Selective separation of virus proteins and double-stranded RNAs by SDS-KCl precipitation.
The total viral structural polypeptides and the double-stranded genomic RNAs of bluetongue virus can be selectively separated by a single SDS-KCl precipitation step. This simple, rapid and highly reproducible method enables greater than 95% recovery and purity of both viral proteins and dsRNAs within 30 min. The serotypic identity of the separated dsRNAs can be analyzed by SDS-PAGE electrophorogram immediately. After a single phenol/chloroform extraction, the dsRNA can also be used as hybridization probes, templates for molecular cloning and direct RNA sequencing. The SDS-KCl-precipitated viral proteins could be used readily for peptide mapping and as immunogens. Polyclonal and monoclonal antibodies raised against SDS-KCl-precipitated viral structural polypeptides were useful in Western immunoblots.
Li
J K
JK
Department of Biology, Utah State University, Logan 84322-5500.
Johnson
T
T
Yang
Y Y
YY
Shore
V
V
D015152:Blotting, Northern; D015153:Blotting, Western; D001820:Bluetongue virus / Q000032:analysis; D011232:Chemical Precipitation; D004591:Electrophoresis, Polyacrylamide Gel; D012089:Mammalian orthoreovirus 3 / Q000032:analysis; D011189:Potassium Chloride; D012330:RNA, Double-Stranded / Q000302:isolation & purification*; D012367:RNA, Viral / Q000302:isolation & purification*; D012967:Sodium Dodecyl Sulfate; D014764:Viral Proteins / Q000302:isolation & purification*
D012330:RNA, Double-Stranded; D012367:RNA, Viral; D014764:Viral Proteins; D012967:Sodium Dodecyl Sulfate; D011189:Potassium Chloride