title: Prokaryotic expression of CP gene of Fritillary virus Y infecting Thunberg fritillary and antiserum preparation.
abstract: OBJECTIVE
  To prepare antiserum against Fritillary virus Y (FVY) CP for detecting FVY and study serological relationships with other viruses.
  
  
  METHODS
  Specific primer was designed according to Genbank (accession: AM039800) to amplify CP gene of FVY infecting Thunberg fritillary. Sequence relationship with other potyviruses was made by Blast. The CP gene was inserted into pSBET and expressed in Escherichia coli BL21 (DE3) plys E strain. The object protein was purified by 12% SDS-PAGE firstly and subsequently 5% - 20% gradient SDS-PAGE. The antiserum against the CP was raised in mouse and its specificity was confirmed by Western blot analysis. The reactivity of the antiserum produced to FVY CP was tested by Western blot against the over-expressed coat proteins of 17 potyviruses. The ability to combine with nature FVY particles was confirmed by ELISA analysis.
  
  
  RESULTS
  It shared 81.2% nucleotide acids identities with TrVY (Tricyrtis virus Y, AY 864850) CP gene, 68.1% with SMV-P (Soybean mosaic virus Pinellia strain, AJ507388. 2) CP gene and 67.2% with ZYMV (Zucchini yellow mosaic virus Luan isolate) CP gene. The prepared antiserum was special to FVY CP, also reacted moderately to the expressed CP of SMV-P (Soybean mosaic virus Pinellia strain) and weakly to that of ZYMV (Zucchini yellow mosaic virus Luan isolate).
  
  
  CONCLUSIONS
  The antibody could combine to nature FVY particles and the antiserum is suitable for FVY detection by ELISA in large scale.
authors.affiliation: Western Anhui University, Anhui Provincial Laboratory of Biomimetic Sensor and Detecting Technology, Liu'an 237012, China. weichuanbao@sina.com
authors.forename: Chuan-Bao
authors.initials: CB
authors.lastname: Wei
authors.forename: Yang-Yang
authors.initials: YY
authors.lastname: Wei
authors.forename: Yu
authors.initials: Y
authors.lastname: Yang
authors.forename: Shi-Liang
authors.initials: SL
authors.lastname: Liu
authors.forename: Hao-Yu
authors.initials: HY
authors.lastname: Hu
authors.forename: Yue
authors.initials: Y
authors.lastname: He
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D036022:Capsid Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D002681:China; D003001:Cloning, Molecular; D017931:DNA Primers; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D031410:Fritillaria / Q000821:virology*; D015870:Gene Expression; D007106:Immune Sera / Q000276:immunology / Q000302:isolation & purification*; D051379:Mice; D008969:Molecular Sequence Data; D010802:Phylogeny; D030021:Pinellia / Q000821:virology; D016133:Polymerase Chain Reaction; D017800:Potyvirus / Q000145:classification / Q000235:genetics* / Q000276:immunology; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D013025:Glycine max / Q000821:virology
chemical_list: D036022:Capsid Proteins; D017931:DNA Primers; D007106:Immune Sera; D011994:Recombinant Proteins