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https://www.readbyqxmd.com/read/29431738/nanopore-sequencing-and-assembly-of-a-human-genome-with-ultra-long-reads
#1
Miten Jain, Sergey Koren, Karen H Miga, Josh Quick, Arthur C Rand, Thomas A Sasani, John R Tyson, Andrew D Beggs, Alexander T Dilthey, Ian T Fiddes, Sunir Malla, Hannah Marriott, Tom Nieto, Justin O'Grady, Hugh E Olsen, Brent S Pedersen, Arang Rhie, Hollian Richardson, Aaron R Quinlan, Terrance P Snutch, Louise Tee, Benedict Paten, Adam M Phillippy, Jared T Simpson, Nicholas J Loman, Matthew Loose
We report the sequencing and assembly of a reference genome for the human GM12878 Utah/Ceph cell line using the MinION (Oxford Nanopore Technologies) nanopore sequencer. 91.2 Gb of sequence data, representing ∼30× theoretical coverage, were produced. Reference-based alignment enabled detection of large structural variants and epigenetic modifications. De novo assembly of nanopore reads alone yielded a contiguous assembly (NG50 ∼3 Mb). We developed a protocol to generate ultra-long reads (N50 > 100 kb, read lengths up to 882 kb)...
January 29, 2018: Nature Biotechnology
https://www.readbyqxmd.com/read/29416032/high-contiguity-arabidopsis-thaliana-genome-assembly-with-a-single-nanopore-flow-cell
#2
Todd P Michael, Florian Jupe, Felix Bemm, S Timothy Motley, Justin P Sandoval, Christa Lanz, Olivier Loudet, Detlef Weigel, Joseph R Ecker
The handheld Oxford Nanopore MinION sequencer generates ultra-long reads with minimal cost and time requirements, which makes sequencing genomes at the bench feasible. Here, we sequence the gold standard Arabidopsis thaliana genome (KBS-Mac-74 accession) on the bench with the MinION sequencer, and assemble the genome using typical consumer computing hardware (4 Cores, 16 Gb RAM) into chromosome arms (62 contigs with an N50 length of 12.3 Mb). We validate the contiguity and quality of the assembly with two independent single-molecule technologies, Bionano optical genome maps and Pacific Biosciences Sequel sequencing...
February 7, 2018: Nature Communications
https://www.readbyqxmd.com/read/29386054/myoblast-fusion-confusion-the-resolution-begins
#3
REVIEW
Srihari C Sampath, Srinath C Sampath, Douglas P Millay
The fusion of muscle precursor cells is a required event for proper skeletal muscle development and regeneration. Numerous proteins have been implicated to function in myoblast fusion; however, the majority are expressed in diverse tissues and regulate numerous cellular processes. How myoblast fusion is triggered and coordinated in a muscle-specific manner has remained a mystery for decades. Through the discovery of two muscle-specific fusion proteins, Myomaker and Myomerger-Minion, we are now primed to make significant advances in our knowledge of myoblast fusion...
January 31, 2018: Skeletal Muscle
https://www.readbyqxmd.com/read/29382965/development-and-optimization-of-a-cell-associated-challenge-model-for-mycoplasma-hyorhinis-in-7-week-old-cesarean-derived-colostrum-deprived-pigs
#4
Brian Martinson, F Chris Minion, Dianna Jordan
Mycoplasma hyorhinis (MHR) causes polyserositis and lameness in grower pigs. While herd-specific vaccines for this bacterium are being marketed, there are currently no licensed, commercially available vaccines for MHR. The objective of this study was to develop a challenge model in cesarean-derived, colostrum-deprived (CDCD) pigs using cell-associated MHR that results in both severe pericarditis and lameness, in order to evaluate suitable vaccine candidates. We investigated administering MHR to 7-week-old pigs over 3 d using 3 different routes compared to administering MHR on a single day using 1 of 3 routes...
