Colony hybridizations using nylon membranes and RNA probes--an improved method for screening bacterial clones containing bluetongue virus genome.
Bluetongue virus (BTV) total genomic and isolated individual segment dsRNAs end-labeled with 32P were successfully used as probes in colony hybridization to detect clones of BTV genomic material. The RNA probes were highly specific for cloned BTV genomic material. DNA probes, however, gave false positive results. DNA from bacterial clones was fixed to nylon and nitrocellulose membranes. The hybridized nylon membranes could be stripped of probe and reprobed at least 6 times without loss of signal strength.
Unger
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RE
Department of Pharmacology, University of California, Davis 95616.
Chuang
R Y
RY
Chuang
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LF
Osburn
B I
BI
Dol
R H
RH
D000818:Animals; D001419:Bacteria / Q000032:analysis / Q000235:genetics*; D001820:Bluetongue virus / Q000302:isolation & purification*; D002478:Cells, Cultured; D003001:Cloning, Molecular; D003101:Collodion; D004279:DNA, Viral / Q000032:analysis* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D008567:Membranes, Artificial; D009693:Nucleic Acid Hybridization; D009757:Nylons; D010761:Phosphorus Radioisotopes; D012330:RNA, Double-Stranded / Q000302:isolation & purification; D012367:RNA, Viral / Q000302:isolation & purification; D012087:Reoviridae / Q000302:isolation & purification*; D014162:Transfection
D004279:DNA, Viral; D008567:Membranes, Artificial; D009757:Nylons; D010761:Phosphorus Radioisotopes; D012330:RNA, Double-Stranded; D012367:RNA, Viral; D003101:Collodion