Cationic Polymer-Mediated CRISPR/Cas9 Plasmid Delivery for Genome Editing.

Delivery of CRISPR (clustered regularly interspaced short palindromic repeats)/CRISPR-associated protein-9 (Cas9) represents a major hurdle for successful clinical translation of genome editing tools. Owing to the large size of plasmids that encode Cas9 and single-guide RNA (sgRNA), genome editing efficiency mediated by current delivery carriers is still unsatisfactory to meet the requirement for its real applications. Herein, cationic polymer polyethyleneimine-beta-cyclodextrin (PC), known to be efficient for small plasmid transfection, is reported to likewise mediate efficient delivery of plasmid encoding Cas9 and sgRNA. Whereas PC can condense and encapsulate large plasmids at high N/P ratio, the delivery of plasmid results in efficient editing at two genome loci, namely, hemoglobin subunit beta (19.1%) and rhomboid 5 homolog 1 (RHBDF1) (7.0%). Sanger sequencing further confirms the successful genome editing at these loci. This study defines a new strategy for the delivery of the large plasmid encoding Cas9/sgRNA for efficient genome editing.