Extended Data Fig. 2: Delivery of ABE using Lipofectamine 3000.
(a) Analysis of ABE editing of rd12 reporter cells transfected with plasmids encoding ABE NG variants and guide RNA targeting Rpe65 rd12. Two biological replicates with 2 analytical replicates each, mean ± s.d. (b,c) ABE RNPs were delivered to rd12 reporter cells using Lipofectamine 3000. Efficiency of delivery was analyzed using (b) fluorescence microscopy (100 nM, 22.4 µg ml-1 RNP shown here) and (c) flow cytometry. Scale bar: 100 µm. Two biological replicates with 2 analytical replicates each, mean ± s.d. (d) Rescue of RPE65 protein expression in NIH/3T3-rd12 cells by delivery of ABE and guide RNA plasmids (p), or 100 nM ABE RNP using Lipofectamine 3000 (RNP). Mouse retinal pigment epithelium (RPE) extract was used as a positive control. M = molecular weight marker; N = untreated cells. (e) NGS analysis of editing of NIH/3T3-rd12 cells treated with ABE. Two biological replicates with at least 3 analytical replicates each, mean ± s.d. (f,g) Rescue of visual function in rd12 mice injected with 25 µM ABE8e RNP (5.6 µg per eye), 23.6 µM ABE8e RNP with 50%(v/v) Lipofectamine 3000 (4.5 µg per eye), or 23.6 µM ABE8e RNP with 10%(v/v) Lipofectamine 3000. Injected material additionally contained 25% (w/v) sucrose. At least 8 eyes, mean ± s.d, Kruskal-Wallis test with Dunn’s multiple comparisons test, **P < 0.01. Uncropped blots are available within Source data.
