Cotyledons from 8-day-old seedlings of Lotus corniculatus were transformed with a gene encoding the avian influenza (H5N1) hemagglutinin (HA) under the control of the CaMV35S promoter along with matrix attachment regions using Agrobacterium-mediated transformation. The HA gene was detected in genomic DNA from transformed plants by PCR and Southern blotting. Immunoblot analysis of the plant-derived protein revealed the presence of the avian influenza (H5N1) HA. The ELISA results showed that the maximum expression level of the HA protein is up to 0.00786% of the total soluble protein. This is the first report of successful expression of the avian influenza (H5N1) HA in transgenic L. corniculatus, cultivar Leo.