The distribution of genes on human chromosomes as studied by in situ nick translation

Genome ◽  
1992 ◽  
Vol 35 (5) ◽  
pp. 890-894 ◽  
Author(s):  
J. de la Torre ◽  
A. T. Sumner ◽  
J. Gosalvez ◽  
L. Stuppia
Keyword(s):  
Cpg Islands ◽  
Dnase I ◽  
Human Chromosomes ◽  
Nick Translation ◽  
X Chromosomes ◽  
Dnase I Hypersensitivity ◽  
In Situ Nick Translation ◽  
Inactive X Chromosome ◽  
Active Genes

We have studied the distribution of potentially active genes on human chromosomes, using two methods: DNAse I hypersensitivity and restriction enzyme – nick translation with enzymes sensitive to methylation of CpG doublets. DNAse hypersensitivity is known to be associated with potentially active genes, and, when the reaction is detected by "in situ" nick translation, produces an R-banding pattern. Digestion of chromosomes with HpaII or CfoI, both of which should preferentially cut unmethylated sequences in the CpG islands associated with the majority of genes, also produces R-banding patterns. Deviations are attributable to overdigestion of the chromosomes, leading to extraction of DNA and loss of the specific sites that were to be detected. Contrary to the results of a number of previous workers, we have failed to demonstrate any differences between the DNAse I hypersensitivity or the degree of methylation of the active and inactive X chromosomes in metaphases from females.Key words: human chromosomes, gene distribution, DNAse I hypersensitivity, in situ nick translation, R-banding, CpG islands, DNA methylation, inactive X chromosome.

Patterns of DNase I sensitivity in the holocentric chromosomes of the aphid Megoura viciae

Genome ◽  
1998 ◽  
Vol 41 (2) ◽  
pp. 169-172 ◽  
Author(s):  
Gian Carlo Manicardi ◽  
Mauro Mandrioli ◽  
Davide Bizzaro ◽  
Umberto Bianchi
Keyword(s):  
Dnase I ◽  
Holocentric Chromosomes ◽  
Telomeric Region ◽  
Nick Translation ◽  
In Situ Nick Translation ◽  
Embryo Cells ◽  
Dnase I Sensitivity ◽  
Megoura Viciae ◽  
Active Genes

Using the in situ nick translation technique, we looked for the presence of DNase I sensitive sites in Megoura viciae chromosomes, to study the distribution of active or potentially active genes in aphids, a group of insects possessing holocentric chromosomes. Cytological preparations obtained by the spreading of embryo cells were treated in situ with increasing concentrations (ranging from 5 to 200 ng/mL) of DNase I. At DNase I concentrations below 50 ng/mL, only one hypersensitive site was observed, and this was located on a telomeric region of the X chromosome that contains transcriptionally active nucleolar organizing regions, as assayed by silver staining. Interestingly, at intermediate concentrations of DNase, the incorporation of biotinylated nucleotide occurred uniformly throughout all chromosomes, whereas at concentrations above 100 ng/mL, a C-like banding pattern was produced. Our data differ from results obtained with mammalian, frog, and grasshopper chromosomes, where it was found that DNase I nicking is concentrated at the distal regions of all chromosomes.Key words: aphids, holocentric chromosomes, DNase I sensitivity, nick translation.

A new banding pattern of human chromosomes by in situ nick translation using ECO RI and biotin-dUTP

2008 ◽  
Vol 28 (2) ◽  
pp. 173-176 ◽  
Author(s):  
Jörn Bullerdiek ◽  
Jürgen Dittmer ◽  
Angelika Faehre ◽  
Sabine Bartnitzke ◽  
Volker Kasche ◽  
...  
Keyword(s):  
Banding Pattern ◽  
Human Chromosomes ◽  
Nick Translation ◽  
In Situ Nick Translation ◽  
Eco Ri

In situ nick-translation distinguishes between active and inactive X chromosomes

Nature ◽  
1983 ◽  
Vol 304 (5921) ◽  
pp. 88-90 ◽  
Author(s):  
Bat-Sheva Kerem ◽  
Ruth Goitein ◽  
Carmelit Richler ◽  
Menashe Marcus ◽  
Howard Cedar
Keyword(s):  
Nick Translation ◽  
X Chromosomes ◽  
In Situ Nick Translation

Distribution of TaqI sites along human chromosomes revealed by in situ enzyme-nick translation

Genome ◽  
1993 ◽  
Vol 36 (1) ◽  
pp. 202-205 ◽  
Author(s):  
I. Tagarro ◽  
J. J. Gonzalez-Aguilera ◽  
A. M. Fernandez-Peralta ◽  
G. F. de Stefano ◽  
L. Ferrucci
Keyword(s):  
Complete Information ◽  
Human Chromosomes ◽  
Nick Translation ◽  
In Situ Nick Translation ◽  
Translation Procedure

We have studied the relative richness of TaqI sites along human chromosomes by means of a nonradioactive in situ enzyme-nick translation procedure. Regions with a higher content of these sequences are shown to be the noncentromeric heterochromatin blocks, whereas within euchromatin, terminal R-bands are the domains more enriched in these sites. Results obtained suggest that the method of performing enzyme digestions using time as a variable, and then in situ nick translation, provides much more complete information about the distribution of enzyme sequences along chromosomes than standard enzyme digestions.Key words: TaqI, nick translation, biotin, T-bands, heterochromatin.

scholarly journals In situ nick translation at the electron microscopic level: a tool for studying the location of DNAse I-sensitive regions within the cell.

