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SMRT sequencing

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https://www.readbyqxmd.com/read/28181326/single-molecule-rna-sequencing-uncovers-trans-splicing-and-improves-annotations-in-anopheles-stephensi
#1
X Jiang, A B Hall, J K Biedler, Z Tu
Single molecule real-time (SMRT) sequencing has recently been used to obtain full-length cDNA sequences that improve genome annotation and reveal RNA isoforms. Here, we used one such method called isoform sequencing from Pacific Biosciences (PacBio) to sequence a cDNA library from the Asian malaria mosquito Anopheles stephensi. More than 600 000 full-length cDNAs, referred to as reads of insert, were identified. Owing to the inherently high error rate of PacBio sequencing, we tested different approaches for error correction...
February 9, 2017: Insect Molecular Biology
https://www.readbyqxmd.com/read/28159629/a-quantitative-smrt-cell-sequencing-method-for-ribosomal-amplicons
#2
Bethan M Jones, Adam B Kustka
Advances in sequencing technologies continue to provide unprecedented opportunities to characterize microbial communities. For example, the Pacific Biosciences Single Molecule Real-Time (SMRT) platform has emerged as a unique approach harnessing DNA polymerase activity to sequence template molecules, enabling long reads at low costs. With the aim to simultaneously classify and enumerate in situ microbial populations, we developed a quantitative SMRT (qSMRT) approach that combines exogenous standard addition to quantify ribosomal amplicons derived from environmental samples...
January 31, 2017: Journal of Microbiological Methods
https://www.readbyqxmd.com/read/28152050/draft-genome-sequence-of-karnal-bunt-pathogen-tilletia-indica-of-wheat-provides-insights-into-the-pathogenic-mechanisms-of-quarantined-fungus
#3
Anil Kumar, Vishakha Pandey, Manoj Singh, Dinesh Pandey, M S Saharan, Soma S Marla
Karnal bunt disease in wheat is caused by hemibiotrophic fungus, Tilletia indica that has been placed as quarantine pest in more than 70 countries. Despite its economic importance, little knowledge about the molecular components of fungal pathogenesis is known. In this study, first time the genome sequence of T. indica has been deciphered for unraveling the effectors' functions of molecular pathogenesis of Karnal bunt disease. The T. indica genome was sequenced employing hybrid approach of PacBio Single Molecule Real Time (SMRT) and Illumina HiSEQ 2000 sequencing platforms...
2017: PloS One
https://www.readbyqxmd.com/read/28143951/single-molecule-sequencing-of-the-drosophila-serrata-genome
#4
Scott L Allen, Emily K Delaney, Artyom Kopp, Stephen F Chenoweth
Long read sequencing technology promises to greatly enhance de novo assembly of genomes for non-model species. Although the high error rates of long reads have been a stumbling block, sequencing at high coverage permits the self-correction of many errors. Here we sequence and de novo assemble the genome of Drosophila serrata, a species from the montium subgroup that has been well studied for latitudinal clines, sexual selection, and gene expression but which lacks a reference genome. Using 11 PacBio SMRT cells, we generated 12 Gbp of raw sequence data comprising approximately 65x whole genome coverage...
January 30, 2017: G3: Genes—Genomes—Genetics
https://www.readbyqxmd.com/read/28138840/pacbio-for-haplotyping-in-gene-families
#5
Wei Zhang, Joachim Messing
The throughput and read length provided by Pacific Bioscience (PacBio) Single Molecule Real Time (SMRT) sequencing platform makes it feasible to construct contiguous, non-chimeric sequences. This is especially useful for genes with repetitive sequences in their gene bodies in gene families. We illustrate the use of PacBio to sequence and assemble hundreds of transcripts of gluten gene families from different cultivars of wheat using sequence from a single SMRT cell. To this end, we barcoded amplicons from different cultivars, then pooled these into one library for sequencing...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28124151/the-helicobacter-pylori-methylome-roles-in-gene-regulation-and-virulence
#6
Rebecca Gorrell, Terry Kwok
The methylome is defined as a map of DNA methylation patterns at single-base resolution. DNA methylation in bacteria was first discovered as a function of restriction-modification (R-M) systems. R-M systems in Helicobacter pylori, like those in other bacteria, are important host-specificity determinants that provide protection against foreign DNA. Moreover, the gene regulatory role of the methyltransferase (Mtase) unit of various Helicobacter pylori R-M systems is being increasingly recognized. Recent advances in the application of single-molecule real-time (SMRT) DNA sequencing to analyse DNA methylation have revealed for the first time comprehensive pictures of the genome-wide distribution of methylation sites in various strains of H...
