E. coli O138 is one of the enterotoxigenic Escherichia coli, causing the postweaning diarrhea and edema disease of weaned pigs. The O-antigen gene cluster of E. coli O138 was sequenced and found to contain the genes rmlB-DAC and gne, gna for the biosynthesis of nucleotide sugars dTDP-rhamnose and UDP-GalNAcA, respectively, genes encoding for O unit flippase(wzx), O-antigen polymerase(wzy) and 3 potential transferase genes. The possible biosynthesis pathway for rare UDP-GalNAcA was proposed. Two genes specific to E. coli O138 were identified. This work provides the basis for a sensitive test by PCR for the rapid detection of E. coli O138. Phylogenetic tree for Gne and GalE proteins was generated and comparisons were made among different strains, and results revealed that these proteins are similar in the second structure, and Gne of E. coli O138 was identified by bioinformatics.