Artificial restriction enzymes can be generated by fusing a natural or engineered DNA-binding domain to a nuclease domain (often the cleavage domain of the type IIS restriction enzyme FokI). Such artificial restriction enzymes can target large DNA sites (up to 36 bp) and can be engineered to bind to desired DNA sequences. Zinc finger nucleases are the most commonly used artificial restriction enzymes and are generally used in genetic engineering applications, but can also be used for more standard gene cloning applications. Other artificial restriction enzymes are based on the DNA binding domain of TAL effectors.
In 2013, a new technology CRISPR-Cas9, based on a prokaryotic viral defense system, was engineered for editing the genome, and it was quickly adopted in laboratories. For more detail, read CRISPR (Clustered regularly interspaced short palindromic repeats).
In 2017, a group from University of Illinois reported using an Argonaute protein taken from Pyrococcus furiosus (PfAgo) along with guide DNA to edit DNA in vitro as artificial restriction enzymes.
Artificial ribonucleases that act as restriction enzymes for RNA have also been developed. A PNA-based system, called a PNAzyme, has a Cu(II)-2,9-dimethylphenanthroline group that mimics ribonucleases for specific RNA sequence and cleaves at a non-base-paired region (RNA bulge) of the targeted RNA formed when the enzyme binds the RNA. This enzyme shows selectivity by cleaving only at one site that either does not have a mismatch or is kinetically preferred out of two possible cleavage sites.