Expression of cauliflower mosaic virus ORF II in a baculovirus system.
The cauliflower mosaic virus ORF II encoding the aphid transmission factor (ATF) was mutagenized to introduce a BamHI restriction site upstream from the initiation codon and then cloned into an eukaryotic viral expression vector (Autographa californica nuclear polyhedrosis virus). All recombinant viruses tested in Spodoptera frugiperda (SF21) cells expressed a protein of about 18 kD which comigrated in PAGE with ATF from infected plants. Western blotting using an oligopeptide antiserum to ATF confirmed the identity of the 18-kD protein from infected cells as the product of the ORF II sequences (P18). Subcellular fractionation of cells infected with the recombinant AcMNPV demonstrated that the expressed P18 accumulated intracellularly in an insoluble form. Antiserum was produced in rabbit against the partially purified P18 expressed in SF21 cells. When used to immunogold label ultrathin sections of cauliflower mosaic virus (CaMV)-infected turnip tissue, this antiserum was shown to be highly specific, labelling only the electronlucent inclusion bodies (containing P18) and not other plant cellular components.
Espinoza
A M
AM
Cellular and Molecular Biology Research Center (CIBCM), University of Costa Rica, San José.
Usmany
M
M
Pirone
T P
TP
Harvey
M
M
Woolston
C J
CJ
Medina
V
V
Vlak
J M
JM
Hull
R
R
D000818:Animals; D016367:Baculoviridae / Q000235:genetics*; D001483:Base Sequence; D005591:Chemical Fractionation; D003001:Cloning, Molecular; D005822:Genetic Vectors / Q000235:genetics*; D007150:Immunohistochemistry; D008969:Molecular Sequence Data; D009029:Mosaic Viruses / Q000235:genetics*; D009036:Moths / Q000235:genetics / Q000382:microbiology; D016366:Open Reading Frames / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D014508:Urea; D014764:Viral Proteins / Q000096:biosynthesis / Q000235:genetics
D011994:Recombinant Proteins; D014764:Viral Proteins; D014508:Urea