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KMID : 0613820180280040478
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Journal of Life Science
2018 Volume.28 No. 4 p.478 ~ p.482
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Use of the Synthetic Gene Encoding the Truncated Human Rotavirus VP8* Protein in Escherichia coli for Production of Vaccine Candidates or Development of Diagnostic Antibodies
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Kim Sang-Rae
Lee Bheong-Uk
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Abstract
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Human rotavirus is a causative agent of acute diarrhea among children. The artificial gene encoding the truncated VP8* protein of human rotavirus A (serotype 1 strain WA) was synthesized according to the Escherichia coli codon preference. The synthetic VP8* gene also possessed the NdeI and HindIII restriction sites for the convenient in-frame cloning for translation and a 6-histidine tag at C-terminus for Ni+ affinity purification. Molecular weight of the truncated VP8* protein deduced from the nucleotide sequences of the artificial gene was a 19.7-kDa. This synthetic VP8* DNA fragment was inserted into the pT7-7 expression vector and transformed into E. coli BL21 (DE3). Transformants harboring the synthetic gene encoding the VP8* protein was induced by supplement of a final concentration of 0.05 mM ITPG at 20℃. Protein crude extract from the E. coli transformants was subjected to Western blotting with the mouse anti-rotavirus capsid antibody, showing ~20-kDa VP8* protein band. The truncated Vp8* protein band was also observed by Western blotting using the rabbit polyclonal antibody serum made against the truncated VP8* protein. This study suggested that the synthetic gene could be used as an easy way to produce the antigenic vaccine candidate for control of virus-associated diseases or to develop antibodies for diagnostic purpose.
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KEYWORD
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Codon preference, Escherichia coli, rotavirus VP8*, synthetic gene
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