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. 2014:546:21-45.
doi: 10.1016/B978-0-12-801185-0.00002-7.

Targeted genome editing in human cells using CRISPR/Cas nucleases and truncated guide RNAs

Yanfang Fu  1 , Deepak Reyon  1 , J Keith Joung  2
Affiliations

Targeted genome editing in human cells using CRISPR/Cas nucleases and truncated guide RNAs

Yanfang Fu et al. Methods Enzymol. 2014.

Abstract

CRISPR RNA-guided nucleases have recently emerged as a robust genome-editing platform that functions in a wide range of organisms. To reduce off-target effects of these nucleases, we developed and validated a modified system that uses truncated guide RNAs (tru-gRNAs). The use of tru-gRNAs leads to decreases in off-target effects and does not generally compromise the on-target efficiencies of these genome-editing nucleases. In this chapter, we describe guidelines for identifying potential tru-gRNA target sites and protocols for measuring the on-target efficiencies of CRISPR RNA-guided nucleases in human cells.

Keywords: CRISPR RNA-guided nucleases; CRISPR/Cas nucleases; Cas9; Genome editing; Off-target effects; Off-target mutations; Truncated guide RNAs.

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