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Single-cell genomics

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https://www.readbyqxmd.com/read/28531161/development-of-optimized-inhibitor-rnas-allowing-multisite-targeting-of-the-hcv-genome
#1
Cristina Romero-López, Thomas Lahlali, Beatriz Berzal-Herranz, Alfredo Berzal-Herranz
Engineered multivalent drugs are promising candidates for fighting infection by highly variable viruses, such as HCV. The combination into a single molecule of more than one inhibitory domain, each with its own target specificity and even a different mechanism of action, results in drugs with potentially enhanced therapeutic properties. In the present work, the anti-HCV chimeric inhibitor RNA HH363-10, which has a hammerhead catalytic domain and an aptamer RNA domain, was subjected to an in vitro selection strategy to isolate ten different optimised chimeric inhibitor RNAs...
May 22, 2017: Molecules: a Journal of Synthetic Chemistry and Natural Product Chemistry
https://www.readbyqxmd.com/read/28530246/short-single-stranded-dna-degradation-products-augment-the-activation-of-toll-like-receptor-9
#2
Jelka Pohar, Duško Lainšček, Karolina Ivičak-Kocjan, Miša-Mojca Cajnko, Roman Jerala, Mojca Benčina
Toll-like receptors encounter a diversity of degradation products in endosomes. TLR7 and TLR8 have been shown to be activated by RNA degradation products. Here we show that although TLR9 requires single-stranded DNA longer than 20 nucleotides for a robust response, TLR9 activation is augmented by CpG-containing oligodeoxyribonucleotides (sODNs) as short as 2 nucleotides, which, by themselves, do not induce activation in cell cultures, as well as in mice. sODNs also activate human TLR9 in combination with ODNs containing a single CpG motif that by themselves do not activate human TLR9...
May 22, 2017: Nature Communications
https://www.readbyqxmd.com/read/28529337/dynamic-chromatin-technologies-from-individual-molecules-to-epigenomic-regulation-in-cells
#3
REVIEW
Olivier Cuvier, Beat Fierz
The establishment and maintenance of chromatin states involves multiscale dynamic processes integrating transcription factor and multiprotein effector dynamics, cycles of chemical chromatin modifications, and chromatin structural organization. Recent developments in genomic technologies are emerging that are enabling a view beyond ensemble- and time-averaged properties and are revealing the importance of dynamic chromatin states for cell fate decisions, differentiation and reprogramming at the single-cell level...
May 22, 2017: Nature Reviews. Genetics
https://www.readbyqxmd.com/read/28529033/high-throughput-characterization-of-hiv-1-reservoir-reactivation-using-a-single-cell-in-droplet-pcr-assay
#4
Robert W Yucha, Kristen S Hobbs, Emily Hanhauser, Louise E Hogan, Wildaliz Nieves, Mehmet O Ozen, Fatih Inci, Vanessa York, Erica A Gibson, Cassandra Thanh, Hadi Shafiee, Rami El Assal, Maja Kiselinova, Yvonne P Robles, Helen Bae, Kaitlyn S Leadabrand, ShuQi Wang, Steven G Deeks, Daniel R Kuritzkes, Utkan Demirci, Timothy J Henrich
Reactivation of latent viral reservoirs is on the forefront of HIV-1 eradication research. However, it is unknown if latency reversing agents (LRAs) increase the level of viral transcription from cells producing HIV RNA or harboring transcriptionally-inactive (latent) infection. We therefore developed a microfluidic single-cell-in-droplet (scd)PCR assay to directly measure the number of CD4(+) T cells that produce unspliced (us)RNA and multiply spliced (ms)RNA following ex vivo latency reversal with either an histone deacetylase inhibitor (romidepsin) or T cell receptor (TCR) stimulation...
May 4, 2017: EBioMedicine
https://www.readbyqxmd.com/read/28528577/single-step-pcr-based-genetic-sex-determination-of-rat-tissues-and-cells
#5
Pramod Dhakal, Michael J Soares
The advent of genome editing strategies has expanded the range of animal models available for gene manipulation and renewed research interest in the rat. Gender is a key variable for in vivo gene function analyses. Here, we present a simple PCR-based method to determine genetic sex in the rat.
