Zanthoxylum armatum DC. is an important economic tree species. Prickle is a type of trichome with special morphology, and there are a lot of prickles on the leaves of Z. armatum, which seriously restricts the development of Z. armatum industry. In this study, the leaves of Z. armatum cv. Zhuye (ZY) and its budding variety ‘Rongchangwuci’ (WC) (A less prickly mutant variety) at different developmental stages were used as materials, and the transcriptome sequencing data were analyzed. A total of 96,931 differentially expressed genes (DEGs) were identified among the samples, among which 1560 were candidate DEGs that might be involved in hormone metabolism. The contents of JA, auxin and CK phytohormones in ZY leaves were significantly higher than those in WC leaves. Combined with weighted gene co-expression network analysis, eight genes (MYC, IAA, ARF, CRE/AHK, PP2C, ARR-A, AOS and LOX) were identified, including 25 transcripts, which might affect the metabolism of the three hormones and indirectly participate in the formation of prickles. Combining with the proteins successfully reported in other plants to regulate trichome formation, ZaMYB86, a transcription factor of R2R3 MYB family, was identified through local Blast and phylogenetic tree analysis, which might regulate prickle formation of Z. armatum. Overexpression of ZaMYB86 in mutant A. thaliana resulted in the reduction of trichomes in A. thaliana leaves, which further verified that ZaMYB86 was involved in the formation of pickles. Yeast two-hybrid results showed that ZaMYB86 interacted with ZaMYB5. Furthermore, ZaMYB5 was highly homologous to AtMYB5, a transcription factor that regulated trichomes development, in MYB DNA binding domain. Taken together, these results indicated that ZaMYB86 and ZaMYB5 act together to regulate the formation of prickles in Z. armatum. Our findings provided a new perspective for revealing the molecular mechanism of prickly formation.