THE USE OF DIG-LABELED CRNA PROBES FOR THE DETECTION OF CYMBIDIUM MOSAIC POTEXVIRUS (CYMMV) AND ODONTOGLOSSUM RINGSPOT TOBAMOVIRUS (ORSV) IN ORCHIDS

DIG-labelled sense and antisense cRNA probes were synthesized from cDN A clones of CymMV and ORSV for virus detection in infected plants. A s lot-blot hybridization assay was developed using either crude leaf ext racts or total RNA from infected leaves. The assay could detect 50 and 250 pg of purified CymMV and ORSV RNA, respectively. As little as 30 mg of Nicotiana benthamiana infected leaves was sufficient to provide positive detection. CymMV and ORSV were detected at 3125 and 625 times dilution of leaf extracts, respectively. The DIG-labelled cRNA probes are stable for more than a year. This method is sensitive, reliable a nd suitable for large-scale routine testing of plant viruses. By using the two DIG-labelled cRNA probes in situ, CymMV and ORSV were localiz ed in systemically infected leaves and stems of N. benthamiana and orc hids. (C) 1998 Elsevier Science B.V. All rights reserved.