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Expression and Production of Polyclonal Antibodies against Recombinant Coat Protein of Peanut bud necrosis virus
Sivaprasad Y, Bhaskara Reddy BV, Sujitha A and Sai Gopal DVR
In vitro gene expression strategy was used for the production of polyclonal antiserum to the coat protein (CP) of Peanut bud necrosis virus (PBNV). The GBNV CP gene from peanut isolate was cloned into pQE-30UA expression vector and transformed into Escherichia coli (M15) cells. Expression of the CP gene of GBNV was induced in vitro and recombinant protein (~34 KDa) was purified and used for immunization of rabbits to produce the GBNV-specific polyclonal antiserum. The antiserum had a titre of 1:5000 in an indirect Enzyme Linked Immunosorbent Assay (ELISA) and reacted specifically in Western blot. The resulting antiserum was used to develop an Immunocapture Reverse Transcription-Polymerase Chain Reaction (IC-RT-PCR) assay and compared its sensitivity levels with ELISA for detection of GBNV isolates. The recombinant antiserum successfully detected natural infection of GBNV in economically important crops and weed hosts from South India