second host cell environment contains active versions of PepA and ArgR or ArcA, and comprises a site specific recombinase selected from XerC and XerD.
Dependencies are inferred from the claim text.