May 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/28527384/embracing-the-enemy-the-diversification-of-microbial-gene-repertoires-by-phage-mediated-horizontal-gene-transfer
#6
REVIEW
Marie Touchon, Jorge A Moura de Sousa, Eduardo Pc Rocha
Bacteriophages and archaeal viruses contribute, through lysogenic conversion or transduction, to the horizontal transfer of genetic material between microbial genomes. Recent genomics, metagenomics, and single cell studies have shown that lysogenic conversion is widespread and provides hosts with adaptive traits often associated with biotic interactions. The quantification of the evolutionary impact of transduction has lagged behind and requires further theoretical and experimental work. Nevertheless, recent studies suggested that generalized transduction plays a role in the transfer of antibiotic resistance genes and in the acquisition of novel genes during intra-specific bacterial competition...
May 17, 2017: Current Opinion in Microbiology
https://www.readbyqxmd.com/read/28527117/non-viral-and-viral-delivery-systems-for-crispr-cas9-technology-in-the-biomedical-field
#7
REVIEW
Zhi-Yao He, Ke Men, Zhou Qin, Yang Yang, Ting Xu, Yu-Quan Wei
The clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein 9 (CRISPR-Cas9) system provides a novel genome editing technology that can precisely target a genomic site to disrupt or repair a specific gene. Some CRISPR-Cas9 systems from different bacteria or artificial variants have been discovered or constructed by biologists, and Cas9 nucleases and single guide RNAs (sgRNA) are the major components of the CRISPR-Cas9 system. These Cas9 systems have been extensively applied for identifying therapeutic targets, identifying gene functions, generating animal models, and developing gene therapies...
May 2, 2017: Science China. Life Sciences
https://www.readbyqxmd.com/read/28526731/validated-bayesian-differentiation-of-causative-and-passenger-mutations
#8
Frederick R Cross, Michal Breker, Kristi Lieberman
In many contexts, the problem arises of determining which of many candidate mutations is the most likely to be causative for some phenotype. It is desirable to have a way to evaluate this probability that relies as little as possible on previous knowledge, to avoid bias against discovering new genes or functions. We are isolating mutants with blocked cell cycle progression in Chlamydomonas, and determining mutant genome sequences. Due to the intensity of UV mutagenesis required for efficient mutant collection, the mutants contain multiple mutations altering coding sequence...
May 19, 2017: G3: Genes—Genomes—Genetics
https://www.readbyqxmd.com/read/28524867/fundamental-differences-in-diversity-and-genomic-population-structure-between-atlantic-and-pacific-prochlorococcus
#9
Nadav Kashtan, Sara E Roggensack, Jessie W Berta-Thompson, Maor Grinberg, Ramunas Stepanauskas, Sallie W Chisholm
The Atlantic and Pacific Oceans represent different biogeochemical regimes in which the abundant marine cyanobacterium Prochlorococcus thrives. We have shown that Prochlorococcus populations in the Atlantic are composed of hundreds of genomically, and likely ecologically, distinct coexisting subpopulations with distinct genomic backbones. Here we ask if differences in the ecology and selection pressures between the Atlantic and Pacific are reflected in the diversity and genomic composition of their indigenous Prochlorococcus populations...
May 19, 2017: ISME Journal
https://www.readbyqxmd.com/read/28523967/comparative-assessment-of-dna-methylation-patterns-between-reduced-representation-bisulfite-sequencing-and-sequenom-epityper-methylation-analysis
#10
Aniruddha Chatterjee, Erin C Macaulay, Antonio Ahn, Jackie L Ludgate, Peter A Stockwell, Rob J Weeks, Matthew F Parry, Timothy J Foster, Ingrid M Knarston, Michael R Eccles, Ian M Morison
AIM: Validation of sequencing-based DNA methylation data is an important step for meaningful translation of findings. However, there has been limited assessment of different platforms to validate methylation data from next generation sequencing. METHODS: We performed a comparative methylation analysis between the genome-wide platform of reduced representation bisulfite sequencing with a targeted, Sequenom EpiTyper platform (four genes were analyzed in 15 cell lines covering 52 CpG sites)...
