Articles
RECENT PROGRESS IN CHRYSANTHEMUM VIROID DETECTION
Previous results have shown the sensitivity and the reliability of this technique and pointed out the importance of sampling.
The electrophoretic assay can be applied to checking of very small samples and consequently is especially appropriate for indexing of mother plants including in vitro plantlets.
We demonstrate here that PASGE can also be applied to mass indexing with some modifications mainly consisting in the elimination of cell nucleic acids.
So, after the usual LiCl treatment, the supernatant is treated by cellulose (Biorad cellex 410) in the presence of 30 % ethanol under conditions where the viroid RNA is adsorbed while DNA fractions and single stranded RNAs which are not adsorbed can be easily eliminated.
Special attention must be paid to the interpretation of the electrophoretic patterns.
There are, in healthy or viroid infected Chrysanthemum plants, 2 small RNAs different from the 9 S cell RNA usually described.
By contrast with this 9 S RNA which can be easily differentiated by its migration from the viroid RNA in 5 % polyacrylamide gels, the other cell RNAs exhibit under these conditions a similar behaviour.
Because of their very low concentration they are never observed when small samples are investigated ; in mass indexing, when nucleic acids from large samples are investigated, these small additionnal cell RNAs can be detected and mistaken for viroid RNA.
Then PASGE can be routinely applied to mass indexing and is very useful for investigating natural infections or reinfections ; it appears very interesting in addition to the normal PASGE previously described, which is appropriate for individual testing especially for the production of mother plants.