The efficacy and the mutation spectrum of genome editing methods can vary substantially depending on the targeted sequence. A simple, quick assay to accurately characterize and quantify the induced mutations is therefore needed. Here we present TIDE, a method for this purpose that requires only a pair of PCR reactions and two standard capillary sequencing runs. The sequence traces are then analyzed by a specially developed decomposition algorithm that identifies the major induced mutations in the projected editing site and accurately determines their frequency in a cell population. This method is cost-effective and quick, and it provides much more detailed information than current enzyme-based assays. An interactive web tool for automated decomposition of the sequence traces is available. TIDE greatly facilitates the testing and rational design of genome editing strategies.

Easy quantitative assessment of genome editing by sequence trace decomposition

Amendola M;
2014-01-01

Abstract

The efficacy and the mutation spectrum of genome editing methods can vary substantially depending on the targeted sequence. A simple, quick assay to accurately characterize and quantify the induced mutations is therefore needed. Here we present TIDE, a method for this purpose that requires only a pair of PCR reactions and two standard capillary sequencing runs. The sequence traces are then analyzed by a specially developed decomposition algorithm that identifies the major induced mutations in the projected editing site and accurately determines their frequency in a cell population. This method is cost-effective and quick, and it provides much more detailed information than current enzyme-based assays. An interactive web tool for automated decomposition of the sequence traces is available. TIDE greatly facilitates the testing and rational design of genome editing strategies.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11369/434223
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