SplitAx: A novel method to assess the function of engineered nucleases.

ABSTRACT: Engineered nucleases have been used to generate knockout or reporter cell lines and a range of animal models for human disease. These new technologies also hold great promise for therapeutic genome editing. Current methods to evaluate the activity of these nucleases are time consuming, require extensive optimization and are hampered by readouts with low signals and high background. We have developed a simple and easy to perform method (SplitAx) that largely addresses these issues and provides a readout of nuclease activity. The assay involves splitting the N-terminal (amino acid 1-158) coding region of GFP and an out-of-frame of C-terminal region with a nuclease binding site sequence. Following exposure to the test nuclease, cutting and repair by error prone non-homologous end joining (NHE

SUBMITTER: Axton RA 

PROVIDER: S-EPMC5315338 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature