Table 1.
Detection and efficiency of CRISPR/Cas9-mediated indel mutation at Ramp2 locus at blastocyst stage
| Experiment no. | Site injected 1 | Cas9 mRNA: R2 gRNA (ng/μL) | No. zygotes injected | No. blastocysts/no. zygotes cultures (%) | No. mutant blastocysts/no. blastocysts tested (%) 2 |
|---|---|---|---|---|---|
| 1 |
cyto |
100:50 |
25 |
19/21 (90) |
6/21 (32) |
| 2 |
pn/cyto |
100:50 |
15 |
8/10 (80) |
2/8 (25) |
| 3 | pn/cyto | 200:50 | 24 | 8/17 (47) | 8/8 (100)3 |
1Injection was performed toward cyto: cytoplasm; pn/cyto pronuclei and cytoplasm.
2Mutated blastocysts were identified by both T7EndonucleaseI digestion and sequencing.
3Representative results from experiment 3 are shown in Figure 2B and C.