1991 ◽  
Vol 39 (6) ◽  
pp. 871-874 ◽  
Author(s):  
M Thiry
Keyword(s):  
Ultrastructural Level ◽  
Dnase I ◽  
Electron Microscopic Level ◽  
Nick Translation ◽  
Dna Polymerase I ◽  
Electron Microscopic ◽  
Thin Sections ◽  
In Situ Nick Translation ◽  
Polymerase I

The in situ nick translation method was adapted to the ultrastructural level, to study the location of DNAse I-sensitive sequences within the cell. Ultra-thin sections of Lowicryl-embedded cells were incubated in a medium containing DNAse I, DNA polymerase I, and all four deoxyribonucleotides, some being biotinylated. The nick-translated sites were then visualized by an indirect immunogold labeling technique. The resulting labeling pattern is closely dependent on the DNAse I concentration in the nick-translation medium. The method reveals with great precision the specific DNAse I-sensitive regions within the nucleus. This technique can be used to discriminate between active and inactive regions of interphase chromatin.

Analysis of inactive X chromosome structure by in situ nick translation

Chromosoma ◽  
1985 ◽  
Vol 92 (3) ◽  
pp. 209-213 ◽  
Author(s):  
Karen A. Dyer ◽  
Donald Riley ◽  
Stanley M. Gartler
Keyword(s):  
X Chromosome ◽  
Chromosome Structure ◽  
Nick Translation ◽  
In Situ Nick Translation ◽  
Inactive X Chromosome

Ultrastructural banding induced by DraI or HaeIII progressive digestion and in situ nick translation on human chromosomes

Genome ◽  
1994 ◽  
Vol 37 (6) ◽  
pp. 950-956
Author(s):  
Rosalba Del Coco ◽  
Liborio Stuppia ◽  
Caterina Cinti ◽  
Rita Peila ◽  
Nadir Mario Maraldi
Keyword(s):  
Electron Microscopy ◽  
Conformational Changes ◽  
Restriction Enzymes ◽  
Human Chromosomes ◽  
Nick Translation ◽  
Metaphase Chromosomes ◽  
In Situ Nick Translation ◽  
Transmission Electron ◽  
Dna Loss

Fixed human metaphase chromosomes were progressively digested with DraI or HaeIII restriction enzymes, submitted to in situ nick translation, and observed by transmission electron microscopy to obtain further information on the localization of the endonuclease target sequences and on the conformational changes in chromosomal bands. This approach allows us to detect specific nick translation patterns, namely, G-banding or R-like banding after short DraI and HaeIII endonuclease digestion, respectively. Intermediate banding recognizable as C-negative banding and G + C banding are induced by longer HaeIII digestion, before the C-positive banding. These patterns appear to depend both on different target sites of the employed endonucleases and on the DNA loss at different digestion times.Key words: human chromosomes, in situ nick translation, DraI banding, HaeIII banding, electron microscopy.

scholarly journals Detection of DNA fragmentation in apoptosis: application of in situ nick translation to cell culture systems and tissue sections.

1993 ◽  
Vol 41 (7) ◽  
pp. 1023-1030 ◽  
Author(s):  
R Gold ◽  
M Schmied ◽  
G Rothe ◽  
H Zischler ◽  
H Breitschopf ◽  
...  
Keyword(s):  
Cell Death ◽  
Dna Fragmentation ◽  
Specific Cell ◽  
Nick Translation ◽  
Proliferating Cells ◽  
Cell Surface Antigens ◽  
Simultaneous Application ◽  
Tissue Sections ◽  
In Situ Nick Translation

Since DNA fragmentation is a key feature of programmed cell death (PCD) and also occurs in certain stages of necrosis, we have adapted the methodology of in situ nick-translation (ISNT) to detect DNA fragmentation on a single-cell level. We first established the technique for cell preparations. Apoptosis was induced by gamma-irradiation on freshly isolated rat thymocytes. After fixation procedures, ISNT was performed by overnight incubation either with fluorescein-12-dUTP or with digoxigenin-labeled 11-dUTP and DNA polymerase I. The enzymatic incorporation of labeled nucleotides at sites of DNA fragmentation was detected by flow cytometry either directly or indirectly with fluorescein-conjugated anti-digoxigenin. The quantitative results demonstrated close correlation with morphological essays for apoptosis, DNA gel electrophoresis, and ISNT. Proliferating cells determined by bromodeoxyuridine immunofluorescence were not labeled by ISNT. Immunocytochemistry for cell surface antigens in combination with ISNT allowed the identification of specific cell types undergoing PCD. Furthermore, the simultaneous application of photolabeling techniques with ethidium monoazide and ISNT led to the identification of DNA fragmentation in cells with still intact membranes. Extending ISNT to tissue sections of paraformaldehyde-fixed, paraffin-embedded material reliably revealed labeling of cells with typical morphological features of apoptosis. However, this technique was not useful in detecting early stages of necrotic cell death.

Sign in / Sign up

Export Citation Format

Share Document