Family-1 UDP-glycosyltransferases, designated as UGTs, glycosylate key cellular metabolites into compounds with improved stability, solubility and transport to regulate plant growth and development in various metabolic pathways. However, little is known about the molecular and functional characteristics of UGT family in an aquatic plant and how it contributes to the Nelumbo nucifera ecotypic differentiation. Therefore, we performed a genome-wide investigation of 108 candidate UGT genes distributing on all eight lotus chromosomes, encompassing 16 distinct phylogenetic groups (A-P). These UGT groups feature different molecular weights, subcellular localizations and evolutionary rates. Intra-specific collinearity analysis further suggests that the expansion of the UGT family in lotus is mainly driven by tandem duplication and whole-genome duplication. Further, expression analysis of 54 RNA-seq samples from ten different lotus tissues suggests that expression of most UGT family members exhibits tissue or developmental-stage specificity, and UGTs expressed in reproductive organs (tissues) evolve much faster than those expressed in vegetative organs. More interestingly, key UGT genes being differentially regulated between temperate and tropical lotus (ecotypes) were uncovered during the seedling stage, indicating that UGTs associated with hormone synthesis and transport likely contribute to early differentiation between lotus ecotypes. Overall, our systematic expression and evolutionary analyses not only characterize the expression, functional diversity and evolutionary dynamics of the UGT family in an aquatic plant but also provide insight into the roles of the UGT family underlying the differentiation of the lotus ecotypes.