Back to results
US 8,263,395 B2Granted
Recombinant adenoviruses preparation and adenovirus banks
AssigneeAventis Pharma S.A.
InventorJean-Jacques Robert
Life of this patent
Estimating the term…
Abstract
The invention concerns compositions and methods for preparing recombinant adenoviruses. The resulting adenoviruses can be used for transferring and/or expressing genes in cells, in vitro, ex vivo or in vivo, or also in functional genomics. More particularly, the invention concerns in particular efficient methods for producing adenovirus banks and the use of said banks in functional genomics. The invention also concerns plasmids used for constructing said adenoviruses.
Claims17
A method for producing adenoviruses, which comprises:
- a) constructing a shuttle plasmid, which comprises a first truncated recombinant adenoviral genome which contains at least one heterologous nucleic acid,
- b) bringing this shuttle plasmid into contact with a parent plasmid, which comprises a second truncated recombinant adenoviral genome which is complementary to the first genome, making it possible, by means of one homologous recombination step, to produce a final plasmid which comprises a complete recombinant adenoviral genome, and
- c) excising the complete linear recombinant adenoviral genome from the final plasmid and introducing it into an encapsidating cell line in order to produce the recombinant adenoviruses which incorporate the complete recombinant adenoviral genome,
- said shuttle plasmid and parent plasmid both comprising an ITR sequence bordered by a restriction site not present on the adenoviral genome.
1 more element
A kit which comprises:
- a shuttle plasmid comprising a first truncated adenovirus genome, and a second parent plasmid comprising a second truncated adenovirus genome, with the two plasmids being able to produce, by means of one homologous recombination step between the two truncated adenovirus genomes, a final plasmid which comprises a complete linear recombinant adenovirus genome which is flanked by one or more restriction sites which is/are not present in said genome, said shuttle plasmid and parent plasmid both comprising an ITR sequence bordered by a restriction site not present on the adenoviral genome.
A method for producing adenoviruses, which comprises:
- a) constructing a shuttle plasmid, which comprises a first truncated recombinant adenoviral genome which contains at least one heterologous nucleic acid,
- b) bringing this shuttle plasmid into contact with a parent plasmid, which comprises a second truncated recombinant adenoviral genome which is complementary to the first genome, making it possible, by means of one homologous recombination step, to produce a final plasmid which comprises a complete recombinant adenoviral genome, and
- c) excising the complete linear recombinant adenoviral genome from the final plasmid and introducing it into an encapsidating cell line in order to produce the recombinant adenoviruses which incorporate the complete recombinant adenoviral genome,
- said shuttle plasmid and said parent plasmid both comprising an identical adenoviral homology region, both comprising an origin of replication and a different selection marker for selecting each element and both comprising an ITR sequence bordered by a restriction site not present on the adenoviral genome, and one or other comprising an encapsidation sequence.
1 more element
A kit which comprises:
- a shuttle plasmid comprising a first truncated adenovirus genome, and a parent plasmid comprising a second truncated adenovirus genome, with the two plasmids being able to produce, by means of one homologous recombination step between the two truncated adenovirus genomes, a final plasmid which comprises a complete linear recombinant adenovirus genome which is flanked by one or more restriction sites which is/are not present in said genome, said shuttle plasmid and said parent plasmid both comprising an identical adenoviral homology region, both comprising an origin of replication and a different selection marker for selecting each element and both comprising an ITR sequence bordered by a restriction site not present on the adenoviral genome, and one or other comprising an encapsidation sequence.
Dependencies are inferred from the claim text.