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Nanopore sequencing

Mélanie V Trudel, Antony T Vincent, Sabrina A Attéré, Myriam Labbé, Nicolas Derome, Alexander I Culley, Steve J Charette
The bacterium Aeromonas salmonicida subsp. salmonicida is a common pathogen in fish farms worldwide. Since the antibiotic resistance of this bacterial species is on the increase, it is important to have a broader view on this issue. In the present study, we tested the presence of known plasmids conferring multi-drug resistance as well as antibiotic resistance genes by a PCR approach in 100 Canadian A. salmonicida subsp. salmonicida isolates. Our study highlighted the dominance of the conjugative pSN254b plasmid, which confers multi-drug resistance...
October 18, 2016: Scientific Reports
James Wilson, Leila Sloman, Zhiren He, Aleksei Aksimentiev
An inexpensive, reliable method for protein sequencing is essential to unraveling the biological mechanisms governing cellular behavior and disease. Current protein sequencing methods suffer from limitations associated with the size of proteins that can be sequenced, the time, and the cost of the sequencing procedures. Here, we report the results of all-atom molecular dynamics simulations that investigated the feasibility of using graphene nanopores for protein sequencing. We focus our study on the biologically significant phenylalanine-glycine repeat peptides (FG-nups)-parts of the nuclear pore transport machinery...
July 19, 2016: Advanced Functional Materials
Dongmei Xi, Jizhen Shang, Enguo Fan, Jin-Mao You, Shusheng Zhang, Hua Wang
The accurate discrimination of microRNAs (miRNAs) with highly similar sequences would greatly facilitate the screening and early diagnosis of diseases. In the present work, a locked nucleic acid (LNA)-modified probe was designed and used for α-hemolysin (α-HL) nanopore to selectively and specifically identify miRNAs. The hybridization of the LNA probe with the target miRNAs generated unique long-lived signals in the nanopore thus facilitated an accurate discrimination of miRNAs with similar sequences, even a single-nucleotide difference...
October 13, 2016: Analytical Chemistry
P Benjamin Stranges, Mirkó Palla, Sergey Kalachikov, Jeff Nivala, Michael Dorwart, Andrew Trans, Shiv Kumar, Mintu Porel, Minchen Chien, Chuanjuan Tao, Irina Morozova, Zengmin Li, Shundi Shi, Aman Aberra, Cleoma Arnold, Alexander Yang, Anne Aguirre, Eric T Harada, Daniel Korenblum, James Pollard, Ashwini Bhat, Dmitriy Gremyachinskiy, Arek Bibillo, Roger Chen, Randy Davis, James J Russo, Carl W Fuller, Stefan Roever, Jingyue Ju, George M Church
Scalable, high-throughput DNA sequencing is a prerequisite for precision medicine and biomedical research. Recently, we presented a nanopore-based sequencing-by-synthesis (Nanopore-SBS) approach, which used a set of nucleotides with polymer tags that allow discrimination of the nucleotides in a biological nanopore. Here, we designed and covalently coupled a DNA polymerase to an α-hemolysin (αHL) heptamer using the SpyCatcher/SpyTag conjugation approach. These porin-polymerase conjugates were inserted into lipid bilayers on a complementary metal oxide semiconductor (CMOS)-based electrode array for high-throughput electrical recording of DNA synthesis...
October 11, 2016: Proceedings of the National Academy of Sciences of the United States of America
Crescenzio Francesco Minervini, Cosimo Cumbo, Paola Orsini, Claudia Brunetti, Luisa Anelli, Antonella Zagaria, Angela Minervini, Paola Casieri, Nicoletta Coccaro, Giuseppina Tota, Luciana Impera, Annamaria Giordano, Giorgina Specchia, Francesco Albano
BACKGROUND: The assessment of TP53 mutational status is becoming a routine clinical practice for chronic lymphocytic leukemia patients (CLL). A broad spectrum of molecular techniques has been employed so far, including both direct Sanger sequencing and next generation sequencing. Oxford Nanopore Technologies recently released the MinION an USB-interfaced sequencer. In this paper we report our experience, with the MinION technology for the detection of the TP53 gene mutation in CLL patients...
October 10, 2016: Diagnostic Pathology
G C A da Hora, N L Archilha, J L S Lopes, D M Müller, K Coutinho, R Itri, T A Soares
Antimicrobial peptides (AMPs) are cationic peptides that kill bacteria with a broad spectrum of action, low toxicity to mammalian cells and exceptionally low rates of bacterial resistance. These features have led to considerable efforts in developing AMPs as an alternative antibacterial therapy. In vitro studies have shown that AMPs interfere with membrane bilayer integrity via several possible mechanisms, which are not entirely understood. We have performed the synthesis, membrane lysis measurements, and biophysical characterization of a novel hybrid peptide...
