The mature surface layer (S-layer) protein SlpC of mosquitocidal Bacillus sphaericus C3-41 comprises amino acids 31–1,176 and could recrystallize in vitro. The Nterminal SLH domain is responsible for binding function. Deletion of this part, S-layer proteins could not bind to the cell wall sacculi. To investigate the self-assembly ability of SlpC from B. sphaericus, nine truncations were constructed and their self-assembly properties were compared with the recombinant mature S-layer protein rSlpC31–1,176. The results showed that rSbsC31–1,176 and truncations rSlpC211–1,176, rSlpC278–1,176, rSlpC31–1,100, and rSlpC31–1,050 could assemble into multilayer cylinder structures, while N-terminal truncations rSlpC338–1,176, rSlpC438–1,176, and rSlpC498–1,176 mainly showed monolayer cylinders in recombinant Escherichia coli BL21 (DE3) cells. Growth phase analysis of the self-assembly process revealed that rSlpC498–1,176 mainly formed monolayer cylinders in the early stage (0.5 and 1 h induction of expression), but few double-layer or multilayer cylinders were also found with the cells growing, while rSlpC31–1,176 could formed multilayer cylinders in all the growth stage in the E. coli cells. It is concluded that the deletion of the C-terminal 126 aa or the N-terminal 497 aa did not interfere with the self-assembly process, the fragment(amino acids 278 to 337) is essential for the multilayer cylinder formation in E. coli BL21 (DE3) cells in the early stage and the fragment (amino acids 338 to 497) is related to monolayer cylinder formation. The information is important for further studies on the assembly mechanism of S-layer proteins and forms a basis for further studies concerning surface display and nanobiotechnology.