Re-engineered RNA-Guided FokI-Nucleases for Improved Genome Editing in Human Cells.
Mol Ther
; 25(2): 342-355, 2017 02 01.
Article
en En
| MEDLINE
| ID: mdl-28153087
ABSTRACT
Clustered regularly interspaced palindromic repeats (CRISPR)/Cas9 enables us to generate targeted sequence changes in the genomes of cells and organisms. However, off-target effects have been a persistent problem hampering the development of therapeutics based on CRISPR/Cas9 and potentially confounding research results. Efforts to improve Cas9 specificity, like the development of RNA-guided FokI-nucleases (RFNs), usually come at the cost of editing efficiency and/or genome targetability. To overcome these limitations, we engineered improved chimeras of RFNs that enable higher cleavage efficiency and provide broader genome targetability, while retaining high fidelity for genome editing in human cells. Furthermore, we developed a new RFN ortholog derived from Staphylococcus aureus Cas9 and characterize its utility for efficient genome engineering. Finally, we demonstrate the feasibility of RFN orthologs to functionally hetero-dimerize to modify endogenous genes, unveiling a new dimension of RFN target design opportunities.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Desoxirribonucleasas de Localización Especificada Tipo II
/
Ingeniería de Proteínas
/
ARN Guía de Kinetoplastida
/
Edición Génica
Tipo de estudio:
Prognostic_studies
Límite:
Humans
Idioma:
En
Revista:
Mol Ther
Asunto de la revista:
BIOLOGIA MOLECULAR
/
TERAPEUTICA
Año:
2017
Tipo del documento:
Article
País de afiliación:
Singapur