Expression of cholera toxin B subunit in transgenic tomato plants.
Cholera toxin, secreted by Vibrio cholerae, consists of A and B subunits. The latter binds to G(M1)-ganglioside receptors as a pentamer (approximately 55 kDa). Tomato plants were transformed with the gene encoding cholera toxin B subunit (ctxB) along with an endoplasmic reticulum retention signal (SEKDEL) under the control of the CaMV 35S promoter via Agrobacterium-mediated transformation. PCR and Southern analysis confirmed the presence of the ctxB gene in transformed tomato plants. Northern analysis showed the presence of the ctxB-specific transcript. Immunoblot assays of the plant-derived protein extract showed the presence of cholera toxin subunit B (CTB) with mobility similar to purified CTB from V. cholerae. Both tomato leaves and fruits expressed CTB at levels up to 0.02 and 0.04% of total soluble protein, respectively. The G(M1)-ELISA showed that the plant-derived CTB bound specifically to G(M1)-ganglioside receptor, suggesting that it retained its native pentameric form. This study forms a basis for exploring the utility of CTB to develop tomato-based edible vaccines against cholera.
Jani
Dewal
D
Department of Plant Molecular Biology, University of Delhi, New Delhi, India.
Meena
Laxman Singh
LS
Rizwan-ul-Haq
Quazi Mohammad
QM
Singh
Yogendra
Y
Sharma
Arun K
AK
Tyagi
Akhilesh K
AK
D016960:Agrobacterium tumefaciens / Q000235:genetics*; D015152:Blotting, Northern; D015139:Blotting, Southern; D015153:Blotting, Western; D002772:Cholera Toxin / Q000235:genetics* / Q000378:metabolism; D017931:DNA Primers / Q000737:chemistry; D004721:Endoplasmic Reticulum / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005677:G(M1) Ganglioside / Q000378:metabolism; D018506:Gene Expression Regulation, Plant; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D018551:Solanum lycopersicum / Q000235:genetics* / Q000378:metabolism; D018515:Plant Leaves / Q000737:chemistry; D030821:Plants, Genetically Modified; D010957:Plasmids; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000378:metabolism
D017931:DNA Primers; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; C034993:ganglioside receptor; D005677:G(M1) Ganglioside; D002772:Cholera Toxin