We expressed the B subunit of enterotoxigenic Escherichia coli heat-labile enterotoxin (LTB) encoded by a synthetic codon-optimized gene in carrot. An Agrobacterium-mediated transformation method was used. Thirty independent transgenic lines were regenerated via somatic embryogenesis after 6 months in culture and were transferred to a greenhouse. GM1-ELISA assay was used to assess LTB protein content in mature taproots. Some transgenic lines expressed LTB up to 0.3% of the total soluble protein, which is tenfold higher than the expression levels reported earlier using the native bacterial gene in plants. Immunological assay confirmed proper assembly of the pentameric complex and in vitro activity of the recombinant LTB protein, suggesting that it can be functional in prevention of diarrhea.