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Expression of a cholera toxin B subunit in transgenic lettuce (Lactuca sativa L.) using Agrobacterium-mediated transformation system

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Abstract

To increase expression level of cholera toxin B subunit (CTB) in lettuce plants, synthetic CTB (sCTB) gene based on the optimized codon usage was fused with an endoplasmic reticulum retention signal, KDEL. The sCTB gene was introduced into a plant expression vector and transformed to lettuce plants using Agrobacterium-mediated transformation system. As a selection marker, a bialaphos resistance (bar) gene that encodes phosphinothricin acetyltransferase (PAT), conferring tolerance to the herbicide phosphinothricin (PPT), was used. PCR amplification of genomic DNA confirmed the presence of the sCTB gene in the transgenic lettuce plants. Expressions of mRNA and protein of sCTB were observed by Northern and Western blot analyses, respectively. The sCTB synthesized in the transgenic lettuce showed strong affinity for GM1-ganglioside suggesting that the sCTB conserved the antigenic sites for binding and proper folding of pentameric CTB structure. The expression level of CTB was relatively high, reaching total soluble protein (TSP) levels of 0.24% in transgenic lettuce.

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Abbreviations

BA:

Benzyl aminopurin

NAA:

Naphthalene acetic acid

GM1 :

Galactosyl-N-acetylgalactosamyl-sialyl-galactosylglucosylceramide

PPT:

Phosphinothricin

sCTB:

Synthetic cholera toxin B subunit

TSP:

Total soluble protein

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Acknowledgments

This work was supported by a Korean Research Foundation Grant (KRT-2004-005-F00025).

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Authors and Affiliations

  1. Division of Biological Sciences and the Research Center of Bioactive Materials, Chonbuk National University, Jeonju, Chonbuk, 561-756, Korea

    Young-Sook Kim, Bang-Geul Kim, Tae-Geum Kim, Tae-Jin Kang & Moon-Sik Yang

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  1. Young-Sook Kim
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  2. Bang-Geul Kim
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  3. Tae-Geum Kim
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  4. Tae-Jin Kang
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  5. Moon-Sik Yang
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Correspondence to Moon-Sik Yang.

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Kim, YS., Kim, BG., Kim, TG. et al. Expression of a cholera toxin B subunit in transgenic lettuce (Lactuca sativa L.) using Agrobacterium-mediated transformation system. Plant Cell Tiss Organ Cult 87, 203–210 (2006). https://doi.org/10.1007/s11240-006-9156-5

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  • DOI: https://doi.org/10.1007/s11240-006-9156-5

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