The CHORI-212 bacterial artificial chromosome (BAC) library was constructed by cloning EcoRI/EcoRI partially digested DNA into the pTARBAC2.1 vector. The library has an average insert size of 161 kb, and provides 10.6-fold coverage of the channel catfish haploid genome. Screening of 32 genes using overgo or cDNA probes indicated that this library had a good representation of the genome as all tested genes existed in the library. We previously reported sequencing of approximately 25,000 BAC ends that generated 20,366 high-quality BAC end sequences (BES) and identified a large number of sequences similar to known genes using BLASTX searches. In this work, particular attention was given to identification of BAC mate pairs with known genes from both ends. When identified, comparative genome analysis was conducted to determine syntenic regions of the catfish genome with the genomes of zebrafish and Tetraodon. Of the 141 mate pairs with known genes from channel catfish, conserved syntenies were identified in 34 (24.1%), with 30 conserved in the zebrafish genome and 14 conserved in the Tetraodon genome. Additional analysis of three of the 34 conserved syntenic groups by direct sequencing indicated conserved gene contents in all three species. This indicates that comparative genome analysis may provide shortcuts to genome analysis in catfish, especially for short genomic regions once the conserved syntenies are identified.