title: Nonradioactive, photobiotin-labelled DNA probes for the routine diagnosis of barley yellow dwarf virus.
abstract: Photobiotin was used to prepare biotinylated, nonradioactive nucleic acid probes for the detection of the RNA of barley yellow dwarf virus (BYDV) in plant extracts. A 1.7-kb cDNA of the PAV isolate of BYDV in the plasmid pUC8 vector was biotinylated and then used intact or as sonicated double-stranded DNA fragments. Simple methods were developed for the preparation of partially purified nucleic acid extracts of cereals and their spotting, after formaldehyde treatment, onto nitrocellulose membranes. After hybridization, biotin-labelled DNA bound to BYDV RNA on the nitrocellulose was detected with an avidin-alkaline phosphatase conjugate. BYDV RNA was readily detected with a sensitivity similar to that found with the same probe labelled with 32P by nick translation. Healthy plant extracts gave colourless spots.
authors: Department of Biochemistry, University of Adelaide, Australia.
authors: N
authors: N
authors: Habili
authors: J L
authors: JL
authors: McInnes
authors: R H
authors: RH
authors: Symons
mesh_terms: D000345:Affinity Labels; D001386:Azides; D001710:Biotin / Q000031:analogs & derivatives*; D004274:DNA, Recombinant; D004279:DNA, Viral / Q000235:genetics*; D005557:Formaldehyde; D001467:Hordeum / Q000382:microbiology; D009693:Nucleic Acid Hybridization; D010942:Plant Viruses / Q000235:genetics*; D012367:RNA, Viral / Q000032:analysis*
chemical_list: D000345:Affinity Labels; D001386:Azides; D004274:DNA, Recombinant; D004279:DNA, Viral; D012367:RNA, Viral; D005557:Formaldehyde; D001710:Biotin; C045549:photobiotin