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Development of Methods Derived from Iodine-Induced Specific Cleavage for Identification and Quantitation of DNA Phosphorothioate Modifications

DNA phosphorothioate (PT) modification is a novel modification that occurs on the DNA backbone, which refers to a non-bridging phosphate oxygen replaced by sulfur. This exclusive DNA modification widely distributes in bacteria but has not been found in eukaryotes to date. PT modification renders DNA...

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Autores principales: Zhu, Sucheng, Zheng, Tao, Kong, Lingxin, Li, Jinli, Cao, Bo, DeMott, Michael S., Sun, Yihua, Chen, Ying, Deng, Zixin, Dedon, Peter C., You, Delin
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7692671/
https://www.ncbi.nlm.nih.gov/pubmed/33126637
http://dx.doi.org/10.3390/biom10111491
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author Zhu, Sucheng
Zheng, Tao
Kong, Lingxin
Li, Jinli
Cao, Bo
DeMott, Michael S.
Sun, Yihua
Chen, Ying
Deng, Zixin
Dedon, Peter C.
You, Delin
author_facet Zhu, Sucheng
Zheng, Tao
Kong, Lingxin
Li, Jinli
Cao, Bo
DeMott, Michael S.
Sun, Yihua
Chen, Ying
Deng, Zixin
Dedon, Peter C.
You, Delin
author_sort Zhu, Sucheng
collection PubMed
description DNA phosphorothioate (PT) modification is a novel modification that occurs on the DNA backbone, which refers to a non-bridging phosphate oxygen replaced by sulfur. This exclusive DNA modification widely distributes in bacteria but has not been found in eukaryotes to date. PT modification renders DNA nuclease tolerance and serves as a constitute element of bacterial restriction–modification (R–M) defensive system and more biological functions are awaiting exploration. Identification and quantification of the bacterial PT modifications are thus critical to better understanding their biological functions. This work describes three detailed methods derived from iodine-induced specific cleavage-an iodine-induced cleavage assay (ICA), a deep sequencing of iodine-induced cleavage at PT site (ICDS) and an iodine-induced cleavage PT sequencing (PT-IC-Seq)-for the investigation of PT modifications. Using these approaches, we have identified the presence of PT modifications and quantized the frequency of PT modifications in bacteria. These characterizations contributed to the high-resolution genomic mapping of PT modifications, in which the distribution of PT modification sites on the genome was marked accurately and the frequency of the specific modified sites was reliably obtained. Here, we provide time-saving and less labor-consuming methods for both of qualitative and quantitative analysis of genomic PT modifications. The application of these methodologies will offer great potential for better understanding the biology of the PT modifications and open the door to future further systematical study.
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spelling pubmed-76926712020-11-28 Development of Methods Derived from Iodine-Induced Specific Cleavage for Identification and Quantitation of DNA Phosphorothioate Modifications Zhu, Sucheng Zheng, Tao Kong, Lingxin Li, Jinli Cao, Bo DeMott, Michael S. Sun, Yihua Chen, Ying Deng, Zixin Dedon, Peter C. You, Delin Biomolecules Article DNA phosphorothioate (PT) modification is a novel modification that occurs on the DNA backbone, which refers to a non-bridging phosphate oxygen replaced by sulfur. This exclusive DNA modification widely distributes in bacteria but has not been found in eukaryotes to date. PT modification renders DNA nuclease tolerance and serves as a constitute element of bacterial restriction–modification (R–M) defensive system and more biological functions are awaiting exploration. Identification and quantification of the bacterial PT modifications are thus critical to better understanding their biological functions. This work describes three detailed methods derived from iodine-induced specific cleavage-an iodine-induced cleavage assay (ICA), a deep sequencing of iodine-induced cleavage at PT site (ICDS) and an iodine-induced cleavage PT sequencing (PT-IC-Seq)-for the investigation of PT modifications. Using these approaches, we have identified the presence of PT modifications and quantized the frequency of PT modifications in bacteria. These characterizations contributed to the high-resolution genomic mapping of PT modifications, in which the distribution of PT modification sites on the genome was marked accurately and the frequency of the specific modified sites was reliably obtained. Here, we provide time-saving and less labor-consuming methods for both of qualitative and quantitative analysis of genomic PT modifications. The application of these methodologies will offer great potential for better understanding the biology of the PT modifications and open the door to future further systematical study. MDPI 2020-10-28 /pmc/articles/PMC7692671/ /pubmed/33126637 http://dx.doi.org/10.3390/biom10111491 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Zhu, Sucheng
Zheng, Tao
Kong, Lingxin
Li, Jinli
Cao, Bo
DeMott, Michael S.
Sun, Yihua
Chen, Ying
Deng, Zixin
Dedon, Peter C.
You, Delin
Development of Methods Derived from Iodine-Induced Specific Cleavage for Identification and Quantitation of DNA Phosphorothioate Modifications
title Development of Methods Derived from Iodine-Induced Specific Cleavage for Identification and Quantitation of DNA Phosphorothioate Modifications
title_full Development of Methods Derived from Iodine-Induced Specific Cleavage for Identification and Quantitation of DNA Phosphorothioate Modifications
title_fullStr Development of Methods Derived from Iodine-Induced Specific Cleavage for Identification and Quantitation of DNA Phosphorothioate Modifications
title_full_unstemmed Development of Methods Derived from Iodine-Induced Specific Cleavage for Identification and Quantitation of DNA Phosphorothioate Modifications
title_short Development of Methods Derived from Iodine-Induced Specific Cleavage for Identification and Quantitation of DNA Phosphorothioate Modifications
title_sort development of methods derived from iodine-induced specific cleavage for identification and quantitation of dna phosphorothioate modifications
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7692671/
https://www.ncbi.nlm.nih.gov/pubmed/33126637
http://dx.doi.org/10.3390/biom10111491
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