January 2018: Canadian Journal of Veterinary Research, Revue Canadienne de Recherche Vétérinaire
https://www.readbyqxmd.com/read/29374775/serogroup-level-resolution-of-the-super-7-shiga-toxin-producing-escherichia-coli-using-nanopore-single-molecule-dna-sequencing
#5
Adam Peritz, George C Paoli, Chin-Yi Chen, Andrew G Gehring
DNA sequencing and other DNA-based methods are now broadly used for detection and identification of bacterial foodborne pathogens. For the identification of foodborne bacterial pathogens, taxonomic assignments must be made to the species or even subspecies level. Long-read DNA sequencing provides finer taxonomic resolution than short-read sequencing. Here, we demonstrate the potential of long-read shotgun sequencing obtained from the Oxford Nanopore Technologies (ONT) MinION single-molecule sequencer, in combination with the Basic Local Alignment Search Tool (BLAST) with custom sequence databases, for foodborne pathogen identification...
January 27, 2018: Analytical and Bioanalytical Chemistry
https://www.readbyqxmd.com/read/29325570/de-novo-assembly-of-the-complex-genome-of-nippostrongylus-brasiliensis-using-minion-long-reads
#6
David Eccles, Jodie Chandler, Mali Camberis, Bernard Henrissat, Sergey Koren, Graham Le Gros, Jonathan J Ewbank
BACKGROUND: Eukaryotic genome assembly remains a challenge in part due to the prevalence of complex DNA repeats. This is a particularly acute problem for holocentric nematodes because of the large number of satellite DNA sequences found throughout their genomes. These have been recalcitrant to most genome sequencing methods. At the same time, many nematodes are parasites and some represent a serious threat to human health. There is a pressing need for better molecular characterization of animal and plant parasitic nematodes...
January 11, 2018: BMC Biology
https://www.readbyqxmd.com/read/29325009/efficient-generation-of-complete-sequences-of-mdr-encoding-plasmids-by-rapid-assembly-of-minion-barcoding-sequencing-data
#7
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/29321680/cryptic-transmission-of-st405-escherichia-coli-carrying-bla-ndm-4-in-hospital
#8
Xiaoxia Zhang, Yu Feng, Weilong Zhou, Alan McNally, Zhiyong Zong
Three carbapenem-resistant Escherichia coli were recovered from rectal swabs of different patients in a tertiary hospital and were found carrying bla NDM-4, an uncommon bla NDM variant. Genome sequences of the isolates were obtained using Illumina technology and the long-read MinION sequencer. The isolates belonged to ST405 and phylogenetic group D, a globally distributed lineage associated with antimicrobial resistance. In addition to bla NDM-4, the three isolates carried 14 known resistance genes including the extended-spectrum β-lactamase gene bla CTX-M-15...
January 10, 2018: Scientific Reports
https://www.readbyqxmd.com/read/29315344/culture-independent-analysis-of-liver-abscess-using-nanopore-sequencing
#9
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/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/29237003/illumina-short-read-and-minion-long-read-wgs-to-characterize-the-molecular-epidemiology-of-an-ndm-1-serratia-marcescens-outbreak-in-romania
#12
H T T Phan, N Stoesser, I E Maciuca, F Toma, E Szekely, M Flonta, A T M Hubbard, L Pankhurst, T Do, T E A Peto, A S Walker, D W Crook, D Timofte
Background and Objectives: Serratia marcescens is an emerging nosocomial pathogen, and the carbapenemase blaNDM has been reported in several surveys in Romania. We aimed to investigate the molecular epidemiology of S. marcescens in two Romanian hospitals over 2010-15, including a neonatal NDM-1 S. marcescens outbreak. Methods: Isolates were sequenced using Illumina technology together with carbapenem-non-susceptible NDM-1-positive and NDM-1-negative Klebsiella pneumoniae and Enterobacter cloacae to provide genomic context...
