the nucleoplasm, must be equally active, but no obvious Norwich NR4 7UH, UK ultrastructure has yet been associated with them, except by fluorescence in situ hybridization (Highett et al., 1993a). Active rDNA transcription complexes have been visualized Summary by a hypotonic spreading technique first devised by Miller Incorporation by RNA polymerases of BrUTP into both and Beatty (1969) for the amplified rDNA found in Xenopus plant root tissue and isolated plant nuclei as a method for oocytes, and since applied by others to many different cell localization of the sites of transcription has been used. In types and organisms. These spread preparations show the this paper pea root tissue was used, and under the condiaxis of the gene associated with many (50-100) attached tions employed, nearly all the incorporation occurs in the RNA polymerase I molecules, together with nascent RNA nucleolus, and thus must be catalysed by RNA polymerase transcripts radiating away from the polymerases. Often the I. Immunofluorescence and confocal microscopy shows nascent RNAs show a terminal knob, assumed to be an that incorporation occurs in a pattern consisting of many RNP complex, and the increasing length of transcripts small foci distributed widely through the dense fibrillar along the genes has given rise to the description of the component of the nucleoli. Immunogold labelling using transcription complexes as 'Christmas trees'. Various comsilver-enhanced Nanogold probe at the electron microponents involved in the cleavage of the external transcribed scopic level confirms the sites of transcription as small spacer (ETS), such as fibrillarin, have been shown to be foci approximately 200nm in diameter. Simultaneous present in the terminal knobs (Mougey et al., 1993). fluorescence in situ hybridization with a probe to the Until recently, however, there has been little progress in external transcribed spacer (ETS) region of the pre-rRNA determining the organization of transcription and proshows that the structures revealed by this probe and the cessing in situ within the nucleolus. Conventional thin BrUTP immunofluorescence labelling are very similar. A section electron microscopy shows the nucleolus as a probe to the transcribed portion of the rDNA (18S) also densely stained structure, within which differently strucshows a good correlation to the sites of BrUTP incorporatured regions can often be discerned: fibrillar centres (FCs) tion within the nucleolus. On the other hand a probe to are small, lightly stained regions, typically approximately the non-transcribed intergenic spacer region (NTS) shows spherical and less than 1 µm in diameter; these are often very little coincidence with the sites of BrUTP incorporasurrounded by densely staining material-the dense fibriltion, and double fluorescence in situ labelling with both lar component (DFC); the rest of the volume of the nucleolus 18S and NTS probes confirms this difference in localization.contains densely packed particles, assumed to be preThese results suggest that most...