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https://www.readbyqxmd.com/read/29026658/resolving-plasmid-structures-in-enterobacteriaceae-using-the-minion-nanopore-sequencer-assessment-of-minion-and-minion-illumina-hybrid-data-assembly-approaches
#1
Sophie George, Louise Pankhurst, Alasdair Hubbard, Antonia Votintseva, Nicole Stoesser, Anna E Sheppard, Amy Mathers, Rachel Norris, Indre Navickaite, Chloe Eaton, Zamin Iqbal, Derrick W Crook, Hang T T Phan
This study aimed to assess the feasibility of using the Oxford Nanopore Technologies (ONT) MinION long-read sequencer in reconstructing fully closed plasmid sequences from eight Enterobacteriaceae isolates of six different species with plasmid populations of varying complexity. Species represented were Escherichia coli, Klebsiella pneumoniae, Citrobacter freundii, Enterobacter cloacae, Serratia marcescens and Klebsiella oxytoca, with plasmid populations ranging from 1-11 plasmids with sizes of 2-330 kb. Isolates were sequenced using Illumina (short-read) and ONT's MinION (long-read) platforms, and compared with fully resolved PacBio (long-read) sequence assemblies for the same isolates...
August 2017: Microbial Genomics
https://www.readbyqxmd.com/read/29026655/sharing-of-carbapenemase-encoding-plasmids-between-enterobacteriaceae-in-uk-sewage-uncovered-by-minion-sequencing
#2
Catherine Ludden, Sandra Reuter, Kim Judge, Theodore Gouliouris, Beth Blane, Francesc Coll, Plamena Naydenova, Martin Hunt, Alan Tracey, Katie L Hopkins, Nicholas M Brown, Neil Woodford, Julian Parkhill, Sharon J Peacock
Dissemination of carbapenem resistance among pathogenic Gram-negative bacteria is a looming medical emergency. Efficient spread of resistance within and between bacterial species is facilitated by mobile genetic elements. We hypothesized that wastewater contributes to the dissemination of carbapenemase-producing Enterobacteriaceae (CPE), and studied this through a cross-sectional observational study of wastewater in the East of England. We isolated clinically relevant species of CPE in untreated and treated wastewater, confirming that waste treatment does not prevent release of CPE into the environment...
July 2017: Microbial Genomics
https://www.readbyqxmd.com/read/29021151/rapid-nanopore-sequencing-of-plasmids-and-resistance-gene-detection-in-clinical-isolates
#3
Jamie K Lemon, Pavel P Khil, Karen M Frank, John P Dekker
Recent advances in nanopore sequencing technology have led to a substantial increase in throughput and sequence quality. Together, these improvements may permit real-time benchtop genomic sequencing and antimicrobial resistance gene detection in clinical isolates. Here, we evaluated workflows and turnaround times for a benchtop long-read sequencing approach in the clinical microbiology laboratory using the Oxford Nanopore Technologies MinION sequencer. We performed genomic and plasmid sequencing of three clinical isolates with both MinION and Illumina MiSeq, using different library preparation methods (2D and rapid 1D) with the goal of antimicrobial resistance gene detection...
October 11, 2017: Journal of Clinical Microbiology
https://www.readbyqxmd.com/read/28992056/a-world-of-opportunities-with-nanopore-sequencing
#4
Richard M Leggett, Matthew D Clark
Oxford Nanopore Technologies' MinION sequencer was launched in pre-release form in 2014 and represents an exciting new sequencing paradigm. The device offers multi-kilobase reads and a streamed mode of operation that allows processing of reads as they are generated. Crucially, it is an extremely compact device that is powered from the USB port of a laptop computer, enabling it to be taken out of the lab and facilitating previously impossible in-field sequencing experiments to be undertaken. Many of the initial publications concerning the platform focused on provision of tools to access and analyse the new sequence formats and then demonstrating the assembly of microbial genomes...
August 23, 2017: Journal of Experimental Botany
https://www.readbyqxmd.com/read/28983066/high-quality-de-novo-genome-assembly-of-the-dekkera-bruxellensis-yeast-using-nanopore-minion-sequencing
#5
Téo Fournier, Jean-Sébastien Gounot, Kelle Freel, Corinne Cruaud, Arnaud Lemainque, Jean-Marc Aury, Patrick Wincker, Joseph Schacherer, Anne Friedrich
Genetic variation in natural populations represents the raw material for phenotypic diversity. Species-wide characterization of genetic variants is crucial to have a deeper insight into the genotype-phenotype relationship. With the advent of new sequencing strategies and more recently the release of long-read sequencing platforms, it is now possible to explore the genetic diversity of any nonmodel organisms, representing a fundamental resource for biological research. In the frame of population genomic surveys, a first step is to obtain the complete sequence and high-quality assembly of a reference genome...
