Sensitive detection of cucumber mosaic cucumovirus RNA with digoxigenin-labeled cRNA probe

To nonradioactively detect cucumber mosaic cucumovirus (CMV) RNA with high sensitivity, we tested the use of CMV RNA-specific digoxigenin (DIG)-labele d ribonucleic acid probe, which was in vitro transcribed, in dot-blot, nort hern blot and microplate hybridization. The probe could detect up to 10 fg of CMV RNA by dot-blot hybridization with a chemiluminescent detection. Nor thern blot analysis showed that the riboprobe hybridizes to all CMV genomic RNAs in the total RNAs extracted from the inoculated leaves of Japanese ra dish without nonspecific reactions against mock-inoculated samples. Further more, accumulation of CMV RNA in the inoculated leaves of bottle gourd coul d be monitored with dot-blot and microplate hybridization. Thus, we demonst rated that the hybridization methods with nonradioactive riboprobe are appl icable to highly sensitive and specific detection of CMV RNA not only in pu rified virion but also in total RNA extracted from host plants infected wit h CMV.