. Author manuscript; available in PMC: 2021 Mar 13.

Published in final edited form as: Curr Opin Genet Dev. 2020 Jul 18;65:160–168. doi: 10.1016/j.gde.2020.05.040

Table 1. Potential genome editing strategies for next generation NHP models.

G: generation, KO: knockout, CNV: copy number variation, LD: large deletion, DSBs: DNA double strand breaks, RNP: ribonucleoprotein, HDR: homology-directed repair.

	Types	Methods	Pros	Cons	New improvements
1^st Generation	KO	Indel	Very efficient	Unpredictable outcomes Unexpected truncated protein	Predictable algorithms [65] Entire gene deletion [66]
1^st Generation	CNV (LD)	2-DSBs	Efficient	Unpredictable outcomes	Bridging donor with RNP [67]
2^nd Generation	SNV	HDR	Precise	Variable efficiency Indel formation Needs donor DNA	RNP [34] Donor tethering [68] HDR enhancers [69]
		Base Editing	Precise Efficient No DSB	Deaminase DNA and RNA off-target mutations C>T/G>A, A>G/T>C only Bystander editing Limited target window	New variants without deaminase off-target [39] PAMless Cas9 [70]
		Prime Editing	Precise Efficient Any SNVs Few DSB	Not tested in animals yet Needs pegRNA and nicking sgRNA optimization	None yet
3rd Generation	Humanized	HDR	Precise Up to several kb	Variable efficiency Indel formation Needs donor DNA	RNP [34] Donor tethering and 2-cell stage injection [71] HDR enhancers [72]
3rd Generation	Humanized	Prime Editing	Precise Efficient Few DSB	Needs optimization Not tested in animals yet Currently, up to ~40bp	None yet