High-frequency genome editing using ssDNA oligonucleotides with zinc-finger nucleases

Abstract

Single-stranded oligonucleotides are used as donor templates for zinc-finger nucleases to create targeted genomic deletions and mutations in mammalian cell lines. Zinc-finger nucleases (ZFNs) have enabled highly efficient gene targeting in multiple cell types and organisms. Here we describe methods for using simple ssDNA oligonucleotides in tandem with ZFNs to efficiently produce human cell lines with three distinct genetic outcomes: (i) targeted point mutation, (ii) targeted genomic deletion of up to 100 kb and (iii) targeted insertion of small genetic elements concomitant with large genomic deletions.