LINEAR DONOR CONSTRUCTS FOR TARGETED INTEGRATION

摘要

Disclosed herein are linear donor molecules comprising homology arms of 50-750 base pairs (e.g., 50-100 base pairs) flanking one or more sequences of interest. The donor molecules and/or compositions comprising these molecules can be used in methods for targeted integration of an exogenous sequence into a specified region of interest in the genome of a cell.

主权项

1. A method for homology-dependent targeted integration of a sequence of interest into a region of interest in the genome of a cell, the method comprising the steps of:
(a) expressing a fusion protein in the cell, the fusion protein comprising a DNA-binding domain and cleavage domain or a cleavage half-domain, wherein the DNA-binding domain has been engineered to bind to a target site in the region of interest; (b) contacting the cell with a donor polynucleotide comprising a linear donor nucleic acid molecule comprising homology arms of between 50 and 100 base pairs and a sequence of interest, wherein the homology arms flank the sequence of interest, wherein binding of the fusion protein to the target site cleaves the genome of the cell in the region of the interest, thereby resulting in homology-dependent targeted integration of the sequence of interest into the genome of the cell.