7365AB, a recessive genetic male sterility (RGMS) system, is controlled by BnMs3 in Brassica napus which encodes a Tic40 protein required for tapetum development.However, the role of BnMs3 in rapeseed anther development is still largely unclear.In this research, cytological analysis revealed that anther development of Bnms3 has the defects in the transition of tapetum to the secretory type, callose degradation, and pollen wall formation.A total of 76 down-regulated unigenes in Bnms3, several of which are associated with tapetum development, callose degeneration, and pollen development, were isolated by suppression subtractive hybridization (SSH) combined with macroarray.Reverse genetics was applied by means of Arabidopsis insertional mutant lines to characterize the function of these unigenes, and it revealed MSR02 is only required for transport of sporopollenin precursors through the plasma membrane (PM) of tapetum.The real-time PCR data have further verified that BnMs3 plays primary role in tapetal differentiation through affecting the expression of a few key transcription factors, participates in tapetal degradation by modulating expression of cysteine protease genes, and influences microspore separation via manipulating the expression of BnA6 and BnMSR66 related to callose degradation and that of BnQRT1 and BnQRT3 required for the primary cell wall degradation of the pollen mother cell (PMC) as well.Moreover, BnMs3 takes part in pollen wall formation by affecting the expression of a series of genes involved in biosynthesis and transport of sporopollenin precursors.All the above results suggest that BnMs3 participates in tapetum development, microspore release, and pollen wall formation in B.napus.