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, Henry

doi:10.1039/c8sc05228j

Cited by 41 publications

(49 citation statements)

References 39 publications

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“…Theh eight of the MspA NP requires % 15 nucleotides to span from the NP restriction to bulk solvent, where Phi29 DNAP is located. [28] Although the 82-mer DNAs (DNA2-4 in Figure 1b and Table S1) would be just long enough to reach out of the pore,the reduced electrophoretic force (fewer charges remaining in the pore) would allow the DNAt oe scape the pore too quickly by Brownian motion. Hence 92-mer DNAs (DNA-5 and DNA-6, see Figure 1c for synthesis and Table S1 for precise structures) bearing a1 0- ( Table S1), contain different O 6 -CMG sites within the sequence.…”

Section: Angewandte Chemiementioning

confidence: 99%

Nanopore Sequencing Accurately Identifies the Mutagenic DNA Lesion O⁶‐Carboxymethyl Guanine and Reveals Its Behavior in Replication

et al. 2019

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O6‐carboxymethylguanine (O6‐CMG) is a highly mutagenic alkylation product of DNA, triggering transition mutations relevant to gastrointestinal cancer. However, precise localization of a single O6‐CMG with conventional sequencing platforms is challenging. Here nanopore sequencing (NPS), which directly senses single DNA bases according to their physiochemical properties, was employed to detect O6‐CMG. A unique O6‐CMG signal was observed during NPS and a single‐event call accuracy of >95 % was achieved. Moreover, O6‐CMG was found to be a replication obstacle for Phi29 DNA polymerase (Phi29 DNAP), suggesting this lesion could cause DNA sequencing biases in next generation sequencing (NGS) approaches.

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Section: Angewandte Chemiementioning

confidence: 99%

Nanopore Sequencing Accurately Identifies the Mutagenic DNA Lesion O⁶‐Carboxymethyl Guanine and Reveals Its Behavior in Replication

et al. 2019

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“…The sequencing library was constructed by thermal annealing from three separate strands: the chimeric template, the primer and the blocker ( Fig. 1a , Table S1, Methods 1) 28, 29 .…”

Section: Direct Mirna Sequencing Using Nipssmentioning

confidence: 99%

“…As reported, NIPSS was carried out with a mutant Mycobacterium smegmatis porin A (MspA) nanopore (Methods 2) , atop of which a wildtype (WT) phi29 DNA polymerase (DNAP) served as the ratcheting enzyme during sequencing 28 . During NIPSS (Methods 3) , the sequencing library complex, which is bound with the phi29 DNAP, was first electrophoretically dragged into the nanopore to unzip the blocker strand mechanically.…”

Section: Direct Mirna Sequencing Using Nipssmentioning

confidence: 99%

“…S1, Table S1) . Since this initiation, miRNA sequencing signals with a read-length of ∼15 nucleotides are expected until the abasic spacer reaches the polymerase synthesis site 28 . Unless otherwise stated, all NIPSS assays described in this paper were carried out by following this configuration.…”

Section: Direct Mirna Sequencing Using Nipssmentioning

confidence: 99%

“…Nanopore Induced Phase-Shift Sequencing (NIPSS), which is a variant of nanopore sequencing, was recently developed as a universal strategy to sequence analytes other than long stretches of DNA 25 or RNA 27 . Though NIPSS is limited by its short read-length of 15 nucleotides, the concept has been verified by sequencing different 2’-deoxy-2’-fluoroarabinonucleic acid (FANA) strands 28 . Mature miRNAs, measuring ∼22 nucleotides (nt) in length, are ideal analytes for NIPSS.…”

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Direct microRNA sequencing using Nanopore Induced Phase-Shift Sequencing (NIPSS)