2017: Current Topics in Microbiology and Immunology
https://www.readbyqxmd.com/read/28078358/major-histocompatibility-complex-haplotyping-and-long-amplicon-allele-discovery-in-cynomolgus-macaques-from-chinese-breeding-facilities
#7
Julie A Karl, Michael E Graham, Roger W Wiseman, Katelyn E Heimbruch, Samantha M Gieger, Gaby G M Doxiadis, Ronald E Bontrop, David H O'Connor
Very little is currently known about the major histocompatibility complex (MHC) region of cynomolgus macaques (Macaca fascicularis; Mafa) from Chinese breeding centers. We performed comprehensive MHC class I haplotype analysis of 100 cynomolgus macaques from two different centers, with animals from different reported original geographic origins (Vietnamese, Cambodian, and Cambodian/Indonesian mixed-origin). Many of the samples were of known relation to each other (sire, dam, and progeny sets), making it possible to characterize lineage-level haplotypes in these animals...
January 11, 2017: Immunogenetics
https://www.readbyqxmd.com/read/28077634/characterization-of-hcv-envelope-diversification-from-acute-to-chronic-infection-using-smrt-sequencing-within-a-sexually-transmitted-hepatitis-c-virus-cluster
#8
Cynthia K Y Ho, Jayna Raghwani, Sylvie Koekkoek, Richard H Liang, Jan T M Van der Meer, Marc Van Der Valk, Menno De Jong, Oliver G Pybus, Janke Schinkel, Richard Molenkamp
: In contrast to other available next generation sequencing platforms, Pacbio Single Molecule, Real-Time (SMRT) sequencing has the advantage of generating long reads, albeit with a relatively higher error rate in unprocessed data. Using this platform we longitudinally sampled and sequenced the hepatitis C virus (HCV) envelope genome region (1680 nt) from individuals belonging to a cluster of sexually-transmitted cases. All five subjects were HIV-1 coinfected and infected with a closely related strain of HCV genotype 4d...
January 11, 2017: Journal of Virology
https://www.readbyqxmd.com/read/28068902/bacterial-microbiota-of-kazakhstan-cheese-revealed-by-single-molecule-real-time-smrt-sequencing-and-its-comparison-with-belgian-kalmykian-and-italian-artisanal-cheeses
#9
Jing Li, Yi Zheng, Haiyan Xu, Xiaoxia Xi, Qiangchuan Hou, Shuzhen Feng, Laga Wuri, Yanfei Bian, Zhongjie Yu, Lai-Yu Kwok, Zhihong Sun, Tiansong Sun
BACKGROUND: In Kazakhstan, traditional artisanal cheeses have a long history and are widely consumed. The unique characteristics of local artisanal cheeses are almost completely preserved. However, their microbial communities have rarely been reported. The current study firstly generated the Single Molecule, Real-Time (SMRT) sequencing bacterial diversity profiles of 6 traditional artisanal cheese samples of Kazakhstan origin, followed by comparatively analyzed the microbiota composition between the current dataset and those from cheeses originated from Belgium, Russian Republic of Kalmykia (Kalmykia) and Italy...
January 9, 2017: BMC Microbiology
https://www.readbyqxmd.com/read/28068217/manual-curation-and-reannotation-of-the-genomes-of-clostridium-difficile-630%C3%AE-erm-and-clostridium-difficile-630
#10
Henning Dannheim, Thomas Riedel, Meina Neumann-Schaal, Boyke Bunk, Isabel Schober, Cathrin Spröer, Cynthia Maria Chibani, Sabine Gronow, Heiko Liesegang, Jörg Overmann, Dietmar Schomburg
We resequenced the genome of Clostridium difficile 630Δerm (DSM 28645), a model strain commonly used for the generation of insertion mutants. The genome sequence was obtained by a combination of single-molecule real-time (SMRT) and Illumina sequencing technology. Detailed manual curation and comparison to the previously published genomic sequence revealed sequence differences including inverted regions and the presence of plasmid pCD630. Manual curation of our previously deposited genome sequence of the parental strain 630 (DSM 27543) led to an improved genome sequence...