October 5, 2017: G3: Genes—Genomes—Genetics
https://www.readbyqxmd.com/read/28977016/on-site-dna-barcoding-by-nanopore-sequencing
#6
Michele Menegon, Chiara Cantaloni, Ana Rodriguez-Prieto, Cesare Centomo, Ahmed Abdelfattah, Marzia Rossato, Massimo Bernardi, Luciano Xumerle, Simon Loader, Massimo Delledonne
Biodiversity research is becoming increasingly dependent on genomics, which allows the unprecedented digitization and understanding of the planet's biological heritage. The use of genetic markers i.e. DNA barcoding, has proved to be a powerful tool in species identification. However, full exploitation of this approach is hampered by the high sequencing costs and the absence of equipped facilities in biodiversity-rich countries. In the present work, we developed a portable sequencing laboratory based on the portable DNA sequencer from Oxford Nanopore Technologies, the MinION...
2017: PloS One
https://www.readbyqxmd.com/read/28961793/covert-dissemination-of-carbapenemase-producing-klebsiella-pneumoniae-kpc-in-a-successfully-controlled-outbreak-long-and-short-read-whole-genome-sequencing-demonstrate-multiple-genetic-modes-of-transmission
#7
Jessica Martin, Hang T T Phan, Jacqueline Findlay, Nicole Stoesser, Louise Pankhurst, Indre Navickaite, Nicola De Maio, David W Eyre, Giles Toogood, Nicolas M Orsi, Andrew Kirby, Nicola Young, Jane F Turton, Robert L R Hill, Katie L Hopkins, Neil Woodford, Tim E A Peto, A Sarah Walker, Derrick W Crook, Mark H Wilcox
Background: Carbapenemase-producing Enterobacteriaceae (CPE), including KPC-producing Klebsiella pneumoniae (KPC -Kpn ), are an increasing threat to patient safety. Objectives: To use WGS to investigate the extent and complexity of carbapenemase gene dissemination in a controlled KPC outbreak. Materials and methods: Enterobacteriaceae with reduced ertapenem susceptibility recovered from rectal screening swabs/clinical samples, during a 3 month KPC outbreak (2013-14), were investigated for carbapenemase production, antimicrobial susceptibility, variable-number-tandem-repeat profile and WGS [short-read (Illumina), long-read (MinION)]...
August 7, 2017: Journal of Antimicrobial Chemotherapy
https://www.readbyqxmd.com/read/28961779/nanopore-sequencing-enables-near-complete-de-novo-assembly-of-saccharomyces-cerevisiae-reference-strain-cen-pk113-7d
#8
Alex N Salazar, Arthur R Gorter de Vries, Marcel van den Broek, Melanie Wijsman, Pilar de la Torre Cortés, Anja Brickwedde, Nick Brouwers, Jean-Marc G Daran, Thomas Abeel
The haploid Saccharomyces cerevisiae strain CEN.PK113-7D is a popular model system for metabolic engineering and systems biology research. Current genome assemblies are based on short-read sequencing data scaffolded based on homology to strain S288C. However, these assemblies contain large sequence gaps, particularly in subtelomeric regions, and the assumption of perfect homology to S288C for scaffolding introduces bias. In this study, we obtained a near-complete genome assembly of CEN.PK113-7D using only Oxford Nanopore Technology's MinION sequencing platform...
September 13, 2017: FEMS Yeast Research
https://www.readbyqxmd.com/read/28928943/investigation-of-chimeric-reads-using-the-minion
#9
Ruby White, Christophe Pellefigues, Franca Ronchese, Olivier Lamiable, David Eccles
Following a nanopore sequencing run of PCR products of three amplicons less than 1kb, an abundance of reads failed quality control due to template/complement mismatch. A BLAST search demonstrated that some of the failed reads mapped to two different genes -- an unexpected observation, given that PCR was carried out separately for each amplicon. A further investigation was carried out specifically to search for chimeric reads, using separate barcodes for each amplicon and trying two different ligation methods prior to sample loading...
