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Sequence-specific microscopic visualization of DNA methylation status at satellite repeats in individual cell nuclei and chromosomes

Methylation-specific fluorescence in situ hybridization (MeFISH) was developed for microscopic visualization of DNA methylation status at specific repeat sequences in individual cells. MeFISH is based on the differential reactivity of 5-methylcytosine and cytosine in target DNA for interstrand compl...

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Autores principales: Li, Yufeng, Miyanari, Yusuke, Shirane, Kenjiro, Nitta, Hirohisa, Kubota, Takeo, Ohashi, Hirofumi, Okamoto, Akimitsu, Sasaki, Hiroyuki
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2013
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3799461/
https://www.ncbi.nlm.nih.gov/pubmed/23990328
http://dx.doi.org/10.1093/nar/gkt766
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author Li, Yufeng
Miyanari, Yusuke
Shirane, Kenjiro
Nitta, Hirohisa
Kubota, Takeo
Ohashi, Hirofumi
Okamoto, Akimitsu
Sasaki, Hiroyuki
author_facet Li, Yufeng
Miyanari, Yusuke
Shirane, Kenjiro
Nitta, Hirohisa
Kubota, Takeo
Ohashi, Hirofumi
Okamoto, Akimitsu
Sasaki, Hiroyuki
author_sort Li, Yufeng
collection PubMed
description Methylation-specific fluorescence in situ hybridization (MeFISH) was developed for microscopic visualization of DNA methylation status at specific repeat sequences in individual cells. MeFISH is based on the differential reactivity of 5-methylcytosine and cytosine in target DNA for interstrand complex formation with osmium and bipyridine-containing nucleic acids (ICON). Cell nuclei and chromosomes hybridized with fluorescence-labeled ICON probes for mouse major and minor satellite repeats were treated with osmium for crosslinking. After denaturation, fluorescent signals were retained specifically at satellite repeats in wild-type, but not in DNA methyltransferase triple-knockout (negative control) mouse embryonic stem cells. Moreover, using MeFISH, we successfully detected hypomethylated satellite repeats in cells from patients with immunodeficiency, centromeric instability and facial anomalies syndrome and 5-hydroxymethylated satellite repeats in male germ cells, the latter of which had been considered to be unmethylated based on anti-5-methylcytosine antibody staining. MeFISH will be suitable for a wide range of applications in epigenetics research and medical diagnosis.
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spelling pubmed-37994612013-10-21 Sequence-specific microscopic visualization of DNA methylation status at satellite repeats in individual cell nuclei and chromosomes Li, Yufeng Miyanari, Yusuke Shirane, Kenjiro Nitta, Hirohisa Kubota, Takeo Ohashi, Hirofumi Okamoto, Akimitsu Sasaki, Hiroyuki Nucleic Acids Res Methods Online Methylation-specific fluorescence in situ hybridization (MeFISH) was developed for microscopic visualization of DNA methylation status at specific repeat sequences in individual cells. MeFISH is based on the differential reactivity of 5-methylcytosine and cytosine in target DNA for interstrand complex formation with osmium and bipyridine-containing nucleic acids (ICON). Cell nuclei and chromosomes hybridized with fluorescence-labeled ICON probes for mouse major and minor satellite repeats were treated with osmium for crosslinking. After denaturation, fluorescent signals were retained specifically at satellite repeats in wild-type, but not in DNA methyltransferase triple-knockout (negative control) mouse embryonic stem cells. Moreover, using MeFISH, we successfully detected hypomethylated satellite repeats in cells from patients with immunodeficiency, centromeric instability and facial anomalies syndrome and 5-hydroxymethylated satellite repeats in male germ cells, the latter of which had been considered to be unmethylated based on anti-5-methylcytosine antibody staining. MeFISH will be suitable for a wide range of applications in epigenetics research and medical diagnosis. Oxford University Press 2013-10 2013-08-28 /pmc/articles/PMC3799461/ /pubmed/23990328 http://dx.doi.org/10.1093/nar/gkt766 Text en © The Author(s) 2013. Published by Oxford University Press. http://creativecommons.org/licenses/by/3.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Methods Online
Li, Yufeng
Miyanari, Yusuke
Shirane, Kenjiro
Nitta, Hirohisa
Kubota, Takeo
Ohashi, Hirofumi
Okamoto, Akimitsu
Sasaki, Hiroyuki
Sequence-specific microscopic visualization of DNA methylation status at satellite repeats in individual cell nuclei and chromosomes
title Sequence-specific microscopic visualization of DNA methylation status at satellite repeats in individual cell nuclei and chromosomes
title_full Sequence-specific microscopic visualization of DNA methylation status at satellite repeats in individual cell nuclei and chromosomes
title_fullStr Sequence-specific microscopic visualization of DNA methylation status at satellite repeats in individual cell nuclei and chromosomes
title_full_unstemmed Sequence-specific microscopic visualization of DNA methylation status at satellite repeats in individual cell nuclei and chromosomes
title_short Sequence-specific microscopic visualization of DNA methylation status at satellite repeats in individual cell nuclei and chromosomes
title_sort sequence-specific microscopic visualization of dna methylation status at satellite repeats in individual cell nuclei and chromosomes
topic Methods Online
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3799461/
https://www.ncbi.nlm.nih.gov/pubmed/23990328
http://dx.doi.org/10.1093/nar/gkt766
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