Fig. 3

a Binary vector T-DNA architecture, OsU3-p – rice U3 promoter; gRNA – chimeric guide RNA; OsU3-t – rice U3 terminator; ZmUbi1-p – maize Ubiquitin 1 promoter; zCas9 – maize codon-optimized Streptococcus pyogenes cas9 endonuclease gene; Nos-t – 3′-signal of Agrobacterium tumefaciens nopaline synthase gene; E9-t – pea Ribulose-1,5-Bisphosphate Carboxylase Small Subunit (rbcS) E9; HPT – hygromycin phosphotransferase, 2x35S-p – doubled enhanced CaMV 35S promoter. b Mutation detection in T0, Sanger chromatogram of intact (WT) and mutated (plant 16) Nud50 target motif. The sequence with colored background indicates the target motif with the PAM being also marked by a red frame. Arrows indicate fragments deleted from the WT sequence and the corresponding ligation points in the mutant alleles of the biallelic plant 16. c Examples of mutated alleles of the NUD gene found in T0 population after mutagenesis directed by the Nud45 and Nud50 gRNAs. Target motifs are underlined, the PAM is marked grey. d Phenotype of grain harvested from mutant plant (left) in comparison to wild-type ‘Golden Promise’ grain (right)