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Live‐cell CRISPR imaging in plants reveals dynamic telomere movements
Elucidating the spatiotemporal organization of the genome inside the nucleus is imperative to our understanding of the regulation of genes and non‐coding sequences during development and environmental changes. Emerging techniques of chromatin imaging promise to bridge the long‐standing gap between s...
Autores principales: | , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
John Wiley and Sons Inc.
2017
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5599988/ https://www.ncbi.nlm.nih.gov/pubmed/28509419 http://dx.doi.org/10.1111/tpj.13601 |
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author | Dreissig, Steven Schiml, Simon Schindele, Patrick Weiss, Oda Rutten, Twan Schubert, Veit Gladilin, Evgeny Mette, Michael F. Puchta, Holger Houben, Andreas |
author_facet | Dreissig, Steven Schiml, Simon Schindele, Patrick Weiss, Oda Rutten, Twan Schubert, Veit Gladilin, Evgeny Mette, Michael F. Puchta, Holger Houben, Andreas |
author_sort | Dreissig, Steven |
collection | PubMed |
description | Elucidating the spatiotemporal organization of the genome inside the nucleus is imperative to our understanding of the regulation of genes and non‐coding sequences during development and environmental changes. Emerging techniques of chromatin imaging promise to bridge the long‐standing gap between sequencing studies, which reveal genomic information, and imaging studies that provide spatial and temporal information of defined genomic regions. Here, we demonstrate such an imaging technique based on two orthologues of the bacterial clustered regularly interspaced short palindromic repeats (CRISPR)–CRISPR associated protein 9 (Cas9). By fusing eGFP/mRuby2 to catalytically inactive versions of Streptococcus pyogenes and Staphylococcus aureus Cas9, we show robust visualization of telomere repeats in live leaf cells of Nicotiana benthamiana. By tracking the dynamics of telomeres visualized by CRISPR–dCas9, we reveal dynamic telomere movements of up to 2 μm over 30 min during interphase. Furthermore, we show that CRISPR–dCas9 can be combined with fluorescence‐labelled proteins to visualize DNA–protein interactions in vivo. By simultaneously using two dCas9 orthologues, we pave the way for the imaging of multiple genomic loci in live plants cells. CRISPR imaging bears the potential to significantly improve our understanding of the dynamics of chromosomes in live plant cells. |
format | Online Article Text |
id | pubmed-5599988 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | John Wiley and Sons Inc. |
record_format | MEDLINE/PubMed |
spelling | pubmed-55999882017-10-02 Live‐cell CRISPR imaging in plants reveals dynamic telomere movements Dreissig, Steven Schiml, Simon Schindele, Patrick Weiss, Oda Rutten, Twan Schubert, Veit Gladilin, Evgeny Mette, Michael F. Puchta, Holger Houben, Andreas Plant J Featured Article (Technical Advance) Elucidating the spatiotemporal organization of the genome inside the nucleus is imperative to our understanding of the regulation of genes and non‐coding sequences during development and environmental changes. Emerging techniques of chromatin imaging promise to bridge the long‐standing gap between sequencing studies, which reveal genomic information, and imaging studies that provide spatial and temporal information of defined genomic regions. Here, we demonstrate such an imaging technique based on two orthologues of the bacterial clustered regularly interspaced short palindromic repeats (CRISPR)–CRISPR associated protein 9 (Cas9). By fusing eGFP/mRuby2 to catalytically inactive versions of Streptococcus pyogenes and Staphylococcus aureus Cas9, we show robust visualization of telomere repeats in live leaf cells of Nicotiana benthamiana. By tracking the dynamics of telomeres visualized by CRISPR–dCas9, we reveal dynamic telomere movements of up to 2 μm over 30 min during interphase. Furthermore, we show that CRISPR–dCas9 can be combined with fluorescence‐labelled proteins to visualize DNA–protein interactions in vivo. By simultaneously using two dCas9 orthologues, we pave the way for the imaging of multiple genomic loci in live plants cells. CRISPR imaging bears the potential to significantly improve our understanding of the dynamics of chromosomes in live plant cells. John Wiley and Sons Inc. 2017-07-14 2017-08 /pmc/articles/PMC5599988/ /pubmed/28509419 http://dx.doi.org/10.1111/tpj.13601 Text en © 2017 The Authors The Plant Journal published by John Wiley & Sons Ltd and Society for Experimental Biology. This is an open access article under the terms of the Creative Commons Attribution (http://creativecommons.org/licenses/by/4.0/) License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Featured Article (Technical Advance) Dreissig, Steven Schiml, Simon Schindele, Patrick Weiss, Oda Rutten, Twan Schubert, Veit Gladilin, Evgeny Mette, Michael F. Puchta, Holger Houben, Andreas Live‐cell CRISPR imaging in plants reveals dynamic telomere movements |
title | Live‐cell CRISPR imaging in plants reveals dynamic telomere movements |
title_full | Live‐cell CRISPR imaging in plants reveals dynamic telomere movements |
title_fullStr | Live‐cell CRISPR imaging in plants reveals dynamic telomere movements |
title_full_unstemmed | Live‐cell CRISPR imaging in plants reveals dynamic telomere movements |
title_short | Live‐cell CRISPR imaging in plants reveals dynamic telomere movements |
title_sort | live‐cell crispr imaging in plants reveals dynamic telomere movements |
topic | Featured Article (Technical Advance) |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5599988/ https://www.ncbi.nlm.nih.gov/pubmed/28509419 http://dx.doi.org/10.1111/tpj.13601 |
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