December 8, 2017: Journal of Antimicrobial Chemotherapy
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/29221894/efficacy-of-an-inactivated-mycoplasma-hyorhinis-vaccine-in-pigs
#14
Brian Martinson, Whitney Zoghby, Kenneth Barrett, Lawrence Bryson, Rodney Christmas, F C Minion, Jeremy Kroll
Lameness and polyserositis in pigs caused by Mycoplasma hyorhinis are generally treated with antibiotics and may require multiple doses. The costs of these antibiotics combined with economic losses from culling and reduced feed conversion due to lameness are hardships to the swine producer. In this study we have demonstrated efficacy of an inactivated M. hyorhinis vaccine administered to three-week old caesarian-derived colostrum-deprived piglets. Three doses of vaccine (high, medium, and low) were evaluated and compared to a placebo control...
December 5, 2017: Vaccine
https://www.readbyqxmd.com/read/29186559/phyloscanner-inferring-transmission-from-within-and-between-host-pathogen-genetic-diversity
#15
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
#16
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/29182147/rapid-re-identification-of-human-samples-using-portable-dna-sequencing
#17
Sophie Zaaijer, Assaf Gordon, Daniel Speyer, Robert Piccone, Simon Cornelis Groen, Yaniv Erlich
DNA re-identification is used for a broad suite of applications, ranging from cell line authentication to forensics. However, current re-identification schemes suffer from high latency and limited access. Here, we describe a rapid, inexpensive, and portable strategy to robustly re-identify human DNA called 'MinION sketching'. MinION sketching requires as few as 3 min of sequencing and 60-300 random SNPs to re-identify a sample enabling near real-time applications of DNA re-identification. Our method capitalizes on the rapidly growing availability of genomic reference data for cell lines, tissues in biobanks, and individuals...
November 28, 2017: ELife
https://www.readbyqxmd.com/read/29177090/completing-bacterial-genome-assemblies-with-multiplex-minion-sequencing
#18
Ryan R Wick, Louise M Judd, Claire L Gorrie, Kathryn E Holt
Illumina sequencing platforms have enabled widespread bacterial whole genome sequencing. While Illumina data is appropriate for many analyses, its short read length limits its ability to resolve genomic structure. This has major implications for tracking the spread of mobile genetic elements, including those which carry antimicrobial resistance determinants. Fully resolving a bacterial genome requires long-read sequencing such as those generated by Oxford Nanopore Technologies (ONT) platforms. Here we describe our use of the ONT MinION to sequence 12 isolates of Klebsiella pneumoniae on a single flow cell...
October 2017: Microbial Genomics
https://www.readbyqxmd.com/read/29163399/minion-nanopore-sequencing-enables-correlation-between-resistome-phenotype-and-genotype-of-coliform-bacteria-in-municipal-sewage
#19
Yu Xia, An-Dong Li, Yu Deng, Xiao-Tao Jiang, Li-Guan Li, Tong Zhang
Wastewater treatment plants (WWTPs) functioned as the intersection between the human society and nature environment, are receiving increasingly more attention on risk assessment of the acquisition of environmental antibiotic resistance genes (ARGs) by pathogenetic populations during treatment. However, because of the general lack of robust resistome profiling methods, genotype, and resistance phenotype is still poorly correlated in human pathogens of sewage samples. Here we applied MinION sequencing to quantify the resistance genes of multiple antibiotic resistant (MAR) coliform bacteria, a common indicator for human enteric pathogens in sewage samples...
2017: Frontiers in Microbiology
https://www.readbyqxmd.com/read/29136087/the-utility-and-perspectives-of-ngs-based-methods-in-bsl-3-and-bsl-4-laboratory-sequencing-and-analysis-strategies
#20
Tomasz Wolkowicz
Modern diagnostics is in general based on molecular biology methods. Nowadays sequencing-based methods, especially whole genome sequencing, are becoming increasingly important. Implementation of such methods into routine diagnostic of highly dangerous pathogens, like Bacillus anthracis, Francisella tularensis, Yersinia pestis, Ebola virus, MERS, Lassa virus etc. would be very helpful. The best diagnostic strategy would be the metagenomic sequencing directly from the clinical sample. Implementation of majority of currently available WGS platforms inside the BSL-3 or 4 laboratory is impractical because of the size of the equipment and time consuming wet lab part (e...
November 10, 2017: Briefings in Functional Genomics
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