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https://www.readbyqxmd.com/read/28441604/digital-quantitative-analysis-of-microrna-in-single-cell-based-on-ligation-depended-polymerase-colony-polony
#1
Hui Wang, Honghong Wang, Xinrui Duan, Chenghui Liu, Zhengping Li
The ability to dissect cell-to-cell variations of microRNA (miRNA) expression with single-cell resolution has become a powerful tool to investigate the regulatory function of miRNAs in biological processes and the pathogenesis of miRNA-related diseases. Herein, we have developed a novel scheme for digital detection of miRNA in single cell by using the ligation-depended DNA polymerase colony (polony). Firstly, two simply designed target-specific DNA probes were ligated by using individual miRNA as the template...
April 7, 2017: Biosensors & Bioelectronics
https://www.readbyqxmd.com/read/28422522/molecular-genetic-characterization-of-a-chinese-family-with-severe-split-hand-foot-malformation
#2
Lihua Cao, Wei Yang, Shusen Wang, Chen Chen, Xue Zhang, Yang Luo
AIMS: Split hand/foot malformation (SHFM) is a congenital limb malformation characterized by underdeveloped or absent central digital rays, clefts of the hands and feet, and variable syndactyly of the remaining digits. SHFM is a genetically heterogeneous disease; the aim of this study was to identify pathogenic variations in a Chinese family with SHFM. MATERIALS AND METHODS: Haplotype analyses with microsatellite markers covering the five SHFM loci were performed to localize the causative locus...
April 19, 2017: Genetic Testing and Molecular Biomarkers
https://www.readbyqxmd.com/read/28414286/-pcr-analysis-of-the-absolute-number-of-copies-of-human-chromosome-18-transcripts-in-liver-and-hepg2-cells
#3
Y Y Kiseleva, K G Ptitsyn, O V Tikhonova, S P Radko, L K Kurbatov, I V Vakhrushev, V G Zgoda, E A Ponomarenko, A V Lisitsa, A I Archakov
Using reverse transcription in conjunction with the quantitative real-time PCR or digital droplet PCR, the transcriptome profiling of human chromosome 18 has been carried out in liver hepatocytes and hepatoblastoma cells (HepG2 cell line) in terms of the absolute number of each transcript per cell. The transcript abundance varies within the range of 0.006 to 9635 and 0.011 to 4819 copies per cell for HepG2 cell line and hepatocytes, respectively. The expression profiles for genes of chromosome 18 in hepatocytes and HepG2 cells were found to significantly correlate: the Spearman's correlation coefficient was equal to 0...
March 2017: Biomedit︠s︡inskai︠a︡ Khimii︠a︡
https://www.readbyqxmd.com/read/28414163/droplet-digital-pcr-ddpcr-vs-quantitative-real-time-pcr-qpcr-approach-for-detection-and-quantification-of-merkel-cell-polyomavirus-mcpyv-dna-in-formalin-fixed-paraffin-embedded-ffpe-cutaneous-biopsies
#4
Rosaria Arvia, Mauro Sollai, Federica Pierucci, Carmelo Urso, Daniela Massi, Krystyna Zakrzewska
BACKGROUND: Merkel cell polyomavirus (MCPyV) is associated with Merkel cell carcinoma and high viral load in the skin was proposed as a risk factor for the occurrence of this tumour. MCPyV DNA was detected, with lower frequency, in different skin cancers but since the viral load was usually low, the real prevalence of viral DNA could be underestimated. OBJECTIVE: To evaluate the performance of two assays (qPCR and ddPCR) for MCPyV detection and quantification in formalin fixed paraffin embedded (FFPE) tissue samples...
April 14, 2017: Journal of Virological Methods
https://www.readbyqxmd.com/read/28397990/germline-transformation-of-the-western-corn-rootworm-diabrotica-virgifera-virgifera
#5
F Chu, W Klobasa, P Wu, S Pinzi, N Grubbs, S Gorski, Y Cardoza, M D Lorenzen
The western corn rootworm (WCR), a major pest of maize, is notorious for rapidly adapting biochemically, behaviourally and developmentally to a variety of control methods. Despite much effort, the genetic basis of WCR adaptation remains a mystery. Since transformation-based applications such as transposon tagging and enhancer trapping have facilitated genetic dissection of model species such as Drosophila melanogaster, we developed a germline-transformation system for WCR in an effort to gain a greater understanding of the basic biology of this economically important insect...