January 9, 2017: Journal of Medical Microbiology
https://www.readbyqxmd.com/read/28061749/organelle_pba-a-pipeline-for-assembling-chloroplast-and-mitochondrial-genomes-from-pacbio-dna-sequencing-data
#11
Aboozar Soorni, David Haak, David Zaitlin, Aureliano Bombarely
BACKGROUND: The development of long-read sequencing technologies, such as single-molecule real-time (SMRT) sequencing by PacBio, has produced a revolution in the sequencing of small genomes. Sequencing organelle genomes using PacBio long-read data is a cost effective, straightforward approach. Nevertheless, the availability of simple-to-use software to perform the assembly from raw reads is limited at present. RESULTS: We present Organelle-PBA, a Perl program designed specifically for the assembly of chloroplast and mitochondrial genomes...
January 7, 2017: BMC Genomics
https://www.readbyqxmd.com/read/28056367/bacterial-community-structure-in-simultaneous-nitrification-denitrification-and-organic-matter-removal-process-treating-saline-mustard-tuber-wastewater-as-revealed-by-16s-rrna-sequencing
#12
Jiale Wang, Benzhou Gong, Wei Huang, Yingmu Wang, Jian Zhou
A simultaneous nitrification, denitrification and organic matter removal (SNDOR) process in sequencing batch biofilm reactor (SBBR) was established to treat saline mustard tuber wastewater (MTWW) in this study. An average COD removal efficiency of 86.48% and total nitrogen removal efficiency of 86.48% were achieved at 30gNaClL(-1) during 100days' operation. The underlying mechanisms were investigated by PacBio SMRT DNA sequencing (V1-V9) to analyze the microbial community structures and its variation from low salinity at 10gNaClL(-1) to high salinity at 30gNaClL(-1)...
March 2017: Bioresource Technology
https://www.readbyqxmd.com/read/27998939/a-pipeline-for-local-assembly-of-minisatellite-alleles-from-single-molecule-sequencing-data
#13
Denye Ogeh, Richard Badge
MOTIVATION: The advent of Next Generation Sequencing (NGS) has led to the generation of enormous volumes of short read sequence data, cheaply and in reasonable time scales. Nevertheless, the quality of genome assemblies generated using NGS technologies has been greatly affected, compared to those generated using Sanger DNA sequencing. This is largely due to the inability of short read sequence data to scaffold repetitive structures, creating gaps, inversions and rearrangements and resulting in assemblies that are, at best, draft forms...
December 20, 2016: Bioinformatics
https://www.readbyqxmd.com/read/27997733/global-identification-of-alternative-splicing-via-comparative-analysis-of-smrt-and-illumina-based-rna-seq-in-strawberry
#14
Yongping Li, Cheng Dai, Chungen Hu, Zhongchi Liu, Chunying Kang
Alternative splicing (AS) is a key post-transcriptional regulatory mechanism, yet little is known about its roles in fruit crops. Here, AS was globally analyzed in the wild strawberry Fragaria vesca genome with RNA-seq data derived from different stages of fruit development. The AS landscape was characterized and compared between the single-molecule, real-time (SMRT) and Illumina RNA-seq platform. While SMRT has a lower sequencing depth, it identifies more genes undergoing AS (57.67% of detected multiexon genes) when it is compared with Illumina (33...
December 20, 2016: Plant Journal: for Cell and Molecular Biology
https://www.readbyqxmd.com/read/27997543/cell-cycle-constraints-and-environmental-control-of-local-dna-hypomethylation-in-%C3%AE-proteobacteria
#15
Silvia Ardissone, Peter Redder, Giancarlo Russo, Antonio Frandi, Coralie Fumeaux, Andrea Patrignani, Ralph Schlapbach, Laurent Falquet, Patrick H Viollier
Heritable DNA methylation imprints are ubiquitous and underlie genetic variability from bacteria to humans. In microbial genomes, DNA methylation has been implicated in gene transcription, DNA replication and repair, nucleoid segregation, transposition and virulence of pathogenic strains. Despite the importance of local (hypo)methylation at specific loci, how and when these patterns are established during the cell cycle remains poorly characterized. Taking advantage of the small genomes and the synchronizability of α-proteobacteria, we discovered that conserved determinants of the cell cycle transcriptional circuitry establish specific hypomethylation patterns in the cell cycle model system Caulobacter crescentus...
