Bioscience Evidence 2025, Vol.15, No.5, 228-236 http://bioscipublisher.com/index.php/be 232 5.3 Key outcomes: reduction in immunogenic epitopes, retention of acceptable dough properties The contents of α-, γ- and ω -gliolysis proteins in these CRISPR mutants decreased significantly. In some materials, the total amount of gluten decreased by up to 97%, and the immune epitopes also decreased significantly (Guzmán-López et al., 2021; Yu et al., 2023; Sanchez-Leon et al., 2024). Meanwhile, the edited flour performed normally in terms of dough performance and baking quality, with no obvious problems (Yu et al., 2023). 5.4 Clinical/functional assessment: immunoreactivity tests with celiac patient sera Serum tests using patients with celiac disease revealed that the immune reactivity of edited wheat was significantly reduced, with some materials showing a decrease of up to 47 times (Yu et al., 2023). In addition, its safety was verified by studies using animal models and T-cell activation experiments (Jouanin et al., 2020; Yu et al., 2023). 5.5 Broader implications: toward gluten-safe wheat lines for commercial production The multiple editing of CRISPR/Cas9 provides a way to cultivate wheat with low immunogenicity or even "gluten-free". Some editing strains have achieved no transgenic residue and are commercially viable (Liang et al., 2017; Jouanin et al., 2020; Sanchez-Leon et al., 2024). In the future, if high-throughput screening and precise epitope editing are combined, it is expected to obtain "celiac disease-safe" wheat that is both safe and has good processing performance (Jouanin et al., 2019; 2020; Guzmán-López et al., 2021; Yu et al., 2023). 6 Challenges and Trade-Offs 6.1 Maintaining dough quality and baking performance with reduced gluten Gluten is very important for wheat dough and baking quality. Reducing gluten, especially glycolin, often leads to a decrease in dough elasticity and poor extensibility, affecting the structure and taste of foods such as bread (Gil-Humanes et al., 2014; Sharma et al., 2020). Some studies have obtained low-alcohol-soluble protein wheat using RNAi and found that its dough stability and taste are close to those of common wheat, but it still needs to be optimized to meet industrial demands (Gil-Humanes et al., 2014; Wang et al., 2020). In addition, adding excipients such as low-ester pectin, dietary fiber or polyphenols can partially make up for the deficiency of dough network and improve rheology and texture (Girard and Awika, 2020; Cui et al., 2022; Li et al., 2023). 6.2 Balancing nutritional value (protein composition changes) Low-gluten protein may alter protein composition, influencing the proportion of amino acids and nutritional value. Some low-alcohol-soluble protein wheat strains have increased contents of essential amino acids such as lysine and higher nutritional value (Gil-humanes et al., 2014). However, some studies have found that when gluten decreases, the expression of other storage proteins (such as high-molecular-weight gluten subunits) may increase. Therefore, attention should be paid to overall protein quality and digestibility (Gil-Humanes et al., 2014; Nye-Wood et al., 2021). 6.3 Agronomic performance and yield stability Gluten genes are closely related to agronomic traits such as wheat grain development and yield. Some improved strains showed no significant changes in yield and grain quality after reducing gluten (Landolfi et al., 2021; Buczek, 2024). However, some studies have pointed out that if the gluten level drops too much, it may affect indicators such as grain filling and 1000-grain weight. Therefore, a balance needs to be struck between reducing immunogenicity and maintaining high and stable yields (Wang et al., 2020; Buczek, 2024). 6.4 Regulatory and biosafety issues for gene-edited crops Gene editing technologies such as CRISPR/Cas are not considered genetically modified in some countries, but policies are not consistent worldwide. Some countries require strict safety assessment and label management, which may affect the promotion of new varieties (Wang et al., 2020; Nye-Wood et al., 2021). At the same time, attention should also be paid to the off-target effects and ecological risks that may be brought about by gene editing.
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