Genome Editing without Nucleases
AssigneeThe Board of Trustees of the Leland Stanford Junior University
InventorsAdi Barzel, Mark A. Kay
Life of this patent
Estimating the term…
Abstract
Methods and compositions are provided for editing the genome of a cell without the use of an exogenously supplied nuclease. Aspects of the methods include contacting a cell with a targeting vector comprising nucleic acid sequence to be integrated into the target locus, where the cell is not also contacted with a nuclease. In addition, reagents, devices and kits thereof that find use in practicing the subject methods are provided.
Claims23
A method for the targeted integration of a transgene into the genome of a cell in the absence of an exogenously provided nuclease, the method comprising:
- contacting a cell with a recombinant viral vector, the recombinant viral vector comprising:
- i. a polynucleotide comprising a first nucleic acid sequence and a second nucleic acid sequence, wherein the first nucleic acid sequence encodes the transgene;
- and the second nucleic acid sequence is positioned 5′ or 3′ to the first nucleic acid sequence and promotes the production of two independent gene products upon integration into the target integration site in the genome of the cell;
- ii. a third nucleic acid sequence positioned 5′ to the polynucleotide and comprising sequence that is substantially homologous to genomic sequence 5′ of a target integration site in the genome of the cell;
- and
- iii. a fourth nucleic acid sequence positioned 3′ of the polynucleotide and comprising sequence that is substantially homologous to genomic sequence 3′ of a target integration site in the genome of the cell;
- wherein the cell is not contacted with a nuclease or nucleic acid encoding a nuclease.
A recombinant viral vector for integrating a transgene into a target integration site in the genome of the cell, comprising:
- a polynucleotide cassette comprising a first nucleic acid sequence and a second nucleic acid sequence, wherein the first nucleic acid sequence comprises the transgene;
- and the second nucleic acid sequence is positioned 5′ or 3′ to the first nucleic acid sequence and promotes the production of two independent gene products upon integration into the target integration site in the genome of the cell;
- a third nucleic acid sequence positioned 5′ to the polynucleotide cassette and comprising sequence that is substantially homologous to genomic sequence 5′ of the target integration site in the genome of the cell;
- and
- a fourth nucleic acid sequence positioned 3′ of the polynucleotide cassette and comprising sequence that is substantially homologous to genomic sequence 3′ of the target integration site in the genome of the cell.
Dependencies are inferred from the claim text.