Detection Of dsRNA Research Articles

Detection of plum pox virus by isolation of double‐stranded ribonucleic acid (dsRNA)

Double‐stranded RNA (dsRNA) associated with plum pox virus (PPV) in Nicotiana clevelandii and Prunus domestica has been isolated. While dsRNA was detected in N. clevelandii in considerable amounts by electrophoresis, only small amounts were found in P. domestica. This may be due to viscous substances in the leaves of this woody host. Different PPV strains (NAT ‐ not aphid‐transmissible; AT ‐ aphid‐transmissible) showed specific patterns in electrophoresis gels. When PPV was assayed in N. clevelandii by dsRNA detection or by standard ELISA or ISEM, all three methods were found to be efficient, with none being superior. ELISA, as a simple and fast routine method, is still the method of choice. DsRNA detection will be suitable for plant disease agents undetectable by ELISA and ISEM.

Detection of dsRNA in Particles of Vicia Cryptic Virus and in Vicia faba Tissues and Protoplasts

SUMMARY Isometric particles of vicia cryptic virus (VCV), purified from Vicia faba seedlings infected through seed, had a buoyant density in CsCl of 1.37 g/ml. Analysis by PAGE showed that VCV particles contained three species of dsRNA with mol. wt. of 1.37 × 106, 1.26 × 106 and 1.21 × 106. These dsRNA species were also detected in extracts from leaves, stems, roots, flowers, seeds and mesophyll protoplasts of V. faba; dsRNA was readily detected in extracts from one leaflet (about 0.6 g). Slower-migrating species of dsRNA occurred in many leaf extracts but were not obtained from VCV particles. Electrophoresis of dsRNA extracts in gels that are then stained with silver appears to be a simple and reliable method for detecting ‘cryptic’ viruses. Using this method, VCV dsRNA was detected in 18 cultivars of V. faba, but not in cultivars Beryl and Minica which were thought from previous work to be free from VCV. Neither VCV particles nor VCV dsRNA were detected in VCV-free individual V. faba plants inoculated with VCV mechanically or by dodder. VCV was not detected in leaves of plants affected by cytoplasmic male sterility or in a maintainer line for this condition.

Immunochemical detection of ds-RNA in healthy and virus-infected plants and specific detection of viral ds-RNA by hybridization to labelled complementary DNA

Double-stranded (ds) RNA was detected in nucleic acid preparations from virus-free leaf tissues of several plant species by radial double-diffusion tests with antiserum raised against polyinosinic : polycytidylic acid [poly(I) : poly(C)]. No significant differences in the concentrations of ds-RNA were detected by such tests in virus-free French bean leaves and those infected with tobacco mosaic virus or tobacco ringspot virus. However, virus-specific ds-RNAs as well as genomic RNAs of both viruses were readily detected and estimated quantitatively by hybridization to 3H-labelled complementary DNA probes. It is concluded that it is not feasible to screen plant material infected with single-stranded RNA viruses by immunochemical tests for virus-associated ds-RNAs.