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Different sensitivity of DNA in situ in interphase and metaphase chromatin to heat denaturation
Heat denaturation of DNA in situ, in unbroken cells, was studied in relation to the cell cycle. DNA in metaphase cells denatured at lower temperatures (8 degrees-10 degrees C lower) than DNA in interphase cells. Among interphase cells, small differences between G1, S, and G2 cells were observed at t...
Formato: | Texto |
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Lenguaje: | English |
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The Rockefeller University Press
1977
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2109887/ https://www.ncbi.nlm.nih.gov/pubmed/16017 |
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collection | PubMed |
description | Heat denaturation of DNA in situ, in unbroken cells, was studied in relation to the cell cycle. DNA in metaphase cells denatured at lower temperatures (8 degrees-10 degrees C lower) than DNA in interphase cells. Among interphase cells, small differences between G1, S, and G2 cells were observed at temperatures above 90 degrees C. The difference between metaphase and interphase cells increased after short pretreatment with formaldehyde, decreased when cells were heated in the presence of 1 mM MgCl2, and was abolished by cell pretreatment with 0.5 N HCl. The results suggest that acid-soluble constituents of chromatin confer local stability to DNA and that the degree of stabilization is lower in metaphase chromosomes than in interphase nuclei. These in situ results remain in contrast to the published data showing no difference in DNA denaturation in chromatin isolated from interphase and metaphase cells. It is likely that factors exist which influence the stability of DNA in situ are associated with the super-structural organization of chromatin in intact nuclei and which are lost during chromatin isolation and solubilization. Since DNA denaturation is assayed after cell cooling, there is also a possibility that the extent of denatured DNA may be influenced by some factors that control strand separation and DNA reassociation. The different stainability of interphase vs. metaphase cells, based on the difference in stability of DNA, offers a method for determining mitotic indices by flow cytofluorometry, and a possible new parameter for sorting cells in metaphase. |
format | Text |
id | pubmed-2109887 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 1977 |
publisher | The Rockefeller University Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-21098872008-05-01 Different sensitivity of DNA in situ in interphase and metaphase chromatin to heat denaturation J Cell Biol Articles Heat denaturation of DNA in situ, in unbroken cells, was studied in relation to the cell cycle. DNA in metaphase cells denatured at lower temperatures (8 degrees-10 degrees C lower) than DNA in interphase cells. Among interphase cells, small differences between G1, S, and G2 cells were observed at temperatures above 90 degrees C. The difference between metaphase and interphase cells increased after short pretreatment with formaldehyde, decreased when cells were heated in the presence of 1 mM MgCl2, and was abolished by cell pretreatment with 0.5 N HCl. The results suggest that acid-soluble constituents of chromatin confer local stability to DNA and that the degree of stabilization is lower in metaphase chromosomes than in interphase nuclei. These in situ results remain in contrast to the published data showing no difference in DNA denaturation in chromatin isolated from interphase and metaphase cells. It is likely that factors exist which influence the stability of DNA in situ are associated with the super-structural organization of chromatin in intact nuclei and which are lost during chromatin isolation and solubilization. Since DNA denaturation is assayed after cell cooling, there is also a possibility that the extent of denatured DNA may be influenced by some factors that control strand separation and DNA reassociation. The different stainability of interphase vs. metaphase cells, based on the difference in stability of DNA, offers a method for determining mitotic indices by flow cytofluorometry, and a possible new parameter for sorting cells in metaphase. The Rockefeller University Press 1977-04-01 /pmc/articles/PMC2109887/ /pubmed/16017 Text en This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/4.0/). |
spellingShingle | Articles Different sensitivity of DNA in situ in interphase and metaphase chromatin to heat denaturation |
title | Different sensitivity of DNA in situ in interphase and metaphase chromatin to heat denaturation |
title_full | Different sensitivity of DNA in situ in interphase and metaphase chromatin to heat denaturation |
title_fullStr | Different sensitivity of DNA in situ in interphase and metaphase chromatin to heat denaturation |
title_full_unstemmed | Different sensitivity of DNA in situ in interphase and metaphase chromatin to heat denaturation |
title_short | Different sensitivity of DNA in situ in interphase and metaphase chromatin to heat denaturation |
title_sort | different sensitivity of dna in situ in interphase and metaphase chromatin to heat denaturation |
topic | Articles |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2109887/ https://www.ncbi.nlm.nih.gov/pubmed/16017 |