USING NONRADIOACTIVE PROBES ON PLANTS - A FEW EXAMPLES

Due to costs in using and disposing of radiochemicals and to health co nsiderations, we have been developing applications which include non-i sotopic detection of DNA and proteins using chemiluminescence. Our maj or interests are in the detection of viral nucleic: acids and in the a nalysis of transgenic plants. Generally, probes were labelled with dig oxigenin, either by the random priming method or by PCR, and then dete cted with CSPD or CDP-Star. We routinely use a tissue blotting protoco l for diagnosing TYLCV, a plant virus becoming a pest in the Mediterra nean region. Test results were comparable with those using the same ra diolabelled probe. When total nucleic acids are extracted from the pla nt samples and used in dot-blot or Southern blot assays, viral DNAs ar e promptly detected by chemiluminescence. In transgenic plants, chemil uminescence was used to detect the transgene on genomic Southern blots , the transgenic mRNAs on Northern blots, and the transgenic protein o n Western blots. In Southern and Northern blots, the quality of the re sults obtained was usually satisfactory, but not as good as with a rad iolabelled probe, the main problem being the signal-to-background rati o. Our goal is now to improve the quality of results in demanding appl ications such as genomic Southern blots, by reducing the background on membranes. (C) 1998 John Wiley & Sons, Ltd.