Extended Data Fig. 9: In vitro and in vivo RNA editing activities of the REPAIR variants.
From: Structural insights into RNA-guided RNA editing by the Cas13b–ADAR2 complex
(a) SDS–PAGE analysis of dPspCas13b–ADAR2DD and its variants. The proteins were visualized with CBB. All experiments were performed at least three times. (b) Western blot showing the expression levels of the WT dPspCas13b–ADAR2DD and the V2 variant in human cells. The two proteins were detected using an anti-FLAG antibody. The samples were normalized based on β-actin expression levels. All experiments were performed at least three times. (c) Design of the V4 dPspCas13b–ADAR2DD variant and crRNAs with varying guide lengths (GL) and mismatch distances (MD). (d) Preferences of the WT dPspCas13b–ADAR2DD and the V4 variant for editing positions. RNA editing activities were examined in human cells, using different crRNAs targeting the exogenous Cluc (W85X) transcript, the endogenous STAT1 (Y701C) transcript, or the endogenous CTNNB1 (T41A) transcript. Editing rates for A nucleotides in the target transcripts are displayed. Data are shown as mean ± S.D. (n = 4 biological replicates). The target A positions and the complementary guide regions are outlined in red and black, respectively. T, targeting guide; NT, nontargeting guide.
