Acta Entomologica Sinica ›› 2019, Vol. 62 ›› Issue (7): 823-829.doi: 10.16380/j.kcxb.2019.07.005

• RESEARCH PAPERS • Previous Articles     Next Articles

Preparation of the polyclonal antibody of capsid protein VP1 of Himetobi P virus in the small brown planthopper, Laodelphax striatellus (Hemiptera: Delphacidae) and its application in the virus detection

PIAO Jun1,#, XU Chun-Ling1,2,#, PIAO Jing-Ai1, ZHOU Yi-Jun2, LI Shuo2,*   

  1. (1. School of Life Science, Liaoning Normal University, Dalian, Liaoning 116081, China; 2. Institute of Plant Protection, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, China)
  • Online:2019-07-20 Published:2019-07-09

Abstract: 【Aim】 In our previous work, the interaction between rice stripe virus (RSV) and Himetobi P virus (HiPV) of the small brown planthopper (SBPH), Laodelphax striatellus, was found. This study aims to prepare the polyclonal antibody (PAb) of the capsid protein VP1 of HiPV and to assess its application in the detection of HiPV, so as to provide a technical basis for further studying the interaction of HiPV-RSV and HiPV-SBPH. 【Methods】 VP1 gene of HiPV was amplified from SBPH adult via RT-PCR, and subcloned into the prokaryotic expression vector pET-32a to construct the vector pET-VP1. The recombinant plasmid was transformed into Escherichia coli BL21 (DE3), and the recombinant protein was obtained by IPTG induction and purified by Ni2+-NTA affinity chromatography. The antibody was prepared by immunizing New Zealand white rabbit with the purified recombinant protein. 【Results】 A 774 bp product of capsid protein gene VP1 of HiPV was amplified from SBPH, and a 47.5 kD VP1 fusion protein was obtained through prokaryotic expression and purification. The purified VP1 recombinant protein was used to immunize New Zealand white rabbit, and the PAb was prepared. The titer of PAb was 1∶819 200 in indirect ELISA, and the PAb could react specifically with HiPV VP1 but not with SBPH proteins. Using PAb against HiPV, methods of Western blot and immunocapture-RT-PCR (IC-RT-PCR) for the detection of HiPV in single adult of SBPH were established. The detection results showed that HiPV was prevalent in the RSV-infected and uninfected SBPH high-affinity populations. 【Conclusion】 HiPV can be detected specifically in SBPH using the prepared VP1 polyclonal antibody. This study provides a technical support for the rapid detection of HiPV and the study of HiPV-RSV and HiPV-SBPH interaction.

Key words: Laodelphax striatellus, Himetobi P virus, VP1 gene, prokaryotic expression, polyclonal antibody, virus detection