Patent ID: 9005973
Filing Date: 2015-04-14
Classification: C07K,C12N

Abstract:
1. A method of non-homologous end joining (NHEJ)-mediated targeted integration of a linear nucleic acid molecule into the genome of a mammalian cell, the method comprising: introducing a linear nucleic acid molecule into the mammalian cell, the linear nucleic acid molecule comprising a double-stranded sequence of interest having first and second ends and first and second single-stranded nucleotides between 1 and 10 nucleotides in length at the first and second ends of the double-stranded sequence; and creating a double-stranded break using a nuclease in an endogenous locus of the genome of the mammalian cell such that the linear nucleic acid molecule is integrated at the site of the double-stranded break and further wherein the nuclease comprises a TALE domain and the first and second single-stranded nucleotides are 100% complementary to the overhangs of the double-stranded break in the mammalian genome created by the nuclease; and annealing the first and second single-stranded nucleotides of the donor to the overhangs of the double-stranded break in the mammalian genome, thereby directly integrating the donor at the site of double-stranded break by NHEJ-mediated targeted integration.