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https://www.readbyqxmd.com/read/28448034/using-a-fluorescent-pcr-capillary-gel-electrophoresis-technique-to-genotype-crispr-cas9-mediated-knockout-mutants-in-a-high-throughput-format
#1
Muhammad Khairul Ramlee, Jing Wang, Alice M S Cheung, Shang Li
The development of programmable genome-editing tools has facilitated the use of reverse genetics to understand the roles specific genomic sequences play in the functioning of cells and whole organisms. This cause has been tremendously aided by the recent introduction of the CRISPR/Cas9 system-a versatile tool that allows researchers to manipulate the genome and transcriptome in order to, among other things, knock out, knock down, or knock in genes in a targeted manner. For the purpose of knocking out a gene, CRISPR/Cas9-mediated double-strand breaks recruit the non-homologous end-joining DNA repair pathway to introduce the frameshift-causing insertion or deletion of nucleotides at the break site...
April 8, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/28447785/graphene-microcapsule-arrays-for-combinatorial-electron-microscopy-and-spectroscopy-in-liquids
#2
Alexander Yulaev, Hongxuan Guo, Evgheni Strelcov, Lei Chen, Ivan V Vlassiouk, Andrei Kolmakov
Atomic-scale thickness, molecular impermeability, low atomic number, and mechanical strength make graphene an ideal electron-transparent membrane for material characterization in liquids and gases with scanning electron microscopy and spectroscopy. Here, we present a novel sample platform made of an array of thousands of identical isolated graphene-capped micro-channels with high aspect ratio. A combination of a global wide field of view with high resolution local imaging of the array allows for high throughput in situ studies as well as for combinatorial screening of solutions, liquid interfaces and immersed samples...
April 27, 2017: ACS Applied Materials & Interfaces
https://www.readbyqxmd.com/read/28446788/resolution-enhanced-fourier-ptychographic-microscopy-based-on-high-numerical-aperture-illuminations
#3
Jiasong Sun, Chao Zuo, Liang Zhang, Qian Chen
High-resolution and wide field-of-view (FOV) microscopic imaging plays a central role in diverse applications such as high-throughput screening and digital pathology. However, conventional microscopes face inherent trade-offs between the spatial resolution and FOV, which are fundamental limited by the space-bandwidth product (SBP) of the optical system. The resolution-FOV tradeoff can be effectively decoupled in Fourier ptychography microscopy (FPM), however, to date, the effective imaging NA achievable with a typical FPM system is still limited to the range of 0...
April 26, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28446772/large-scale-snp-screenings-identify-markers-linked-with-gcrv-resistant-traits-through-transcriptomes-of-individuals-and-cell-lines-in-ctenopharyngodon-idella
#4
Zhiwei Liao, Quanyuan Wan, Xueying Shang, Jianguo Su
Grass carp (Ctenopharyngodon idella) is an important economic species in freshwater aquaculture and its industry has been confined due to variety degeneration and frequent diseases. Marker-assisted selection is a feasible method for selective breeding of new varieties. Transcriptome data have greatly facilitated high-throughput single nucleotide polymorphism (SNP) marker discovery and phenotype association study. In this study, we gained a total of 25,981 and 5,775 high quality SNPs in two transcriptomes from individuals and cell lines, respectively...
April 26, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28446504/feasibility-of-ultra-high-throughput-functional-screening-of-melanoma-biopsies-for-discovery-of-novel-cancer-drug-combinations
#5
Adam A Friedman, Yun Xia, Lorenzo Trippa, Long P Le, Vivien Igras, Dennie T Frederick, Jennifer A Wargo, Kenneth K Tanabe, Donald P Lawrence, Donna S Neuberg, Keith T Flaherty, David Fisher
Purpose: Successful development of targeted therapy combinations for cancer patients depends on first discovering such combinations in predictive preclinical models. Stable cell lines and mouse xenograft models can have genetic and phenotypic drift and may take too long to generate to be useful as a personalized medicine tool. <p>Experimental Design: To overcome these limitations, we have used a platform of ultra-high-throughput functional screening of primary biopsies preserving both cancer and stroma cell populations from melanoma patients to nominate such novel combinations from a library of  thousands of drug combinations in a patient-specific manner within days of biopsy...
