Triticeae Genomics and Genetics, 2024, Vol.15, No.2, 77-87 http://cropscipublisher.com/index.php/lgg 83 non-prolamin allergens (Juhász et al., 2018; Hussain et al., 2022). These genomic resources have enabled researchers to pinpoint specific genetic variations and expression patterns linked to wheat allergies, thereby offering new avenues for developing hypoallergenic wheat varieties. 6.2 Molecular breeding for reduced allergenicity Molecular breeding techniques, including marker-assisted selection (MAS) and genomic selection (GS), have been employed to develop wheat varieties with reduced allergenicity. By leveraging the extensive genomic data available, breeders can now select for traits that minimize the presence of allergenic proteins. For example, RNA interference (RNAi) has been used to silence the γ-gliadin genes, which are major sensitizing allergens in wheat-dependent exercise-induced anaphylaxis (WDEIA). This approach has shown promise in reducing the immunogenicity of wheat, although the complexity of immune responses among individuals suggests that further refinement is needed (Altenbach et al., 2015). Additionally, the integration of omics technologies, such as proteomics and metabolomics, has provided deeper insights into the molecular mechanisms underlying wheat allergenicity, thereby enhancing the effectiveness of molecular breeding strategies (Alotaibi et al., 2020). 6.3 Genetic engineering and CRISPR/Cas9 technology The CRISPR/Cas9 genome editing technology has revolutionized the field of genetic engineering, providing precise and efficient tools to modify the wheat genome to reduce allergenicity. CRISPR/Cas9 has been successfully used to target and edit specific genes associated with allergenic proteins. For example, combining CRISPR/Cas9 with microspore technology can rapidly generate homozygous mutant plants, thereby accelerating the breeding process (Bhowmik et al., 2018). Bhowmik et al. (2018) demonstrated the ability of CRISPR/Cas9 technology to introduce both exogenous and endogenous gene mutations in wheat microspores, providing strong evidence of how genetic engineering and CRISPR/Cas9 technology can accelerate the breeding process (Figure 3). Advances in CRISPR technology, such as base editing and prime editing, have further increased the precision and scope of gene modification, paving the way for the development of low-allergenicity wheat varieties (Li et al., 2021; Jogam et al., 2021). Figure 3 by Bhowmik et al. (2018) shows the results of editing the endogenous ubiquitin gene (TaUbiL1) inwheat microspores using the CRISPR/Cas9 system. These images and charts display various insertion and deletion mutations, demonstrating the introduction of mutations through non-homologous end joining (NHEJ) repair following double-strand breaks. This result further proves the high efficiency of the CRISPR/Cas9 system in wheat gene editing, particularly in the application of endogenous gene editing. Utilizing Figure 3, the potential application of CRISPR/Cas9 technology in improving wheat traits, enhancing resistance, and increasing adaptability can be effectively demonstrated. In summary, the integration of genomic approaches, molecular breeding, and cutting-edge genetic engineering techniques like CRISPR/Cas9 holds great promise for mitigating wheat allergies and sensitivities. These advancements not only contribute to the development of safer wheat products but also enhance our overall understanding of the genetic bases of wheat allergenicity. 7 Implications for Wheat Breeding and Food Industry 7.1 Breeding strategies for allergen-free wheat Breeding strategies aimed at developing allergen-free wheat focus on reducing or eliminating the presence of immunogenic gluten proteins, particularly gliadins and glutenins, which are implicated in celiac disease (CD) and non-celiac wheat sensitivity (NCWS). Recent studies have shown that domestication and breeding have contributed to a decrease in the content of gliadins in cultivated Triticeae species compared to their wild ancestors (Ozuna and Barro, 2018). This reduction in gliadins, which are rich in CD immunogenic epitopes, suggests that selective breeding can be an effective strategy for developing wheat varieties with lower allergenic potential. Additionally, the identification of specific alleles in wild wheat relatives, such as Triticum urartu, which exhibit diverse and potentially beneficial traits for wheat improvement, provides a valuable genetic resource for breeding programs (Talini et al., 2019).
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