Fig. 3

Characterization of the PHS phenotype in the complementation lines (COM-1 and COM-2) and OsAAH CRISPR knockout mutants (Osaah-1 and Osaah-2). A DNA sequencing peak chromatograms of the mutated site produced by EMS mutagenesis in the OsAAH gene. Red arrowheads indicate heterozygous point mutations in the complementation lines. B Photographs of seed germination in harvested panicles at 30 DAP from WT, phs39 mutant, and complementation transgenic lines at 7 days of imbibition. Scale bars = 3 cm. C Time-course germination rate of seeds in panicles harvested at 30 DAP from WT, phs39 mutant and complementation transgenic lines. Data are means ± SD (n = 3). D Construction of two Osaah mutants using CRISPR/Cas9 technology. Red dashes represent the deleted bases, and the inserted bases are indicated by red uppercase letters. The number represents the number of changed bases. E Western blot analysis showed that OsAAH protein was detected in WT seeds but not in the phs39 and Osaah mutants by anti-OsAAH antibody. F Photographs of seed germination in panicles harvested at 30 DAP from WT and Osaah mutants at 7 days of imbibition. Scale bar = 3 cm. G Time-course germination rates of seeds in panicles harvested at 30 DAP from WT and Osaah mutants. Data are means ± SD (n = 3)