April 26, 2017: Clinical Cancer Research: An Official Journal of the American Association for Cancer Research
https://www.readbyqxmd.com/read/28445253/micro-rna-screening-and-prediction-model-construction-for-diagnosis-of-salt-sensitive-essential-hypertension
#6
Han Qi, Zheng Liu, Bin Liu, Han Cao, Weiping Sun, Yuxiang Yan, Ling Zhang
Commonly used tests for diagnosis of salt-sensitive hypertension (SSH) are complex and time-consuming, so new methods are required. Many studies have demonstrated roles for miRNAs in hypertension; however, the diagnostic value of miRNAs has yet to be determined for human SSH. In this study, we examined miRNA expression profiles by initial high-throughput miRNA sequencing of samples from patients with salt-sensitive and salt-resistant hypertension (SSH and SRH, respectively; n = 6, both groups), followed by validation by quantitative real-time polymerase chain reaction (qRT-PCR) in a larger cohort (n = 91)...
April 2017: Medicine (Baltimore)
https://www.readbyqxmd.com/read/28445128/3-dimensional-micropillar-drug-screening-identifies-fgfr2-iiic-overexpression-as-a-potential-target-in-metastatic-giant-cell-tumor
#7
Seung Tae Kim, Jusun Kim, Sumin Shin, Sun Young Kim, Dongwoo Lee, Bosung Ku, Yong Sung Shin, Jhingook Kim, Jeeyun Lee
We established two patient derived tumor cells (PDCs) from right and left pulmonary metastatic lesions respectively of a patient with giant cell tumor. At that time, patient-derived tumor cells from right and left surgical specimens were collected and cultured. High-throughput screening (HTS) for 24 drugs was conducted with a micropillar/microwell chip platform using giant cell tumor PDCs. Using 6 doses per drug in 6 replicates for giant cell tumor PDCs, the dose response curves and corresponding IC50 values were calculated from the scanned images using the S+ Chip Analyzer...
April 6, 2017: Oncotarget
https://www.readbyqxmd.com/read/28445076/image-based-phenotypic-screening-with-human-primary-t-cells-using-one-dimensional-imaging-cytometry-with-self-tuning-statistical-gating-algorithms
#8
Steve S Wang, Daniel J Ehrlich
The parallel microfluidic cytometer (PMC) is an imaging flow cytometer that operates on statistical analysis of low-pixel-count, one-dimensional (1D) line scans. It is highly efficient in data collection and operates on suspension cells. In this article, we present a supervised automated pipeline for the PMC that minimizes operator intervention by incorporating multivariate logistic regression for data scoring. We test the self-tuning statistical algorithms in a human primary T-cell activation assay in flow using nuclear factor of activated T cells (NFAT) translocation as a readout and readily achieve an average Z' of 0...
April 1, 2017: SLAS Discovery
https://www.readbyqxmd.com/read/28445069/targeted-dereplication-of-microbial-natural-products-by-high-resolution-ms-and-predicted-lc-retention-time
#9
Justine Chervin, Marc Stierhof, Ming Him Tong, Doe Peace, Kine Østnes Hansen, Dagmar Solveig Urgast, Jeanette Hammer Andersen, Yi Yu, Rainer Ebel, Kwaku Kyeremeh, Veronica Paget, Gabriela Cimpan, Albert Van Wyk, Hai Deng, Marcel Jaspars, Jioji N Tabudravu
A new strategy for the identification of known compounds in Streptomyces extracts that can be applied in the discovery of natural products is presented. The strategy incorporates screening a database of 5555 natural products including 5098 structures from Streptomyces sp., using a high-throughput LCMS data processing algorithm that utilizes HRMS data and predicted LC retention times (tR) as filters for rapid identification of known compounds in the natural product extract. The database, named StrepDB, contains for each compound the structure, molecular formula, molecular mass, and predicted LC retention time...
April 26, 2017: Journal of Natural Products
https://www.readbyqxmd.com/read/28444691/maldi-ms-profiling-of-serum-o-and-n-glycosylation-in-cog5-cdg
#10
Angelo Palmigiano, Rosaria Ornella Bua, Rita Barone, Daisy Rymen, Luc Régal, Nicolas Deconinck, Carlo Dionisi-Vici, Cheuk-Wing Fung, Domenico Garozzo, Jaak Jaeken, Luisa Sturiale
Congenital disorders of glycosylation (CDG) are due to defective glycosylation of glycoconjugates. COG-CDG are genetic diseases due to defects of the conserved oligomeric Golgi (COG) complex subunits 1-8 causing N- and O-glycan processing abnormalities. In COG-CDG, IEF separation of undersialylated glycoforms of serum transferrin and apolipoprotein C-III (apoC-III) allows to detect N- and O-glycosylation defects respectively. COG5-CDG (COG5 subunit deficiency) is a multisystem disease with dysmorphic features, intellectual disability of variable degree, seizures, acquired microcephaly, sensory defects and autistic behavior...
