Extended Data Fig. 1: Delivery of PE RNP into rd12 reporter cells.
(a) Analysis of delivery of 500 nM PE2 RNP (149 µg ml-1), with epegRNA targeting Rpe65 rd12, into the rd12 reporter cells using 2% (w/v) sucrose or 10% (w/v) sucrose, and with or without added cell-penetrating peptide 6xHis-CM18-PTD4; or using Lipofectamine 3000 (L3000). In parallel, the rd12 reporter cells were transfected with PE2 and rd12 epegRNA plasmids (right-most panel). The cells were analyzed using fluorescence microscopy and flow cytometry. Faint-yellow cells that were modified with PE2 RNP are highlighted with yellow circles. Scale bar: 100 µm. (b) Delivery of PE2 RNP using 10% sucrose and 6xHis-CM18-PTD4 peptide, quantified using flow cytometry. Two biological replicates with two analytical replicates each, mean ± s.d. One-way ANOVA with Tukey’s multiple comparisons test. ****P < 0.0001; ns, P > 0.05. (c) Comparison of efficiency of delivery of PE2 with Lipofectamine 3000 as RNP, and as a pair of PE and epegRNA plasmids. *P < 0.05; ****P < 0.0001. One-way ANOVA with Tukey’s multiple comparisons test. NE = minus enzyme control with Lipofectamine 3000 alone. (d) Titration of rd12 reporter cells with increasing PE RNP and constant L3000, quantified by flow cytometry. Four biological replicates with 2 analytical replicates each, mean ± s.d.
