PROCEDURE FOR WHOLE-MOUNT FLUORESCENCE IN-SITU HYBRIDIZATION OF INTERPHASE NUCLEI ON ARABIDOPSIS-THALIANA

A procedure for whole mount fluorescence in situ hybridization (FISH) on plant tissue is reported. The technique was demonstrated on seedlin gs and flowers of Arabidopsis thaliana L. with rDNA as a probe, labell ed, both for direct and indirect detection. It was found that fixation in 1% formaldehyde yielded the best results with respect to morpholog y and hybridization efficiency. The combination of whole mount FISH an d confocal scanning laser microscopy allowed the nuclear localization of the rDNA loci in all tissues of both seedlings and flowers. Direct labelling yielded the best signal-to-noise ratio, especially in the ap ical zones of the seedlings. The technique was further illustrated on seedlings of A, thaliana in double labelling experiments with rDNA and a tandemly repeated, 500 bp sequence of A. thaliana. Although nuclei in all tissues in the seedling exhibited both signals, hybridization e fficiency for both signals was reduced in the dense, apical zones as c ompared with single labelling experiments with rDNA.