Orthogonal Cas9-Cas9 chimeras provide a versatile platform for genome editing

Nat Commun. 2018 Nov 19;9(1):4856. doi: 10.1038/s41467-018-07310-x.


The development of robust, versatile and accurate toolsets is critical to facilitate therapeutic genome editing applications. Here we establish RNA-programmable Cas9-Cas9 chimeras, in single- and dual-nuclease formats, as versatile genome engineering systems. In both of these formats, Cas9-Cas9 fusions display an expanded targeting repertoire and achieve highly specific genome editing. Dual-nuclease Cas9-Cas9 chimeras have distinct advantages over monomeric Cas9s including higher target site activity and the generation of predictable precise deletion products between their target sites. At a therapeutically relevant site within the BCL11A erythroid enhancer, Cas9-Cas9 nucleases produced precise deletions that comprised up to 97% of all sequence alterations. Thus Cas9-Cas9 chimeras represent an important tool that could be particularly valuable for therapeutic genome editing applications where a precise cleavage position and defined sequence end products are desirable.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Bacterial Proteins / genetics*
  • Bacterial Proteins / metabolism
  • Base Sequence*
  • CRISPR-Cas Systems*
  • Carrier Proteins / genetics
  • Carrier Proteins / metabolism
  • Endonucleases / genetics*
  • Endonucleases / metabolism
  • Gene Editing / methods*
  • Genetic Engineering
  • Genome, Human
  • HEK293 Cells
  • Humans
  • Jurkat Cells
  • K562 Cells
  • Mutant Chimeric Proteins / genetics*
  • Mutant Chimeric Proteins / metabolism
  • Neisseria meningitidis / enzymology
  • Neisseria meningitidis / genetics
  • Nuclear Proteins / genetics
  • Nuclear Proteins / metabolism
  • Repressor Proteins
  • Sequence Deletion*
  • Staphylococcus aureus / enzymology
  • Staphylococcus aureus / genetics


  • BCL11A protein, human
  • Bacterial Proteins
  • Carrier Proteins
  • Mutant Chimeric Proteins
  • Nuclear Proteins
  • Repressor Proteins
  • Endonucleases