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https://www.readbyqxmd.com/read/27892470/vertical-magnetic-separation-of-circulating-tumor-cells-for-somatic-genomic-alteration-analysis-in-lung-cancer-patients
#1
Chang Eun Yoo, Jong-Myeon Park, Hui-Sung Moon, Je-Gun Joung, Dae-Soon Son, Hyo-Jeong Jeon, Yeon Jeong Kim, Kyung-Yeon Han, Jong-Mu Sun, Keunchil Park, Donghyun Park, Woong-Yang Park
Efficient isolation and genetic analysis of circulating tumor cells (CTCs) from cancer patients' blood is a critical step for clinical applications using CTCs. Here, we report a novel CTC-isolation method and subsequent genetic analysis. CTCs from the blood were complexed with magnetic beads coated with antibodies against the epithelial cell adhesion molecule (EpCAM) and separated vertically on a density-gradient medium in a modified well-plate. The recovery rate of model CTCs was reasonable and the cell purity was enhanced dramatically when compared to those parameters obtained using a conventional magnetic isolation method...
November 28, 2016: Scientific Reports
https://www.readbyqxmd.com/read/27878529/association-between-copy-number-variation-on-metabolic-phenotypes-and-hdl-c-levels-in-patients-with-polycystic-ovary-syndrome
#2
Birgit Knebel, Stefan Lehr, Onno E Janssen, Susanne Hahn, Sylvia Jacob, Ulrike Nitzgen, Dirk Müller-Wieland, Jorg Kotzka
Polygenic diseases with a broad phenotypic spectrum, such as polycystic ovary syndrome (PCOS), present a particular challenge in terms of identifying the underlying genetic mechanisms, nevertheless genetic variants have impact on the individual phenotype. We aimed to determine if next to genetic variations like SNPs further mechanisms might play a role in the pathogenesis of PCOS. We examined the effect of copy-number variations (CNVs) on metabolic phenotypes in PCOS. The intragenic rs1244979, rs2815752 in NEGR1 gene, and rs780094 in GCKR gene were genotyped and CNVs were determined by droplet digital polymerase chain reaction (ddPCR) in PCOS patients (n = 153) and controls without metabolic syndrome (n = 142)...
November 22, 2016: Molecular Biology Reports
https://www.readbyqxmd.com/read/27878440/cognitive-dysfunction-in-patients-with-spinocerebellar-ataxia-type-6
#3
Itaru Tamura, Asako Takei, Shinsuke Hamada, Michio Nonaka, Yoshiko Kurosaki, Fumio Moriwaka
The aim of this study was to assess the cognitive functions of patients with spinocerebellar ataxia type 6 (SCA6). We examined 13 patients with genetically confirmed SCA6 and 13 healthy control subjects matched for age, years of education, global cognitive status, and intellectual ability. We administered verbal memory (word recall and word recognition), executive function (digit span, category and letter fluency, Frontal Assessment Battery, and Trail Making Test-A and B), and visuospatial construction (figure copying) tests...
November 22, 2016: Journal of Neurology
https://www.readbyqxmd.com/read/27869597/digital-pcr-for-quantifying-norovirus-in-oysters-implicated-in-outbreaks-france
#4
David Polo, Julien Schaeffer, Nelly Fournet, Jean-Claude Le Saux, Sylvain Parnaudeau, Catherine McLeod, Françoise S Le Guyader
Using samples from oysters clearly implicated in human disease, we quantified norovirus levels by using digital PCR. Concentrations varied from 43 to 1,170 RNA copies/oyster. The analysis of frozen samples from the production area showed the presence of norovirus 2 weeks before consumption.
December 2016: Emerging Infectious Diseases
https://www.readbyqxmd.com/read/27865449/characterizing-nuclear-and-mitochondrial-dna-in-spent-embryo%C3%A2-culture-media-genetic-contamination-identified
#5
Elizabeth R Hammond, Brent C McGillivray, Sophie M Wicker, John C Peek, Andrew N Shelling, Peter Stone, Larry W Chamley, Lynsey M Cree
OBJECTIVE: To characterize nuclear and mitochondrial DNA (mtDNA) in spent culture media from normally developing blastocysts to determine whether it could be used for noninvasive genetic assessment. DESIGN: Prospective embryo cohort study. SETTING: Academic center and private in vitro fertilization (IVF) clinic. PATIENT(S): Seventy patients undergoing intracytoplasmic sperm injection (ICSI) and 227 blastocysts. INTERVENTION(S): Culture media assessment, artificial blastocoele fluid collapse and DNA analysis using digital polymerase chain reaction (dPCR), long-range PCR, quantitative PCR (qPCR), and DNA fingerprinting...
