Objective The aim of the stydy was to explore the role of RAB27B in cervical cancer stem cells and explore new therapeutic targets for patients with advanced and recurrent disease. Methods The si ⁃NC and si⁃RAB27B fragments were successfully transferred into cervical cancer SP cells by cell transfection；Western blotting was used to detect the expression of target gene protein before and after RAB27B knockout；Real ⁃time fluorescence quantitative polymerase chain reaction was used to detected the mRNA expression of target gene in cells； in vivo functional experiments to detect cell proliferation，invasion and clone formation before and after RAB27B knockout；immunofluorescence was used to detect cellular localization of RAB27B and SFRP1；enzyme ⁃linked immunosorbent assay was used to detect SFRP1 secretion. Results After knocking out RAB27B，the protein and mRNA expressions of RAB27B in cervical cancer SP cells were significantly decreased；After RAB27B knockout，the proliferation，invasion and clone formation abilities of cervical cancer SP cells were significantly decreased. There was obvious co⁃localization between RAB27B and SFRP1 in cervical cancer SP cells after knock⁃ ing out RAB27B ；the content of SFRP1 secreted by cervical cancer SP cells was also significantly decreased. Conclusion RAB27B could promote the proliferation，invasion and clone formation of cervical cancer stem cells， and might work throughregulating the expression of SFRP1.