Two cDNA libraries were constructed from the gut of European corn borer (ECB, Ostrinia nubilalis) larvae. A total of 9,000 random clones from both the libraries were sequenced. These sequences were assembled into 2,451 clusters including 1,787 singletons (unique sequences) and 664 contiguous sequences (contigs). Among these sequences, 78% encoded for putative proteins which shared significant sequence similarity with known sequences in GenBank with E-values ≤ 10-5. These sequences includes many genes encoding for various digestive enzymes such as chymotrypsin-like proteases, trypsin like-proteases, elastases, cysteine proteases, carboxypeptidases, aminopeptidases, and cathepsin L-like proteases. These putative proteins were further analyzed and sorted into different categories of molecular functions, biological processes and cellular components. This work provides foundation for the future research to use microarray technology for analyzing the global changes of gene expression in response to Bt protoxins/toxins as well as the genetic difference between Bt resistant and susceptible strains of ECB.