May 19, 2017: Epigenomics
https://www.readbyqxmd.com/read/28523561/technologies-for-deciphering-epigenomic-dna-patterns
#11
Sebastian Moran
DNA methylation, consisting on the covalent addition of a methyl group in cytosines, plays a vital role for the development and correct functioning of cells. It constitutes a mechanism by which cell genome is regulated, allowing from a common genome of an individual to obtain all the different cell types that constitute the individual. Nowadays, we understand how the epigenetic machinery works; however, this critical mechanism might promote the appearance of certain diseases if dysregulated, thus the importance of studying the epigenetic patterns on both normal and disease tissues...
2017: Advances in Experimental Medicine and Biology
https://www.readbyqxmd.com/read/28523557/single-cell-genomics-unravels-brain-cell-type-complexity
#12
Amy Guillaumet-Adkins, Holger Heyn
The brain is the most complex tissue in terms of cell types that it comprises, to the extent that it is still poorly understood. Single cell genome and transcriptome profiling allow to disentangle the neuronal heterogeneity, enabling the categorization of individual neurons into groups with similar molecular signatures. Herein, we unravel the current state of knowledge in single cell neurogenomics. We describe the molecular understanding of the cellular architecture of the mammalian nervous system in health and in disease; from the discovery of unrecognized cell types to the validation of known ones, applying these state-of-the-art technologies...
2017: Advances in Experimental Medicine and Biology
https://www.readbyqxmd.com/read/28523015/transcription-factors-in-fungi-tfome-dynamics-three-major-families-and-dual-specificity-tfs
#13
Ekaterina Shelest
Transcription factors (TFs) are essential regulators of gene expression in a cell; the entire repertoire of TFs (TFome) of a species reflects its regulatory potential and the evolutionary history of the regulatory mechanisms. In this work, I give an overview of fungal TFs, analyze TFome dynamics, and discuss TF families and types of particular interest. Whole-genome annotation of TFs in more than 200 fungal species revealed ~80 families of TFs that are typically found in fungi. Almost half of the considered genomes belonged to basidiomycetes and zygomycetes, which have been underrepresented in earlier annotations due to dearth of sequenced genomes...
2017: Frontiers in Genetics
https://www.readbyqxmd.com/read/28522818/efficient-modification-of-%C3%AE-dna-substrates-for-single-molecule-studies
#14
Yoori Kim, Armando de la Torre, Andrew A Leal, Ilya J Finkelstein
Single-molecule studies of protein-nucleic acid interactions frequently require site-specific modification of long DNA substrates. The bacteriophage λ is a convenient source of high quality long (48.5 kb) DNA. However, introducing specific sequences, tertiary structures, and chemical modifications into λ-DNA remains technically challenging. Most current approaches rely on multi-step ligations with low yields and incomplete products. Here, we describe a molecular toolkit for rapid preparation of modified λ-DNA...
May 18, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28522803/precise-genome-wide-base-editing-by-the-crispr-nickase-system-in-yeast
#15
Atsushi Satomura, Ryosuke Nishioka, Hitoshi Mori, Kosuke Sato, Kouichi Kuroda, Mitsuyoshi Ueda
The CRISPR/Cas9 system has been applied to efficient genome editing in many eukaryotic cells. However, the bases that can be edited by this system have been limited to those within the protospacer adjacent motif (PAM) and guide RNA-targeting sequences. In this study, we developed a genome-wide base editing technology, "CRISPR Nickase system" that utilizes a single Cas9 nickase. This system was free from the limitation of editable bases that was observed in the CRISPR/Cas9 system, and was able to precisely edit bases up to 53 bp from the nicking site...
May 18, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28522359/a-novel-specialized-single-linkage-clustering-algorithm-for-taxonomically-ordered-data
#16
Markus Schmidt, Arne Kutzner, Klaus Heese
Similarities among ortholog genes for a given set of species S can be expressed by alignment matrices, where each matrix cell results from aligning a gene transcript against the genome of a species within S. Gene clusters can be computed by using single-linkage clustering in time n*m, where n denotes the number of ortholog genes and m denotes the number of inspected assemblies. Our approach can break the O(n*m) complexity of single-linkage clustering by exploiting an order among species that results from an in-order traversal of a given phylogenetic tree...
