Abstract: To assess the effect of NtJAZ15 on tobacco glandular trichome development, the molecular mechanism underlying its involvement in trichome development was investigated. The NtJAZ15 gene was cloned from K326, and its bioinformatics analysis and tissue expression patterns were examined. By generating NtJAZ15 knockout and overexpression mutants, the regulatory roles of NtJAZ15 in glandular trichome formationwere analyzed. Results indicated that both NtJAZ15-1 and NtJAZ15-2 were highly conserved across different tobacco varieties, exhibiting typical TIFY and Jas domains characteristic of the JAZ family. NtJAZ15 was found to be expressed in all tissues, with NtJAZ15-2 exhibiting higher expression levels compared to NtJAZ15-1 in all tissues. Interestingly, the density of long and short glandular trichomes increased in NtJAZ15 knockout mutants, while the opposite effect was observed in overexpressed mutants. Furthermore, qRT-PCR analysis showed that NtJAZ15 significantly affected the expression of NtWo, NtCycB2, NtHD9 and NtHD12, but had no significant effect on the expression of NtMIXTA. These findings together suggest that NtJAZ15 negatively regulates the occurrence of long and short glandular trichome , and NtWo, NtCycB2, NtHD9 and NtHD12 may participate in the regulatory network involving NtJAZ15.