Table 1 i-GONAD in various mouse strains
From: i-GONAD: a robust method for in situ germline genome engineering using CRISPR nucleases
Type of genome editing | Concentration of CRISPR components | Electroporator used | No. of female mice treated | Strain | Locus | No. of pregnant mice | No. G0 pups/fetuses obtained | No. of G0 pups/fetuses with modified allele (%) | No. of G0 pups/fetuses with intended allele (%) |
|---|---|---|---|---|---|---|---|---|---|
Knock-out | Cas9 protein (1 mg/ml) Integrated DNA Technologies (IDT) | NEPA21 | 7 | C3H/HeSlc (inbred) | Tyr | 3 | 13 | 12 (92%) | 12 (92%) both alleles |
Tyr-crRNA (30 μM) FASMAC | |||||||||
tracrRNA (30 μM) FASMAC | |||||||||
Knock-out | Cas9 protein (1 mg/ml) IDT | NEPA21 | 6 | C3H/HeSlc (inbred) | Kit | 4 | 16 | 11 (69%) | 7 (44%) both alleles |
Kit-crRNA (30 μM) FASMAC | 4 (25%) one allele | ||||||||
tracrRNA (30 μM) FASMAC | |||||||||
Knock-out | Cas9 protein (1 mg/ml) IDT | NEPA21 | 11 | C57BL/6NCrSlc (inbred) | Tyr | 0 | 0 | 0 (0%) | 0 (0%) |
Tyr-crRNA (30 μM) FASMAC | |||||||||
tracrRNA (30 μM) FASMAC | |||||||||
Knock-out | Cas9 protein (1 mg/ml) IDT | NEPA21 | 7 | C57BL/6NCrSlc (inbred) | Kit | 2 | 3 | 3 (100%) | 1 (33%) both alleles |
Kit-crRNA (30 μM) FASMAC | 2 (67%) one allele | ||||||||
tracrRNA (30 μM) FASMAC | |||||||||
Knock-out | Cas9 protein (1 mg/ml) IDT | NEPA21 | 10 | DBA/2CrSlc (inbred) | Tyr | 4 | 6 | 6 (100%) | 6 (100%) both alleles |
Tyr-crRNA (30 μM) FASMAC | |||||||||
tracrRNA (30 μM) FASMAC | |||||||||
Knock-out | Cas9 protein (1 mg/ml) IDT | NEPA21 | 4 | B6D2F1/Slc (hybrid) | Tyr | 4 | 23 | 20 (87%) | 19 (83%) both alleles |
Tyr-crRNA (30 μM) FASMAC | 1 (4%) one allele | ||||||||
tracrRNA (30 μM) FASMAC | |||||||||
Knock-out | Cas9 protein (1 mg/ml) IDT | NEPA21 | 4 | B6D2F1/Slc (female) x C57BL/6NCrSlc (male) (hybrid) | Tyr | 4 | 19 | 18 (95%) | 15 (79%) both alleles |
Tyr-crRNA (30 μM) FASMAC | 3 (16%) one allele | ||||||||
tracrRNA (30 μM) FASMAC | |||||||||
ssODN knock-in | Cas9 protein (1 mg/ml) IDT | NEPA21 | 5 | C57BL/6NCrl (inbred) | Cdkn1a | 2 | 13 | 5 (38%) | 4 (31%) |
crRNA-p21 (30 μM) IDT | |||||||||
tracrRNA (30 μM) IDT | |||||||||
ssODN-p21 (1 μg/μl) Eurofins Genomics | |||||||||
ssODN knock-in | Cas9 protein (1 mg/ml) IDT | NEPA21 | 4 | C57BL/6NCrl (inbred) | Cdkn2a | 2 | 10 | 7 (70%) | 4 (40%) |
crRNA-p16/p19 (30 μM) IDT | |||||||||
tracrRNA (30 μM) IDT | |||||||||
ssODN-p16/p19 (1 μg/μl) Eurofins Genomics | |||||||||
ssODN knock-in | Cas9 protein (1 mg/ml) IDT | CUY21EDIT II | 4 | BALB/cAJcl (inbred) | Tyr | 2 | 6 | 3 (50%) | 1 (17%) |
crRNA-ICR-tyr (30 μM) IDT | |||||||||
tracrRNA (30 μM) IDT | |||||||||
ssODN-tyr (2 μg/μl) IDT |