P.24. Serological detection of Watermelon bud necrosis virus by polyclonal antiserum to virus nucleocapsid protein
Abstract
Watermelon bud necrosis virus (WBNV) has emerged as a serious pathogen of cucurbits in India. Its natural infection has been detected by using polyclonal antiserum to nucleocapsid protein (NP). Serological diagnosis, however, is limited by the difficulty in producing quality polyclonal antiserum to NP, which is difficult to purify in reasonable quantities. In-vitro expression strategy was thus followed for the production of polyclonal antiserum to NP of WBNV. The WBNV NP gene from watermelon isolate was cloned into 6x His-tagged UA cloning vector and expressed in Escherichia coli [M15] cells. The fusion protein was detected in insoluble fraction and was purified by using Ni-NTA agarose resin. The purified 6x His-fusion protein (∼32 kDa) reacted with antiserum to NP of Watermelon silver mottle virus and was used for immunisation to produce a high titre polyclonal antiserum. The antiserum (1:4000 dilution) successfrully detected the natural infection of WBNV as well as Groundnut bud necrosis virus in a wide range of hosts from different locations.
