DETECTION OF DOUBLE-STRANDED-RNA (DSRNA) IN CRUDE EXTRACTS OF VIRUS-INFECTED PLANTS BY INDIRECT ELISA

An antiserum against polyinosinic polycytidylic acid (In-Cn) was used to detect double-stranded RNA (dsRNA) by indirect ELISA (ELISA-I). DsR NA from cucumber mosaic virus (CMV) and plum pox virus (PPV)-infected plants was detected using different types of extracts. The pH of the e xtraction buffer was very important in dsRNA detection, the highest op tical density values being obtained at pH 6 or in aqueous extracts. Ex tracts heated at 80 degrees C for 2 min showed increased optical densi ty values compared with unheated extracts. DsRNA from Nicotiana bentha mina plants infected with each of six PPV isolates was readily detecte d by ELISA-I 50 days after inoculation ELISA values then obtained with the In-Cn antiserum were generally higher than those obtained by doub le antibody sandwich ELISA using an antiserum to virus coat protein. P urified dsRNA from the same infected plants showed no visible band, bu t it produced a fluorescent background when analysed by polyacrylamide gel electrophoresis.