Fluram induces species-dependent C and G bands in mammalian chromosomes, revealing heterogeneous distribution of chromosomal proteins

Genome ◽  
1991 ◽  
Vol 34 (5) ◽  
pp. 772-776 ◽  
Author(s):  
T. Cuéllar ◽  
J. Gosálvez ◽  
P. Del Castillo ◽  
J. C. Stockert
Keyword(s):  
Banding Pattern ◽  
Chromosome Banding ◽  
Chromosome Region ◽  
Centromeric Heterochromatin ◽  
Primary Amines ◽  
Metaphase Chromosomes ◽  
Heterogeneous Distribution ◽  
Chromosomal Proteins ◽  
Differential Reactivity ◽  
Chromosome Regions

Fluram (Fluorescamine; 4-phenylspiro(furan-2(3H),1′-phthalan)-3,3′-dione) is a fluorogenic reagent, which permits the detection of primary amines by forming highly fluorescent pyrrolinone derivatives. This reagent has been used on methanol – acetic acid fixed metaphase chromosomes of mouse and man and proved to be very effective in differentiating chromosome regions in both genomes. Mouse centromeric heterochromatin is highly reactive, showing intense fluorescence in all centromeric regions, whereas human chromosomes show no fluorescence in such regions. In addition, a G-like banding pattern is also obtained in both types of chromosomes. The differential reactivity of each chromosome region showed by this method demonstrates a heterogeneous distribution of chromosome proteins, resulting in a chromosome banding pattern, which is in this case species dependent.Key words: cytogenetics, chromosome banding, mammalian chromosomes, fluorescence microscopy, Fluram.

scholarly journals Light and scanning electron microscopy of the same human metaphase chromosomes

1985 ◽  
Vol 77 (1) ◽  
pp. 143-153
Author(s):  
C.J. Harrison ◽  
E.M. Jack ◽  
T.D. Allen ◽  
R. Harris
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Electron Microscope ◽  
Scanning Electron Microscope ◽  
Banding Pattern ◽  
Light Microscope ◽  
Chromosome Banding ◽  
Metaphase Chromosomes ◽  
Scanning Electron

A technique has been developed to examine the same G-banded human metaphase chromosomes, first in the light microscope and then in the scanning electron microscope (SEM). A structural involvement in chromosome banding was confirmed by a positional correlation between the G-positive bands observed in the light microscope and the circumferential grooves between the quaternary coils of the metaphase chromosomes, observed in the SEM. In further support of this the regions between the grooves showed a positional relationship with the G-negative or reverse (R) bands. The examination of slightly extended metaphase chromosomes in the light microscope demonstrated that the G-banding pattern corresponded to that described by the Paris nomenclature for metaphase chromosomes. The arrangement of the circumferential grooves of the same chromosomes, observed in the SEM, was shown to relate to that described by the Paris nomenclature for prometaphase chromosomes. Therefore, using the SEM it is possible to demonstrate the details of prometaphase banding in metaphase chromosomes.

INSECT CHROMOSOME BANDING: TECHNIQUE FOR G- AND Q-BANDING PATTERN IN THE MOSQUITO AEDES ALBOPICTUS

1975 ◽  
Vol 17 (2) ◽  
pp. 241-244 ◽  
Author(s):  
Gerald E. Steiniger ◽  
Asit B. Mukherjee
Keyword(s):  
Aedes Albopictus ◽  
Banding Pattern ◽  
Chromosome Banding ◽  
Banding Technique ◽  
Modified Technique ◽  
Metaphase Chromosomes ◽  
Somatic Metaphase

A modified technique is described for the production of clear G- and Q-bands of somatic metaphase chromosomes of the mosquito, Aedes albopictus Skuse.

The effect of the chromosome banding techniques on the histone and nonhistone proteins of isolated chromatin

1982 ◽  
Vol 60 (3) ◽  
pp. 328-337 ◽  
Author(s):  
Gary D. Burkholder ◽  
Laurel L. Duczek
Keyword(s):  
Chromosome Banding ◽  
Polyacrylamide Gel Electrophoresis ◽  
Mitotic Chromosomes ◽  
Specific Chromosome ◽  
Nonhistone Proteins ◽  
Chromosomal Proteins ◽  
Different Types ◽  
Protein Alterations ◽  
Chromosome Regions ◽  
Qualitative Changes

