Eggplant is one of the most important vegetables worldwide. Prickles on the leaves, stems and fruit calyxes of eggplant may cause difficulties during cultivation, harvesting and transportation, and therefore is an undesirable agronomic trait. However, limited knowledge about molecular mechanisms of prickle morphogenesis has hindered the genetic improvement of eggplant. In this study, we performed the phenotypic characterization and transcriptome analysis on prickly and prickleless eggplant genotypes to understand prickle development at the morphological and molecular levels. Morphological analysis revealed that eggplant prickles were multicellular, lignified and layered organs. Comparative transcriptome analysis identified key pathways and hub genes involved in the cell cycle as well as flavonoid biosynthetic, photosynthetic, and hormone metabolic processes during prickle development. Interestingly, genes associated with flavonoid biosynthesis were up-regulated in developing prickles, and genes associated with photosynthesis were down-regulated in developing and matured prickles. It was also noteworthy that several development-related transcription factors such as bHLH, C2H2, MYB, TCP and WRKY were specifically down- or up-regulated in developing prickles. Furthermore, four genes were found to be differentially expressed within the Pl locus interval. This study provides new insights into the regulatory molecular mechanisms underlying prickle morphogenesis in eggplant, and the genes identified might be exploited in breeding programs to develop prickleless eggplant cultivars.