Modification of starch traits in commercial wheat through TaWaxy gene editing

Amylose content (AC) is a key determinant of wheat quality, and the TaWaxy gene determined amylose synthesis with a dose-dependent effect on AC. In this study, the TaWOX5 gene, which significantly enhances wheat transformation efficiency, was combined with CRISPR/SpCas9 system to generate TaWaxy mutants in a commercial winter wheat Jimai 22. Seven transgene-free mutant types were produced, compared to only three transgene-free mutants in the spring wheat variety Ningchun 4. The TaWaxy mutants from the two varieties showed decreased ACs ranging from 0 to 19.05 %. Results demonstrated that the Waxy-B1 protein has the most significant effect on amylose synthesis. The mutants with TaWaxy-abd, TaWaxy-ab, and TaWaxy-bd alleles showed waxy wheat trait. Interestingly, the TaWaxy-b mutant from Jimai 22 exhibited a waxy trait, unlike the TaWaxy-b mutant from Ningchun 4. Transmission electron microscope and scanning electron microscopy showed increased B-type starch granules in mutant grains. The mutants displayed varying effects on bread, cake, cookie, and noodle quality. All mutants showed decreased quality in bread and cake production, while TaWaxy-ad-JM and TaWaxy-b-NC mutants showed improved noodle and cookie quality. The generated mutants provide optimized optimized amylose content, enhancing noodle and biscuit quality as a practical alternative to blending.