Supplementary Figure 9: CRISPR/Cas9-engineered mutations in FAB result in weakly fasciated plants resembling fab mutants.

(a) Schematic illustrating two guide RNAs (sgRNAs; red arrows) targeting the FAB (SlCLV1) coding sequence. Cas9/sgRNA1/sgRNA2 were expressed from the same construct. Black arrows indicate the PCR primers used to evaluate mutation type and efficiency. (b) PCR genotyping of eight T₀ CR-fab plants showing predominantly chimeric plants with a range of indel mutations, including the desired deletion. One plant (CR-fab-8) was homozygous for the desired deletion. PCR for the presence of Cas9 is shown along with evaluation for fasciation. All eight CR-fab plants were fasciated and exhibited similar severity. (c) Representative inflorescence from a CR-fab plant (CR-fab-3) showing branching (red arrowheads) and weakly fasciated flowers like fab (Fig. 1b). Images of a representative flower and fruit are shown (insets). Scale bar, 1 cm. (d) Quantification and statistical comparisons of floral organ numbers from WT and CR-fab flowers. Data were collected from three plants with confirmed mutations in all sequenced alleles (e; see also Online Methods). Data are means ± s.d., n ≥ 10. A two-tailed, two-sample t test was performed, and significant differences are represented by black asterisks: ***P < 0.0001. (e) CR-fab alleles identified by cloning and sequencing PCR products from the FAB targeted region from three fasciated T₀ plants. All alleles carried indel mutations (blue dashed lines and letters), and one of each representative allele identified from sequencing at least eight cloned PCR products is shown. CR-fab-2 and CR-fab-3 are biallelic, and CR-fab-6 is chimeric. Red font highlights sgRNA target sites, and black rectangles indicate PAM sequences; a, allele.