November 16, 2016: Fertility and Sterility
https://www.readbyqxmd.com/read/27863426/quantification-of-tumor-derived-cell-free-dna-cfdna-by-digital-pcr-digpcr-in-cerebrospinal-fluid-of-patients-with-brafv600-mutated-malignancies
#6
Parisa Momtaz, Elena Pentsova, Omar Abdel-Wahab, Eli Diamond, David Hyman, Taha Merghoub, Daoqi You, Billel Gasmi, Agnes Viale, Paul B Chapman
Tumor-derived cell free DNA (cfDNA) can be detected in plasma. We hypothesized that mutated BRAF V600 cfDNA could be quantified in the cerebrospinal fluid (CSF) of patients with central nervous system (CNS) metastases. We collected CSF from patients with BRAF V600E or K-mutated melanoma (N=8) or BRAF V600E mutated Erdheim-Chester Disease (ECD) (N=3) with suspected central nervous system (CNS) involvement on the basis of neurological symptoms (10/11), MRI imaging (8/11), or both. Tumor-derived cfDNA was quantified by digital PCR in the CSF of 6/11 patients (range from 0...
November 16, 2016: Oncotarget
https://www.readbyqxmd.com/read/27861535/a-segmental-copy-number-loss-of-the-sfmbt1-gene-is-a-genetic-risk-for-shunt-responsive-idiopathic-normal-pressure-hydrocephalus-inph-a-case-control-study
#7
Hidenori Sato, Yoshimi Takahashi, Luna Kimihira, Chifumi Iseki, Hajime Kato, Yuya Suzuki, Ryosuke Igari, Hiroyasu Sato, Shingo Koyama, Shigeki Arawaka, Toru Kawanami, Masakazu Miyajima, Naoyuki Samejima, Shinya Sato, Masahiro Kameda, Shinya Yamada, Daisuke Kita, Mitsunobu Kaijima, Isao Date, Yukihiko Sonoda, Takamasa Kayama, Nobumasa Kuwana, Hajime Arai, Takeo Kato
Little is known about genetic risk factors for idiopathic normal pressure hydrocephalus (iNPH). We examined whether a copy number loss in intron 2 of the SFMBT1 gene could be a genetic risk for shunt-responsive, definite iNPH. Quantitative and digital PCR analyses revealed that 26.0% of shunt-responsive definite iNPH patients (n = 50) had such a genetic change, as compared with 4.2% of the healthy elderly (n = 191) (OR = 7.94, 95%CI: 2.82-23.79, p = 1.8 x 10-5) and 6.3% of patients with Parkinson's disease (n = 32) (OR = 5...
2016: PloS One
https://www.readbyqxmd.com/read/27858717/diagnostic-value-of-a-tablet-based-drawing-task-for-discrimination-of-patients-in-the-early-course-of-alzheimer-s-disease-from-healthy-individuals
#8
Stephan Müller, Oliver Preische, Petra Heymann, Ulrich Elbing, Christoph Laske
There is a considerable delay in the diagnosis of dementia, which may reduce the effectiveness of available treatments. Thus, it is of great interest to develop fast and easy to perform, non-invasive and non-expensive diagnostic measures for the early detection of cognitive impairment and dementia. Here we investigate movement kinematics between 20 patients with early dementia due to Alzheimer's disease (eDAT), 30 patients with amnestic MCI (aMCI), and 20 cognitively healthy control (HC) individuals while copying a three-dimensional house using a digitizing tablet...
November 14, 2016: Journal of Alzheimer's Disease: JAD
https://www.readbyqxmd.com/read/27844142/efficiency-and-sensitivity-of-the-digital-droplet-pcr-for-the-quantification-of-antibiotic-resistance-genes-in-soils-and-organic-residues
#9
Laura Cavé, Elisabeth Brothier, Danis Abrouk, Panignimyandé Salomon Bouda, Edmond Hien, Sylvie Nazaret
Droplet digital PCR (ddPCR) allows absolute quantification and tolerance to inhibitors and has been proposed as the method of choice to overcome limitations of qPCR. The aim of this study was to evaluate ddPCR and qPCR performances to detect low copy number and copy number variation of antibiotic resistance genes (sul1 and qnrB genes encoding for resistance to sulfonamides and quinolones, respectively) using bacterial genomic DNA (gDNA) and metagenomic DNA extracted from soil and organic residue samples. With gDNA, qPCR showed a better range of quantification but the lower limit of quantification was at 15 copies of qnrB target vs...