April 11, 2017: Insect Molecular Biology
https://www.readbyqxmd.com/read/28393575/using-circulating-cell-free-dna-to-monitor-personalized-cancer-therapy
#6
Michael Oellerich, Ekkehard Schütz, Julia Beck, Philipp Kanzow, Piers N Plowman, Glen J Weiss, Philip D Walson
High-quality genomic analysis is critical for personalized pharmacotherapy in patients with cancer. Tumor-specific genomic alterations can be identified in cell-free DNA (cfDNA) from patient blood samples and can complement biopsies for real-time molecular monitoring of treatment, detection of recurrence, and tracking resistance. cfDNA can be especially useful when tumor tissue is unavailable or insufficient for testing. For blood-based genomic profiling, next-generation sequencing (NGS) and droplet digital PCR (ddPCR) have been successfully applied...
April 10, 2017: Critical Reviews in Clinical Laboratory Sciences
https://www.readbyqxmd.com/read/28347708/analytical-validation-of-a-reverse-transcriptase-droplet-digital-pcr-rt-ddpcr-for-quantitative-detection-of-infectious-hematopoietic-necrosis-virus
#7
Peng Jia, Maureen K Purcell, Guang Pan, Jinjin Wang, Shifu Kan, Yin Liu, Xiaocong Zheng, Xiujie Shi, Junqiang He, Li Yu, Qunyi Hua, Tikang Lu, Wensheng Lan, James R Winton, Ningyi Jin, Hong Liu
Infectious hematopoietic necrosis virus (IHNV) is an important pathogen of salmonid fishes. A validated universal reverse transcriptase quantitative PCR (RT-qPCR) assay that can quantify levels of IHNV in fish tissues has been previously reported. In the present study, we adapted the published set of IHNV primers and probe for use in a reverse-transcriptase droplet digital PCR (RT-ddPCR) assay for quantification of the virus in fish tissue samples. The RT-ddPCR and RT-qPCR assays detected 13 phylogenetically diverse IHNV strains, but neither assay produced detectable amplification when RNA from other fish viruses was used...
March 24, 2017: Journal of Virological Methods
https://www.readbyqxmd.com/read/28347285/whole-exome-sequencing-and-digital-pcr-identified-a-novel-compound-heterozygous-mutation-in-the-nphp1-gene-in-a-case-of-joubert-syndrome-and-related-disorders
#8
Shingo Koyama, Hidenori Sato, Manabu Wada, Toru Kawanami, Mitsuru Emi, Takeo Kato
BACKGROUND: Joubert syndrome and related disorders (JSRD) is a clinically and genetically heterogeneous condition with autosomal recessive or X-linked inheritance, which share a distinctive neuroradiological hallmark, the so-called molar tooth sign. JSRD is classified into six clinical subtypes based on associated variable multiorgan involvement. To date, 21 causative genes have been identified in JSRD, which makes genetic diagnosis difficult. CASE PRESENTATION: We report here a case of a 28-year-old Japanese woman diagnosed with JS with oculorenal defects with a novel compound heterozygous mutation (p...
March 27, 2017: BMC Medical Genetics
https://www.readbyqxmd.com/read/28347134/evaluating-droplet-digital-polymerase-chain-reaction-for-the-quantification-of-human-genomic-dna-lifting-the-traceability-fog
#9
Margaret C Kline, David L Duewer
Digital polymerase chain reaction (dPCR) end point platforms directly estimate the number of DNA target copies per reaction partition, λ, where the partitions are fixed-location chambers (cdPCR) or aqueous droplets floating in oil (ddPCR). For use in the certification of target concentration in primary calibrant certified reference materials (CRMs), both λ and the partition volume, V, must be metrologically traceable to some accessible reference system, ideally, the International System of Units (SI). The fixed spatial distribution of cdPCR chambers enables real-time monitoring of PCR amplification...
April 6, 2017: Analytical Chemistry
https://www.readbyqxmd.com/read/28345028/self-powered-integrated-microfluidic-point-of-care-low-cost-enabling-simple-chip
#10
Erh-Chia Yeh, Chi-Cheng Fu, Lucy Hu, Rohan Thakur, Jeffrey Feng, Luke P Lee
Portable, low-cost, and quantitative nucleic acid detection is desirable for point-of-care diagnostics; however, current polymerase chain reaction testing often requires time-consuming multiple steps and costly equipment. We report an integrated microfluidic diagnostic device capable of on-site quantitative nucleic acid detection directly from the blood without separate sample preparation steps. First, we prepatterned the amplification initiator [magnesium acetate (MgOAc)] on the chip to enable digital nucleic acid amplification...
