GENETIC TRANSFORMATION AND HYBRIDIZATION Transgenic tomatoes express an antigenic polypeptide containing epitopes of the diphtheria, pertussis and tetanus exotoxins, encoded by a synthetic gene Ruth Elena Soria-Guerra Æ Sergio Rosales-Mendoza Æ Criso ´ foro Ma ´ rquez-Mercado Æ Rube ´n Lo ´ pez-Revilla Æ Rosalba Castillo-Collazo Æ A ´ ngel Gabriel Alpuche-Solı´s Received: 12 September 2006 / Revised: 21 December 2006 / Accepted: 11 January 2007 Ó Springer-Verlag 2007 Abstract A current priority of vaccinology is the development of multicomponent vaccines that protect against several pathogens. The diphtheria–pertussis– tetanus (DPT) vaccine prevents the symptoms of three serious and often fatal diseases due to the exotoxins produced by Corynebacterium diphteriae, Bordetella pertussis and Clostridium tetani. We are attempting to develop an edible DPT multicomponent vaccine in plants, based on the fusion of protective exotoxin epitopes encoded by synthetic genes. By means of Agrobacterium mediated transformation we generated transgenic tomatoes with a plant-optimised synthetic gene encoding a novel polypeptide containing two adjuvant and six DPT immunoprotective exotoxin epitopes joined by peptide linkers. In transformed to- mato plants, integration of the synthetic DPT (sDPT) gene detected by PCR was confirmed by Southern blot, and specific transcripts of the expected molecular size were detected by RT-PCR. Expression of the putative polypeptide encoded by the sDPT gene was detected by immunoassay with specific antibodies to the diph- theria, pertussis and tetanus exotoxins. The sDPT gene is therefore integrated, transcribed and translated as the expected recombinant sDPT multiepitope poly- peptide in transgenic tomatoes that constitute a po- tential edible vaccine. Keywords Diphtheria Á Whooping cough Á Tetanus Á Synthetic genes Á Transgenic plants Abbreviations DPT Diphtheria–pertussis–tetanus RT-PCR Reverse-transcriptase polymerase chain reaction MS Murashige and Skoog medium IAA Indole-3-acetic acid NAA Naphthalene acetic acid BAP 6-Benzylaminopurine Introduction The diphtheria–pertussis–tetanus (DPT) vaccine, used to immunize against diphtheria, pertussis (whooping cough) and tetanus has dramatically cut down the risks of these diseases and the consequent deaths in children (Gerathy 1984; Hinman and Koplan 1984). Commer- cially available DPT vaccines are composed of toxoids of Corynebacterium diphteriae and Clostridium tetani; the pertussis portion is composed of killed Bordetella pertussis bacteria, whose toxins are responsible for the Communicated by P. Lakshmanan. R. E. Soria-Guerra Á S. Rosales-Mendoza Á R. Lo ´ pez-Revilla Á R. Castillo-Collazo Á A ´ . G. Alpuche-Solı´s (&) Divisio ´n de Biologı´a Molecular, Instituto Potosino de Investigacio ´n Cientı´fica y Tecnolo ´ gica, Camino a la Presa San Jose ´ 2055, 78216 San Luis Potosı´, S.L.P., Mexico e-mail: alpuche@ipicyt.edu.mx C. Ma ´ rquez-Mercado Departamento de Biologı´a Celular, CINVESTAV-IPN, Av. IPN 2508, 07360 Me ´ xico, D.F., Mexico Present Address: C. Ma ´ rquez-Mercado FES-Cuautitla ´n UNAM, Km 2.5 Carretera Cuautitla ´ n-Teoloyucan, 54700 Me ´ xico, Mexico 123 Plant Cell Rep DOI 10.1007/s00299-007-0306-y