title: Detection of plant viruses in mixed infection by a macroarray-assisted method.
abstract: The protocol for a simple, sensitive, and specific method using a cDNA macroarray to detect multiple viruses is provided. The method can be used even at the production sites for crops, which need a reliable routine diagnosis for mixed infection of plant viruses. The method consists of three steps: RNA extraction, duplex RT-PCR, and "microtube hybridization" (MTH). Biotinylated cDNA probes are prepared using RT-PCR and used to hybridize a nylon membrane containing target viral cDNAs by MTH. Positive signals can be visualized by colorimetric reaction and judged by eyes. We here demonstrate this method to detect asparagus viruses (Asparagus virus 1 and Asparagus virus 2) from latently infected asparagus plants.
authors.affiliation: Horticultural Science, Research Faculty of Agriculture, Hokkaido University, Kita 9 Nishi 9, Kita-ku, Sapporo, Hokkaido, 060-8589, Japan, hana@res.agr.hokudai.ac.jp.
authors.forename: Hanako
authors.initials: H
authors.lastname: Shimura
authors.forename: Kazuyoshi
authors.initials: K
authors.lastname: Furuta
authors.forename: Chikara
authors.initials: C
authors.lastname: Masuta
mesh_terms: D001710:Biotin / Q000737:chemistry; D060085:Coinfection; D003124:Colorimetry / Q000379:methods; D017931:DNA Primers; D015342:DNA Probes / Q000737:chemistry; D018076:DNA, Complementary / Q000737:chemistry; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation / Q000379:methods*; D010935:Plant Diseases / Q000821:virology*; D010942:Plant Viruses / Q000235:genetics / Q000302:isolation & purification* / Q000472:pathogenicity; D010957:Plasmids; D012367:RNA, Viral / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods
chemical_list: D017931:DNA Primers; D015342:DNA Probes; D018076:DNA, Complementary; D012367:RNA, Viral; D001710:Biotin