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Bulletin of Botanical Research ›› 2013, Vol. 33 ›› Issue (1): 73-79.doi: 10.7525/j.issn.1673-5102.2013.01.012

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Prokaryotic Expression of TFMV,FVY and LMoV CP Gene,Antiserum Preparation and Virus Detection

WU Qi-Yao;WEI Chuan-Bao*;LI Jin-Hua   

  1. 1.School of Resources and Environmental Engineering, Anhui University, Hefei 230601;2.Western Anhui University Plant Biotechnology Training Centre of Anhui Province, Lu’an 237012
  • Received:1900-01-01 Revised:1900-01-01 Online:2013-01-20 Published:2013-01-20
  • Contact: WEI Chuan-Bao
  • Supported by:

Abstract: According to Genbank, specific primers were designed to amplify the CP genes of TFMV, FVY and LMoV infecting Fritillaria thunbergii Miq. and the CP sequences were analyzed. Then the CP genes were inserted into pSBET vector and expressed in Escherichia coli BL21(DE3)plys E strain. The object proteins were purified by 12% SDS-PAGE firstly and subsequently 5%-20% gradient SDS-PAGE. The antiserum against the CPs was raised in mouse. The specificity and serological relationship were confirmed by Western blot analysis and the ability to combine with nature virus particles was confirmed by ELISA analysis. Western blot, indirect ELISA and Dot-ELISA techniques were used for the detection of viruses infecting F.thunbergii. The results indicated that the antiserum is specific to its CP and no acrossing-reaction with others. It could combine with nature virus particles. Indirect ELISA and Dot-ELISA techniques are suitable for detecting three kinds of viruses.

Key words: Fritillaria thumbergii Miq., virus, prokaryotic expression, antiserum preparation, virus detection

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