The effect of a variety of chromosome banding techniques on the histone and nonhistone proteins of isolated, fixed, air-dried chromatin has been studied. Chromatin preparations were exposed to G-banding (SSC, urea, NaCl–urea, or trypsin), R-banding (Earle's BSS), and C-banding (NaOH or Ba(OH)2) treatments, and the proteins extracted from, and those remaining in the posttreatment chromatin, were examined by SDS polyacrylamide gel electrophoresis. The results indicate that: (i) each banding treatment produces specific quantitative and qualitative changes in the proteins of chromatin; (ii) the diverse treatments producing the same type of chromosome banding have both common and unique effects on the chromosomal proteins; and (iii) the treatments producing different types of chromosome banding have substantially different effects on the chromosomal proteins. It is not known whether these protein alterations are side effects occurring coincidentally with chromosome banding, or whether they are directly involved in mechanisms of banding. Results (ii) and (iii) suggest the latter possibility is more likely. If the protein alterations occur in specific chromosome regions, they may well influence the appearance of bands on mitotic chromosomes.

scholarly journals Chromosome banding pattern in a polymorphic population of Sorex araneus from northeastern Finland

Hereditas ◽  
2009 ◽  
Vol 76 (2) ◽  
pp. 305-314 ◽  
Author(s):  
LIISA HALKKA ◽  
O. HALKKA ◽  
U. SKARÉN ◽  
VERONICA SÖDERLUND
Keyword(s):  
Banding Pattern ◽  
Chromosome Banding ◽  
Sorex Araneus

scholarly journals Fine Structure and Staining Behaviour of Heterochromatic Segments in Two Plants

1974 ◽  
Vol 14 (3) ◽  
pp. 505-521
Author(s):  
L. F. LACOUR ◽  
B. WELLS
Keyword(s):  
Fine Structure ◽  
Light Microscope ◽  
Mitotic Cycle ◽  
The Other ◽  
Masking Effect ◽  
Root Tips ◽  
Metaphase Chromosomes ◽  
Differential Reactivity ◽  
Scilla Sibirica ◽  
Electron Microscopes

With the use of the light and electron microscopes, the chromosomes of Fritillaria lanceolata and Scilla sibirica are shown to differ in respect of the heterochromatin they contain. In root meristems of the former, the heterochromatic regions (H-segments) were recognizable at all phases of the mitotic cycle by their slighter opacity to electrons than that of euchromatic parts. This was due both to less tight packing of the chromatin fibrils and lower opacity of the fibrils themselves, even though both had the same diameter, about 3 nm. In root tips of the Scilla, the heterochromatin was invariably of similar opacity to euchromatin and thus only recognizable at telophase and interphase as large chromocentres. The opacity to electrons in the heterochromatin of the 2 species, was at all times closely paralleled by the staining behaviour seen with the light microscope in sections (0.07-0.5 µm in thickness) stained with toluidine blue. The disparity in the Fritillaria, as seen in sections with the light microscope, in respect of the stainability of the hetero- and euchromatin, was masked in Feulgen squash preparations of root tips from plants grown at 18-20 °C; at metaphase by the thickness of the chromosomes and at interphase by the density of the chromocentres. When, on the other hand, the plants were grown for 4 days at 2 °C, the masking effect of thickness was circumvented in metaphase chromosomes by differential super-contraction in euchromatin. The implications of these findings in respect to previous interpretations of the differential reactivity of H-segments to low temperature, as well as the phenomenon of enhanced and reduced fluorescence in these segments with fluorochromes are discussed.

scholarly journals Electron microscopy of gold-labeled human and equine chromosomes.

1989 ◽  
Vol 37 (9) ◽  
pp. 1443-1447 ◽  
Author(s):  
P E Messier ◽  
R Drouin ◽  
C L Richer
Keyword(s):  
Electron Microscopy ◽  
Chromosome Banding ◽  
Protein A ◽  
Gold Complex ◽  
Chromosome Identification ◽  
Chromosomal Dna ◽  
Metaphase Chromosomes ◽  
Sister Chromatids ◽  
Antigen Localization ◽  
Chromatin Reorganization

We present an immunochemical technique for the detection of 5-bromo-2'-deoxyuridine (BrdU) incorporated discontinuously into the chromosomal DNA. A monoclonal anti-BrdU antibody and a protein A-gold complex were used to produce chromosome banding of human and equine chromosomes, specific for electron microscopy (EM). Well-defined bands, symmetry of sister chromatids, concordance between homologues, and band patterns similar to those observed by light microscopy facilitate chromosome identification and karyotyping. From prophase to late metaphase, chromosomes condense and bands appear to fuse. The fusion appears to be owing to chromatin reorganization. Our results underline the value of using immunogold reagents, which are ideal probes for antigen localization on chromosomes.

scholarly journals A New Fluorescence Reaction in DNA Cytochemistry: Microscopic and Spectroscopic Studies on the Aromatic Diamidino Compound M&B 938

