Targeted gene modification by parvoviral vectors
First Claim
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1. An in vitro method of producing a vertebrate cell having a genetic modification at a preselected genomic target locus, the method comprising:
- (i) transducing into the vertebrate cell an effective amount of a recombinant parvovirus particle genome that comprises;
a) a targeting construct which comprises a DNA sequence which is substantially identical to the genomic target locus except for the modification being introduced, wherein the modification being introduced is flanked by regions substantially identical to the genomic target locus, said regions being at least 100 nucleotides in length; and
b) at least one parvoviral inverted terminal repeat (ITR) flanking the targeting construct; and
(ii) determining if the genomic target locus has the introduced genetic modification, wherein the targeting construct facilitates homologous pairing between the targeting construct and the genomic target locus resulting in the genetic modification being introduced into the genomic target locus.
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Abstract
This invention provides methods for obtaining targeted gene modification in vertebrate cells using parvoviral vectors, including adeno-associated virus (AAV). The parvoviral vectors used in the methods of the invention are capable of targeting a specific genetic modification to a preselected target locus in a cellular genome by homologous pairing.
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Citations
34 Claims
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1. An in vitro method of producing a vertebrate cell having a genetic modification at a preselected genomic target locus, the method comprising:
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(i) transducing into the vertebrate cell an effective amount of a recombinant parvovirus particle genome that comprises; a) a targeting construct which comprises a DNA sequence which is substantially identical to the genomic target locus except for the modification being introduced, wherein the modification being introduced is flanked by regions substantially identical to the genomic target locus, said regions being at least 100 nucleotides in length; and b) at least one parvoviral inverted terminal repeat (ITR) flanking the targeting construct; and (ii) determining if the genomic target locus has the introduced genetic modification, wherein the targeting construct facilitates homologous pairing between the targeting construct and the genomic target locus resulting in the genetic modification being introduced into the genomic target locus. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34)
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Specification