Improving the Agronomic Performance of High-Amylose Durum Wheat.

High-amylose wheat has garnered significant attention from the food industry for its potential to produce low-glycaemic food products. It is well-established that there is a direct correlation between the amylose content in flour and the amount of resistant starch (RS) in foods. Recently, some research initiatives have successfully produced high-amylose durum wheat by targeting key enzymes in the amylopectin biosynthesis pathway, though this has resulted in a reduction in seed weight. This study aimed to develop durum wheat genotypes with enhanced nutritional and agronomic traits by pyramiding mutations in the SSIIa genes and the GW2-A1 null allele. A cross between Svevo SSIIa and Kronos GW2-A1 was performed, and marker-assisted selection (MAS) strategies were employed to identify ten sister lines (GW2-A1/SSIIa). Biochemical analyses revealed that the GW2-A1/SSIIa genotypes exhibited significantly higher amylose and resistant starch (5-10-fold) levels compared to Svevo and GW2-A1 controls. Phenotypic analyses demonstrated that GW2-A1/SSIIa lines showed a 50% increase in hundred-grain weight (HGW) and improved grain size parameters compared to Svevo SSIIa, though these values remained lower than Svevo and Kronos GW2-A1. Yield per plot increased by 67% compared to Svevo SSIIa but was 30-40% lower than Svevo and Kronos GW2-A1. Gene expression analysis revealed upregulation of key starch biosynthesis genes (Susy2, UGPase) in GW2-A1/SSIIa lines, suggesting compensatory mechanisms for reduced starch content. Downregulation of TPS7 indicated potential limitations in trehalose-6-phosphate biosynthesis, which may influence starch accumulation. This study demonstrates that combining SSIIa and GW2-A1 null mutations can mitigate yield losses associated with high-amylose genotypes while maintaining elevated levels of resistant starch and dietary fiber.