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https://www.readbyqxmd.com/read/27913434/a-study-of-tp53-rna-splicing-illustrates-pitfalls-of-rna-seq-methodology
#1
Sunali Mehta, Peter Tsai, Annette Lasham, Hamish Campbell, Roger Reddel, Antony Braithwaite, Cristin Print
TP53 undergoes multiple RNA-splicing events, resulting in at least nine mRNA transcripts encoding at least 12 functionally different protein isoforms. Antibodies specific to p53 protein isoforms have proven difficult to develop, thus researchers must rely on the transcript information to infer isoform abundance. In this study, we used deep RNA-seq, droplet digital PCR (ddPCR), and real-time quantitative reverse transcriptase PCR (RT-qPCR) from nine human cell lines and RNA-seq data available for tumors in The Cancer Genome Atlas to analyze TP53 splice variant expression...
October 20, 2016: Cancer Research
https://www.readbyqxmd.com/read/27908228/detection-of-mutations-in-mitochondrial-dna-by-droplet-digital-pcr
#2
J K Sofronova, Y Y Ilinsky, K E Orishchenko, E G Chupakhin, E A Lunev, I O Mazunin
Mutations in mitochondrial DNA (mtDNA) may result in various pathological processes. Detection of mutant mtDNAs is a problem for diagnostic practice that is complicated by heteroplasmy - a phenomenon of the inferring presence of at least two allelic variants of the mitochondrial genome. Also, the level of heteroplasmy largely determines the profile and severity of clinical manifestations. Here we discuss detection of mutations in heteroplasmic mtDNA using up-to-date methods that have not yet been introduced as routine clinical assays...
October 2016: Biochemistry. Biokhimii︠a︡
https://www.readbyqxmd.com/read/27903892/methylation-sensitive-enrichment-of-minor-dna-alleles-using-a-double-strand-dna-specific-nuclease
#3
Yibin Liu, Chen Song, Ioannis Ladas, Mariana Fitarelli-Kiehl, G Mike Makrigiorgos
Aberrant methylation changes, often present in a minor allelic fraction in clinical samples such as plasma-circulating DNA (cfDNA), are potentially powerful prognostic and predictive biomarkers in human disease including cancer. We report on a novel, highly-multiplexed approach to facilitate analysis of clinically useful methylation changes in minor DNA populations. Methylation Specific Nuclease-assisted Minor-allele Enrichment (MS-NaME) employs a double-strand-specific DNA nuclease (DSN) to remove excess DNA with normal methylation patterns...
November 29, 2016: Nucleic Acids Research
https://www.readbyqxmd.com/read/27902802/identification-and-characterization-of-quantitative-trait-loci-for-shattering-in-rice-landrace-jiucaiqing-from-taihu-lake-valley-china
#4
Jinping Cheng, Yongqi He, Chengfang Zhan, Bin Yang, Enshun Xu, Hongsheng Zhang, Zhoufei Wang
Easy shattering reduces yield from grain loss during rice ( L.) harvest. We characterized a nonshattering rice landrace Jiucaiqing from Taihu Lake valley in China. The breaking tensile strength (BTS; grams force, gf) of the grain pedicel was measured using a digital force gauge to evaluate the degree of shattering at 0, 7, 14, 21, 28, and 35 d after heading (DAH). The BTS of Jiucaiqing did not significantly decrease with increasing DAH, maintaining a level of 152.2 to 195.9 gf, while that of IR26 decreased greatly during 0 to 14 DAH and finally stabilized at ∼100 gf...
November 2016: Plant Genome
https://www.readbyqxmd.com/read/27901057/digital-gene-expression-profiling-analysis-of-duodenum-transcriptomes-in-sd-rats-administered-ferrous-sulfate-or-ferrous-glycine-chelate-by-gavage
#5
Zhao Zhuo, Shenglin Fang, Qiaoling Hu, Danping Huang, Jie Feng
The absorption of different iron sources is a trending research topic. Many studies have revealed that organic iron exhibits better bioavailability than inorganic iron, but the concrete underlying mechanism is still unclear. In the present study, we examined the differences in bioavailability of ferrous sulfate and ferrous glycinate in the intestines of SD rats using Illumina sequencing technology. Digital gene expression analysis resulted in the generation of almost 128 million clean reads, with expression data for 17,089 unigenes...
November 30, 2016: Scientific Reports
https://www.readbyqxmd.com/read/27899805/circulating-tumour-dna-analysis-demonstrates-spatial-mutational-heterogeneity-that-coincides-with-disease-relapse-in-myeloma
#6
S Mithraprabhu, T Khong, M Ramachandran, A Chow, D Klarica, L Mai, S Walsh, D Broemeling, A Marziali, M Wiggin, J Hocking, A Kalff, B Durie, A Spencer
Mutational characterisation in multiple myeloma (MM) currently relies on bone marrow (BM) biopsy, which fails to capture the putative spatial and genetic heterogeneity of this multi-focal disease. Analysis of plasma (PL)-derived circulating free tumour DNA (ctDNA) as an adjunct to BM biopsy, for mutational characterisation and tracking disease progression, was evaluated. Paired BM MM cell DNA and ctDNA from 33 relapsed [RR] and 15 newly diagnosed [ND] patients were analysed for KRAS, NRAS, BRAF and TP53 mutations using the OnTarget™ Mutation Detection (OMD) platform...
