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Direct sequencing of 2′-deoxy-2′-fluoroarabinonucleic acid (FANA) using nanopore-induced phase-shift sequencing (NIPSS)

2′-deoxy-2′-fluoroarabinonucleic acid (FANA), which is one type of xeno-nucleic acid (XNA), has been intensively studied in molecular medicine and synthetic biology because of its superior gene-silencing and catalytic activities. Although urgently required, FANA cannot be directly sequenced by any e...

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Autores principales: Yan, Shuanghong, Li, Xintong, Zhang, Panke, Wang, Yuqin, Chen, Hong-Yuan, Huang, Shuo, Yu, Hanyang
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Royal Society of Chemistry 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6429604/
https://www.ncbi.nlm.nih.gov/pubmed/30996894
http://dx.doi.org/10.1039/c8sc05228j
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author Yan, Shuanghong
Li, Xintong
Zhang, Panke
Wang, Yuqin
Chen, Hong-Yuan
Huang, Shuo
Yu, Hanyang
author_facet Yan, Shuanghong
Li, Xintong
Zhang, Panke
Wang, Yuqin
Chen, Hong-Yuan
Huang, Shuo
Yu, Hanyang
author_sort Yan, Shuanghong
collection PubMed
description 2′-deoxy-2′-fluoroarabinonucleic acid (FANA), which is one type of xeno-nucleic acid (XNA), has been intensively studied in molecular medicine and synthetic biology because of its superior gene-silencing and catalytic activities. Although urgently required, FANA cannot be directly sequenced by any existing platform. Nanopore sequencing, which identifies a single molecule analyte directly from its physical and chemical properties, shows promise for direct XNA sequencing. As a proof of concept, different FANA homopolymers show well-distinguished pore blockage signals in a Mycobacterium smegmatis porin A (MspA) nanopore. By ligating FANA with a DNA drive-strand, direct FANA sequencing has been demonstrated using phi29 DNA polymerase by Nanopore-Induced Phase Shift Sequencing (NIPSS). When bound with an FANA template, the phi29 DNA polymerase shows unexpected reverse transcriptase activity when monitored in a single molecule assay. Following further investigations into the ensemble, phi29 DNA polymerase is shown to be a previously unknown reverse transcriptase for FANA that operates at room temperature, and is potentially ideal for nanopore sequencing. These results represent the first direct sequencing of a sugar-modified XNA and suggest that phi29 DNA polymerase could act as a promising enzyme for sustained sequencing of a wide variety of XNAs.
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spelling pubmed-64296042019-04-17 Direct sequencing of 2′-deoxy-2′-fluoroarabinonucleic acid (FANA) using nanopore-induced phase-shift sequencing (NIPSS) Yan, Shuanghong Li, Xintong Zhang, Panke Wang, Yuqin Chen, Hong-Yuan Huang, Shuo Yu, Hanyang Chem Sci Chemistry 2′-deoxy-2′-fluoroarabinonucleic acid (FANA), which is one type of xeno-nucleic acid (XNA), has been intensively studied in molecular medicine and synthetic biology because of its superior gene-silencing and catalytic activities. Although urgently required, FANA cannot be directly sequenced by any existing platform. Nanopore sequencing, which identifies a single molecule analyte directly from its physical and chemical properties, shows promise for direct XNA sequencing. As a proof of concept, different FANA homopolymers show well-distinguished pore blockage signals in a Mycobacterium smegmatis porin A (MspA) nanopore. By ligating FANA with a DNA drive-strand, direct FANA sequencing has been demonstrated using phi29 DNA polymerase by Nanopore-Induced Phase Shift Sequencing (NIPSS). When bound with an FANA template, the phi29 DNA polymerase shows unexpected reverse transcriptase activity when monitored in a single molecule assay. Following further investigations into the ensemble, phi29 DNA polymerase is shown to be a previously unknown reverse transcriptase for FANA that operates at room temperature, and is potentially ideal for nanopore sequencing. These results represent the first direct sequencing of a sugar-modified XNA and suggest that phi29 DNA polymerase could act as a promising enzyme for sustained sequencing of a wide variety of XNAs. Royal Society of Chemistry 2019-01-23 /pmc/articles/PMC6429604/ /pubmed/30996894 http://dx.doi.org/10.1039/c8sc05228j Text en This journal is © The Royal Society of Chemistry 2019 http://creativecommons.org/licenses/by-nc/3.0/ This article is freely available. This article is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported Licence (CC BY-NC 3.0)
spellingShingle Chemistry
Yan, Shuanghong
Li, Xintong
Zhang, Panke
Wang, Yuqin
Chen, Hong-Yuan
Huang, Shuo
Yu, Hanyang
Direct sequencing of 2′-deoxy-2′-fluoroarabinonucleic acid (FANA) using nanopore-induced phase-shift sequencing (NIPSS)
title Direct sequencing of 2′-deoxy-2′-fluoroarabinonucleic acid (FANA) using nanopore-induced phase-shift sequencing (NIPSS)
title_full Direct sequencing of 2′-deoxy-2′-fluoroarabinonucleic acid (FANA) using nanopore-induced phase-shift sequencing (NIPSS)
title_fullStr Direct sequencing of 2′-deoxy-2′-fluoroarabinonucleic acid (FANA) using nanopore-induced phase-shift sequencing (NIPSS)
title_full_unstemmed Direct sequencing of 2′-deoxy-2′-fluoroarabinonucleic acid (FANA) using nanopore-induced phase-shift sequencing (NIPSS)
title_short Direct sequencing of 2′-deoxy-2′-fluoroarabinonucleic acid (FANA) using nanopore-induced phase-shift sequencing (NIPSS)
title_sort direct sequencing of 2′-deoxy-2′-fluoroarabinonucleic acid (fana) using nanopore-induced phase-shift sequencing (nipss)
topic Chemistry
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6429604/
https://www.ncbi.nlm.nih.gov/pubmed/30996894
http://dx.doi.org/10.1039/c8sc05228j
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