May 15, 2017: Journal of Theoretical Biology
https://www.readbyqxmd.com/read/28521784/functional-regression-method-for-whole-genome-eqtl-epistasis-analysis-with-sequencing-data
#17
Kelin Xu, Li Jin, Momiao Xiong
BACKGROUND: Epistasis plays an essential rule in understanding the regulation mechanisms and is an essential component of the genetic architecture of the gene expressions. However, interaction analysis of gene expressions remains fundamentally unexplored due to great computational challenges and data availability. Due to variation in splicing, transcription start sites, polyadenylation sites, post-transcriptional RNA editing across the entire gene, and transcription rates of the cells, RNA-seq measurements generate large expression variability and collectively create the observed position level read count curves...
May 18, 2017: BMC Genomics
https://www.readbyqxmd.com/read/28515430/a-microfluidic-chip-for-screening-individual-cancer-cells-via-eavesdropping-on-autophagy-inducing-crosstalk-in-the-stroma-niche
#18
Hacer Ezgi Karakas, Junyoung Kim, Juhee Park, Jung Min Oh, Yongjun Choi, Devrim Gozuacik, Yoon-Kyoung Cho
Autophagy is a cellular homeostatic mechanism where proteins and organelles are digested and recycled to provide an alternative source of building blocks and energy to cells. The role of autophagy in cancer microenvironment is still poorly understood. Here, we present a microfluidic system allowing monitoring of the crosstalk between single cells. We used this system to study how tumor cells induced autophagy in the stromal niche. Firstly, we could confirm that transforming growth factor β1 (TGFβ1) secreted from breast tumor cells is a paracrine mediator of tumor-stroma interaction leading to the activation of autophagy in the stroma component fibroblasts...
May 17, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28515316/dna-damage-induced-degradation-of-exo1-limits-dna-end-resection-to-ensure-accurate-dna-repair
#19
Nozomi Tomimatsu, Bipasha Mukherjee, Janelle Louise Harris, Francesca Ludovica Boffo, Molly Hardebeck, Patrick Ryan Potts, Kum Kum Khanna, Sandeep Burma
End resection of DNA double-strand breaks (DSBs) to generate 3'-single-stranded DNA facilitates DSB repair via error-free homologous recombination (HR) while stymieing repair by the error-prone non-homologous end joining (NHEJ) pathway. Activation of DNA end resection involves phosphorylation of the 5' to 3' exonuclease EXO1 by the phosphoinositide 3-kinase-like kinases ATM and ATR, and by the cyclin-dependent kinases 1 and 2. After activation, EXO1 must also be restrained in order to prevent over-resection which is known to hamper optimal HR and trigger global genomic instability...
May 17, 2017: Journal of Biological Chemistry
https://www.readbyqxmd.com/read/28515305/cell-cycle-dependent-expression-of-aav2-rep-in-hsv-1-co-infections-gives-rise-to-a-mosaic-of-cells-replicating-either-aav2-or-hsv-1
#20
Francesca D Franzoso, Michael Seyffert, Rebecca Vogel, Artur Yakimovich, Bruna de Andrade Pereira, Anita F Meier, Sereina O Sutter, Kurt Tobler, Bernd Vogt, Urs F Greber, Hildegard Büning, Mathias Ackermann, Cornel Fraefel
Adeno-associated virus 2 (AAV2) depends for productive replication on the simultaneous presence of a helper virus such as herpes simplex virus type 1 (HSV-1). At the same time, AAV2 efficiently blocks the replication of HSV-1, which would eventually limit its own replication by diminishing the helper virus reservoir. This discrepancy begs the question how AAV2 and HSV-1 can co-exist in a cell population. Here we show that in co-infected cultures, AAV2 DNA replication takes place almost exclusively in S/G2 cells, while HSV-1 DNA replication is restricted to G1...
May 17, 2017: Journal of Virology
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