Journal
NUCLEIC ACIDS RESEARCH
Volume 44, Issue 9, Pages -Publisher
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkw064
Keywords
-
Categories
Funding
- Research Grants Council of Hong Kong [CUHK478812, CUHK14102214, CUHK14104614, HKUST T13-607/12R]
- National Natural Science Foundation of China [NSFC 31171433]
- National Basic Research Program of China [973-Program] [2015CB964700]
- Shenzhen SZSIA foundation [JCYJ20140425184428469]
- CUHK graduate school scholarship
- National Basic Research Program of China
Ask authors/readers for more resources
CRISPR/Cas9-induced site-specific DNA double-strand breaks (DSBs) can be repaired by homology-directed repair (HDR) or non-homologous end joining (NHEJ) pathways. Extensive efforts have been made to knock-in exogenous DNA to a selected genomic locus in human cells; which, however, has focused on HDR-based strategies and was proven inefficient. Here, we report that NHEJ pathway mediates efficient rejoining of genome and plasmids following CRISPR/Cas9-induced DNA DSBs, and promotes high-efficiency DNA integration in various human cell types. With this homology-independent knock-in strategy, integration of a 4.6 kb promoterless ires-eGFP fragment into the GAPDH locus yielded up to 20% GFP+ cells in somatic LO2 cells, and 1.70% GFP+ cells in human embryonic stem cells (ESCs). Quantitative comparison further demonstrated that the NHEJ-based knock-in is more efficient than HDR-mediated gene targeting in all human cell types examined. These data support that CRISPR/Cas9-induced NHEJ provides a valuable new path for efficient genome editing in human ESCs and somatic cells.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available