Detection of rice tungro bacilliform virus gene products in vivo.
To study the products of the open reading frames (ORFs) of rice tungro bacilliform virus in rice plants the sequences containing ORFs I (encoding a 24-kDa protein, P24) and IV (P46) and the protease and polymerase (reverse transcriptase+RNaseH) domains of ORF III were cloned into a pGEX expression vector. The proteins, which were C-terminal fusions to glutathione S-transferase, were expressed in Escherichia coli and antisera were raised against them which, together with an antiserum against virus particles, was used to probe blots of proteins from infected and uninoculated plants and from virus preparations. The P24 antiserum detected virus-specific proteins of 74, 60, and 52 kDa, which are much bigger than expected. These proteins were found in virus preparations and immunogold labeling suggested that they might be internal in the particles. Virus-specific proteins of 33, 37, 62, and > 150 kDa were revealed by antiserum to virus particles. The antiserum to the protease revealed proteins of 13.5, 37, and 68 kDa both in extracts from infected plants and in purified virus preparations. This antiserum decorated intact virus particles as did the particle antiserum. The polymerase domain antiserum reacted with products of 56, 65, and 68 kDa in extracts from infected plants but not in virus particles. The antiserum to the ORF IV product did not detect any bands in either infected plant extracts or virus preparations. The significance of these products is discussed.
Hay
J
J
Department of Virology, John Innes Centre, Norwich Research Park, Colney, United Kingdom.
Grieco
F
F
Druka
A
A
Pinner
M
M
Lee
S C
SC
Hull
R
R
D016282:Aspartic Acid Endopeptidases / Q000235:genetics; D001483:Base Sequence; D015153:Blotting, Western; D004926:Escherichia coli; D007106:Immune Sera; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D012275:Oryza / Q000821:virology*; D010935:Plant Diseases; D010942:Plant Viruses / Q000201:enzymology / Q000235:genetics*; D016133:Polymerase Chain Reaction; D012194:RNA-Directed DNA Polymerase / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D016914:Ribonuclease H / Q000235:genetics; D014764:Viral Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification
D007106:Immune Sera; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; D012194:RNA-Directed DNA Polymerase; D016914:Ribonuclease H; D016282:Aspartic Acid Endopeptidases