November 14, 2016: Applied Microbiology and Biotechnology
https://www.readbyqxmd.com/read/27837339/her2-mrna-transcript-quantitation-in-breast-cancer
#10
K Meehan, B Clynick, B Mirzai, P Maslen, J M Harvey, W N Erber
PURPOSE: The human epidermal growth factor receptor 2 (HER2) status in breast cancer is important for prognostic prediction and the determination of optimal treatment. Current methods rely on protein expression, as determined by immunohistochemistry (IHC), as well as gene amplification as determined by in situ hybridisation (ISH). We explored whether quantitative droplet digital PCR (ddPCR) can be used for the detection and absolute quantitation of HER2 mRNA. METHODS: Digital droplet PCR (ddPCR) was performed for HER2 mRNA on 178 formalin-fixed paraffin-embedded (FFPE) breast cancer specimens...
November 11, 2016: Clinical & Translational Oncology
https://www.readbyqxmd.com/read/27825302/arabidopsis-thaliana-population-analysis-reveals-high-plasticity-of-the-genomic-region-spanning-msh2-at3g18530-and-at3g18535-genes-and-provides-evidence-for-nahr-driven-recurrent-cnv-events-occurring-in-this-location
#11
Agnieszka Zmienko, Anna Samelak-Czajka, Piotr Kozlowski, Maja Szymanska, Marek Figlerowicz
BACKGROUND: Intraspecies copy number variations (CNVs), defined as unbalanced structural variations of specific genomic loci, ≥1 kb in size, are present in the genomes of animals and plants. A growing number of examples indicate that CNVs may have functional significance and contribute to phenotypic diversity. In the model plant Arabidopsis thaliana at least several hundred protein-coding genes might display CNV; however, locus-specific genotyping studies in this plant have not been conducted...
November 8, 2016: BMC Genomics
https://www.readbyqxmd.com/read/27822863/quantitative-dna-analysis-using-droplet-digital-pcr
#12
Rolf H A M Vossen, Stefan J White
Droplet digital PCR (ddPCR) is based on the isolated amplification of thousands of individual DNA molecules simultaneously, with each molecule compartmentalized in a droplet. The presence of amplified product in each droplet is indicated by a fluorescent signal, and the proportion of positive droplets allows the precise quantification of a given sequence. In this chapter we briefly outline the basis of ddPCR, and describe two different applications using the Bio-Rad QX200 system: genotyping copy number variation and quantification of Illumina sequencing libraries...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/27806739/validation-of-droplet-digital-pcr-for-the-detection-and-absolute-quantification-of-borrelia-dna-in-ixodes-scapularis-ticks
#13
Jenny L King, Ashley D Smith, Elizabeth A Mitchell, Michael S Allen
We evaluated the QX200 Droplet Digital PCR (ddPCR™, Bio-Rad) system and protocols for the detection of the tick-borne pathogens Borrelia burgdorferi and Borrelia miyamotoi in Ixodes scapularis nymphs and adults collected from North Truro, Massachusetts. Preliminary screening by nested PCR determined positive infection levels of 60% for B. burgdorferi in these ticks. To investigate the utility of ddPCR as a screening tool and to calculate the absolute number of bacterial genome copies in an infected tick, we adapted previously reported TaqMan®-based qPCR assays for ddPCR...
November 3, 2016: Parasitology
https://www.readbyqxmd.com/read/27802737/vertical-wicking-tester-for-monitoring-water-transportation-behavior-in-fibrous-assembly
#14
Pratibha Singh, Arobindo Chatterjee, Subrata Ghosh
An instrument based on the principle of change of resistance of fibrous assembly on wetting has been developed for precise monitoring of the water transportation behaviour in the fibrous assemblies. The conducting probes sense the change in resistance of a dry fibrous assembly on wetting. This change in resistance generates analog signals which trigger an amplifying circuit. This circuit produces an enlarged copy of the received signals which are further converted to digital signals by a Darlington pair and are encoded to measurable quantity with the help of a microcontroller...
October 2016: Review of Scientific Instruments
https://www.readbyqxmd.com/read/27800678/precise-quantitation-of-microrna-in-a-single-cell-with-droplet-digital-pcr-based-on-ligation-reaction
#15
Hui Tian, Yuanyuan Sun, Chenghui Liu, Xinrui Duan, Wei Tang, Zhengping Li
MicroRNA (miRNA) analysis in a single cell is extremely important because it allows deep understanding of the exact correlation between the miRNAs and cell functions. Herein, we wish to report a highly sensitive and precisely quantitative assay for miRNA detection based on ligation-based droplet digital polymerase chain reaction (ddPCR), which permits the quantitation of miRNA in a single cell. In this ligation-based ddPCR assay, two target-specific oligonucleotide probes can be simply designed to be complementary to the half-sequence of the target miRNA respectively, which avoids the sophisticated design of reverse transcription and provides high specificity to discriminate single-base difference among miRNAs with simple operations...
