A Watermelon mosaic virus (WMV)-specific, polyclonal antiserum was raised in rabbits by immunization with purified capsid protein expressed and purified from Escherichia coli cells. The cp gene was RT-PCR-amplified using specific oligonucleotides, and viral RNA extracted from concentrated particles as a template. The amplified fragment was cloned in pBLUESCRIPT KS+ and fully sequenced in order to confirm its identity and integrity. The fragment was subsequently subcloned into the pRSET-A expression vector. Recombinant plasmids were used to induce the expression of the capsid protein in E. coli BL21::DE3. The protein was purified by affinity chromatography using a Ni+-NTA resin, from E. coli total protein extract. The purified protein was quantified and used for the immunization of rabbits. The antiserum was tested for its sensitivity and specificity in Western blot, DAS-ELISA, immunodiffusion and indirect ELISA assays. All tests indicated that the antiserum raised from in vitro-expressed capsid protein is highly specific for the detection of WMV in plant extracts, with no interspecific, heterologous reactions being observed.
