Comparison of dgRNA efficiency between injection of pre-assembled RNP and a dgRNA/Cas9 mRNA mixture. (A) Selected crRNAs from the first set of 51 crRNAs were tested for their activity through injection of 3 fmol dgRNA with 200 pg HiFi Cas9 mRNA. Indel frequencies (in %) assessed with ICE were compared to those obtained with HiFi Cas9 RNP, using bar graphs. Means of replicates are shown with standard error bars. The experimental dataset for dgRNA/Cas9 mRNA mixture is shown in Supplementary Table S9. HiFi Cas9 RNP data were adopted from Figure 2. (B) A second set of 27 crRNAs were tested for their activity through injection of 3 fmol dgRNA with 200 pg HiFi Cas9 mRNA. Indel frequencies (in %) assessed with ICE were compared to those obtained with HiFi Cas9 RNP using bar graphs. Means of replicates are shown with standard error bars. The experimental dataset for dgRNA/Cas9 mRNA mixture is shown in Supplementary Table S9. HiFi Cas9 RNP data were adopted from Supplementary Figure S4. (C−E) Performance of CRISPR-kp on efficiency of 27 crRNAs in dgRNA/HiFi Cas9 mRNA format in comparison with CRISPRscan and Rule Set 2 scores. Indel frequencies (in %) obtained with dgRNA/HiFi Cas9 mRNA were compared to scores obtained with CRISPR-kp (C), CRISPRscan (D) and Rule Set 2 (Doench/Fusi 2016) (E) using scatter plots. Spearman correlation coefficients (rs) are indicated in each panel.
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