EXPRESSION OF NUCLEOCAPSID PROTEIN OF CAPSICUM CHLOROSIS VIRUS IN A PROKARYOTIC SYSTEM AND ITS APPLICATION IN VIRUS DETECTION

X. Rao, Y. Li, Z. Wu, Y. Liu, W. Wang

Abstract


Capsicum chlorosis virus (CaCV), which belongs to the Watermelon silver mottle virus (WSMoV) serogroup of the genus Tospovirus, can seriously harm economically important crops. A CaCV strain from Phalaenopsis orchids (CaCV-pha) was identified by RT-PCR. The amino acid sequence of the CaCV-pha nucleocapsid protein (NP) shared 92.0% to 97.1% identity with comparable proteins of other CaCV isolates, with which it was phylogentically related. Since the identification of CaCV-infected crops requires the availability of sensitive diagnostic tools, a prokaryotic expression system was used to obtain a recombinant CaCV-pha NP for antiserum production. The CaCV-pha NP gene was subcloned into the pET-28a (+) vector and transformed into Escherichia coli Rosetta (DE3) cells for expression. After purification, the recombinant protein was injected into rabbits to generate a polyclonal antiserum which, in Western blot tests reacted strongly with the recombinant protein used as antigen. The availability of an anti-CaCV-NP serum will facilitates field surveys for, and further research on CaCV.

Keywords


Tospovirus;recombinant protein;polyclonal antiserum;diagnosis;indirect ELISA

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DOI: http://dx.doi.org/10.4454/JPP.V96I2.032

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EDIZIONI ETS, Pisa, Italy