December 2016: PLoS Genetics
https://www.readbyqxmd.com/read/27966108/membrane-fusion-and-infection-of-the-influenza-hemagglutinin
#16
Sean T Smrt, Justin L Lorieau
The influenza virus is a major health concern associated with an estimated 5000 to 30,000 deaths every year (Reed et al. 2015) and a significant economic impact with the development of treatments, vaccinations and research (Molinari et al. 2007). The entirety of the influenza genome is comprised of only eleven coding genes. An enormous degree of variation in non-conserved regions leads to significant challenges in the development of inclusive inhibitors for treatment. The fusion peptide domain of the influenza A hemagglutinin (HA) is a promising candidate for treatment since it is one of the most highly conserved sequences in the influenza genome (Heiny et al...
December 14, 2016: Advances in Experimental Medicine and Biology
https://www.readbyqxmd.com/read/27964731/use-of-single-molecule-sequencing-for-comparative-genomics-of-an-environmental-and-a-clinical-isolate-of-clostridium-difficile-ribotype-078
#17
Katherine R Hargreaves, Anisha M Thanki, Bethany R Jose, Marco R Oggioni, Martha R J Clokie
BACKGROUND: How the pathogen Clostridium difficile might survive, evolve and be transferred between reservoirs within the natural environment is poorly understood. Some ribotypes are found both in clinical and environmental settings. Whether these strains are distinct from each another and evolve in the specific environments is not established. The possession of a highly mobile genome has contributed to the genetic diversity and ongoing evolution of C. difficile. Interpretations of genetic diversity have been limited by fragmented assemblies resulting from short-read length sequencing approaches and by a limited understanding of epigenetic regulation of diversity...
December 13, 2016: BMC Genomics
https://www.readbyqxmd.com/read/27940952/hapcut2-robust-and-accurate-haplotype-assembly-for-diverse-sequencing-technologies
#18
Peter Edge, Vineet Bafna, Vikas Bansal
Many tools have been developed for haplotype assembly - the reconstruction of individual haplotypes using reads mapped to a reference genome sequence. Due to increasing interest in obtaining haplotype-resolved human genomes, a range of new sequencing protocols and technologies have been developed to enable the reconstruction of whole-genome haplotypes. However, existing computational methods designed to handle specific technologies do not scale well on data from different protocols. We describe a new algorithm, HapCUT2, that extends our previous method (HapCUT) to handle multiple sequencing technologies...
December 9, 2016: Genome Research
https://www.readbyqxmd.com/read/27924023/methsmrt-an-integrative-database-for-dna-n6-methyladenine-and-n4-methylcytosine-generated-by-single-molecular-real-time-sequencing
#19
Pohao Ye, Yizhao Luan, Kaining Chen, Yizhi Liu, Chuanle Xiao, Zhi Xie
DNA methylation is an important type of epigenetic modifications, where 5- methylcytosine (5mC), 6-methyadenine (6mA) and 4-methylcytosine (4mC) are the most common types. Previous efforts have been largely focused on 5mC, providing invaluable insights into epigenetic regulation through DNA methylation. Recently developed single-molecule real-time (SMRT) sequencing technology provides a unique opportunity to detect the less studied DNA 6mA and 4mC modifications at single-nucleotide resolution. With a rapidly increased amount of SMRT sequencing data generated, there is an emerging demand to systematically explore DNA 6mA and 4mC modifications from these data sets...
January 4, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/27904456/methylome-analysis-of-two-xanthomonas-spp-using-single-molecule-real-time-sequencing
#20
Hoon Je Seong, Hye-Jee Park, Eunji Hong, Sung Chul Lee, Woo Jun Sul, Sang-Wook Han
Single-molecule real-time (SMRT) sequencing allows identification of methylated DNA bases and methylation patterns/motifs at the genome level. Using SMRT sequencing, diverse bacterial methylomes including those of Helicobacter pylori, Lactobacillus spp., and Escherichia coli have been determined, and previously unreported DNA methylation motifs have been identified. However, the methylomes of Xanthomonas species, which belong to the most important plant pathogenic bacterial genus, have not been documented. Here, we report the methylomes of Xanthomonas axonopodis pv...
December 2016: Plant Pathology Journal
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