Genomic editing using CRISPR/Cas is widely used in recent years for its relative simple mechanism. However, it is also restricted by the recognition of necessary sequence called PAM site. For most common enzyme of this system, SpCas9, recognizes “NGG” sequence for activation of digestion. (N refers to any nucleotide)
In order to expand the capacity of CRISPR/Cas system to achieve much extensive genomic editing, Benjamin P. Kleinstiver from Massachusetts General Hospital modified the spCas9 enzyme into different versions (by mutating specific amino acids, creating new function sites) for recognizing more types of PAMs, such as SpG for recognizing “NGN” PAMs, and spRY for recognizing “NRN and NYN” PAMs. (R refers to A and G, Y refers to C and T) However, with more targeting sites for CRISPR/Cas system, comes higher “off-target” chance. The feasibility of these new enzymes still needs to be tested before being applied to aiming previously inaccessible genetic diseases.
Walton, R., Christie, K., Whittaker, M. and Kleinstiver, B., 2020. Unconstrained genome targeting with near-PAMless engineered CRISPR-Cas9 variants. Science, p.eaba8853.