April 25, 2017: Journal of Mass Spectrometry: JMS
https://www.readbyqxmd.com/read/28443514/compound-libraries-recent-advances-and-their-applications-in-drug-discovery
#11
Zhen Gong, Guoping Hu, Qiang Li, Zhiguo Liu, Fei Wang, Xuejin Zhang, Jian Xiong, Peng Li, Yan Xu, Rujian Ma, Shuhui Chen, Jian Li
Hit identification is the starting point of small-molecule drug discovery and is therefore very important to the pharmaceutical industry. One of the most important approaches to identify a new hit is to screen a compound library using an in vitro assay. High-throughput screening has made great contributions to drug discovery since the 1990s but requires expensive equipment and facilities, and its success depends on the size of the compound library. Recent progress in the development of compound libraries has provided more efficient ways to identify new hits for novel drug targets, thereby helping to promote the development of the pharmaceutical industry, especially for first-in-class drugs...
April 25, 2017: Current Drug Discovery Technologies
https://www.readbyqxmd.com/read/28440957/the-role-of-high-throughput-screening-in-ecotoxicology-and-engineered-nanomaterials
#12
REVIEW
Andrew Barrick, Amélie Châtel, Mélanie Bruneau, Catherine Mouneyrac
The field of environmental toxicology developed as a result of growing concerns about anthropogenic influences on the environment and how to ameliorate ecological impact. Many governmental bodies are beginning to emphasize prevention rather than mitigation when addressing novel products, leading to more of a focus on identifying potential toxicity prior to release. With the exponential advances in their development and sale, novel metamaterials and biotechnology are set to dramatically outpace the capabilities of current testing strategies...
April 25, 2017: Environmental Toxicology and Chemistry
https://www.readbyqxmd.com/read/28440616/discovery-and-characterization-of-a-eukaryotic-initiation-factor-4a-3-selective-inhibitor-that-suppresses-nonsense-mediated-mrna-decay
#13
Misa Iwatani-Yoshihara, Masahiro Ito, Yoshihiro Ishibashi, Hideyuki Oki, Toshio Tanaka, Daisuke Morishita, Takashi Ito, Hiromichi Kimura, Yasuhiro Imaeda, Samuel A Aparicio, Atsushi Nakanishi, Tomohiro Kawamoto
Eukaryotic initiation factor 4A-3 (eIF4A3) is an Asp-Glu-Ala-Asp (DEAD) box-family adenosine triphosphate (ATP)-dependent RNA helicase. Subtypes eIF4A1 and eIF4A2 are required for translation initiation, but eIF4A3 participates in the exon junction complex (EJC) and functions in RNA metabolism including nonsense-mediated RNA decay (NMD). No small molecules for NMD inhibition via selective inhibition of eIF4A3 have been discovered. Here we identified allosteric eIF4A3 inhibitors from a high-throughput screening campaign...
April 25, 2017: ACS Chemical Biology
https://www.readbyqxmd.com/read/28440476/exploration-of-mir-1202-and-mir-196a-in-human-endometrial-cancer-based-on-high-throughout-gene-screening-analysis
#14
Hong Chen, Yujuan Fan, Wensheng Xu, Junying Chen, Yugang Meng, Di Fang, Jingran Wang
Altered microRNA (miRNA) expression has been reported to participate in the pathogenesis of several human diseases, and particularly cancer. The present study examined the involvement of various miRNAs in the pathophysiology of endometrial cancer (EC) and atypical endometrial hyperplasia (AEH). We performed a high-throughput analysis of the miRNAs (miRNA microarray) found in samples of endometrial tissue obtained from 45 patients; among whom, 15 patients were diagnosed with EC, 15 patients were diagnosed with AEH, and the remainder were healthy donors...
