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Nucleic Acids Research, 2003, Vol. 31, No. 4 1174-1179
© 2003 Oxford University Press

Expression of tetanus toxin Fragment C in tobacco chloroplasts

John S. Tregoning1,2, Peter Nixon2, Hiroshi Kuroda1, Zora Svab1, Simon Clare2, Frances Bowe2, Neil Fairweather2, Jimmy Ytterberg3, Klaas J. van Wijk3, Gordon Dougan2 and Pal Maliga*,1

1 Waksman Institute, Rutgers University, 190 Frelinghuysen Road, Piscataway, NJ 08854-8020, USA, 2 Department of Biological Sciences and Centre for Molecular Microbiology and Infection, Imperial College of Science and Technology, Exhibition Road, London SW7 2AY, UK and 3 Department of Plant Biology, Emerson Building, Tower Road, Cornell University, Ithaca, NY 14853, USA

*To whom correspondence should be addressed. Tel: +1 732 445 5329; Fax: +1 732 445 5735; Email: maliga{at}waksman.rutgers.edu

Fragment C (TetC) is a non-toxic 47 kDa polypeptide fragment of tetanus toxin that can be used as a subunit vaccine against tetanus. Expression of TetC in Escherichia coli and yeast was dependent on the availability of synthetic genes that were required to improve translation efficiency and stabilize the mRNA. To explore the feasibility of producing TetC in tobacco leaves, we attempted expression of both the bacterial high-AT (72.3% AT) and the synthetic higher-GC genes (52.5% AT) in tobacco chloroplasts. We report here that the bacterial high-AT mRNA is stable in tobacco chloroplasts. Significant TetC accumulation was obtained from both genes, 25 and 10% of total soluble cellular protein, respectively, proving the versatility of plastids for expression of unmodified high-AT and high-GC genes. Mucosal immunization of mice with the plastid- produced TetC induced protective levels of TetC antibodies. Thus, expression of TetC in chloroplasts provides a potential route towards the development of a safe, plant-based tetanus vaccine for nasal and oral applications.


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