November 1, 2016: Analytical Chemistry
https://www.readbyqxmd.com/read/27784328/draft-genome-of-the-leopard-gecko-eublepharis-macularius
#16
Zijun Xiong, Fang Li, Qiye Li, Long Zhou, Tony Gamble, Jiao Zheng, Ling Kui, Cai Li, Shengbin Li, Huanming Yang, Guojie Zhang
BACKGROUND: Geckos are among the most species-rich reptile groups and the sister clade to all other lizards and snakes. Geckos possess a suite of distinctive characteristics, including adhesive digits, nocturnal activity, hard, calcareous eggshells, and a lack of eyelids. However, one gecko clade, the Eublepharidae, appears to be the exception to most of these 'rules' and lacks adhesive toe pads, has eyelids, and lays eggs with soft, leathery eggshells. These differences make eublepharids an important component of any investigation into the underlying genomic innovations contributing to the distinctive phenotypes in 'typical' geckos...
October 26, 2016: GigaScience
https://www.readbyqxmd.com/read/27784303/event-specific-detection-of-transgenic-potato-av43-6-g7-using-real-time-and-digital-pcr-methods
#17
Hongwei Gao, Xiaofan Yu, Tingting Deng, Min Sun, Xizhi Xiao, Xin Huang, Ying Chen, Ronggui Li
BACKGROUND: The isolation of unknown DNA sequences flanked by known sequences is an important task in the event-specific detection of GMOs. None of event-specific detection method was developed based on the junction sequence of an exogenous integrant in the transgenic potato AV43-6-G7. RESULTS: The flanking sequence between the exogenous fragment and recombinant chromosome of this potato was successfully acquired through exogenous gene 5'-RACE. The event-specific primers and Taqman probe were designed to amplify fragments spanning the exogenous DNA and potato genomic DNA...
October 27, 2016: BMC Biotechnology
https://www.readbyqxmd.com/read/27769252/genomic-copy-number-variation-association-study-in-caucasian-patients-with-nonsyndromic-cryptorchidism
#18
Yanping Wang, Jin Li, Thomas F Kolon, Alicia Olivant Fisher, T Ernesto Figueroa, Ahmad H BaniHani, Jennifer A Hagerty, Ricardo Gonzalez, Paul H Noh, Rosetta M Chiavacci, Kisha R Harden, Debra J Abrams, Deborah Stabley, Cecilia E Kim, Katia Sol-Church, Hakon Hakonarson, Marcella Devoto, Julia Spencer Barthold
BACKGROUND: Copy number variation (CNV) is a potential contributing factor to many genetic diseases. Here we investigated the potential association of CNV with nonsyndromic cryptorchidism, the most common male congenital genitourinary defect, in a Caucasian population. METHODS: Genome wide genotyping were performed in 559 cases and 1772 controls (Group 1) using Illumina HumanHap550 v1, HumanHap550 v3 or Human610-Quad platforms and in 353 cases and 1149 controls (Group 2) using the Illumina Human OmniExpress 12v1 or Human OmniExpress 12v1-1...
October 21, 2016: BMC Urology
https://www.readbyqxmd.com/read/27763786/application-of-droplet-digital-pcr-for-estimating-vector-copy-number-states-in-stem-cell-gene-therapy
#19
Huan-Ting Lin, Takashi Okumura, Yukinori Yatsuda, Satoru Ito, Hiromitsu Nakauchi, Makoto Otsu
Stable gene transfer into target cell populations via integrating viral vectors is widely used in stem cell gene therapy (SCGT). Accurate vector copy number (VCN) estimation has become increasingly important. However, existing methods of estimation such as real-time quantitative PCR are more restricted in practicality, especially during clinical trials, given the limited availability of sample materials from patients. This study demonstrates the application of an emerging technology called droplet digital PCR (ddPCR) in estimating VCN states in the context of SCGT...
October 2016: Human Gene Therapy Methods
https://www.readbyqxmd.com/read/27760227/reproducibility-of-digital-pcr-assays-for-circulating-tumor-dna-analysis-in-advanced-breast-cancer
#20
Sarah Hrebien, Ben O'Leary, Matthew Beaney, Gaia Schiavon, Charlotte Fribbens, Amarjit Bhambra, Richard Johnson, Isaac Garcia-Murillas, Nicholas Turner
Circulating tumor DNA (ctDNA) analysis has the potential to allow non-invasive analysis of tumor mutations in advanced cancer. In this study we assessed the reproducibility of digital PCR (dPCR) assays of circulating tumor DNA in a cohort of patients with advanced breast cancer and assessed delayed plasma processing using cell free DNA preservative tubes. We recruited a cohort of 96 paired samples from 71 women with advanced breast cancer who had paired blood samples processed either immediately or delayed in preservative tubes with processing 48-72 hours after collection...
2016: PloS One
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