Editing of the starch synthase IIa gene led to transcriptomic and metabolomic changes and high amylose starch in barley

In this study, a range of barley allelic mutants lost ADPG binding structure ofstarch synthaseIIa(SSIIa) were created through targeted mutagenesis ofSSIIaby RNA-guided Cas9. The transcriptomic and qRT-PCR results showed the increased mRNA expression ofHvGBSSIand the decreasedHvSSIIaandHvSBEIlevels inssIIamutant grains, which were consistent with the expressions of GBSSI, SSS and SBE enzymatic activities, respectively. However, the increased expressions ofHvSSIcannot effectively compensate for the loss ofHvSSIIa. The metabolic pathway analysis showed that the mutation of SSIIa led to increased ADP-glucose synthesis in barley grains. ThessIIamutant grains had two and six times amylose, and RS contents in control grains, respectively, and significantly changed starch structure and functions compared to the controls. No metabolite changes could compensate for the decrease of starchbiosynthesis in the ssIIa null mutant.