1997 ◽  
Vol 45 (1) ◽  
pp. 97-105 ◽  
Author(s):  
Juan C. Stockert ◽  
Clara I. Trigoso ◽  
Teresa Cuéllar ◽  
José L. Bella ◽  
José A. Lisanti
Keyword(s):  
Specific Binding ◽  
Linear Regression Analysis ◽  
Spectroscopic Studies ◽  
Centromeric Heterochromatin ◽  
Dna Ladder ◽  
Metaphase Chromosomes ◽  
Distamycin A ◽  
Ehrlich Ascites ◽  
Fluorescence Reaction ◽  
High Fluorescence

We describe the fluorescence properties and cytochemical applications of the aromatic diamidine M&B 938. Treatment of cell smears (chicken blood, Ehrlich ascites tumor, rat bone marrow, mouse mast cells, and Trypanosoma cruzi epimastigotes) with aqueous solutions of M&B 938 (0.5–1 μg/ml at pH 6–7; UV excitation) induced bright bluish-white fluorescence in DNA-containing structures (interphase and mitotic chromatin, AT-rich kinetoplast DNA of T. cruzi), which was abolished by previous DNA extraction. DNA was the unique fluorescent polyanion after staining with M&B 938 at neutral or alkaline pH, other polyanions such as RNA and heparin showing no emission. M&B 938-stained mouse metaphase chromosomes revealed high fluorescence of the AT-rich centromeric heterochromatin, and strong emission of heterochromatin in human chromosomes 1, 9, 15, 16, and Y was found after distamycin A counterstaining. On agarose gel electrophoresis, M&B 938-stained DNA markers appeared as fluorescent bands. The 1.635-kBP fragment from DNA ladder revealed a higher emission value than that expected from linear regression analysis. Spectroscopic studies showed bathochromic and hyperchromic shifts in the absorption spectrum of M&B 938 complexed with DNA, as well as strong enhancement of fluorescence at 420 nm. In the presence of poly(dA)-poly(dT), the emission of M&B 938 was 4.25-fold higher than with DNA; no fluorescence was observed with poly(dG)-poly(dC). Experimental results and considerations of the chemical structure suggest that the minor groove of AT regions of DNA could be the specific binding site for M&B 938, which shows interesting properties and useful applications as a new DNA fluorochrome.

scholarly journals Mouse satellite DNA, centromere structure, and sister chromatid pairing.

1986 ◽  
Vol 103 (4) ◽  
pp. 1145-1151 ◽  
Author(s):  
L M Lica ◽  
S Narayanswami ◽  
B A Hamkalo
Keyword(s):  
Electron Microscopy ◽  
Sister Chromatid ◽  
Satellite Dna ◽  
Marker Chromosome ◽  
Restriction Enzymes ◽  
Centromeric Heterochromatin ◽  
Mitotic Chromosomes ◽  
Metaphase Chromosomes ◽  
Sister Chromatids ◽  
Mouse Satellite Dna

The experiments described were directed toward understanding relationships between mouse satellite DNA, sister chromatid pairing, and centromere function. Electron microscopy of a large mouse L929 marker chromosome shows that each of its multiple constrictions is coincident with a site of sister chromatid contact and the presence of mouse satellite DNA. However, only one of these sites, the central one, possesses kinetochores. This observation suggests either that satellite DNA alone is not sufficient for kinetochore formation or that when one kinetochore forms, other potential sites are suppressed. In the second set of experiments, we show that highly extended chromosomes from Hoechst 33258-treated cells (Hilwig, I., and A. Gropp, 1973, Exp. Cell Res., 81:474-477) lack kinetochores. Kinetochores are not seen in Miller spreads of these chromosomes, and at least one kinetochore antigen is not associated with these chromosomes when they were subjected to immunofluorescent analysis using anti-kinetochore scleroderma serum. These data suggest that kinetochore formation at centromeric heterochromatin may require a higher order chromatin structure which is altered by Hoechst binding. Finally, when metaphase chromosomes are subjected to digestion by restriction enzymes that degrade the bulk of mouse satellite DNA, contact between sister chromatids appears to be disrupted. Electron microscopy of digested chromosomes shows that there is a significant loss of heterochromatin between the sister chromatids at paired sites. In addition, fluorescence microscopy using anti-kinetochore serum reveals a greater inter-kinetochore distance than in controls or chromosomes digested with enzymes that spare satellite. We conclude that the presence of mouse satellite DNA in these regions is necessary for maintenance of contact between the sister chromatids of mouse mitotic chromosomes.

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