Extended Data Fig. 5: Characterization of mismatch tolerance of gRNA for dCas13 binding efficiency in the CRISPR-TO system.
From: Programmable control of spatial transcriptome in live cells and neurons
a, Schematic showing the design of 30 gRNAs with different mismatches in the spacer of gG1 which targets the endogenous GAPDH mRNA. The mutated nucleotides are shown in black. b, Quantification of percentage of RNA relocalization efficiency and percentage of dCas13 localized on the OMM in individual cells for different gRNAs. % RNA relocalization efficiency was calculated by subtracting the background % of GAPDH mRNA localized on the OMM using gNT from the % of GAPDH mRNA localized on the OMM for each gRNA. Only cells co-expressing dCas13 and MAVS*-PYL1 were analyzed. Black bar, mean. n, cell number. Two-sided, unpaired Student’s t-test between gG1 and individual groups. The dashed line indicates the mean of the gG1 group. c, Comparison of % RNA relocalization efficiency of gRNAs with single mutation (SM), double mutations (DM), and triple mutations (TM) on different ___location of the gRNA spacer. Two-sided, one sample t-test with a hypothetical value 0.1298 (indicated by the dashed line) which is the mean % RNA relocalization efficiency of the gG1 group. d, Comparison of % RNA relocalization efficiency and % dCas13 localized on the OMM between gG1 group and gG12 group which co-expresses gG1 and gG2 together. The number above each plot is the fold change between the two groups. n, number of cells analyzed per group. Black bar, mean. n, cell number. Two-sided, unpaired Student’s t-test.
