Table 2 Comparison of Cas9–HE and Cas9 in obtaining targeted transgene integration into the rat Rosa26 and Il22bp loci

From: CtIP fusion to Cas9 enhances transgene integration by homology-dependent repair

Cas9 form

Target locus

KI fragments

Dose Cas9 mRNA/sgRNA/donor DNA (ng/µl)

Eggs injected (survival rate %)

Eggs transferred

E14 embryos (% transferred)

Indels + (% of E14)

Donor integration+ (% of E14)

HDR+ (% of E14)

Cas9–HE

Rosa26

CAG-GFP

50/10/2

216 (75%)

154

37 (24%)

29 (78%)

5 (13%)

3 (8%)

Cas9

Rosa26

CAG-GFP

50/10/2

284 (77%)

211

84 (39%)

62 (73%)

2 (2%)

1 (1%)

Cas9–HE

Il22bp

Il22bp-2A-GFP

50/10/2

277 (78%)

217

82 (37%)

45 (55%)

7 (8%)

2 (2%)

Cas9

Il22bp

Il22bp-2A-GFP

50/10/2

286 (80%)

226

91 (40%)

55 (60%)

8 (8%)

4 (4%)

  1. HDR+, HDR-mediated GFP transgene integration was defined as positive in E14 embryos for which both 5′ and 3′ in–out PCRs were detected using primers in the transgene and outside the homology arms (with PCR products confirmed by DNA sequencing, schematic of PCR primer positions in Supplementary Fig. 5); indels+, E14 embryos in which indels were detected by sequencing of PCR amplicons from the targeted genomic region; donor integration, E14 embryos in which PCR amplification of the transgene was positive
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