Single molecule DNA sequencing in microcapillaries

2001 ◽  
Author(s):  
Markus Sauer ◽  
F. Gobel ◽  
K.-T. Han ◽  
C. Zander
Keyword(s):  
Dna Sequencing ◽  
Single Molecule

Direct Read and Quantify Damage Nucleotide from an Oligonucleotide Using a Single Molecule Interface

2019 ◽  
Author(s):  
Jiajun Wang ◽  
Meng-Yin Li ◽  
Jie Yang ◽  
Ya-Qian Wang ◽  
Xue-Yuan Wu ◽  
...  
Keyword(s):  
Dna Sequencing ◽  
Single Molecule ◽  
Genetic Diseases ◽  
High Sensitivity ◽  
Dna Lesions ◽  
Lesion Detection ◽  
The Novel ◽  
Structural Variations ◽  
Dna Lesion ◽  
Base Lesion

DNA lesion such as metholcytosine(<sup>m</sup>C), 8-OXO-guanine(<sup>O</sup>G), inosine(I) <i>etc</i> could cause the genetic diseases. Identification of the varieties of lesion bases are usually beyond the capability of conventional DNA sequencing which is mainly designed to discriminate four bases only. Therefore, lesion detection remain challenge due to the massive varieties and less distinguishable readouts for minor structural variations. Moreover, standard amplification and labelling hardly works in DNA lesions detection. Herein, we designed a single molecule interface from the mutant K238Q Aerolysin, whose confined sensing region shows the high compatible to capture and then directly convert each base lesion into distinguishable current readouts. Compared with previous single molecule sensing interface, the resolution of the K238Q Aerolysin nanopore is enhanced by 2-order. The novel K238Q could direct discriminate at least 3 types (<sup>m</sup>C, <sup>O</sup>G, I) lesions without lableing and quantify modification sites under mixed hetero-composition condition of oligonucleotide. Such nanopore could be further applied to diagnose genetic diseases at high sensitivity.

scholarly journals Next-Generation Sequencing: From Basic Research to Diagnostics

2009 ◽  
Vol 55 (4) ◽  
pp. 641-658 ◽  
Author(s):  
Karl V Voelkerding ◽  
Shale A Dames ◽  
Jacob D Durtschi
Keyword(s):  
Next Generation Sequencing ◽  
Dna Sequencing ◽  
Single Molecule ◽  
Basic Research ◽  
Clinical Diagnostics ◽  
Massively Parallel ◽  
Next Generation ◽  
New Paradigm ◽  
The Impact ◽  
Generation Sequencing

Abstract Background: For the past 30 years, the Sanger method has been the dominant approach and gold standard for DNA sequencing. The commercial launch of the first massively parallel pyrosequencing platform in 2005 ushered in the new era of high-throughput genomic analysis now referred to as next-generation sequencing (NGS). Content: This review describes fundamental principles of commercially available NGS platforms. Although the platforms differ in their engineering configurations and sequencing chemistries, they share a technical paradigm in that sequencing of spatially separated, clonally amplified DNA templates or single DNA molecules is performed in a flow cell in a massively parallel manner. Through iterative cycles of polymerase-mediated nucleotide extensions or, in one approach, through successive oligonucleotide ligations, sequence outputs in the range of hundreds of megabases to gigabases are now obtained routinely. Highlighted in this review are the impact of NGS on basic research, bioinformatics considerations, and translation of this technology into clinical diagnostics. Also presented is a view into future technologies, including real-time single-molecule DNA sequencing and nanopore-based sequencing. Summary: In the relatively short time frame since 2005, NGS has fundamentally altered genomics research and allowed investigators to conduct experiments that were previously not technically feasible or affordable. The various technologies that constitute this new paradigm continue to evolve, and further improvements in technology robustness and process streamlining will pave the path for translation into clinical diagnostics.

Long-Read, Single Molecule, Real-Time (SMRT) DNA Sequencing for Metagenomic Applications

2015 ◽  
pp. 25-38 ◽  
Author(s):  
Brett Bowman ◽  
Mincheol Kim ◽  
Yong-Joon Cho ◽  
Jonas Korlach
Keyword(s):  
Dna Sequencing ◽  
Real Time ◽  
Single Molecule ◽  
Long Read

2. How to read the book of life

2018 ◽  
pp. 12-27
Author(s):  
John Archibald
Keyword(s):  
Dna Sequencing ◽  
Human Genome ◽  
Single Molecule ◽  
Human Genome Project ◽  
Genome Project ◽  
Chain Termination ◽  
Single Molecule Sequencing ◽  
Semiconductor Sequencing ◽  
The Cost ◽  
The Human Genome Project

For all its biological importance, DNA is a fragile molecule so extracting it is a difficult process. ‘How to read the book of life’ explains the techniques required to sequence DNA. It begins by explaining the techniques developed for protein and RNA sequencing by Frederick Sanger, Robert Holley, and Carl Woese that were then developed further for DNA sequencing. Following the success of the Human Genome Project, the next generation of DNA sequencing was developed in the mid-2000s. Pyrosequencing was capable of generating orders of magnitude more data at a fraction of the cost, but was superceded within a decade by semiconductor sequencing, reversible chain-termination sequencing, and single-molecule sequencing.

scholarly journals Five Closed Salmonella enterica Genome Sequences from a 2017–2018 Multistrain, Multistate Kratom Outbreak

2020 ◽  
Vol 9 (3) ◽  
Author(s):  
Hallie E. Rauch ◽  
Julie Haendiges ◽  
Maria Balkey ◽  
Maria Hoffmann
Keyword(s):  
Dna Sequencing ◽  
Real Time ◽  
Single Molecule ◽  
Salmonella Enterica ◽  
Genome Sequences ◽  
Circular Chromosome ◽  
Content Type ◽  
Single Plasmid

We report here the closed genomes of Salmonella enterica strains from the 2017–2018 multistrain, multistate kratom outbreak using single-molecule real-time DNA sequencing. Four of the genomes consist of one circular chromosome, and the fifth has a circular chromosome and a single plasmid.

scholarly journals Single-molecule detection of deoxyribonucleoside triphosphates in microdroplets

2019 ◽  
Vol 47 (17) ◽  
pp. e101-e101 ◽  
Author(s):  
Boris Breiner ◽  
Kerr Johnson ◽  
Magdalena Stolarek ◽  
Ana-Luisa Silva ◽  
Aurel Negrea ◽  
...  
Keyword(s):  
Dna Sequencing ◽  
Single Molecule ◽  
Dna Polymerases ◽  
Single Molecule Detection ◽  
Synthesis Methods ◽  
Single Nucleotide ◽  
New Approach ◽  
Fluorescent Signal ◽  
Single Molecule Level ◽  
Current Sequence

AbstractA new approach to single-molecule DNA sequencing in which dNTPs, released by pyrophosphorolysis from the strand to be sequenced, are captured in microdroplets and read directly could have substantial advantages over current sequence-by-synthesis methods; however, there is no existing method sensitive enough to detect a single nucleotide in a microdroplet. We have developed a method for dNTP detection based on an enzymatic two-stage reaction which produces a robust fluorescent signal that is easy to detect and process. By taking advantage of the inherent specificity of DNA polymerases and ligases, coupled with volume restriction in microdroplets, this method allows us to simultaneously detect the presence of and distinguish between, the four natural dNTPs at the single-molecule level, with negligible cross-talk.

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