Basal gene expression levels across the bovine gastrointestinal tract (GI) were examined in an attempt to formulate genetic explanations for the differences in function that are known or thought to exist between the various regions. Gene expression along the tract was studied through the random sequencing of a total of 16 412 clones from seven tissue-specific cDNA libraries spanning its length. The expressed sequence tags (ESTs) within each library were clustered to reduce clone redundancy and obtain longer consensus sequences. BLASTN and BLASTX searches against the NCBI human RefSeq databases were used to find putative matches for the bovine sequences and gene ontology assignments were made. Notable similarities and differences in gene expression were observed among the various compartments of the GI tract of the bovine. Many of the prominent transcripts have yet to be reliably identified and the prominence of others may be worthy of further examination. This collection of ESTs represents an important resource for the future construction of a GI tract specific microarray for further gene expression studies.