March 2017: Science Advances
https://www.readbyqxmd.com/read/28337907/single-cell-based-platform-for-copy-number-variation-profiling-through-digital-counting-of-amplified-genomic-dna-fragments
#11
Chunmei Li, Zhilong Yu, Yusi Fu, Yuhong Pang, Yanyi Huang
We develop a novel single-cell-based platform through digital counting of amplified genomic DNA fragments, named multifraction amplification (mfA), to detect the copy number variations (CNVs) in a single cell. Amplification is required to acquire genomic information from a single cell, while introducing unavoidable bias. Unlike prevalent methods that directly infer CNV profiles from the pattern of sequencing depth, our mfA platform denatures and separates the DNA molecules from a single cell into multiple fractions of a reaction mix before amplification...
April 14, 2017: ACS Applied Materials & Interfaces
https://www.readbyqxmd.com/read/28326203/esthetic-rehabilitation-of-anterior-teeth-with-copy-milled-restorations-a-report-of-two-cases
#12
Sapna Rani, Jyoti Devi, Chandan Jain, Parul Mutneja, Mahesh Verma
Digitalization has become part and parcel of contemporary prosthodontics with the probability of most of the procedures being based on the digital techniques in the near future. This digital revolution started in the latter half of the 20th century by converting analog objects/signals into digital bits and bytes. Recent developments in all-ceramic materials and systems of computer-aided designing and computer-aided manufacturing (CAD/CAM), copy milling, and so forth offer excellent esthetics and superb biocompatibility...
2017: Case Reports in Dentistry
https://www.readbyqxmd.com/read/28322802/simple-quantitative-pcr-analysis-for-allelic-pten-loss-in-tumor-progression
#13
Xingping Wu, Ailin Zhu, Xiaowu Pang, Guiqin Xie, Xinbin Gu
We describe a simple method to accurately detect and quantify both Pten mutation and allele-specific loss using allele-specific PCR analysis. Our approach used a heterozygous genomic DNA with one wild-type and one mutant Pten allele as a reference at a single concentration to calculate the percent ratio of the wild-type Pten gene for the detection of allele-specific gene loss. With a standard curve, ratios from PCR data were used to quantitate the wild-type Pten allele copy number loss in tumor specimens. We demonstrate the utility of our approach to calculate allele-specific Pten loss during tumor progression and show that our approach generates quantitative data that are comparable to those obtained from digital droplet PCR...
March 18, 2017: Analytical Biochemistry
https://www.readbyqxmd.com/read/28322208/prevalence-and-pathogen-load-estimates-for-the-fungus-batrachochytrium-dendrobatidis-are-impacted-by-its-dna-copy-number-variation
#14
Eria A Rebollar, Douglas C Woodhams, Brandon LaBumbard, Jos Kielgast, Reid N Harris
The ribosomal gene complex is a multi-copy region that is widely used for phylogenetic analyses of organisms from all 3 domains of life. In fungi, the copy number of the internal transcribed spacer (ITS) is used to detect abundance of pathogens causing diseases such as chytridiomycosis in amphibians and white nose syndrome in bats. Chytridiomycosis is caused by the fungi Batrachochytrium dendrobatidis (Bd) and B. salamandrivorans (Bsal), and is responsible for declines and extinctions of amphibians worldwide...
March 21, 2017: Diseases of Aquatic Organisms
https://www.readbyqxmd.com/read/28319152/a-digital-pcr-method-for-identifying-and-quantifying-adulteration-of-meat-species-in-raw-and-processed-food
#15
Junan Ren, Tingting Deng, Wensheng Huang, Ying Chen, Yiqiang Ge
Meat adulteration is a worldwide concern. In this paper, a new droplet digital PCR (ddPCR) method was developed for the quantitative determination of the presence of chicken in sheep and goat meat products. Meanwhile, a constant (multiplication factor) was introduced to transform the ratio of copy numbers to the proportion of meats. The presented ddPCR method was also proved to be more accurate (showing bias of less than 9% in the range from 5% to 80%) than real-time PCR, which has been widely used in this determination...
