The fat body plays key roles in metabolism and sustenance of growth throughout the life of a silkworm, and thus represents a model tissue for studying development and metamorphosis. Analysis of 18480 ESTs derived from larval and pupal fat body cDNA libraries allowed characterization of gene expression patterns in the fat body during metamorphosis. By Phrap assembling, 6814 estimated transcription units (TUs) were generated, 2673 of which contained 60% of the total ESTs and represented 1366 distinctive known genes. These genes were classified into 14 categories based on their known or putative functions, and most were found to be involved in metabolism and cell defense. Further comparative analysis of the expression profiles revealed that the gene expression patterns in the larval fat body strikingly differed from those in the pupal fat body. From this, we identified a subset of genes potentially associated with metamorphic events such as the formation or destruction of specific tissues, and simultaneously confirmed the stage-specific expression patterns of several identified genes using RT-PCR examination. This work will provide a valuable resource for studying regulatory mechanisms associated with fat body developmental changes and silkworm metamorphosis.