CompoZr® zinc finger nucleases (ZFNs) from Sigma® Life Science enable efficient and affordable gene editing across many species and cell types, including humans, rats, mice, frogs, rabbits, pigs, cattle, C. elegans, and Drosophila.
Recent literature has shown ZFNs mediating increasingly diverse types of in vitro genome modifications, some of which we describe in this tutorial, as well as the potential of in vivo genome editing through the restoration of hemostasis in a mouse model of hemophilia. Simultaneously, advances inCompoZr ZFN production and design have halved the price of both Custom ZFNs and Knockout ZFNs for any gene in the human, mouse or rat genomes, bringing CompoZr ZFNs within the budget of any laboratory.
CompoZr ZFNs work by creating a user-defined double-stranded break (DSB) that is repaired primarily by one of two natural mechanisms—nonhomologous end joining (NHEJ) or homology-directed repair (HDR). NHEJ can create nucleotide insertions or deletions (indels) that lead to frameshifts, subsequent nonsense-mediated decay, and functional gene knockout.
In the presence of a DNA donor plasmid that flanks the DSB, HDR can integrate entire transgenes, fuse reporter genes, introduce or correct a point mutation, or perform many other sophisticated genomic modifications.
Donor plasmid design, while an established process, can require several weeks from initial concept to delivery and costs hundreds of dollars. A recent Nature Methods publication by Chen et al. reports the use of ssDNA oligonucleotides (ssODNs), as opposed to donor plasmids, to derive three unique genetic outcomes—targeted point mutation, targeted gene deletion up to 100 kb using a single CompoZr ZFN, and targeted insertion of genetic elements along with large genomic deletions.
In this article, we describe the modification of a targeted codon in the human RPS6KA3 gene that encodes kinase RSK2, which is implicated in cancer, mental retardation, and psychomotor and skeletal disorders.