Kunitz-type chymotrypsin inhibitor CIb1 of silkworm Bombyx mori is a basic peptide consisting of 62 amino acid residues. To investigate the physiological roles of this peptide, we cloned it for genomic structure analysis. CIb1 cDNA was used as a probe to screen a BAC sub-library. One positive clone containing the upstream sequences was isolated and the sequence result showed that CIb1 gene consists 3 exons spaced by 2 introns. Consensus TATA and CCAAT boxes were identified in the 5'-flanking region. Other binding sites for transcription factors such as NF-kB, GATA, C/EBP, COUP-TF/HNF-4, SRY, and HOXA3 were also detected. Southern blotting suggested a single copy of CIb1 gene in the silkworm genome. Northern blotting indicated that the expression of CIb1 gene is transcriptionally regulated during development and apparently tissue specific. Together our data, the transcription factors putatively responsible for the roles of CIb1 in silkworm hemolymph were discussed.