Methods and compositions for targeted genomic deletion

Abstract

Disclosed herein are compositions and methods for generating chromosomal translocations and targeted deletions of specific lengths and at specific locations the genome of cell.

1 . A method for creating a targeted deletion of specific length and specific borders in a genomic locus in a host cell, the method comprising
 introducing one or more nucleases and a donor polynucleotide into the host cell, wherein   (i) the one or more nucleases each comprise a DNA-binding domain that recognizes a target sequence in the genomic locus and further wherein the one or more nucleases cleave the genomic locus;   (ii) the donor polynucleotide comprises at least one of the target sequences recognized by one of the nucleases and regions of homology to the genomic locus and further wherein the donor polynucleotide includes a deletion relative to the target sequence of specific length and specific borders; and   (iii) the donor polynucleotide is introduced into the genomic locus such that a targeted deletion of specific length and specific borders is generated in the genomic locus.   
     
     
         2 . The method of  claim 1 , wherein the regions of homology are between about 50 and 1500 base pairs in length. 
     
     
         3 . The method of  claim 1  wherein the nuclease is selected from the group consisting of a zinc finger nuclease (ZFN), a meganuclease and combinations thereof. 
     
     
         4 . The method of  claim 1 , wherein the one or more nucleases cleave the genomic locus at two locations. 
     
     
         5 . The method of  claim 1 , wherein between the regions of homology, the donor polynucleotide comprises a sequence selected from the group consisting of a coding sequence, a 2A peptide, an SA site, an IRES, a shRNA molecule, an miRNA molecule, an RNAi and combinations thereof. 
     
     
         6 . The method of  claim 1 , wherein the targeted deletion recapitulates a known structural variant at the genomic locus. 
     
     
         7 . The method of  claim 1 , wherein the host cell is a eukaryotic cell. 
     
     
         8 . The method of  claim 7 , wherein the eukaryotic cell is a mammalian or plant cell. 
     
     
         9 . A method of producing a chromosomal translocation, the method comprising:
 cleaving first and second chromosomes using a one or more nucleases in the presence of a donor polynucleotide, the donor molecule including regions of homology to the cleaved first and second chromosomal fragments such that the first and second cleaved chromosomes are joined and a translocated chromosome is produced.   
     
     
         10 . The method of  claim 9 , wherein the translocation is associated with a disease or disorder. 
     
     
         11 . The method of  claim 10 , wherein the disease is a cancer.