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Detection of spacer precursors formed in vivo during primed CRISPR adaptation
Type I CRISPR-Cas loci provide prokaryotes with a nucleic-acid-based adaptive immunity against foreign DNA. Immunity involves adaptation, the integration of ~30-bp DNA fragments, termed prespacers, into the CRISPR array as spacers, and interference, the targeted degradation of DNA containing a proto...
Autores principales: | , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Nature Publishing Group UK
2019
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6787059/ https://www.ncbi.nlm.nih.gov/pubmed/31601800 http://dx.doi.org/10.1038/s41467-019-12417-w |
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author | Shiriaeva, Anna A. Savitskaya, Ekaterina Datsenko, Kirill A. Vvedenskaya, Irina O. Fedorova, Iana Morozova, Natalia Metlitskaya, Anastasia Sabantsev, Anton Nickels, Bryce E. Severinov, Konstantin Semenova, Ekaterina |
author_facet | Shiriaeva, Anna A. Savitskaya, Ekaterina Datsenko, Kirill A. Vvedenskaya, Irina O. Fedorova, Iana Morozova, Natalia Metlitskaya, Anastasia Sabantsev, Anton Nickels, Bryce E. Severinov, Konstantin Semenova, Ekaterina |
author_sort | Shiriaeva, Anna A. |
collection | PubMed |
description | Type I CRISPR-Cas loci provide prokaryotes with a nucleic-acid-based adaptive immunity against foreign DNA. Immunity involves adaptation, the integration of ~30-bp DNA fragments, termed prespacers, into the CRISPR array as spacers, and interference, the targeted degradation of DNA containing a protospacer. Interference-driven DNA degradation can be coupled with primed adaptation, in which spacers are acquired from DNA surrounding the targeted protospacer. Here we develop a method for strand-specific, high-throughput sequencing of DNA fragments, FragSeq, and apply this method to identify DNA fragments accumulated in Escherichia coli cells undergoing robust primed adaptation by a type I-E or type I-F CRISPR-Cas system. The detected fragments have sequences matching spacers acquired during primed adaptation and function as spacer precursors when introduced exogenously into cells by transformation. The identified prespacers contain a characteristic asymmetrical structure that we propose is a key determinant of integration into the CRISPR array in an orientation that confers immunity. |
format | Online Article Text |
id | pubmed-6787059 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2019 |
publisher | Nature Publishing Group UK |
record_format | MEDLINE/PubMed |
spelling | pubmed-67870592019-10-15 Detection of spacer precursors formed in vivo during primed CRISPR adaptation Shiriaeva, Anna A. Savitskaya, Ekaterina Datsenko, Kirill A. Vvedenskaya, Irina O. Fedorova, Iana Morozova, Natalia Metlitskaya, Anastasia Sabantsev, Anton Nickels, Bryce E. Severinov, Konstantin Semenova, Ekaterina Nat Commun Article Type I CRISPR-Cas loci provide prokaryotes with a nucleic-acid-based adaptive immunity against foreign DNA. Immunity involves adaptation, the integration of ~30-bp DNA fragments, termed prespacers, into the CRISPR array as spacers, and interference, the targeted degradation of DNA containing a protospacer. Interference-driven DNA degradation can be coupled with primed adaptation, in which spacers are acquired from DNA surrounding the targeted protospacer. Here we develop a method for strand-specific, high-throughput sequencing of DNA fragments, FragSeq, and apply this method to identify DNA fragments accumulated in Escherichia coli cells undergoing robust primed adaptation by a type I-E or type I-F CRISPR-Cas system. The detected fragments have sequences matching spacers acquired during primed adaptation and function as spacer precursors when introduced exogenously into cells by transformation. The identified prespacers contain a characteristic asymmetrical structure that we propose is a key determinant of integration into the CRISPR array in an orientation that confers immunity. Nature Publishing Group UK 2019-10-10 /pmc/articles/PMC6787059/ /pubmed/31601800 http://dx.doi.org/10.1038/s41467-019-12417-w Text en © The Author(s) 2019 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. |
spellingShingle | Article Shiriaeva, Anna A. Savitskaya, Ekaterina Datsenko, Kirill A. Vvedenskaya, Irina O. Fedorova, Iana Morozova, Natalia Metlitskaya, Anastasia Sabantsev, Anton Nickels, Bryce E. Severinov, Konstantin Semenova, Ekaterina Detection of spacer precursors formed in vivo during primed CRISPR adaptation |
title | Detection of spacer precursors formed in vivo during primed CRISPR adaptation |
title_full | Detection of spacer precursors formed in vivo during primed CRISPR adaptation |
title_fullStr | Detection of spacer precursors formed in vivo during primed CRISPR adaptation |
title_full_unstemmed | Detection of spacer precursors formed in vivo during primed CRISPR adaptation |
title_short | Detection of spacer precursors formed in vivo during primed CRISPR adaptation |
title_sort | detection of spacer precursors formed in vivo during primed crispr adaptation |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6787059/ https://www.ncbi.nlm.nih.gov/pubmed/31601800 http://dx.doi.org/10.1038/s41467-019-12417-w |
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