2017: PloS One
https://www.readbyqxmd.com/read/28303130/new-approaches-on-quantification-of-campylobacter-jejuni-in-poultry-samples-the-use-of-digital-pcr-and-real-time-pcr-against-the-iso-standard-plate-count-method
#16
Bojan Papić, Mateja Pate, Urška Henigman, Urška Zajc, Igor Gruntar, Majda Biasizzo, Matjaž Ocepek, Darja Kušar
Campylobacteriosis is the most frequently reported bacterial food-borne illness in the European Union and contaminated broiler meat is considered the most important source of infection in humans. The aim of the present study was to evaluate real-time PCR (qPCR) and digital PCR (dPCR) for quantification of Campylobacter jejuni in 75 broiler neck-skin samples collected from a poultry slaughterhouse, and to compare them with the ISO 10272-2 standard plate count method. For qPCR standard curve, C. jejuni-negative neck-skin samples were spiked with C...
2017: Frontiers in Microbiology
https://www.readbyqxmd.com/read/28294696/cerebrospinal-fluid-mtdna-concentration-is-elevated-in-multiple-sclerosis-disease-and-responds-to-treatment
#17
Cyra E Leurs, Petar Podlesniy, Ramon Trullas, Lisanne Balk, Martijn D Steenwijk, Arjan Malekzadeh, Fredrik Piehl, Bernard Mj Uitdehaag, Joep Killestein, Jack van Horssen, C E Teunissen
BACKGROUND: Mitochondrial dysfunction is increasingly recognized as an important feature of multiple sclerosis (MS) pathology and may be relevant for clinical disease progression. However, it is unknown whether mitochondrial DNA (mtDNA) levels in the cerebrospinal fluid (CSF) associate with disease progression and therapeutic response. OBJECTIVES: To evaluate whether CSF concentrations of mtDNA in MS patients can serve as a marker of ongoing neuropathology and may be helpful to differentiate between MS disease subtypes...
March 1, 2017: Multiple Sclerosis: Clinical and Laboratory Research
https://www.readbyqxmd.com/read/28291853/an-in-vitro-evaluation-of-the-loosening-of-different-interchangeable-abutments-in-internal-connection-type-implants
#18
Ji-Man Park, Chang-Hyun Baek, Seong-Joo Heo, Seong-Kyun Kim, Jai-Young Koak, Shin-Koo Kim, Urs C Belser
PURPOSE: The aim of this study was to compare the loosening of interchangeable one-piece abutments connected to internal-connection-type implants after cyclic loading. MATERIALS AND METHODS: Four implant abutment groups (n = 7 in each group) with Straumann tissue-level implants were assessed: Straumann solid abutment (group S), Southern Implants solid abutment (group SI), Implant Direct straight abutment (group ID), and Blue Sky Bio regular platform abutment (group BSB)...
March 2017: International Journal of Oral & Maxillofacial Implants
https://www.readbyqxmd.com/read/28284873/establishing-a-reference-dataset-for-the-authentication-of-spinal-muscular-atrophy-cell-lines-using-str-profiling-and-digital-pcr
#19
Deborah L Stabley, Jennifer Holbrook, Ashlee W Harris, Kathryn J Swoboda, Thomas O Crawford, Katia Sol-Church, Matthew E R Butchbach
Fibroblasts and lymphoblastoid cell lines (LCLs) derived from individuals with spinal muscular atrophy (SMA) have been and continue to be essential for translational SMA research. Authentication of cell lines helps ensure reproducibility and rigor in biomedical research. This quality control measure identifies mislabeling or cross-contamination of cell lines and prevents misinterpretation of data. Unfortunately, authentication of SMA cell lines used in various studies has not been possible because of a lack of a reference...
May 2017: Neuromuscular Disorders: NMD
https://www.readbyqxmd.com/read/28281732/utilisation-of-three-dimensional-printed-heart-models-for-operative-planning-of-complex-congenital-heart-defects
#20
Peter Olejník, Matej Nosal, Tomas Havran, Adriana Furdova, Maros Cizmar, Michal Slabej, Andrej Thurzo, Pavol Vitovic, Martin Klvac, Tibor Acel, Jozef Masura
BACKGROUND: To evaluate the accuracy of the three-dimensional printing of cardiovascular structures.To explore whether utilization of three-dimensional printed heart replicas can improve surgical and catheter interventional planning in patients with complex congenital heart defects. METHODS: Between December 2014 and November 2015, we fabricated n=8 cardiovascular models based on computed tomography data in patients with complex spatial anatomical relationships of cardiovascular structures...
March 10, 2017: Kardiologia Polska
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