Research Projects

Coeliac disease (CD) is a widespread wheat-related autoimmune disorder affecting 1 in 100 individuals worldwide, with an estimated 6 to 8 million people affected. Classical breeding strategies have failed to develop wheat varieties that are CD-safe. Novel genome editing techniques, such as the CRISPR/CAS system, have the potential to develop CD-safe wheat varieties. RNAi-based silencing and CRISPR/CAS9-based knockdown have been shown to reduce gliadin content in hexaploid wheat. However, the high copy number of gliadins and the presence of multiple epitopes across gliadins and glutenins present complexities that must be overcome to achieve CD-safe wheat. A total of 31 coeliac disease epitopes have been identified in wheat, with gliadins being the major causative agents of coeliac disease. In wheat, α-gliadins contain the major and clinically most relevant immunodominant epitopes. Tetraploid wheat (Triticum durum AABB) is less immunogenic than hexaploid wheat, and the researchers hypothesize that targeting A genome-specific α-gliadins in T. durum can reduce immunogenicity for developing CD-safe wheat. The proposed CRISPR/CAS9-based editing of A genome-specific α-gliadins in tetraploid wheat involves designing guide RNAs specific to α-gliadins genes, followed by CRISPR/CAS9-based genome editing. The edited plants will be selected based on screening for target edits through sequencing and gliadin profile on SDS PAGE, 2D PAGE, and RP-HPLC. Immunoblot assays will be performed to assess the immunogenicity of genome-edited plants.