2017: F1000Research
https://www.readbyqxmd.com/read/28911333/profiling-bacterial-communities-by-minion-sequencing-of-ribosomal-operons
#10
Lee J Kerkhof, Kevin P Dillon, Max M Häggblom, Lora R McGuinness
BACKGROUND: An approach utilizing the long-read capability of the Oxford Nanopore MinION to rapidly sequence bacterial ribosomal operons of complex natural communities was developed. Microbial fingerprinting employs domain-specific forward primers (16S rRNA subunit), reverse primers (23S rRNA subunit), and a high-fidelity Taq polymerase with proofreading capabilities. Amplicons contained both ribosomal subunits for broad-based phylogenetic assignment (~ 3900 bp of sequence), plus the intergenic spacer (ITS) region (~ 300 bp) for potential strain-specific identification...
September 15, 2017: Microbiome
https://www.readbyqxmd.com/read/28903726/a-novel-diagnostic-method-for-malaria-using-loop-mediated-isothermal-amplification-lamp-and-minion%C3%A2-nanopore-sequencer
#11
Kazuo Imai, Norihito Tarumoto, Kazuhisa Misawa, Lucky Ronald Runtuwene, Jun Sakai, Kyoko Hayashida, Yuki Eshita, Ryuichiro Maeda, Josef Tuda, Takashi Murakami, Shigefumi Maesaki, Yutaka Suzuki, Junya Yamagishi, Takuya Maeda
BACKGROUND: A simple and accurate molecular diagnostic method for malaria is urgently needed due to the limitations of conventional microscopic examination. In this study, we demonstrate a new diagnostic procedure for human malaria using loop mediated isothermal amplification (LAMP) and the MinION™ nanopore sequencer. METHODS: We generated specific LAMP primers targeting the 18S-rRNA gene of all five human Plasmodium species including two P. ovale subspecies (P...
September 13, 2017: BMC Infectious Diseases
https://www.readbyqxmd.com/read/28855093/metagenomic-arbovirus-detection-using-minion-nanopore-sequencing
#12
Jana Batovska, Stacey E Lynch, Brendan C Rodoni, Tim I Sawbridge, Noel Oi Cogan
With its small size and low cost, the hand-held MinION sequencer is a powerful tool for in-field surveillance. Using a metagenomic approach, it allows non-targeted detection of viruses in a sample within a few hours. This study aimed to determine the ability of the MinION to metagenomically detect and characterise a virus from an infected mosquito. RNA was extracted from an Aedes notoscriptus mosquito infected with Ross River virus (RRV), converted into cDNA and sequenced on the MinION. Bioinformatic analysis of the MinION reads led to detection of full-length RRV, with reads of up to 2...
September 4, 2017: Journal of Virological Methods
https://www.readbyqxmd.com/read/28837856/secure-and-robust-cloud-computing-for-high-throughput-forensic-microsatellite-sequence-analysis-and-databasing
#13
Sarah F Bailey, Melissa K Scheible, Christopher Williams, Deborah S B S Silva, Marina Hoggan, Christopher Eichman, Seth A Faith
Next-generation Sequencing (NGS) is a rapidly evolving technology with demonstrated benefits for forensic genetic applications, and the strategies to analyze and manage the massive NGS datasets are currently in development. Here, the computing, data storage, connectivity, and security resources of the Cloud were evaluated as a model for forensic laboratory systems that produce NGS data. A complete front-to-end Cloud system was developed to upload, process, and interpret raw NGS data using a web browser dashboard...
August 8, 2017: Forensic Science International. Genetics
https://www.readbyqxmd.com/read/28794860/minion-analysis-and-reference-consortium-phase-2-data-release-and-analysis-of-r9-0-chemistry
#14
Miten Jain, John R Tyson, Matthew Loose, Camilla L C Ip, David A Eccles, Justin O'Grady, Sunir Malla, Richard M Leggett, Ola Wallerman, Hans J Jansen, Vadim Zalunin, Ewan Birney, Bonnie L Brown, Terrance P Snutch, Hugh E Olsen
BACKGROUND: Long-read sequencing is rapidly evolving and reshaping the suite of opportunities for genomic analysis. For the MinION in particular, as both the platform and chemistry develop, the user community requires reference data to set performance expectations and maximally exploit third-generation sequencing. We performed an analysis of MinION data derived from whole genome sequencing of Escherichiacoli K-12 using the R9.0 chemistry, comparing the results with the older R7.3 chemistry...
2017: F1000Research
https://www.readbyqxmd.com/read/28794062/high-quality-de-novo-genome-assembly-of-the-dekkera-bruxellensis-yeast-isolate-using-nanopore-minion-sequencing
#15
Téo Fournier, Jean-Sébastien Gounot, Kelle Freel, Corinne Cruaud, Arnaud Lemainque, Jean-Marc Aury, Patrick Wincker, Joseph Schacherer, Anne Friedrich
Genetic variation in natural populations represents the raw material for phenotypic diversity. Species-wide characterization of genetic variants is crucial to have a deeper insight into the genotype-phenotype relationship. With the advent of new sequencing strategies and more recently the release of long-read sequencing platforms, it is now possible to explore the genetic diversity of any non-model organisms, representing a fundamental resource for biological research. In the frame of population genomic surveys, a first step is to obtain the complete sequence and high quality assembly of a reference genome...
