Expression and Production of Polyclonal Antibodies against Recombinant Coat Protein of Peanut bud necrosis virus

Sivaprasad Y Bhaskara Reddy Bv

doi:10.4172/2157-7471.1000s1-002

Abstract

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

Highlights

Peanut bud necrosis virus (PBNV) is one of the re-emerging viral diseases on several economically important crops such as peanut, tomato, chilli, potato, sunflower, black gram, green gram, cowpea, soybean, jute, taro, cotton, carrot, onion, etc., in India
The virus was identified as distinct Tospovirus and named as Groundnut bud necrosis virus (GBNV), which is placed in serogroup IV [1,4]
The coat protein (CP) gene of GBNV isolates was initially targeted for the genetic characterization of GBNV associated with mosaic and necrosis disease of groundnut and other crops in India

Summary

Introduction

Peanut bud necrosis virus (PBNV) is one of the re-emerging viral diseases on several economically important crops such as peanut, tomato, chilli, potato, sunflower, black gram, green gram, cowpea, soybean, jute, taro, cotton, carrot, onion, etc., in India. The bud necrosis disease was distributed in South and South-East Asia [2], and its cause was first identified as Tomato spotted wilt virus (TSWV) in India [3]. The virus was identified as distinct Tospovirus and named as Groundnut bud necrosis virus (GBNV), which is placed in serogroup IV [1,4]. The RNAs are designated L (large), M (medium) and S (small) and have a size of calculated 8.9, 4.8 and 2.9 kb, respectively They are bounded by nucleocapsid (N) protein [5]. The S-RNA encodes NSs protein in the V sense and N protein in the VC sense

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Conclusion