An easy and efficient inducible CRISPR/Cas9 platform with improved specificity for multiple gene targeting.

ABSTRACT: The CRISPR/Cas9 system is a powerful genome editing tool and has been widely used for biomedical research. However, many challenges, such as off-target effects and lack of easy solutions for multiplex targeting, are still limiting its applications. To overcome these challenges, we first developed a highly efficient doxycycline-inducible Cas9-EGFP vector. This vector allowed us to track the cells for uniform temporal control and efficient gene disruption, even in a polyclonal setting. Furthermore, the inducible CRISPR/Cas9 system dramatically decreased off-target effects with a pulse exposure of the genome to the Cas9/sgRNA complex. To target multiple genes simultaneously, we established simple one-step cloning approaches for expression of multiple sgRNAs with improved vectors. By combining

SUBMITTER: Cao J 

PROVIDER: S-EPMC5100567 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature