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https://www.readbyqxmd.com/read/27915180/self-primed-isothermal-amplification-for-genomic-dna-detection-of-human-papillomavirus
#1
Wei Lu, Qingpan Yuan, Zhiliu Yang, Bo Yao
Rolling circle amplification (RCA) is an isothermal amplification technique with high efficiency and perfect accuracy for nucleic acids detection. However, RCA technique suffers the limitation to detect short DNA or RNA molecules. For long nucleic acid molecules, enzymatic restriction as well as heat denaturation process is usually required, which makes the amplification not effective and strictly isothermal. In this article, a simple and efficient one-pot self-primed isothermal amplification (SIA) was developed for detection of genomic DNA directly based on the combination of nicking endonuclease assisted strand displacement amplification (SDA) and exponential RCA...
October 10, 2016: Biosensors & Bioelectronics
https://www.readbyqxmd.com/read/27914932/detection-of-human-immunodeficiency-virus-rnas-in-living-cells-using-spinach-rna-aptamers
#2
Brandon D Burch, Carolina Garrido, David M Margolis
Many techniques currently used to measure HIV RNA production in cells suffer from limitations that include high background signal or the potential to destroy cellular context. Fluorophore-binding RNA aptamers offer the potential for visualizing RNAs directly in living cells with minimal cellular perturbation. We inserted a sequence encoding a fluorophore-binding RNA aptamer, known as Spinach, into the HIV genome such that predicted RNA secondary structures in both Spinach and HIV were preserved. Chimeric HIV-Spinach RNAs were functionally validated in vitro by testing their ability to enhance the fluorescence of a conditional fluorophore (DFHBI), which specifically binds Spinach...
November 30, 2016: Virus Research
https://www.readbyqxmd.com/read/27913418/a-novel-rhamnose-rich-hetero-exopolysaccharide-isolated-from-lactobacillus-paracasei-dg-activates-thp-1-human-monocytic-cells
#3
Silvia Balzaretti, Valentina Taverniti, Simone Guglielmetti, Walter Fiore, Mario Minuzzo, Hansel N Ngo, Judith B Ngere, Sohaib Sadiq, Paul N Humphreys, Andrew P Laws
: Lactobacillus paracasei DG is a bacterial strain with recognized probiotic properties and is used in commercial probiotic products. However, the mechanisms underlying its probiotic properties are mainly unknown. In this study, we tested the hypothesis that the capability of strain DG to interact with the host is, at least partly, associated with its ability to synthesize a surface-associated exopolysaccharide (EPS). Comparative genomics revealed the presence of putative EPS gene clusters in DG genome; accordingly, EPS was isolated from the surface of the bacterium...
December 2, 2016: Applied and Environmental Microbiology
https://www.readbyqxmd.com/read/27912729/genomic-insights-from-whole-genome-sequencing-of-four-clonal-outbreak-campylobacter-jejuni-assessed-within-the-global-c-jejuni-population
#4
Clifford G Clark, Chrystal Berry, Matthew Walker, Aaron Petkau, Dillon O R Barker, Cai Guan, Aleisha Reimer, Eduardo N Taboada
BACKGROUND: Whole genome sequencing (WGS) is useful for determining clusters of human cases, investigating outbreaks, and defining the population genetics of bacteria. It also provides information about other aspects of bacterial biology, including classical typing results, virulence, and adaptive strategies of the organism. Cell culture invasion and protein expression patterns of four related multilocus sequence type 21 (ST21) C. jejuni isolates from a significant Canadian water-borne outbreak were previously associated with the presence of a CJIE1 prophage...
December 3, 2016: BMC Genomics
https://www.readbyqxmd.com/read/27911848/global-analysis-of-genomic-instability-caused-by-dna-replication-stress-in-saccharomyces-cerevisiae
#5
Dao-Qiong Zheng, Ke Zhang, Xue-Chang Wu, Piotr A Mieczkowski, Thomas D Petes
DNA replication stress (DRS)-induced genomic instability is an important factor driving cancer development. To understand the mechanisms of DRS-associated genomic instability, we measured the rates of genomic alterations throughout the genome in a yeast strain with lowered expression of the replicative DNA polymerase δ. By a genetic test, we showed that most recombinogenic DNA lesions were introduced during S or G2 phase, presumably as a consequence of broken replication forks. We observed a high rate of chromosome loss, likely reflecting a reduced capacity of the low-polymerase strains to repair double-stranded DNA breaks (DSBs)...
November 28, 2016: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/27911846/dna-methylome-of-the-20-gigabase-norway-spruce-genome
#6
Israel Ausin, Suhua Feng, Chaowei Yu, Wanlu Liu, Hsuan Yu Kuo, Elise L Jacobsen, Jixian Zhai, Javier Gallego-Bartolome, Lin Wang, Ulrika Egertsdotter, Nathaniel R Street, Steven E Jacobsen, Haifeng Wang
DNA methylation plays important roles in many biological processes, such as silencing of transposable elements, imprinting, and regulating gene expression. Many studies of DNA methylation have shown its essential roles in angiosperms (flowering plants). However, few studies have examined the roles and patterns of DNA methylation in gymnosperms. Here, we present genome-wide high coverage single-base resolution methylation maps of Norway spruce (Picea abies) from both needles and somatic embryogenesis culture cells via whole genome bisulfite sequencing...
November 28, 2016: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/27910942/improved-bi-allelic-modification-of-a-transcriptionally-silent-locus-in-patient-derived-ipsc-by-cas9-nickase
#7
Reto Eggenschwiler, Mohsen Moslem, Mariane Serra Fráguas, Melanie Galla, Oliver Papp, Maximilian Naujock, Ines Fonfara, Ingrid Gensch, Annabell Wähner, Abbas Beh-Pajooh, Claudio Mussolino, Marcel Tauscher, Doris Steinemann, Florian Wegner, Susanne Petri, Axel Schambach, Emmanuelle Charpentier, Toni Cathomen, Tobias Cantz
Homology directed repair (HDR)-based genome editing via selectable long flanking arm donors can be hampered by local transgene silencing at transcriptionally silent loci. Here, we report efficient bi-allelic modification of a silent locus in patient-derived hiPSC by using Cas9 nickase and a silencing-resistant donor construct that contains an excisable selection/counter-selection cassette. To identify the most active single guide RNA (sgRNA)/nickase combinations, we employed a lentiviral vector-based reporter assay to determine the HDR efficiencies in cella...
December 2, 2016: Scientific Reports
https://www.readbyqxmd.com/read/27910009/cancer-cytogenetics-an-introduction
#8
Thomas S K Wan
The Philadelphia chromosome was the first chromosomal abnormality discovered in cancer using the cytogenetics technique in 1960, and was consistently associated with chronic myeloid leukemia. Over the past five decades, innovative technical advances in the field of cancer cytogenetics have greatly enhanced the detection ability of chromosomal alterations, and have facilitated the research and diagnostic potential of chromosomal studies in neoplasms. These developments notwithstanding, chromosome analysis of a single cell is still the easiest way to delineate and understand the relationship between clonal evolution and disease progression of cancer cells...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/27906459/integrative-genomic-analysis-identifies-ancestry-related-expression-quantitative-trait-loci-on-dna-polymerase-%C3%AE-and-supports-the-association-of-genetic-ancestry-with-survival-disparities-in-head-and-neck-squamous-cell-carcinoma
#9
Meganathan P Ramakodi, Karthik Devarajan, Elizabeth Blackman, Denise Gibbs, Danièle Luce, Jacqueline Deloumeaux, Suzy Duflo, Jeffrey C Liu, Ranee Mehra, Rob J Kulathinal, Camille C Ragin
BACKGROUND: African Americans with head and neck squamous cell carcinoma (HNSCC) have a lower survival rate than whites. This study investigated the functional importance of ancestry-informative single-nucleotide polymorphisms (SNPs) in HNSCC and also examined the effect of functionally important genetic elements on racial disparities in HNSCC survival. METHODS: Ancestry-informative SNPs, RNA sequencing, methylation, and copy number variation data for 316 oral cavity and laryngeal cancer patients were analyzed across 178 DNA repair genes...
December 1, 2016: Cancer
https://www.readbyqxmd.com/read/27906190/robust-high-throughput-kinetic-analysis-of-apoptosis-with-real-time-high-content-live-cell-imaging
#10
Jesse D Gelles, Jerry Edward Chipuk
Quantitative and kinetic analyses of apoptotic cell death are integral components of exploring cell biology, measuring cellular stress responses, and performing high-throughput genomic/RNAi/drug screens. Here, we present a detailed method that integrates robust kinetic real-time high-content imaging with Annexin V labelling to provide a highly sensitive, accurate, simple and zero-handling approach to quantify extrinsic and intrinsic inducers of apoptosis. The sensitivity of this non-toxic method outperforms previous high-throughput methodologies using viability dyes or caspase-activation reporters...
December 1, 2016: Cell Death & Disease
https://www.readbyqxmd.com/read/27906046/chromatin-landscapes-and-genetic-risk-in-systemic-lupus
#11
Joyce S Hui-Yuen, Lisha Zhu, Lai Ping Wong, Kaiyu Jiang, Yanmin Chen, Tao Liu, James N Jarvis
BACKGROUND: Systemic lupus erythematosus (SLE) is a multi-system, complex disease in which the environment interacts with inherited genes to produce broad phenotypes with inter-individual variability. Of 46 single nucleotide polymorphisms (SNPs) shown to confer genetic risk for SLE in recent genome-wide association studies, 30 lie within noncoding regions of the human genome. We therefore sought to identify and describe the functional elements (aside from genes) located within these regions of interest...
December 1, 2016: Arthritis Research & Therapy
https://www.readbyqxmd.com/read/27905878/genomic-prediction-using-preselected-dna-variants-from-a-gwas-with-whole-genome-sequence-data-in-holstein-friesian-cattle
#12
Roel F Veerkamp, Aniek C Bouwman, Chris Schrooten, Mario P L Calus
BACKGROUND: Whole-genome sequence data is expected to capture genetic variation more completely than common genotyping panels. Our objective was to compare the proportion of variance explained and the accuracy of genomic prediction by using imputed sequence data or preselected SNPs from a genome-wide association study (GWAS) with imputed whole-genome sequence data. METHODS: Phenotypes were available for 5503 Holstein-Friesian bulls. Genotypes were imputed up to whole-genome sequence (13,789,029 segregating DNA variants) by using run 4 of the 1000 bull genomes project...
December 1, 2016: Genetics, Selection, Evolution: GSE
https://www.readbyqxmd.com/read/27905063/efficient-gene-targeting-in-mouse-zygotes-mediated-by-crispr-cas9-protein
#13
Chris J Jung, Junli Zhang, Elizabeth Trenchard, Kent C Lloyd, David B West, Barry Rosen, Pieter J de Jong
The CRISPR/Cas9 system has rapidly advanced targeted genome editing technologies. However, its efficiency in targeting with constructs in mouse zygotes via homology directed repair (HDR) remains low. Here, we systematically explored optimal parameters for targeting constructs in mouse zygotes via HDR using mouse embryonic stem cells as a model system. We characterized several parameters, including single guide RNA cleavage activity and the length and symmetry of homology arms in the construct, and we compared the targeting efficiency between Cas9, Cas9nickase, and dCas9-FokI...
November 30, 2016: Transgenic Research
https://www.readbyqxmd.com/read/27904811/legionella-shows-a-diverse-secondary-metabolism-dependent-on-a-broad-spectrum-sfp-type-phosphopantetheinyl-transferase
#14
Nicholas J Tobias, Tilman Ahrendt, Ursula Schell, Melissa Miltenberger, Hubert Hilbi, Helge B Bode
Several members of the genus Legionella cause Legionnaires' disease, a potentially debilitating form of pneumonia. Studies frequently focus on the abundant number of virulence factors present in this genus. However, what is often overlooked is the role of secondary metabolites from Legionella. Following whole genome sequencing, we assembled and annotated the Legionella parisiensis DSM 19216 genome. Together with 14 other members of the Legionella, we performed comparative genomics and analysed the secondary metabolite potential of each strain...
2016: PeerJ
https://www.readbyqxmd.com/read/27904648/editing-of-the-urease-gene-by-crispr-cas-in-the-diatom-thalassiosira-pseudonana
#15
Amanda Hopes, Vladimir Nekrasov, Sophien Kamoun, Thomas Mock
BACKGROUND: CRISPR-Cas is a recent and powerful addition to the molecular toolbox which allows programmable genome editing. It has been used to modify genes in a wide variety of organisms, but only two alga to date. Here we present a methodology to edit the genome of Thalassiosira pseudonana, a model centric diatom with both ecological significance and high biotechnological potential, using CRISPR-Cas. RESULTS: A single construct was assembled using Golden Gate cloning...
2016: Plant Methods
https://www.readbyqxmd.com/read/27903987/mapping-heterogeneity-in-patient-derived-melanoma-cultures-by-single-cell-rna-seq
#16
Tobias Gerber, Edith Willscher, Henry Loeffler-Wirth, Lydia Hopp, Dirk Schadendorf, Manfred Schartl, Ulf Anderegg, Gray Camp, Barbara Treutlein, Hans Binder, Manfred Kunz
Recent technological advances in single-cell genomics make it possible to analyze cellular heterogeneity of tumor samples. Here, we applied single-cell RNA-seq to measure the transcriptomes of 307 single cells cultured from three biopsies of three different patients with a BRAF/NRAS wild type, BRAF mutant/NRAS wild type and BRAF wild type/NRAS mutant melanoma metastasis, respectively. Analysis based on self-organizing maps identified sub-populations defined by multiple gene expression modules involved in proliferation, oxidative phosphorylation, pigmentation and cellular stroma...
