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Nanopore sequencing

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https://www.readbyqxmd.com/read/29342277/finding-nemo-hybrid-assembly-with-oxford-nanopore-and-illumina-reads-greatly-improves-the-clownfish-amphiprion-ocellaris-genome-assembly
#1
Mun Hua Tan, Christopher M Austin, Michael P Hammer, Yin Peng Lee, Laurence J Croft, Han Ming Gan
Background: Some of the most widely recognised coral-reef fishes are clownfish or anemonefishes, members of the family Pomacentridae (subfamily: Amphiprioninae). They are popular aquarium species due to their bright colours, adaptability to captivity and fascinating behavior. Their breeding biology (sequential hermaphrodites) and symbiotic mutualism with sea anemones have attracted much scientific interest. Moreover, there are some curious geographic-based phenotypes which warrant investigation...
January 12, 2018: GigaScience
https://www.readbyqxmd.com/read/29340588/antimicrobial-resistant-klebsiella-pneumoniae-carriage-and-infection-in-specialized-geriatric-care-wards-linked-to-acquisition-in-the-referring-hospital
#2
Claire L Gorrie, Mirjana Mirceta, Ryan R Wick, Louise M Judd, Kelly L Wyres, Nicholas R Thomson, Richard A Strugnell, Nigel F Pratt, Jill S Garlick, Kerrie M Watson, Peter C Hunter, Steve A McGloughlin, Denis W Spelman, Adam W J Jenney, Kathryn E Holt
Background: Klebsiella pneumoniae is a leading cause of extended-spectrum beta-lactamase (ESBL) producing hospital-associated infections, for which elderly patients are at increased risk. Methods: We conducted a 1-year prospective cohort study, in which a third of patients admitted to two geriatric wards in a specialized hospital were recruited and screened for carriage of K. pneumoniae by microbiological culture. Clinical isolates were monitored via the hospital laboratory...
January 11, 2018: Clinical Infectious Diseases: An Official Publication of the Infectious Diseases Society of America
https://www.readbyqxmd.com/read/29338842/real-time-nanopore-based-recognition-of-protein-translocation-success
#3
David P Hoogerheide, Philip A Gurnev, Tatiana K Rostovtseva, Sergey M Bezrukov
A growing number of new technologies are supported by a single- or multi-nanopore architecture for capture, sensing, and delivery of polymeric biomolecules. Nanopore-based single-molecule DNA sequencing is the premier example. This method relies on the uniform linear charge density of DNA, so that each DNA strand is overwhelmingly likely to pass through the nanopore and across the separating membrane. For disordered peptides, folded proteins, or block copolymers with heterogeneous charge densities, by contrast, translocation is not assured, and additional strategies to monitor the progress of the polymer molecule through a nanopore are required...
January 12, 2018: Biophysical Journal
https://www.readbyqxmd.com/read/29334379/highly-parallel-direct-rna-sequencing-on-an-array-of-nanopores
#4
Daniel R Garalde, Elizabeth A Snell, Daniel Jachimowicz, Botond Sipos, Joseph H Lloyd, Mark Bruce, Nadia Pantic, Tigist Admassu, Phillip James, Anthony Warland, Michael Jordan, Jonah Ciccone, Sabrina Serra, Jemma Keenan, Samuel Martin, Luke McNeill, E Jayne Wallace, Lakmal Jayasinghe, Chris Wright, Javier Blasco, Stephen Young, Denise Brocklebank, Sissel Juul, James Clarke, Andrew J Heron, Daniel J Turner
Sequencing the RNA in a biological sample can unlock a wealth of information, including the identity of bacteria and viruses, the nuances of alternative splicing or the transcriptional state of organisms. However, current methods have limitations due to short read lengths and reverse transcription or amplification biases. Here we demonstrate nanopore direct RNA-seq, a highly parallel, real-time, single-molecule method that circumvents reverse transcription or amplification steps. This method yields full-length, strand-specific RNA sequences and enables the direct detection of nucleotide analogs in RNA...