August 9, 2017: G3: Genes—Genomes—Genetics
https://www.readbyqxmd.com/read/28775309/rapid-de-novo-assembly-of-the-european-eel-genome-from-nanopore-sequencing-reads
#16
Hans J Jansen, Michael Liem, Susanne A Jong-Raadsen, Sylvie Dufour, Finn-Arne Weltzien, William Swinkels, Alex Koelewijn, Arjan P Palstra, Bernd Pelster, Herman P Spaink, Guido E van den Thillart, Ron P Dirks, Christiaan V Henkel
We have sequenced the genome of the endangered European eel using the MinION by Oxford Nanopore, and assembled these data using a novel algorithm specifically designed for large eukaryotic genomes. For this 860 Mbp genome, the entire computational process takes two days on a single CPU. The resulting genome assembly significantly improves on a previous draft based on short reads only, both in terms of contiguity (N50 1.2 Mbp) and structural quality. This combination of affordable nanopore sequencing and light weight assembly promises to make high-quality genomic resources accessible for many non-model plants and animals...
August 3, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28774990/complete-genome-sequence-of-achromobacter-denitrificans-pr1
#17
Ana C Reis, Kevin Kroll, Margarita Gomila, Boris A Kolvenbach, Philippe F X Corvini, Olga C Nunes
Achromobacter denitrificans strain PR1 was isolated from an enrichment culture able to use sulfamethoxazole as an energy source. Here, we describe the complete genome of this strain sequenced by Illumina MiSeq and Oxford Nanopore MinION.
August 3, 2017: Genome Announcements
https://www.readbyqxmd.com/read/28747151/comparative-analysis-of-targeted-long-read-sequencing-approaches-for-characterization-of-a-plant-s-immune-receptor-repertoire
#18
Michael Giolai, Pirita Paajanen, Walter Verweij, Kamil Witek, Jonathan D G Jones, Matthew D Clark
BACKGROUND: The Oxford Nanopore Technologies MinION™ sequencer is a small, portable, low cost device that is accessible to labs of all sizes and attractive for in-the-field sequencing experiments. Selective breeding of crops has led to a reduction in genetic diversity, and wild relatives are a key source of new genetic resistance to pathogens, usually via NLR immune receptor-encoding genes. Recent studies have demonstrated how crop NLR repertoires can be targeted for sequencing on Illumina or PacBio (RenSeq) and the specific gene conveying pathogen resistance identified...
July 26, 2017: BMC Genomics
https://www.readbyqxmd.com/read/28722025/nanopore-long-read-rnaseq-reveals-widespread-transcriptional-variation-among-the-surface-receptors-of-individual-b-cells
#19
Ashley Byrne, Anna E Beaudin, Hugh E Olsen, Miten Jain, Charles Cole, Theron Palmer, Rebecca M DuBois, E Camilla Forsberg, Mark Akeson, Christopher Vollmers
Understanding gene regulation and function requires a genome-wide method capable of capturing both gene expression levels and isoform diversity at the single-cell level. Short-read RNAseq is limited in its ability to resolve complex isoforms because it fails to sequence full-length cDNA copies of RNA molecules. Here, we investigate whether RNAseq using the long-read single-molecule Oxford Nanopore MinION sequencer is able to identify and quantify complex isoforms without sacrificing accurate gene expression quantification...
July 19, 2017: Nature Communications
https://www.readbyqxmd.com/read/28720805/a-portable-system-for-rapid-bacterial-composition-analysis-using-a-nanopore-based-sequencer-and-laptop-computer
#20
Satomi Mitsuhashi, Kirill Kryukov, So Nakagawa, Junko S Takeuchi, Yoshiki Shiraishi, Koichiro Asano, Tadashi Imanishi
We developed a portable system for 16S rDNA analyses consisting of a nanopore technology-based sequencer, the MinION, and laptop computers, and assessed its potential ability to determine bacterial compositions rapidly. We tested our protocols using a mock bacterial community that contained equimolar 16S rDNA and a pleural effusion from a patient with empyema, for time effectiveness and accuracy. MinION sequencing targeting 16S rDNA detected all 20 of the bacterial species present in the mock bacterial community...
July 18, 2017: Scientific Reports
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