September 26, 2016: Soft Matter
Lei Liu, Hai-Chen Wu
Nanopore sensing is an attractive, label-free approach that can measure single molecules. Although initially proposed for rapid and low-cost DNA sequencing, nanopore sensors have been successfully employed in the detection of a wide variety of substrates. Early successes were mostly achieved based on two main strategies by 1) creating sensing elements inside the nanopore through protein mutation and chemical modification or 2) using molecular adapters to enhance analyte recognition. Over the past five years, DNA molecules started to be used as probes for sensing rather than substrates for sequencing...
September 27, 2016: Angewandte Chemie
K Schmidt, S Mwaigwisya, L C Crossman, M Doumith, D Munroe, C Pires, A M Khan, N Woodford, N J Saunders, J Wain, J O'Grady, D M Livermore
OBJECTIVES: The introduction of metagenomic sequencing to diagnostic microbiology has been hampered by slowness, cost and complexity. We explored whether MinION nanopore sequencing could accelerate diagnosis and resistance profiling, using complicated urinary tract infections as an exemplar. METHODS: Bacterial DNA was enriched from clinical urines (n = 10) and from healthy urines 'spiked' with multiresistant Escherichia coli (n = 5), then sequenced by MinION...
September 25, 2016: Journal of Antimicrobial Chemotherapy
Hengyun Lu, Francesca Giordano, Zemin Ning
The revolution of genome sequencing is continuing after the successful second-generation sequencing (SGS) technology. The third-generation sequencing (TGS) technology, led by Pacific Biosciences (PacBio), is progressing rapidly, moving from a technology once only capable of providing data for small genome analysis, or for performing targeted screening, to one that promises high quality de novo assembly and structural variation detection for human-sized genomes. In 2014, the MinION, the first commercial sequencer using nanopore technology, was released by Oxford Nanopore Technologies (ONT)...
September 17, 2016: Genomics, Proteomics & Bioinformatics
Carl T Wittwer
No abstract text is available yet for this article.
September 16, 2016: Clinical Chemistry
Florian C Rieger, Peter Virnau
We determine knotting probabilities and typical sizes of knots in double-stranded DNA for chains of up to half a million base pairs with computer simulations of a coarse-grained bead-stick model: Single trefoil knots and composite knots which include at least one trefoil as a prime factor are shown to be common in DNA chains exceeding 250,000 base pairs, assuming physiologically relevant salt conditions. The analysis is motivated by the emergence of DNA nanopore sequencing technology, as knots are a potential cause of erroneous nucleotide reads in nanopore sequencing devices and may severely limit read lengths in the foreseeable future...
September 2016: PLoS Computational Biology
Bashir Fotouhi, Vahid Ahmadi, Vahid Faramarzi
We propose novel nano-plasmonic-based structures for rapid sequencing of DNA molecules. The optical properties of DNA nucleotides have notable differences in the ultraviolet (UV) region of light. Using nanopore, bowtie, and bowtie-nanopore compound structures, probable application of the surface plasmon resonance (SPR) in DNA sequencing is investigated by employing the discrete dipole approximation method. The effects of different materials like chromium (Cr), aluminum (Al), rhodium (Rh), and graphene (Gr) are studied...
September 15, 2016: Optics Letters
Alex Smolyanitsky, Boris I Yakobson, Tsjerk A Wassenaar, Eugene Paulechka, Kenneth Kroenlein
We propose an aqueous functionalized molybdenum disulfide nanoribbon suspended over a solid electrode as a capacitive displacement sensor aimed at determining the DNA sequence. The detectable sequencing events arise from the combination of Watson-Crick base-pairing, one of nature's most basic lock-and-key binding mechanisms, with the ability of appropriately sized atomically thin membranes to flex substantially in response to subnanonewton forces. We employ carefully designed numerical simulations and theoretical estimates to demonstrate excellent (79% to 86%) raw target detection accuracy at ∼70 million bases per second and electrical measurability of the detected events...