November 30, 2016: Leukemia: Official Journal of the Leukemia Society of America, Leukemia Research Fund, U.K
https://www.readbyqxmd.com/read/27898590/reactivation-of-simian-immunodeficiency-virus-reservoirs-in-the-brain-of-virally-suppressed-macaques
#7
Lucio Gama, Celina M Abreu, Erin N Shirk, Sarah L Price, Ming Li, Greg M Laird, Kelly A Metcalf Pate, Stephen W Wietgrefe, Shelby L O'Connor, Luiz Pianowski, Ashley T Haase, Carine Van Lint, Robert F Siliciano, Janice E Clements
OBJECTIVE: Resting CD4 T cells have been recognized as the major cell reservoir of latent HIV-1 during antiretroviral therapy (ART). Using an simian immunodeficiency virus (SIV)/macaque model for AIDS and HIV-related neurocognitive disorders we assessed the contribution of the brain to viral latency and reactivation. DESIGN: Pigtailed macaques were dual inoculated with SIVDeltaB670 and SIV17E-Fr and treated with an efficacious central nervous system-penetrant ART...
January 2, 2017: AIDS
https://www.readbyqxmd.com/read/27897454/considerations-for-standardizing-predictive-molecular-pathology-for-cancer-prognosis
#8
Michelangelo Fiorentino, Marina Scarpelli, Antonio Lopez-Beltran, Liang Cheng, Rodolfo Montironi
Molecular tests that were once ancillary to the core business of cyto-histopathology are becoming the third most relevant workload in pathology departments after histopathology/cytopathology and before autopsies. This has resulted from innovations in molecular biology techniques, which have developed at an incredibly fast pace. Areas covered: Most of the current widely used techniques in molecular pathology such as FISH, direct sequencing, pyrosequencing, and allele-specific PCR will be replaced by massive parallel sequencing that will not be considered next generation, but rather, will be considered to be current generation sequencing...
November 29, 2016: Expert Review of Molecular Diagnostics
https://www.readbyqxmd.com/read/27894866/transcriptional-regulation-of-the-sodium-channel-gene-scn5a-by-gata4-in-human-heart
#9
Anna Tarradas, Mel Lina Pinsach-Abuin, Carlos Mackintosh, Oriol Llorà-Batlle, Alexandra Pérez-Serra, Montserrat Batlle, Félix Pérez-Villa, Thomas Zimmer, Ivan Garcia-Bassets, Ramon Brugada, Pedro Beltran-Alvarez, Sara Pagans
Aberrant expression of the sodium channel gene (SCN5A) has been proposed to disrupt cardiac action potential and cause human cardiac arrhythmias, but the mechanisms of SCN5A gene regulation and dysregulation still remain largely unexplored. To gain insight into the transcriptional regulatory networks of SCN5A, we surveyed the promoter and first intronic regions of the SCN5A gene, predicting the presence of several binding sites for GATA transcription factors (TFs). Consistent with this prediction, chromatin immunoprecipitation (ChIP) and sequential ChIP (Re-ChIP) assays show co-occupancy of cardiac GATA TFs GATA4 and GATA5 on promoter and intron 1 SCN5A regions in fresh-frozen human left ventricle samples...
November 25, 2016: Journal of Molecular and Cellular Cardiology
https://www.readbyqxmd.com/read/27892470/vertical-magnetic-separation-of-circulating-tumor-cells-for-somatic-genomic-alteration-analysis-in-lung-cancer-patients
#10
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/27888686/integration-of-isothermal-amplification-methods-in-microfluidic-devices-recent-advances
#11
REVIEW
Maria Chiara Giuffrida, Giuseppe Spoto
The integration of nucleic acids detection assays in microfluidic devices represents a highly promising approach for the development of convenient, cheap and efficient diagnostic tools for clinical, food safety and environmental monitoring applications. Such tools are expected to operate at the point-of-care and in resource-limited settings. The amplification of the target nucleic acid sequence represents a key step for the development of sensitive detection protocols. The integration in microfluidic devices of the most popular technology for nucleic acids amplifications, polymerase chain reaction (PCR), is significantly limited by the thermal cycling needed to obtain the target sequence amplification...
