Fig. 3

Identification of VvCCD8 knockout mutants. a Overview of identification of regenerated plants. b Identification of exogenous T-DNA insertions in regenerated plants by PCR. The specific primers designed for Cas9 gene were used for PCR identification. Only CCD8-sgRNA plants were identified with exogenous T-DNA insertions. Lanes 1–6 represent different individual CCD8-sgRNA plants. The plasmid was used as the positive control (P), while the wild-type genomic DNA was used as the negative control (N). M, DNA marker. The cropped gel image is shown here, and the original, uncropped image is available in Additional file 3: Figure S3. c Phenotypes of VvCCD8 knockout mutants. The shoot branches of VvCCD8 knockout mutants were indicated in black arrows. Scale bars: 0.5 cm. d The branch number of the four VvCCD8 knockout mutants. e Sequencing results of the target sites in the four VvCCD8 knockout mutants. The gene fragments were amplified from each mutant plant and were cloned into pLB vector for Sanger sequencing assay. A number of 20 clonal amplicons for each plant were analyzed. The mutated sequences identified from the mutants were shown. The plant IDs are shown on the left. Mutation types (colored in red) and the corresponding number (indicated in black) of clones were shown on the right. Those undesired sequences were omitted from the analysis. f Mutations of amino acids in mutated sequences shown in e. The altered amino acids are colored in red and the premature stop codons are indicated in red asterisks (*). The number of amino acids (aa) that are not shown in the figure is indicated in parentheses