期刊
NATURE COMMUNICATIONS
卷 7, 期 -, 页码 -出版社
NATURE PUBLISHING GROUP
DOI: 10.1038/ncomms10548
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资金
- National Institutes of Health [R01HL117491, R01HL129778]
- NATIONAL HEART, LUNG, AND BLOOD INSTITUTE [R01HL129778, R01HL117491] Funding Source: NIH RePORTER
Zinc-finger nuclease, transcription activator-like effector nuclease and CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 (CRISPR-associated protein 9) are becoming major tools for genome editing. Importantly, knock-in in several non-rodent species has been finally achieved thanks to these customizable nucleases; yet the rates remain to be further improved. We hypothesize that inhibiting non-homologous end joining (NHEJ) or enhancing homology-directed repair (HDR) will improve the nuclease-mediated knock-in efficiency. Here we show that the in vitro application of an HDR enhancer, RS-1, increases the knock-in efficiency by two- to five-fold at different loci, whereas NHEJ inhibitor SCR7 has minimal effects. We then apply RS-1 for animal production and have achieved multifold improvement on the knock-in rates as well. Our work presents tools to nuclease-mediated knock-in animal production, and sheds light on improving gene-targeting efficiencies on pluripotent stem cells.
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