Comparison of four radioactive probes for the diagnosis of potato spindle tuber viroid by nucleic acid spot hybridization

Abstract

32P-labelled probes prepared by nick-translation, two methods (A and B) of labelling PSTV cDNA in M13 phage DNA, and RNA transcription were compared for their sensitivity in detecting PSTV. Nick-translated and method B probes were the least sensitive. The minimum concentration of PSTV detected by these probes was 100 pg. Method A probe detected 10–20 pg of PSTV whereas RNA transcripts were capable of detecting as little as 0.33 pg of PSTV. RNA transcripts are easy to prepare, and their use in hybridization analysis do not cause background reactions. Sometimes low non-specific signals are produced but they can be eliminated by washing membranes with RNase A. RNA probes are now used for routine detection of PSTV in tuber flesh and sprouts, leaves and true seed of potato at the International Potato Center (CIP).