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This paper deals with the developing of a new simple and repetitive technique to preferentially stain DNA-containing structures at the E.M. level that can be used “en bloc” and on sections. DNA staining techniques are scarce, being the osmium ammine one the only cytochemical method known until now that specifically stains DNA but not the proteins that are complexed with it, having been used to study chromatin ultrastructure. The osmium ammine synthesis is a complex process and, although an improvement has been lately reported, the stain is not comercially available.Glutaraldehyde-fixedAllium cepa L. root meristematic cells,Capsicum annuum L. & Scilla peruviana L. pollen grains were treated with 0.5N NaOH in 4% formaldehyde overnight, rinsed in water and 1% acetic acid, methanol dehydrated, and inmersed in methanol-acetic anhydride (5:1) at 25ºC overnight. Then, the samples were Epon embedded. To perform the technique on sections, some samples were glutaraldehyde or formaldehyde fixed, methanol dehydrated, Lowicryl embedded at -20ºC and ultrathin sections were treated in the same way as in the “en bloc” procedure.

The distribution of non-histone nuclear antigens was studied during early somatic embryogenesis of carrot using nuclear antibodies specific for small nuclear RNPs (snRNPs), nuclear matrix proteins and nucleolar proteins. Immunofluorescence showed that in interphase cells snRNPs and nuclear matrix proteins were localized in the nucleus with a speckled pattern. Immunoelectron microscopy indicated that the snRNPs were irregularly distributed while the nuclear matrix proteins showed a distribution preferentially over the condensed chromatin. None were localized in the nucleolus. The nucleolar proteins were localized in both dense fibrillar and granular components while the fibrillar centres were devoid of gold particles. The localization of these antibodies in plant cell nuclei, in a way comparable to the patterns described in animal cells, indicates the general presence of snRNPs in all eukaryotic nuclei. It further confirms the relation of the nuclear matrix proteins to plant cell proliferation events and the possible role of the nucleolar proteins in both rRNA transcription and preribosome assembly.

The ultrastructure and morphometric features of nucleolar fibrillar centers have been studied in three different plant species over a long period of gametogenesis, extending from premeiosis to after the first postmeiotic mitosis, with respect to variations in nucleolar activity which occur throughout this process. Fibrillar centers represent the structural counterpart of the transcriptionally inactive portions of the nucleolar organizing region (NOR); in some gametogenic stages, no discrete fibrillar centers can be observed and, therefore, the structure of the whole NOR has been studied. Two clearly differentiated types of fibrillar center structure are described, with a close relationship to different stages of nucleolar activity. Stages with high activity show “homogeneous” fibrillar centers, whose chromatin is decondensed, and stages with low activity show “heterogeneous” fibrillar centers with both condensed and decondensed chromatin. These two types had previously been characterized cytochemically by us in somatic cells. Medium-active stages show fibrillar centers “intermediate” between the two main types. The morphometric behavior of fibrillar centers is also related to activity. The number of fibrillar centers per nucleolar section increases with the nucleolar activity, while their size decreases. However, the location of fibrillar centers in the nucleolus does not follow any special pattern, except in middle meiotic prophase I, in which the whole NOR occupies a peripheral position. This is due to a special NOR activity at these stages, whose products are stored as “nucleolar dense bodies.” All these data allow us to conclude that fibrillar centers are highly dynamic structures, extremely sensitive to changes in nucleolar activity., This work has been supported by the CAICYT Grant 3753/79 C2 and by the CSIC Grant 41123

Two suggested techniques1, thermal analysis and infra-red spectrophotometrys are used to characterise the deterioration process caused by ultra-violet radiation on polymeric materials. The suggested mechanism of photo-oxidation is presented. A correlation between the depletion of tracking resistance and the oxidation of the polimeric surface was obtained, which was proportional to aging time. Good agreement was obtained, also, between the infra-red spetra with the thermal analysis of new and aged samples; thus, T.A. and I.R. data are presented as two useful tools for characterising polymeric materials which are aged by U.V. radiation.

Using a high resolution nucleolar silver staining technique, we have studied the distribution of silver grains at interphase and mitosis. The nucleolar fibrillar centers and the fibrillar component, as well as the mitotic nucleolar organizing region (NOR), are argyrophilic. Of the NOR structures, the fibrillar component is transcriptionally active, but the fibrillar centers and the mitotic NORs are not. Treatment with cycloheximide (CHM), which indirectly impairs rRNA synthesis, does not affect the pattern of silver staining. The currently assumed correlation between activity and Ag-NOR staining appears questionable. On the other hand, all Ag-NOR-stained structures contain decondensed chromatin, whereas condensed portions of this chromatin are not stained by silver. In CHM-treated cells there is apparently no chromatin condensation accompanying inhibition of rRNA synthesis., This work has been supported by CSIC grant 41024.

The nucleolus is a highly dinamic organelle. Variations in the synthesis and/or processing of rRNA produces redistribution of the structural nucleolar components, in both physiological and experimental conditions (Goessens 1984, Hadjiolov 1985, Olmedilla et a1. 1987, Risueno and Medina 1986).

Cherry leaf roll virus (CLRV) infection in Nicotiana tabacum cv. Xanthi nc plants induces the expression of, among others, basic and acidic chitinases at 3 d.p.i. (days postinoculation) and 8 d.p.i. in inoculated and uninoculated, systemically infected leaves, respectively. The spatial distribution of the acidictobacco chitinases (PR-P and PR-Q) mRNAs in CLRV-infected tobacco leaves has been studied by tissue printing and Northern (RNA) blot analysis. The mRNA was preferentially detected in and around the concentric necrotic rings caused by the viral infection. In addition, the accumulation pattern of the proteins was very similar although a basal signal was detected in nonsymptomatic tissue in uninoculated, systemically infected leaves. These results indicate a positive association of acidic chitinase gene expression with the necrotic tissue, where the virus is mainly localized, in the CLRV-tobacco interaction.

The effects of Si doping on the growth mode and residual strain of GaN layers grown on Si(111) substrates by plasma-assisted molecular beam epitaxy are studied by Raman scattering and photoluminescence. As the Si concentration increases a progressive decrease of the high-energy E2 mode frequency is observed, together with a redshift of the excitonic emission. Both effects indicate an enhancement of the biaxial tensile strain of thermal origin for increasing doping level, which is confirmed by x-ray diffraction measurements. Beyond Si concentrations of 5×1018 cm−3 both the phonon frequency and the exciton emission energy increase again. This change indicates a partial strain relaxation due to a change in the growth mode.