January 15, 2018: Nature Methods
https://www.readbyqxmd.com/read/29325009/efficient-generation-of-complete-sequences-of-mdr-encoding-plasmids-by-rapid-assembly-of-minion-barcoding-sequencing-data
#5
Ruichao Li, Miaomiao Xie, Ning Dong, Dachuan Lin, Xuemei Yang, Marcus Ho Yin Wong, Edward Wai-Chi Chan, Sheng Chen
Background: Multidrug resistance (MDR)-encoding plasmids are considered major molecular vehicles responsible for transmission of antibiotic resistance genes among bacteria of the same or different species. Delineating the complete sequences of such plasmids could provide valuable insight into the evolution and transmission mechanisms underlying bacterial antibiotic resistance development. However, due to the presence of multiple repeats of mobile elements, complete sequencing of MDR plasmids remains technically complicated, expensive and time-consuming...
January 9, 2018: GigaScience
https://www.readbyqxmd.com/read/29322780/signal-and-noise-in-fet-nanopore-devices
#6
William M Parkin, Marija Drndic
The combination of a nanopore with a local field-effect transistor (FET-nanopore), like a nanoribbon, nanotube, or nanowire, in order to sense single molecules translocating through the pore is promising for DNA sequencing at megahertz bandwidths. Previously, it was experimentally determined that the detection mechanism was due to local potential fluctuations that arise when an analyte enters a nanopore and constricts ion flow through it, rather than the theoretically proposed mechanism of direct charge coupling between the DNA and nanowire...
January 11, 2018: ACS Sensors
https://www.readbyqxmd.com/read/29315344/culture-independent-analysis-of-liver-abscess-using-nanopore-sequencing
#7
Liang Gong, Yao-Ting Huang, Chee-Hong Wong, Wen-Cheng Chao, Zong-Yen Wu, Chia-Lin Wei, Po-Yu Liu
The identification of microbial species has depended predominantly upon culture-based techniques. However, the difficulty with which types of organisms are cultured implies that the grown species may be overrepresented by both cultivation and plate counts. In recent years, culture-independent analysis using high-throughput sequencing has been advocated for use as a point-of-care diagnostic tool. Although it offers a rapid and unbiased survey to characterize the pathogens in clinical specimens, its accuracy is reduced by the high level of contamination of human DNA...
2018: PloS One
https://www.readbyqxmd.com/read/29286470/nanopore-dna-sequencing-for-metagenomic-soil-analysis
#8
P J Cummings, J Olszewicz, K M Obom
This article describes the steps for construction of a DNA library from soil, preparation and use of the nanopore flow cell, and analysis of the DNA sequences identified using computer software. Nanopore DNA sequencing is a flexible technique that allows for rapid microbial genome sequencing to identify bacterial and viral species, to characterize bacterial strains, and to detect genetic mutations that confer resistance to antibiotics. The advantages of nanopore sequencing (NS) for life sciences include its low complexity, reduced cost, and rapid real-time sequencing of purified genomic DNA, PCR amplicons, cDNA samples, or RNA...
December 14, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/29283420/genome-sequencing-and-assembly-by-long-reads-in-plants
#9
REVIEW
Changsheng Li, Feng Lin, Dong An, Wenqin Wang, Ruidong Huang
Plant genomes generated by Sanger and Next Generation Sequencing (NGS) have provided insight into species diversity and evolution. However, Sanger sequencing is limited in its applications due to high cost, labor intensity, and low throughput, while NGS reads are too short to resolve abundant repeats and polyploidy, leading to incomplete or ambiguous assemblies. The advent and improvement of long-read sequencing by Third Generation Sequencing (TGS) methods such as PacBio and Nanopore have shown promise in producing high-quality assemblies for complex genomes...
December 28, 2017: Genes
https://www.readbyqxmd.com/read/29273626/minion-based-long-read-sequencing-and-assembly-extends-the-caenorhabditis-elegans-reference-genome
#10
John R Tyson, Nigel J O'Neil, Miten Jain, Hugh E Olsen, Philip Hieter, Terrance P Snutch
Advances in long read single molecule sequencing have opened new possibilities for 'benchtop' whole genome sequencing. The Oxford Nanopore Technologies MinION is a portable device that uses nanopore technology that can directly sequence DNA molecules. MinION single molecule long sequence reads are well suited for de novo assembly of complex genomes as they facilitate the construction of highly contiguous physical genome maps obviating the need for labor-intensive physical genome mapping. Long sequence reads can also be used to delineate complex chromosomal rearrangements, such as those that occur in tumour cells, that can confound analysis using short reads...
