<p>Rose (<i>Rosa hybrida</i>) is an ornamental crop cultivated worldwide for its economic significance. However, rose prickles pose the risk of injury to workers and can negatively affect flower quality. Consequently, there has been a long-standing demand for the development of new rose varieties with small, delicate, and fewer prickles. Searching for the genes involved in prickle formation is required for rapid and efficient breeding. In this study, we identified myeloblastosis (MYB) group transcription factors that are anticipated to be involved in the formation and color change of rose prickles through RNA sequencing. In addition, we aimed to estimate the influence of selected transcription factors on the physiological characteristics of roses and suggest possibilities. Three types of stems of the rose ‘Pink Beauty’ (prickless stems (PL), stems with small prickles (SP), and stems with large prickles (BP)) generated a total of 71,830,230 raw reads. The average number of trimmed reads was 71,200,618, indicating a high quality. Through differentially expression gene (DEG) analysis, we selected DEGs that were upregulated and downregulated in SP and BP compared to those in PL, focusing on the MYB group. Finally, we compared and analyzed the expression levels of seven DEGs (<i>MYB5, WER, ETC3, MYB4, MYB8, MYB6,</i> and <i>MYB14</i>). In particular, MYB5 and WER are expected to play important roles in the regulation of trichome formation, whereas ETC3, MYB4, and MYB8 are anticipated to have an essential influence on prickle development. Additionally, MYB6 and MYB14 are likely to be involved in pigment biosynthesis and regulation of color changes in rose prickles. This study suggests that transcription factors play an important role in the development of rose prickles and contributes to the understanding of the functions of related genes. Data from this study will serve as a foundation for future research on the formation and development mechanisms of prickles.</p>