Development of elite germplasm with high ɣ-aminobutyric acid (GABA) content by targeted mutation of ZmGAD3 and ZmGABA-TP1 in maize

γ-Aminobutyric acid (GABA) plays a crucial role in human health. Although tomatoes with enhanced GABA levels have been engineered through single-gene editing, GABA-fortified maize has not previously been developed. In this study, CRISPR/Cas9 editing of the biosynthetic gene ZmGAD3 , the degradation pathway gene ZmGABA-TP1-1 , or both genes simultaneously, increased the GABA content of maize kernels 0.59–2.15 fold, 1.28–2.99 fold, or 2.63–4.87 fold, respectively. We thus developed an elite germplasm resource with significantly fortified GABA content and demonstrated that coordinated editing of ZmGAD3 and ZmGABA-TP1-1 could effectively enhance GABA accumulation. This study reveals important insight into the functional regulation of GABA content through maize metabolic engineering.