Genome editing with engineered zinc finger nucleases

Abstract

Reverse genetics in model organisms such as Drosophila melanogaster, Arabidopsis thaliana, zebrafish and rats, efficient genome engineering in human embryonic stem and induced pluripotent stem cells, targeted integration in crop plants, and HIV resistance in immune cells - this broad range of outcomes has resulted from the application of the same core technology: targeted genome cleavage by engineered, sequence-specific zinc finger nucleases followed by gene modification during subsequent repair. Such 'genome editing' is now established in human cells and a number of model organisms, thus opening the door to a range of new experimental and therapeutic possibilities.

Tapping natural reservoirs of homing endonucleases for targeted gene modification

“…While many engineered ZFNs have been used successfully to modify chromosomal target loci in a variety of multicellular eukaryotic organisms (5,39), to date only the homodimeric I-CreI enzyme has been engineered to modify endogenous chromosomal loci (11,12). However, the properties of monomeric LHEs argue for their continued development as genome-modifying enzymes: They are encoded by particularly short open reading frames that are less than 1,000 base pairs in length, and their cleavage specificities are often very high (18)(19)(20)(21)(22).…”

“…S everal types of highly specific DNA recognition and cleavage enzymes, including homing endonucleases (HEs), zinc finger nucleases (ZFNs), and transcription activator-like (TAL) effector nucleases, are being developed for targeted gene modification, ranging from gene disruption to corrective gene therapy (1)(2)(3)(4)(5). Regardless of the identity of the protein scaffold, site-specific gene modification generally requires the creation of individually tailored site-specific endonucleases that generate double-strand breaks (DSBs) at unique chromosomal targets.…”

“…Regardless of the identity of the protein scaffold, site-specific gene modification generally requires the creation of individually tailored site-specific endonucleases that generate double-strand breaks (DSBs) at unique chromosomal targets. These lesions induce intrinsic DNA repair responses in the targeted cells (primarily homologous recombination and nonhomologous end joining) that lead to DNA sequence alterations at the target (3,5).…”

Dissecting ant recognition systems in the age of genomics

“…[42]). Similarly, recent advances have been made in model organisms using genome editing with engineered nucleases [43], such as transcription activator-like effector nucleases [44,45] and zinc finger nucleases [46]. Although these powerful genetic tools have not yet been developed for ants, the rapid pace of development in model organisms suggests that these techniques may soon find their way into studies of social insect biology.…”

Don’t edit the human germ line

“…One of us (F.U.) helped to develop the first genome-editing technology, zinc-finger nucleases 2 (ZFNs), and is now senior scientist at the company developing them, Sangamo BioSciences of Richmond, California. The Alliance for Regenerative Medicine (ARM; in which E.L., M.W.…”