Functional Domain Dissection of Barley ADP-Glucose Transporter HvBT1 and Application of Its Antibody in Germplasm Evaluation for Starch Content

ADP-glucose transporter HvBT1 is a key component of the starch biosynthesis pathway in barley; however, its functional domains and protein-level detection tools remain poorly characterized. In this study, domain dissection and transport activity assays revealed that the full-length HvBT1 exhibited the highest activity, while combinations of two domains restored 65–69% of the activity and single domains maintained only 42–49% of the activity, suggesting that the synergistic action of the three domains contributes to optimal transport function. Based on these findings, the polyclonal antibody against HvBT1 can effectively detect endogenous barley proteins even at a dilution ratio of 1:20,000. This antibody was then applied to evaluate protein expression in 115 early-maturing barley accessions, revealing a significant positive correlation between protein abundance and total starch content (R2 = 0.5345, p < 0.0001), suggesting a potential association that merits further investigation. Our study provides insights into the potential synergistic contribution of the domains in HvBT1 and develops an antibody-based protein evaluation approach for barley germplasm. The findings suggest that HvBT1 protein abundance may serve as a potential marker for preliminary screening of high-starch barley varieties, although further validation in independent populations is warranted.