Table 1.
CRISPR/Cas9 protein-mediated genome editing with ssODN matrix.
| Target locus | Cas9 (µM)/sgRNA (ng/µl)/ssODN (ng/µl) | Poring pulse conditions: voltage/pulse width | No. of viable embryos/No. of total embryos (%)a | Stage of analysis | No. of embryos or pups analyzed/No. of transferred (%)b | No. of NHEJ+ (%)c | No. of KI+ (%) c | No. of NHEJ+ KI+ (%)c | No. of embryos/pups with 3 or more alleles (%)c |
|---|---|---|---|---|---|---|---|---|---|
| Ephx2 | 50*/10/15 | Microinjection | 293/373 (79) | pups | 82/250 (33) | 17 (21) | 12 (15) | 8 (9.8) | 9 (11) |
| 3/150/150 | 300 V/0.5 ms | 85/85 (100) | E14 | 20/63 (32) | 10 (50) | 1 (5)# | 1 (5)# | 4 (20) | |
| 300 V/2.5 ms | 97/134 (72) | E14 | 20/87 (23) | 12 (60) | 5 (25)# | 3 (15)# | 2 (10) | ||
| 6/200/200 | 250 V/0.5 ms | 62/70 (89) | E14 | 5/48 (10) | 0 (0) | 1 (20) | 0 (0) | 0 (0) | |
| 250 V/2.5 ms | 36/54 (67) | E14 | 1/36 (3) | 0 (0) | 0 (0) | 0 (0) | / | ||
| 300 V/0.5 ms | 31/36 (86) | E14 | 3/31 (10) | 1 (33) | 0 (0) | 0 (0) | 0 (0) | ||
| 300 V/2.5 ms | 14/47 (30) | E14 | 0/14 (0) | / | / | / | / | ||
| FlnA | 3/100/20 | Microinjection | 121/144 (84) | pups | 21/111 (19) | 3 (14) | 2 (9.6) | 1 (4.8) | 2 (9.5) |
| 3/150/150 | 300 V/0.5 ms | 100/103 (97) | pups | 12/100 (12) | 1 (8) | 1 (8)# | 0 (0) | 0 (0) | |
| 300 V/2.5 ms | 70/119 (59) | pups | 1†/100 (1.4) | 1† (100) | 0 (0) | 0 (0) | 0 (0) |
*Cas9 mRNA (ng/µl). aCalculated 1 h after electroporation. bCalculated from the number of transferred eggs. cCalculated from the number of analyzed embryos or pups. †Stillborn pup. #Sequences detailed in Fig. 1.