Extended Data Fig. 10: Specificity and efficiency of Plcb4-SaCas9 expression and Fgfr2 gRNA targeting.

a, Schematic representation of the arrangement of mouse Fgfr2 exons and the two targeting gRNAs. b, Schematic representation of the AAV vectors of Plcb4-SaCas9, U6-Scramble gRNA and U6-Fgfr2 gRNAs. c, Sequencing analyses showing an example of insertion/deletions (indels) mediated by SaCas9/Fgfr2 gRNA1 and gRNA2 in mouse Fgfr2 genomic loci after co-expression of SaCas9 and Fgfr2 gRNAs in mouse embryonic fibroblasts. d, Predicted off-target sites of Fgfr2 gRNAs. e, Genomic DNA sequences surrounding the targeting or off-targeting loci were amplified by PCR and ligated to T vector. After transformation, colonies of each group were sequenced and the percentage of colonies showing indels were calculated. f, Sequencing analyses showing examples of indels in Fgfr2 genomic loci of INTACT-enriched Purkinje neuron nuclei after AAV-mediated expression of SaCas9 and Fgfr2 gRNAs. g, Percentage of indels introduced in mouse Fgfr2 genomic loci of Purkinje neurons after AAV-mediated expression of SaCas9 and Fgfr2 gRNAs. h, Representative immunofluorescence images of Fgfr2 and Calbindin or Plcb4 in lobules IV/V in cerebellar sagittal sections of scramble or Fgfr2-KO mice. Scale bar, 100 μm. i, Split UMAP plot visualization of granule neurons, Purkinje neurons, interneurons, glial cells and oligodendrocytes from anterior vermis lobules of scramble and Fgfr2-KO samples under baseline and learned conditions. j, Visualization of molecular signatures of each population by their percentage and average gene expressions. k, Heat map of INTACT-FACS-snRNA-seq data from the anterior vermis lobules of Fgfr2-KO or control mice, presented as normalized Z-scores. l, Scatter plot showing the differential expression indices for each DEG in Aldoc+ or Plcb4+ Purkinje neurons upon Fgfr2-KO in Plcb4+ Purkinje neurons. m, Brown/black module genes were not reduced upon Fgfr2-KO in Plcb4+ cells. p = 1.08e-07 for scramble and p = 2.2e-16 for Fgfr2-KO, two-sided unpaired t-test. n = 2 in biologically independent snRNA-seq. 977 cells, 637 cells in scramble baseline and learned samples. 974 cells, 1514 cells in Fgfr2-KO baseline and learned samples.