Unconstrained genome targeting with near-PAMless engineered CRISPR-Cas9 variants

Cas9 (SpCas9) to eliminate the NGG PAM requirement. We developed a variant named SpG that is capable of targeting an expanded set of NGN PAMs, and we further optimized this enzyme to develop a near-PAMless SpCas9 variant named SpRY (NRN and to a lesser extent NYN PAMs). SpRY nuclease and base-editor variants can target almost all PAMs, exhibiting robust activities on a wide range of sites with NRN PAMs in human cells and lower but substantial activity on those with NYN PAMs. Using SpG and SpRY, we generated previously inaccessible disease-relevant genetic variants, supporting the utility of high-resolution targeting across genome editing applications.

• MQ
  Margaret Q. Landenberger Research Foundation
• NH
  National Heart, Lung, and Blood Institute

  Award: P01-HL142494
• NC
  National Cancer Institute

  Award: R00-CA218870