April 24, 2017: Oncology Reports
https://www.readbyqxmd.com/read/28439871/application-of-chemical-genomics-to-plant-bacteria-communication-a-high-throughput-system-to-identify-novel-molecules-modulating-the-induction-of-bacterial-virulence-genes-by-plant-signals
#15
Elodie Vandelle, Maria Rita Puttilli, Andrea Chini, Giulia Devescovi, Vittorio Venturi, Annalisa Polverari
The life cycle of bacterial phytopathogens consists of a benign epiphytic phase, during which the bacteria grow in the soil or on the plant surface, and a virulent endophytic phase involving the penetration of host defenses and the colonization of plant tissues. Innovative strategies are urgently required to integrate copper treatments that control the epiphytic phase with complementary tools that control the virulent endophytic phase, thus reducing the quantity of chemicals applied to economically and ecologically acceptable levels...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28439865/identification-of-protein-dna-interactions-using-enhanced-yeast-one-hybrid-assays-and-a-semiautomated-approach
#16
Allison Gaudinier, Michelle Tang, Anne-Maarit Bågman, Siobhan M Brady
Yeast one-hybrid assays are an in vitro gene-centered approach to map transcription factor-DNA interactions. Here we describe this method and adaptations to screen for interactions between plant transcriptional regulators and their targets. Of particular note, the use of yeast one-hybrid assays fills in an important gap in available methodologies. When one is interested in a specific biological process of interest, the yeast one-hybrid assay is the only method that allows researchers to identify upstream regulators of the biological process of interest...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28439239/development-of-automated-patch-clamp-technique-to-investigate-cftr-chloride-channel-function
#17
Arnaud Billet, Lionel Froux, John W Hanrahan, Frederic Becq
The chloride (Cl(-)) channel cystic fibrosis transmembrane conductance regulator (CFTR) is defective in cystic fibrosis (CF), and mutation of its encoding gene leads to various defects such as retention of the misfolded protein in the endoplasmic reticulum, reduced stability at the plasma membrane, abnormal channel gating with low open probability, and thermal instability, which leads to inactivation of the channel at physiological temperature. Pharmacotherapy is one major therapeutic approach in the CF field and needs sensible and fast tools to identify promising compounds...
2017: Frontiers in Pharmacology
https://www.readbyqxmd.com/read/28438642/diagnostic-microarray-for-14-water-and-foodborne-pathogens-using-a-flatbed-scanner
#18
Vidya Srinivasan, Robert D Stedtfeld, Dieter M Tourlousse, Samuel W Baushke, Yu Xin, Sarah M Miller, Trinh Pham, Jean-Marie Rouillard, Erdogan Gulari, James M Tiedje, Syed A Hashsham
Parallel detection approaches are of interest to many researchers interested in identifying multiple water and foodborne pathogens simultaneously. Availability and cost-effectiveness are two key factors determining the usefulness of such approaches for laboratories with limited resources. In this study, we developed and validated a high-density microarray for simultaneous screening of 14 bacterial pathogens using an approach that employs gold labeling with silver enhancement (GLS) protocol. In total, 8887 probes (50-mer) were designed using an in-house database of virulence and marker genes (VMGs), and synthesized in quadruplicate on glass slides using an in-situ synthesis technology...
April 21, 2017: Journal of Microbiological Methods
https://www.readbyqxmd.com/read/28438393/use-of-the-entericbio-gastro-panel-ii-in-a-diagnostic-microbiology-laboratory-challenges-and-opportunities
#19
Gary McAuliffe, Liselle Bissessor, Deborah Williamson, Sharon Moore, Janet Wilson, Muriel Dufour, Susan Taylor, Arlo Upton
We sought to verify the performance of the EntericBio real-time Gastro Panel 2 (GP2; Serosep, Ireland) assay targeting Salmonella, Shigella, Campylobacter, verotoxin producing E. coli (VTEC), Giardia, and Cryptosporidium in a high-throughput diagnostic laboratory. Following a technical verification period where 237 faecal samples were tested in parallel, guided culture and discrepancy analysis were performed. A total of 13,223 faecal samples were screened by the GP2 assay. During technical verification, sensitivity, specificity, negative and positive predictive value of the GP2 assay compared with conventional methods were 100% (83-100%), 98% (96...
April 21, 2017: Pathology
https://www.readbyqxmd.com/read/28437628/advanced-biomaterials-and-microengineering-technologies-to-recapitulate-the-stepwise-process-of-cancer-metastasis
#20
REVIEW
Nitish Peela, Danh Truong, Harpinder Saini, Hunghao Chu, Samaneh Mashaghi, Stephanie L Ham, Sunil Singh, Hossein Tavana, Bobak Mosadegh, Mehdi Nikkhah
Cancer is one of the leading causes of death globally according to the World Health Organization. Although improved treatments and early diagnoses have reduced cancer related mortalities, metastatic disease remains a major clinical challenge. The local tumor microenvironment plays a significant role in cancer metastasis, where tumor cells respond and adapt to a plethora of biochemical and biophysical signals from stromal cells and extracellular matrix (ECM) proteins. Due to these complexities, there is a critical need to understand molecular mechanisms underlying cancer metastasis to facilitate the discovery of more effective therapies...
April 21, 2017: Biomaterials
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