November 19, 2016: Biosensors & Bioelectronics
https://www.readbyqxmd.com/read/27881615/superiority-of-digital-rt-pcr-over-real-time-rt-pcr-for-quantitation-of-highly-divergent-human-rhinoviruses
#12
Ruth Hall Sedlak, Thuy Nguyen, Isabel Palileo, Keith R Jerome, Jane Kuypers
Human rhinoviruses (HRV) are comprised of 3 species, represented by over 150 genotypes. As an important human respiratory pathogen, molecular detection is an indispensable tool for diagnosis and surveillance. However, the sequence diversity of HRV genotypes poses challenges for the development of robust molecular methods that detect all genotypes with equal efficiency. This study compares the accuracy of reverse transcription-quantitative PCR (RT-qPCR) and reverse transcription-digital PCR (RT-dPCR) for quantification of HRV RNA using genotype-specific primers and probes and a consensus primer/probe set targeting the 5' non-coding region of HRV...
November 23, 2016: Journal of Clinical Microbiology
https://www.readbyqxmd.com/read/27870699/resistance-training-alters-the-proportion-of-skeletal-muscle-fibers-but-not-brain-neurotrophic-factors-in-young-adult-rats
#13
José Antonio-Santos, Diórginis José S Ferreira, Gizelle L Gomes Costa, Rhowena Jane B Matos, Ana E Toscano, Raul Manhães-de-Castro, Carol G Leandro
Antonio-Santos, J, Ferreira, DJS, Gomes Costa, GL, Matos, RJB, Toscano, AE, Manhães-de-Castro, R, and Leandro, CG. Resistance training alters the proportion of skeletal muscle fibers but not brain neurotrophic factors in young adult rats. J Strength Cond Res 30(12): 3531-3538, 2016-Resistance training (RT) is related to improved muscular strength and power output. Different programs of RT for rats have been developed, but peripheral and central response has not been evaluated directly in the same animal. To test the hypothesis that RT induces central and peripheral adaptations, this study evaluated the effects of a RT on the performance of a weekly maximum overload test, fiber-type typology, and brain neurotrophic factors in young adult rats...
December 2016: Journal of Strength and Conditioning Research
https://www.readbyqxmd.com/read/27869597/digital-pcr-for-quantifying-norovirus-in-oysters-implicated-in-outbreaks-france
#14
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/27866884/microrna-biomarkers-of-pancreatic-injury-in-a-canine-model
#15
Rodney Rouse, Barry Rosenzweig, Katie Shea, Alan Knapton, Sharron Stewart, Lin Xu, Ashok Chockalingam, Leah Zadrozny, Karol Thompson
Pancreas-enriched microRNAs have been experimentally investigated in rodents as candidate serum biomarkers of pancreatic injury with several different acute pancreatic injury models. In the present study, temporal and magnitude responses of exocrine pancreas-enriched miR-216a, miR-216b, and miR-217 and endocrine-enriched miR-375 and miR-148a were measured by droplet digital PCR in serum in a caerulein model of pancreatic injury in the dog. All 5 microRNAs followed a similar time course that mirrored the responses of the conventional serum pancreatic injury biomarkers, amylase and lipase...
November 17, 2016: Experimental and Toxicologic Pathology: Official Journal of the Gesellschaft Für Toxikologische Pathologie
https://www.readbyqxmd.com/read/27866520/-research-advancement-on-egfr-mutation-detection-of-cell-free-dna-and-tumor-cell-in-peripheral-blood-of-patients-with-non-small-cell-lung-cancer
#16
Yu Zhang, Yan Xu, Mengzhao Wang
Non-small cell lung cancer (NSCLC) is the most common type of lung cancer. Epideral growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) are the most important treatments currently for advanced NSCLC patients harboring activating EGFR gene mutations, and achieve significant clinical efficacy. T790M mutation occurs in half of NSCLC patents with acquired EGFR-TKI resistance. Screening for EGFR gene mutations in histological and/or circulating tumor cell or DNA samples of NSCLC patients can identify patients who would have a response to EGFR-TKIs or acquire resistance during the treatment...
November 20, 2016: Zhongguo Fei Ai za Zhi, Chinese Journal of Lung Cancer
https://www.readbyqxmd.com/read/27865784/a-comparison-of-cell-free-dna-isolation-kits-isolation-and-quantification-of-cell-free-dna-in-plasma
#17
Laure Sorber, Karen Zwaenepoel, Vanessa Deschoolmeester, Geert Roeyen, Filip Lardon, Christian Rolfo, Patrick Pauwels
The analysis of cell-free DNA (cfDNA) as a sensitive biomarker for cancer diagnosis and monitoring has resulted in a need for efficient and standardized cfDNA isolation. In this study, we compared the isolation efficiency of the QIAamp circulating nucleic acid kit (QIA) with four other cfDNA isolation kits: the PME free-circulating DNA Extraction Kit (PME), the Maxwell RSC ccfDNA Plasma Kit (RSC), the EpiQuick Circulating Cell-Free DNA Isolation Kit (EQ), and two consecutive versions of the NEXTprep-Mag cfDNA Isolation Kit (NpMV1/2)...
November 17, 2016: Journal of Molecular Diagnostics: JMD
https://www.readbyqxmd.com/read/27865449/characterizing-nuclear-and-mitochondrial-dna-in-spent-embryo%C3%A2-culture-media-genetic-contamination-identified
#18
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
#19
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
#20
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
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