December 22, 2017: Genome Research
https://www.readbyqxmd.com/read/29238890/real-time-detection-of-braf-v600e-mutation-from-archival-hairy-cell-leukemia-ffpe-tissue-by-nanopore-sequencing
#11
Davide Vacca, Valeria Cancila, Alessandro Gulino, Giosuè Lo Bosco, Beatrice Belmonte, Arianna Di Napoli, Ada Maria Florena, Claudio Tripodo, Walter Arancio
The MinION is a miniaturized high-throughput next generation sequencing platform of novel conception. The use of nucleic acids derived from formalin-fixed paraffin-embedded samples is highly desirable, but their adoption for molecular assays is hurdled by the high degree of fragmentation and by the chemical-induced mutations stemming from the fixation protocols. In order to investigate the suitability of MinION sequencing on formalin-fixed paraffin-embedded samples, the presence and frequency of BRAF c.1799T > A mutation was investigated in two archival tissue specimens of Hairy cell leukemia and Hairy cell leukemia Variant...
December 13, 2017: Molecular Biology Reports
https://www.readbyqxmd.com/read/29232619/engineering-of-protein-nanopores-for-sequencing-chemical-or-protein-sensing-and-disease-diagnosis
#12
REVIEW
Shaoying Wang, Zhengyi Zhao, Farzin Haque, Peixuan Guo
Biological systems contain highly-ordered structures performing diverse functions. The elegant structures of biomachines have inspired the development of nanopores as single molecule sensors. Over the years, the utility of nanopores for detecting a wide variety of analytes have rapidly emerged for sensing, sequencing and diagnostic applications. Several protein channels with diverse shapes and sizes, such as motor channels from bacteriophage Phi29, SPP1, T3, and T4, as well as α-hemolysin, MspA, aerolysin, FluA, OmpF/G, CsgG, ClyA, have been continually investigated and developed as nanopores...
December 9, 2017: Current Opinion in Biotechnology
https://www.readbyqxmd.com/read/29224067/multiplexed-targeted-sequencing-for-oxford-nanopore-minion-a-detailed-library-preparation-procedure
#13
Timokratis Karamitros, Gkikas Magiorkinis
MinION is a small form factor sequencer recently retailed by Oxford Nanopore technologies. This lighter-sized USB3.0-interfaced device uses innovative nanotechnology to generate extra-long reads from libraries prepared using only standard molecular biology lab equipment. The flexibility and the portability of the platform makes it ideal for point-of-interest and real-time surveillance applications. However, MinION's limited capacity is not enough for the study of specific targets within larger genomes. Apart from just PCR-amplifying regions of interest, the capture of long reads spanning the edges of known-unknown genomic regions is of great importance for structural studies, such as the identification of mobile elements' integrations sites, bridging over low complexity repetitive regions etc...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29205138/virulence-genes-in-isolates-of-klebsiella-pneumoniae-from-the-uk-during-2016-including-among-carbapenemase-gene-positive-hypervirulent-k1-st23-and-non-hypervirulent-types-st147-st15-and-st383
#14
Jane F Turton, Zoë Payne, Amy Coward, Katie L Hopkins, Jack A Turton, Michel Doumith, Neil Woodford
PURPOSE: Klebsiella pneumoniae is a concern because of its multidrug resistance and the ability of hypervirulent types, especially capsular type K1-clonal complex 23 (K1-CC23), to cause community-acquired, life-threatening infections. Hypervirulent types carry an array of virulence genes including rmpA/rmpA2, coding for capsule up-regulation. We sought to identify isolates carrying these elements among submissions to the UK national reference laboratory during 2016. METHODOLOGY: Virulence elements and carbapenemase genes were sought by PCR or from whole genome sequences...
December 5, 2017: Journal of Medical Microbiology
https://www.readbyqxmd.com/read/29196558/microfluidic-isoform-sequencing-shows-widespread-splicing-coordination-in-the-human-transcriptome
#15
Hagen Tilgner, Fereshteh Jahanbani, Ishaan Gupta, Paul Collier, Eric Wei, Morten Rasmussen, Michael Snyder
Understanding transcriptome complexity is crucial for understanding human biology and disease. Technologies such as Synthetic long-read RNA sequencing (SLR-RNA-seq) delivered 5 million isoforms and allowed assessing splicing coordination. Pacific Biosciences and Oxford Nanopore increase throughput also but require high input amounts or amplification. Our new droplet-based method, sparse isoform sequencing (spISO-seq), sequences 100k-200k partitions of 10-200 molecules at a time, enabling analysis of 10-100 million RNA molecules...
