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Flavonoids are a class of phenolic compounds that are widely distributed in nature. They have a variety of physiological and pharmacological activities. They exist in free form or in the form of glycosides. The glycosylation occurs by glycosyltransferases, which is a common modification of plant secondary metabolites and the last step of their biosynthesis. Glycosylation can increase the diversity of the structure and function of flavonoids, and is currently a research hotspot. Based on the classification of flavonoids, this paper describes and summarizes the biotransformation and characteristics of glycosylation modification of flavonoids with different structural types, and describes the effects of enzymatic glycosylation using different types of glycosyltransferases on the biological activity and function of flavonoids depending on sugar connection position, sugar quantity, and type. This paper provides a reference for the development and application of flavonoids glycosylation and glycosyltransferases and also opens up a new direction for plant breeding.