Expression and assembly of cholera toxin B subunit (CTB) in transgenic carrot (Daucus carota L.).
We expressed the cholera toxin B subunit (CTB) fused to an endoplasmic reticulum retention signal (SEKDEL) in carrot roots using an Agrobacterium-mediated transformation method. Fourteen independent transgenic lines were regenerated via somatic embryogenesis after 6 months of culture. The sCTB gene was detected in the genomic DNA of transgenic carrot by PCR amplification. Expressions and assembly of sCTB protein into oligomeric structures of pentameric size were observed in transgenic plant extracts by Western blot analysis. The sCTB produced by transgenic carrot roots demonstrated strong affinity for GM1-ganglioside, suggesting that the sCTB conserved the antigenic sites for binding and proper folding of the pentameric sCTB structure. The expression level of sCTB comprised approximately 0.48% of total soluble protein (TSP) in root of transgenic carrot.
Kim
Young-Sook
YS
Division of Biological Sciences and the Research Center of Bioactive Materials, Chonbuk National University, Jeonju, Chonbuk, 561-756, South Korea.
Kim
Mi-Young
MY
Kim
Tae-Geum
TG
Yang
Moon-Sik
MS
D001483:Base Sequence; D002772:Cholera Toxin / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D003001:Cloning, Molecular; D018552:Daucus carota / Q000235:genetics* / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005677:G(M1) Ganglioside / Q000378:metabolism; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D018517:Plant Roots / Q000235:genetics / Q000378:metabolism; D030821:Plants, Genetically Modified; D016133:Polymerase Chain Reaction; D055503:Protein Multimerization; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D012231:Rhizobium / Q000235:genetics / Q000378:metabolism; D014170:Transformation, Genetic
D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; C062059:SEKDEL sequence; D005677:G(M1) Ganglioside; D002772:Cholera Toxin