Use of Endonucleases for Inserting Transgenes Into Safe Harbor Loci
AssigneeCellectis
InventorsOlivier DANOS, Aymeric Duclert
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Abstract
The present invention concerns the endonucleases capable of cleaving a target sequence located in a “safe harbor loci”, i.e. a loci allowing safe expression of a transgene. The present invention further concerns the use of such endonucleases for inserting transgenes into a cell, tissue or individual.
Claims21
A variant endonuclease capable of cleaving a target sequence for use in inserting a transgene into the genome of an individual, wherein
- i. said genome comprises a locus comprising said target sequence;
- and
- ii. said target sequence is located at a distance of at most 200 kb from a retroviral insertion site (RIS), wherein said RIS is neither associated with cancer nor with abnormal cell proliferation.
A variant dimeric I-Crel protein comprising two monomers that each comprises a sequence at least 80% identical to SEQ ID NO:
- 1 or SEQ ID NO:
- 42, wherein:
- i. said dimeric I-Crel protein is capable of cleaving a target sequence located within a locus of an individual, said target sequence being located at a distance of at most 200 kb from a retroviral insertion site (RIS), and said RIS being neither associated with cancer nor with abnormal cell proliferation;
- and
- ii. said target sequence does not comprise a sequence of SEQ ID NO:
- 4.
A method for obtaining an endonuclease suitable for inserting a transgene into the genome of an individual, comprising the step of:
- a) selecting, within the genome of said individual, a retroviral insertion site (RIS) that is neither associated with cancer nor with abnormal cell proliferation;
- b) defining a genomic region extending 200 kb upstream and 200 kb downstream of said RIS;
- and
- identifying a wild-type endonuclease or constructing a variant endonuclease capable of cleaving a target sequence located within said genomic region.
Dependencies are inferred from the claim text.