Taiwan Agricultural Research Institute, Council of Agriculture, Executive Yuan--Publication-Journal of Taiwan Agricultural Research

Journal of Taiwan Agricultural Research

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Immunogenicity of Tobamovirus Universal Antibodies Raised Against Expressed Protein Encoded by a Synthetic Polynucleotide Containing Epitope Genes of ToMV, ORSV, and CGMMV

Chou, C. M., C. T. Chen, C. H. Tsai, C. C. Chen, and T. C. Deng. 2018. Immunogenicity of tobamovirus universal antibodies raised against expressed protein encoded by a synthetic polynucleotide containing epitope genes of ToMV, ORSV, and CGMMV. J. Taiwan Agric. Res. 67(3):301–308.

Immunogenicity of Tobamovirus Universal Antibodies Raised Against Expressed Protein Encoded by a Synthetic Polynucleotide Containing Epitope Genes of ToMV, ORSV, and CGMMV

Chien-Ming Chou1, Chun-Tao Chen2, Chin-Hui Tsai3, Chin-Chih Chen1, and Ting-Chin Deng4,*

Abstract

    Using of synthetic peptide immunogens to produce antibodies with universal immunogenicity are a useful tool for diagnosis of plant viral diseases. Addition to the coat protein (CP) gene of Tomato mosaic virus (ToMV), we adopted the CP gene containing the predicted epitope sequences of Odontoglossum ringspot virus (ORSV) and Cucumber green mottle mosaic virus (CGMMV) to assemble into a fused polynucleotide. A total of 1,136 bases of synthetic nucleotides were joined covalently and created a recombinant plasmid cloned in the vector of pUC57 in Escherichia coli. The protein containing peptides encoded by the synthetic polynucleotide was expressed by the vector pET-28a. After purification, this expressed protein was used to immunize a New Zealand rabbit for production of a polyclonal antiserum. By Western blot, such an antiserum reacted with corresponding virus was examined for immunological properties and the coverage of antigenic universe. As the antiserum was applied to indirect enzyme-linked immunosorbent assay (ELISA), every tobamovirus tested in this study was detectable by the universal antibodies, but Hibiscus latent Singapore virus (HLSV) was an exception. A single reagent can detect a variety of tobamoviruses simultaneously; therefore, it can be practically applied to quarantine inspection and productions of Solanaceous seeds, Cucurbitaceous seeds, and some ornamental vegetative propagules.

Key words: Virus, Coat portion, Immunogen, Polyclonal antiserum, Western blot.


* Corresponding author, e-mail: tcde@tari.gov.tw
1 Assistant Research Fellow, Plant Pathology Division, Taiwan Agricultural Research Institute, Taichung, Taiwan, ROC.
2 Associate Technical Specialist, Hsinchu Branch, Bureau of Animal and Plant Health Inspection and Quarantine, Taoyuan, Taiwan, ROC.
3 Research Assistant, Plant Pathology Division, Taiwan Agricultural Research Institute, Taichung, Taiwan, ROC.
4 Research Fellow, Plant Pathology Division, Taiwan Agricultural Research Institute, Taichung, Taiwan, ROC.

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