Rapid and highly efficient mammalian cell engineering via Cas9 protein transfection.
AuthorsLiang X, Potter J, Kumar S, Zou Y, Quintanilla R, Sridharan M, Carte J, Chen W, Roark N, Ranganathan S, Ravinder N, Chesnut JD,
Journal
PubMed ID26003884
'CRISPR-Cas9 systems provide a platform for high efficiency genome editing that are enabling innovative applications of mammalian cell engineering. However, the delivery of Cas9 and synthesis of guide RNA (gRNA) remain as steps that can limit overall efficiency and ease of use. Here we describe methods for rapid synthesis of ... More
Enhanced CRISPR/Cas9-mediated precise genome editing by improved design and delivery of gRNA, Cas9 nuclease, and donor DNA.
AuthorsLiang X, Potter J, Kumar S, Ravinder N, Chesnut JD
JournalJ Biotechnol
PubMed ID27845164
'While CRISPR-based gene knock out in mammalian cells has proven to be very efficient, precise insertion of genetic elements via the cellular homology directed repair (HDR) pathway remains a rate-limiting step to seamless genome editing. Under the conditions described here, we achieved up to 56% targeted integration efficiency with up ... More
Improved delivery of Cas9 protein/gRNA complexes using lipofectamine CRISPRMAX.
AuthorsYu X, Liang X, Xie H, Kumar S, Ravinder N, Potter J, de Mollerat du Jeu X, Chesnut JD,
JournalBiotechnol Lett
PubMed ID26892225
'To identify the best lipid nanoparticles for delivery of purified Cas9 protein and gRNA complexes (Cas9 RNPs) into mammalian cells and to establish the optimal conditions for transfection. Using a systematic approach, we screened 60 transfection reagents using six commonly-used mammalian cell lines and identified a novel transfection reagent (named ... More