December 1, 2017: Genome Research
https://www.readbyqxmd.com/read/29196295/quasi-metagenomics-and-realtime-sequencing-aided-detection-and-subtyping-of-salmonella-enterica-from-food-samples
#16
Ji-Yeon Hyeon, Shaoting Li, David A Mann, Shaokang Zhang, Zhen Li, Yi Chen, Xiangyu Deng
Metagenomics analysis of food samples promises isolation-independent detection and subtyping of foodborne bacterial pathogens in a single workflow. Selective concentration of Salmonella genomic DNA through immunomagnetic separation (IMS) and multiple displacement amplification (MDA) were shown to shorten culture enrichment of Salmonella-spiked raw chicken breast samples by over 12 hours while permitting serotyping and high-fidelity single nucleotide polymorphisms (SNP) typing of the pathogen using short shotgun sequencing reads...
December 1, 2017: Applied and Environmental Microbiology
https://www.readbyqxmd.com/read/29186559/phyloscanner-inferring-transmission-from-within-and-between-host-pathogen-genetic-diversity
#17
Chris Wymant, Matthew Hall, Oliver Ratmann, David Bonsall, Tanya Golubchik, Mariateresa de Cesare, Astrid Gall, Marion Cornelissen, Christophe Fraser
A central feature of pathogen genomics is that different infectious particles (virions, bacterial cells, etc.) within an infected individual may be genetically distinct, with patterns of relatedness amongst infectious particles being the result of both within-host evolution and transmission from one host to the next. Here we present a new software tool, phyloscanner, which analyses pathogen diversity from multiple infected hosts. phyloscanner provides unprecedented resolution into the transmission process, allowing inference of the direction of transmission from sequence data alone...
November 23, 2017: Molecular Biology and Evolution
https://www.readbyqxmd.com/read/29183387/nanopore-sequencing-of-full-length-brca1-mrna-transcripts-reveals-co-occurrence-of-known-exon-skipping-events
#18
Lucy C de Jong, Simone Cree, Vanessa Lattimore, George A R Wiggins, Amanda B Spurdle, Allison Miller, Martin A Kennedy, Logan C Walker
BACKGROUND: Laboratory assays evaluating the effect of DNA sequence variants on BRCA1 mRNA splicing may contribute to classification by providing molecular evidence. However, our knowledge of normal and aberrant BRCA1 splicing events to date has been limited to data derived from assays targeting partial transcript sequences. This study explored the utility of nanopore sequencing to examine whole BRCA1 mRNA transcripts and to provide accurate categorisation of in-frame and out-of-frame splicing events...
November 28, 2017: Breast Cancer Research: BCR
https://www.readbyqxmd.com/read/29183111/remote-activation-of-a-nanopore-for-high-performance-genetic-detection-using-a-ph-taxis-mimicking-mechanism
#19
Yong Wang, Kai Tian, Xiao Du, Ruicheng Shi, Li-Qun Gu
Aerolysin protein pore has been widely used for sensing peptides and proteins. But only a few groups explored this nanopore for nucleic acids detection. The challenge is the extremely low capture efficiency for nucleic acids (>10 bases), which severely lowers the sensitivity of an aerolysin-based genetic biosensor. Here we reported a simple and easy-to-operate approach to non-covalently transform aerolysin into a highly nucleic acids-sensitive nanopore. Through a remote pH-modulation mechanism, we simply lower the pH on one side of the pore, then aerolysin is immediately "activated" and enabled to capture target DNA/RNA efficiently from the opposite side of the pore...
November 28, 2017: Analytical Chemistry
https://www.readbyqxmd.com/read/29182276/identifying-structure-in-short-dna-scaffolds-using-solid-state-nanopores
#20
Eric Beamish, Vincent Tabard-Cossa, Michel Godin
The identification of molecular tags along nucleic acid sequences has many potential applications in bionanotechnology, disease biomarker detection and DNA sequencing. An attractive approach to this end is the use of solid-state nanopores, which can electrically detect molecular substructure and can be integrated into portable lab-on-a-chip sensors. We present here a DNA origami-based approach of molecular assembly in which solid-state nanopores are capable of differentiating 165 bp scaffolds containing zero, one and two dsDNA protrusions...
November 28, 2017: ACS Sensors
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