Zhang

Yan

Guo

et al. 2019

Preprint

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MicroRNAs (miRNAs) are a class of short non-coding RNAs that function in RNA silencing and post-transcriptional gene regulation. Besides their participation in regulating normal physiological activities, specific miRNA types could act as oncogenes, tumor suppressors or metastasis regulators, which are critical biomarkers for cancer. However, direct characterization of miRNA is challenging due to its unique properties such as its low abundance, sequence similarities and short length. Nanopore Induced Phase Shift Sequencing (NIPSS), which is a variant form of nanopore sequencing, could directly sequence any short analytes including miRNA. In practice, NIPSS clearly discriminates between different identities, isoforms and epigenetic variants of model miRNA sequences.This work demonstrates the first report of direct miRNA sequencing, which serves as a complement to existing miRNA sensing routines by the introduction of single molecule resolution. Future engineering of this technique may assist miRNA based early stage diagnosis or inspire novel cancer therapeutics.MicroRNAs (miRNAs) are a group of short, single-stranded, non-coding RNA molecules that act as posttranscriptional gene regulators for a wide variety of physiological processes, including proliferation, differentiation, apoptosis, and immune reactions 1, 2 . On the other hand, aberrant miRNAs expression levels have been shown to be closely related to diverse diseases, such as cancer 3-6 , auto-immune disorders 7 and inflammatory diseases 8 .Conventionally, miRNAs can be characterized by northern blot, quantitative reverse transcription real-time polymerase chain reaction (qRT-PCR) assays or microarrays 9 .Other emerging platforms for miRNA sensing include colorimetry 10 , bioluminescence 11 , enzymatic activity 12 , nanopore sensing 13 and electrochemistry 14 . Unfortunately, these methods provide limited analytical information because miRNA sequences are not directly reported and prior knowledge of the target miRNA sequence is required. MiRNAs function by binding to the 3' untranslated region (3'UTR) of target messengerRNAs 15 . Minor sequence variations, which include trimming, addition or substitution of miRNA sequences, alter its binding affinities to target messenger RNA 15 . As reported, miRNA isoforms (isomiRs) 16 , which were generated by the addition or deletion of one or multiple nucleotides as terminal modifications, have been shown to participate in proliferative diseases 17 and cancer 18 . On the other hand, N6-methyl-adenosine (m6A), which is an epigenetic modification among RNAs, plays important roles in miRNA biogenesis 19, 20 . These functional roles of miRNAs are associated with their specific sequences. Therefore, directly decoding a miRNA sequence along with its chemical modifications becomes critical to correlate its physiological or pathological relevance.MiRNA could be indirectly sequenced by sequencing its complementary DNAs (cDNA) by performing reverse transcription followed with deep sequencing 21 . However, this strategy suffers f...

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Xeno Nucleic Acids as Functional Materials: From Biophysical Properties to Application

Bian,

Pei,

Gao

et al. 2024

Adv Healthcare Materials

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Xeno nucleic acid (XNA) are artificial nucleic acids, in which the chemical composition of the sugar moiety is changed. These modifications impart distinct physical and chemical properties to XNAs, leading to changes in their biological, chemical, and physical stability. Additionally, these alterations influence the binding dynamics of XNAs to their target molecules. Consequently, XNAs find expanded applications as functional materials in diverse fields. This review provides a comprehensive summary of the distinctive biophysical properties exhibited by various modified XNAs and explores their applications as innovative functional materials in expanded fields.

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

Abstract: The first demonstration of direct sequencing of 2′-deoxy-2′-fluoroarabinonucleic acid (FANA) using Nanopore-Induced Phase-Shift Sequencing (NIPSS).

Cited by 41 publications

References 39 publications

Nanopore Sequencing Accurately Identifies the Mutagenic DNA Lesion O⁶‐Carboxymethyl Guanine and Reveals Its Behavior in Replication

Nanopore Sequencing Accurately Identifies the Mutagenic DNA Lesion O⁶‐Carboxymethyl Guanine and Reveals Its Behavior in Replication

Direct microRNA sequencing using Nanopore Induced Phase-Shift Sequencing (NIPSS)

Xeno Nucleic Acids as Functional Materials: From Biophysical Properties to Application

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

Abstract: The first demonstration of direct sequencing of 2′-deoxy-2′-fluoroarabinonucleic acid (FANA) using Nanopore-Induced Phase-Shift Sequencing (NIPSS).

Cited by 41 publications

References 39 publications

Nanopore Sequencing Accurately Identifies the Mutagenic DNA Lesion O6‐Carboxymethyl Guanine and Reveals Its Behavior in Replication

Nanopore Sequencing Accurately Identifies the Mutagenic DNA Lesion O6‐Carboxymethyl Guanine and Reveals Its Behavior in Replication

Direct microRNA sequencing using Nanopore Induced Phase-Shift Sequencing (NIPSS)

Xeno Nucleic Acids as Functional Materials: From Biophysical Properties to Application

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Product

Resources

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Nanopore Sequencing Accurately Identifies the Mutagenic DNA Lesion O⁶‐Carboxymethyl Guanine and Reveals Its Behavior in Replication

Nanopore Sequencing Accurately Identifies the Mutagenic DNA Lesion O⁶‐Carboxymethyl Guanine and Reveals Its Behavior in Replication