Table 2 Overall strategies that produced high intended transgene integration outcomes
Outcome | Strategy | Cas9 variant | Transient GFP+ (%) | Selected GFP+ (%) | Consensus conclusion |
|---|---|---|---|---|---|
High efficiency high fidelity | Long HA | Cas9-SC | 65.4 | 81.1 | Long HA with SC system |
Long HA | Cas9-Gem-SC | 66.5 | 80.7 | ||
Long HA | Cas9es-Gem-SC | 64.4 | 78.9 | ||
Long HA | Cas9es-SC | 68.8 | 78.3 | ||
High efficiency low fidelity | Short HA | Cas9-SC | 64.9 | 29.8 | Blunt end-mediated KI strategies with SC system |
Short HA | Cas9es-Gem-SC | 61.5 | 32.8 | ||
Short HA | Cas9-Gem-SC | 60.8 | 38.4 | ||
Matched end joining | Cas9-Gem-SC | 60.0 | 27.6 | ||
Matched end joining | Cas9es-SC | 57.3 | 18.7 | ||
Short HA | Cas9es-SC | 57.0 | 39.5 | ||
Matched end joining | Cas9es-Gem-SC | 55.3 | 33.9 | ||
Matched end joining | Cas9-SC | 54.5 | 14.4 | ||
Low efficiency high fidelity | Long HA | Cas9es-Gem | 27.7 | 77.0 | Majority overhang DSB mediated KI strategies with Geminin (lesser extent SC) and long HA non-SC |
5ʹ overhang-Pair1 | Cas9D10Aes-Gem | 20.3 | 63.4 | ||
3ʹ overhang-Pair5 | Cas9H840A-Gem-SC | 2.3 | 62.4 | ||
5ʹ overhang-Pair5 | Cas9D10A | 4.3 | 61.1 | ||
3ʹ overhang-Pair5 | Cas9H840Aes-Gem | 19.9 | 59.4 | ||
long HA | Cas9-Gem | 15.5 | 58.7 | ||
5ʹ overhang-Pair1 | Cas9D10Aes-SC | 37.2 | 56.5 | ||
3ʹ overhang-Pair5 | Cas9H840Aes-Gem-SC | 1.3 | 53.6 | ||
3ʹ overhang-Pair5 | Cas9H840A-Gem | 5.9 | 53.5 | ||
3ʹ overhang-Pair1 | Cas9H840Aes | 5.7 | 52.4 |