November 26, 2016: Oncotarget
https://www.readbyqxmd.com/read/27903644/a-reference-data-set-of-5-4-million-phased-human-variants-validated-by-genetic-inheritance-from-sequencing-a-three-generation-17-member-pedigree
#17
Michael A Eberle, Epameinondas Fritzilas, Peter Krusche, Morten Källberg, Benjamin L Moore, Mitchell A Bekritsky, Zamin Iqbal, Han-Yu Chuang, Sean J Humphray, Aaron L Halpern, Semyon Kruglyak, Elliott H Margulies, Gil McVean, David R Bentley
Improvement of variant calling in next-generation sequence data requires a comprehensive, genome-wide catalog of high-confidence variants called in a set of genomes for use as a benchmark. We generated deep, whole-genome sequence data of 17 individuals in a three-generation pedigree and called variants in each genome using a range of currently available algorithms. We used haplotype transmission information to create a phased "Platinum" variant catalog of 4.7 million single-nucleotide variants (SNVs) plus 0...
November 30, 2016: Genome Research
https://www.readbyqxmd.com/read/27902413/methanosarcina-acetivorans-utilizes-a-single-nadph-dependent-thioredoxin-system-and-contains-additional-thioredoxin-homologs-with-distinct-functions
#18
Addison C McCarver, Faith H Lessner, Jose M Soroeta, Daniel J Lessner
The thioredoxin system plays a central role in the intracellular redox maintenance in the majority of cells. The canonical system is comprised of an NADPH-dependent thioredoxin reductase (TrxR) that reduces the disulfide reductase thioredoxin (Trx). Although Trx is encoded in almost all sequenced genomes of methanogens, its incorporation into their unique physiology is not well understood. Methanosarcina acetivorans contains a single TrxR (MaTrxR) and seven Trx (MaTrx1-7) homologs. We previously showed that MaTrxR and at least MaTrx7 comprise a functional NADPH-dependent thioredoxin system...
November 29, 2016: Microbiology
https://www.readbyqxmd.com/read/27901321/copy-number-variants-in-a-population-based-investigation-of-klippel-trenaunay-syndrome
#19
Aggeliki Dimopoulos, Robert J Sicko, Denise M Kay, Shannon L Rigler, Ruzong Fan, Paul A Romitti, Marilyn L Browne, Charlotte M Druschel, Michele Caggana, Lawrence C Brody, James L Mills
Klippel-Trenaunay syndrome (KTS) is a rare congenital vascular disorder that is thought to occur sporadically; however, reports of familial occurrence suggest a genetic component. We examined KTS cases to identify novel, potentially causal copy number variants (CNVs). We identified 17 KTS cases from all live-births occurring in New York (1998-2010). Extracted DNA was genotyped using Illumina microarrays and CNVs were called using PennCNV software. CNVs selected for follow-up had ≥10 single nucleotide polymorphisms (SNPs) and minimal overlap with in-house controls or controls from the Database of Genomic Variants...
November 30, 2016: American Journal of Medical Genetics. Part A
https://www.readbyqxmd.com/read/27901108/survey-of-the-green-picoalga-bathycoccus-genomes-in-the-global-ocean
#20
Thomas Vannier, Jade Leconte, Yoann Seeleuthner, Samuel Mondy, Eric Pelletier, Jean-Marc Aury, Colomban de Vargas, Michael Sieracki, Daniele Iudicone, Daniel Vaulot, Patrick Wincker, Olivier Jaillon
Bathycoccus is a cosmopolitan green micro-alga belonging to the Mamiellophyceae, a class of picophytoplankton that contains important contributors to oceanic primary production. A single species of Bathycoccus has been described while the existence of two ecotypes has been proposed based on metagenomic data. A genome is available for one strain corresponding to the described phenotype. We report a second genome assembly obtained by a single cell genomics approach corresponding to the second ecotype. The two Bathycoccus genomes are divergent enough to be unambiguously distinguishable in whole DNA metagenomic data although they possess identical sequence of the 18S rRNA gene including in the V9 region...
November 30, 2016: Scientific Reports
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