September 27, 2016: ACS Nano
Suyan Qiu, Fusheng Zhao, Oussama Zenasni, Jingting Li, Wei-Chuan Shih
We report label-free small molecule sensing on nanoporous gold disks functionalized with stabilized Guanine-quadruplex (G4) moieties using surface-enhanced Raman spectroscopy (SERS). Utilizing the unique G4 topological structure, target molecules can be selectively captured onto nanoporous gold (NPG) disk surfaces via π-π stacking and electrostatic attractions. Together with high-density plasmonic "hot spots" of NPG disks, the captured molecules produce a remarkable SERS signal. Our strategy represents the first example of the detection of foreign moleules conjugated to non-double helical DNA nanostructures by SERS while providing a new technique to studying the formation and evolution of G4 moieties...
September 13, 2016: ACS Applied Materials & Interfaces
Matei David, L J Dursi, Delia Yao, Paul C Boutros, Jared T Simpson
MOTIVATION: The highly portable Oxford Nanopore MinION sequencer has enabled new applications of genome sequencing directly in the field. However, the MinION currently relies on a cloud computing platform, Metrichor (, for translating locally generated sequencing data into basecalls. RESULTS: To allow offline and private analysis of MinION data, we created Nanocall. Nanocall is the first freely-available, open-source basecaller for Oxford Nanopore sequencing data and does not require an internet connection...
September 10, 2016: Bioinformatics
Carsten Wloka, Natalie Lisa Mutter, Misha Soskine, Giovanni Maglia
Nanopores are used in single-molecule DNA analysis and sequencing. Herein, we show that Fragaceatoxin C (FraC), an α-helical pore-forming toxin from an actinoporin protein family, can be reconstituted in sphingomyelin-free standard planar lipid bilayers. We engineered FraC for DNA analysis and show that the funnel-shaped geometry allows tight wrapping around single-stranded DNA (ssDNA), resolving between homopolymeric C, T, and A polynucleotide stretches. Remarkably, despite the 1.2 nm internal constriction of FraC, double-stranded DNA (dsDNA) can translocate through the nanopore at high applied potentials, presumably through the deformation of the α-helical transmembrane region of the pore...
September 26, 2016: Angewandte Chemie
Ganesh Sivaraman, Rodrigo G Amorim, Ralph H Scheicher, Maria Fyta
Small diamond-like particles, diamondoids, have been shown to effectively functionalize gold electrodes in order to sense DNA units passing between the nanopore-embedded electrodes. In this work, we present a comparative study of Au(111) electrodes functionalized with different derivatives of lower diamondoids. Focus is put on the electronic and transport properties of such electrodes for different DNA nucleotides placed within the electrode gap. The functionalization promotes a specific binding to DNA leading to different properties for the system, which provides a tool set to systematically improve the signal-to-noise ratio of the electronic measurements across the electrodes...
October 14, 2016: Nanotechnology
Andy S Alic, Ignacio Blanquer
Usually, the information known a priori about a newly sequenced organism is limited. Even resequencing the same organism can generate unpredictable output. We introduce MuffinInfo, a FastQ/Fasta/SAM information extractor implemented in HTML5 capable of offering insights into next-generation sequencing (NGS) data. Our new tool can run on any software or hardware environment, in command line or graphically, and in browser or standalone. It presents information such as average length, base distribution, quality scores distribution, k-mer histogram, and homopolymers analysis...
September 2016: Journal of Computational Biology: a Journal of Computational Molecular Cell Biology
P D Gupta
In health care, importance of DNA sequencing has been fully established. Sanger's Capillary Electrophoresis DNA sequencing methodology is time consuming, cumbersome, hence become more expensive. Lately, because of its versatility DNA sequencing became house hold name, and therefore, there is an urgent need of simple, fast, inexpensive, DNA sequencing technology. In the beginning of this century efforts were made, and Nanopore DNA sequencing technology was developed; still it is infancy, nevertheless, it is the futuristic technology...
October 2016: Indian Journal of Clinical Biochemistry: IJCB
Mónika Szabó, Tibor Nagy, Tímea Wilk, Tibor Farkas, Anna Hegyi, Ferenc Olasz, János Kiss
Two IncA/C family plasmids from the 1960s have been sequenced and classified into A/C2 Type 1 group. R16a and IP40a contain novel antibiotic resistance islands and a complete GIsul2 island not previously found in the family. In the 173.1 kb R16a the 29.9 kb ARI is located in a unique backbone position not utilized by ARIs. ARIR16a consists of Tn1, Tn6020, Tn6333 harboring the resistance genes blaTEM-1D, aphA1b and a mer module, respectively, a truncated Tn5393 copy and a gene cluster with unknown function. Plasmid IP40a is 170...
September 6, 2016: Antimicrobial Agents and Chemotherapy
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