[Objectives]  To examine the expression of the Bombyx mori serine proteinase inhibitor 20 gene (BmSerpin-20) in Escherichia coli; and more specifically, to analyze its mRNA and protein expression levels in tissues, and its mRNA expression patterns in fat bodies after exogenous microorganism injection. [Methods]  The BmSerpin-20 gene was inserted into the pET-28a vector. Recombinant plasmids were transfected into E. coli Transetta (DE3), and expression of the BmSerpin-20 protein induced with IPTG. The recombinant BmSerpin-20 protein was employed as an antigen to immunize New Zealand rabbits and to prepare a polyclonal antibody serum. The expression patterns of BmSerpin-20 were analyzed using real-time quantitative PCR and the Western blot technique. [Results]  The BmSerpin-20 protein was successfully expressed in E. coli cells, purified and a polyclonal antibody against BmSerpin-20 prepared. The BmSerpin-20 gene and protein were expressed in the fat body, midgut, hemocyte, malpighian, integument, testis, ovary, with the highest expression in the integument. After treatment of B. mori larvae with nuclear polyhedrosis viruses, Micrococcus luteus, E. coli or Beauveria bassiana, the expression patterns of the BmSerpin-20 gene in the fat body was first down-regulated and then up-regulated. The expression level of BmSerpin-20 differed depending on the microorganism injected and injection time. [Conclusion]  The expression pattern of BmSerpin-20 was successfully studied and may lay a foundation for further research on the function of this gene in the immune system of B. mori.