L. Ferrucci

Chromatin organization and restriction endonuclease activity on human metaphase chromosomes

Human metaphase chromosomes were treated with HaeIII, HindIII, EcoRI, and AluI restriction endonucleases and subsequently stained with either Giemsa or ethidium bromide. The results obtained seemed to suggest that the structural organization of specific chromosome regions can play a primary role in determining the cytological effect after digestion with specific restriction endonucleases.

Alterations induced in mouse chromosomes by restriction endonucleases

Fixed chromosomes of mouse have been treated with Alu I, Eco RII, Hind III or Bam HI restriction endonucleases and subsequently stained with either Giemsa, Ethidium Bromide or Acridine Orange. The results obtained have been discussed in the light of preferential or non-preferential extraction of DNA from specific chromosome areas following enzyme digestion. The possible involvement of a particular structural organization of some classes of heterochromatin has been hypothesized to account for the findings after Alu I or Eco RII treatment. The meaning of the Giemsa banding observed after Hind III or Bam HI digestion has also been considered, in comparison to the different stain responses obtained by using a DNA-specific dye such as Ethidium Bromide.

Ageing of fixed cytological preparations produces degradation of chromosomal DNA

When fixed chromosome preparations were allowed to age for 1-72 h, they became progressively more susceptible to digestion by exonuclease III and by S1 nuclease. Analysis of DNA from these aged preparations on agarose gels showed that the molecular weight of the DNA decreased as ageing progressed. We conclude that DNA in fixed chromosome preparations becomes progressively degraded as the preparations age.

The longitudinal differentiation produced by photo-oxidation and acridine-orange staining in eukaryote chromosomes: role and involvement of DNA base composition

Photo-oxidation and staining with acridine-orange result in a longitudinal green/orange-red/brown-red color differentiation in fixed chromosomes of Drosophila melanogaster, D. virilis, Culiseta longiareolata, and Mus musculus. Such a result has been correlated with specific DNA base sequences situated in specific chromosomal areas. In fact, given the known base composition of satellite DNAs in three of the species we studied, a correspondence seems to exist between molecular hybridization in situ of AT-rich satellite DNAs and the location of specific stain reactions in specific chromosome regions. Moreover, our results support previous findings suggesting preferential guanine destruction as a consequence of photo-oxidation.

Light-induced banding (LIB) in metaphase chromosomes of Culiseta longiareolata (Diptera: Culicidae)

A technique already used on Drosophila polytene chromosomes was employed on metaphase chromosomes of Culiseta Iongiareolata (Diptera: Culicidae). The coincidence between Q-bands and green fluorescence was pointed out. The centromere regions of all chromosome pairs and one telomere of the Y, although demonstrating uniformly stained C-bands, were shown to differentially stain with Coriphosphine-0 after light treatment. A base destruction mechanism, similar to that proposed for the methylene blue-light-chrtimosomal DNA interaction, has been hypothesized for explaining these findings.

The differential banding pattern produced by Actinomycin-D/Acridine-Orange counterstaining in metaphase chromosomes ofDrosophila melanogaster

A longitudinal differentiation is found in fixed metaphase chromosomes ofDrosophila melanogaster after treatment with Actinomycin-D and subsequent Acridine-Orange staining. We postulate that our findings are strictly related to the presence of AT-rich DNA in specific chromosomal areas of this species.

Methylene-blue/coriphosphine-O/acridine-orange, chromosomal DNA and visible light: Interaction and cytological effect in Drosophila melanogaster

Metaphase preparations of Drosophila melanogaster were irradiated with visible light in the presence of either Methylene-blue, Coriphosphine-O or Acridine-Orange. After treatment with any of these three compounds, Coriphosphine-O staining shows an identical green/orange-red/brown-red chromosome pattern. These findings are discussed in the light of previous reports and guanine destruction is hypothesized as representing the most plausible mechanism for explaining the cytological results.

The differential banding pattern produced by Hoechst 33258 and quinachrine dihydrochloride in metaphase chromosomes of Culiseta longiareolata (Diptera: Culicidae)

SUMMARYHoechst 33258 was used for staining metaphase chromosomes of Culiseta longiareolata (Diptera: Culicidae) and the banding produced was compared to that obtained with Quinacrine. The Y telomeric region, which shows C+ banding pattern, is H+ but Q−, while some patacentromeric areas reveal H+/Q+ staining. The results and possible factors which could affect H and Q banding are discussed.

Y chromosome in the sibling speciesAnopheles atroparvus (van Thiel, 1927) andA. labranchiae (Falleroni, 1926) (Diptera: Culicidae): Differential behaviour of the short arm after acid-alkaline treatment and Coriphosphine-O staining

An acid-alkaline treatment followed by Coriphosphine-O staining was used for detecting chromosomal differences between the 2 sibling speciesAnopheles atroparvus (van Thiel) andA. labranchiae (Falleroni) (Diptera: Culicidae). The short arm of the Y chromosome was found to stain differently in the 2 species.

Restriction enzymes induced bands in the cave cricket Dolichopoda schiavazzii (Orthoptera, Rhaphidophoridae): Implications for heterochromatin characterization and satellite DNA distribution

Abstract In this study the activity of some class II restriction endonucleases, RE, on fixed metaphase chromosomes has been tested on two sample populations of the cave cricket Dolichopoda schiavazzii, a species endemic in Tuscany. In particular, the 6‐base cutter PstI was chosen to detect the occurrence and localization on chromosomes of a satellite DNA family (named pDoP102) whose repetition units, analyzed in a previous study, contain the cleavage site of this enzyme. A comparison of the PstI digestion pattern and the C‐banding one, suggests that this satDNA family is mainly localized in certain heterochromatic regions of chromosomes 1, 3, 8, 10 and in the Y chromosome. As a negative control in situ digestion was performed with RE AluI whose cleavage site is not contained in the pDoP102 satDNA sequence. The partial overlap between the PstI and Alul digested regions suggests the occurrence of at least another satDNA family detected by PstI and containing the Alul target. Alul restriction banding also sh...