Blueberry (Vaccinium ashei) is a popular fruit due to its high anthocyanin content. This study aimed to analyze the transcriptome profile of V. ashei cv. ‘Brightwell’ fruits at different stages of development. A total of 314.26 GB of clean data were obtained and de novo assembled into 254 196 unigenes. In comparisons between the early and late stages of fruit ripening, 27 genes (including PAL, CHS, F3H, F3′H, F3′5′H, LDOX, etc.) were found to cover the main steps in the anthocyanin biosynthesis pathway. Most of these genes were highly expressed in the late stage of fruit development, suggesting that anthocyanins mainly accumulate in the late stage. During the late stage of fruit development, most structural and regulatory genes such as F3′5′H and F3′H, which are involved in the anthocyanin biosynthetic pathway, were upregulated, causing the fruit to turn blue. Decreased expression of a large number of chloroplast-related genes during the fruit ripening period could explain why the green fruit color fades over time. Additionally, abscisic acid and ethylene may play positive roles in promoting fruit ripening and anthocyanin accumulation. This research reveals the transcriptomic characteristics of immature and mature fruits and enhances our understanding of the molecular mechanisms of anthocyanin biosynthesis and accumulation in blueberry fruit.