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Quantitative RT-PCR

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https://www.readbyqxmd.com/read/28738328/a-novel-role-for-mir-520a-3p-in-regulating-egfr-expression-in-colorectal-cancer
#1
Rui Zhang, Rui Liu, Chang Liu, Yahan Niu, Jianguo Zhang, Baoliang Guo, Chen-Yu Zhang, Jing Li, Jie Yang, Xi Chen
BACKGROUND/AIMS: MicroRNAs (miRNAs) have been consistently demonstrated to be involved in colorectal cancer as either tumour oncogenes or tumour suppressors. However, the detailed role of miR-520a-3p in colorectal cancer remains poorly understood. METHODS: Quantitative RT-PCR and western blotting assays were used to measure miR-520a-3p and EGFR expression levels in colorectal cancer tissues, respectively. Luciferase reporter assay was employed to validate the direct targeting of EGFR by miR-520a-3p...
July 24, 2017: Cellular Physiology and Biochemistry
https://www.readbyqxmd.com/read/28736583/microrna-profiling-of-ovarian-granulosa-cell-tumours-reveals-novel-diagnostic-and-prognostic-markers
#2
Wei-Tzu Cheng, Roseanne Rosario, Anita Muthukaruppan, Michelle K Wilson, Kathryn Payne, Peter C Fong, Andrew N Shelling, Cherie Blenkiron
BACKGROUND: The aim of this study was to explore the clinical utility of microRNAs (miRNAs) as improved markers of ovarian granulosa cell tumours (GCTs) for cancer diagnosis and prognosis prediction. Current histopathological and genetic markers, such as the presence of a FOXL2 gene mutation to distinguish between the two major subtypes are not wholly accurate and as such novel biomarkers are warranted. METHODS: The miRNA expression profiles of five formalin-fixed, paraffin-embedded (FFPE) adult-GCTs and five juvenile-GCTs were assessed using Affymetrix miRNA 3...
2017: Clinical Epigenetics
https://www.readbyqxmd.com/read/28735486/detection-and-quantification-of-acute-myeloid-leukemia-associated-fusion-transcripts
#3
Jonathan Schumacher, Philippe Szankasi, Todd W Kelley
Real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR)-based detection of abnormal fusion transcripts is an important strategy for the diagnosis and monitoring of patients with acute myeloid leukemia (AML) with t(8;21)(q22;q22); RUNX1-RUNX1T1, inv(16)(p13.1;q22); CBFB-MYH11 or t(15;17)(q22;q12); PML-RARA. In RT-qPCR assays, patient-derived cDNA is subjected to amplification using PCR primers directed against the fusion transcript of interest as well as a reference gene for normalization...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28735081/anti-inflammatory-activity-of-cecropin-a2-from-musca-domestica
#4
Rui-Yang Wei, Jie Bai, Meng-Fei Zhao, Bin Xu, Wen-Jia Li, Feng-Xian Wei, Yan-Yan Xi, Shao-Yu Li
This study aimed to investigate the anti-inflammatory activity of Musca domestica cecropin-A2 (Mdc-A2) toward Staphylococcus aureus (S. aureus) to learn more about their immunological functions. RAW264.7 cells were transfected with recombinant lentiviruses introduce pLEX-Mdc-A2into the RAW264.7 cell line (RAW-Mdc-A2). The RAW264.7 cell line with empty pLEX (RAW-pLEX) was produced in the same manner as a negative control. Real-time quantitative reverse transcription PCR (RT-PCR) was performed to analyze the mRNA expression of TNF-a, IL-1β, NFκB-1 and NFκB-2 in S...
July 19, 2017: Microbial Pathogenesis
https://www.readbyqxmd.com/read/28734156/a-novel-hypoxia-response-element-regulates-oxygen-related-repression-of-tissue-factor-pathway-inhibitor-in-the-breast-cancer-cell-line-mcf-7
#5
Xue Yan Cui, Grethe Skretting, Mari Tinholt, Benedicte Stavik, Anders Erik Astrup Dahm, Kristine Kleivi Sahlberg, Sandip Kanse, Nina Iversen, Per Morten Sandset
BACKGROUND: Hypoxia is one of the most pervasive physiological stresses in solid tumors. We have previously demonstrated that tissue factor (TF) pathway inhibitor (TFPI) expression was transcriptionally repressed by the activation of hypoxia inducible factor (HIF)-1α under hypoxic conditions. However, the role of HIF-2α, also known as endothelial PAS domain-containing protein 1 (EPAS1), on TFPI expression remains unclear. AIM: To explore the role of HIF-2α/EPAS1 in the regulation of TFPI expression under hypoxia in breast cancer cells...
July 14, 2017: Thrombosis Research
https://www.readbyqxmd.com/read/28733893/effects-of-new-generation-tmem16a-inhibitors-on-calcium-activated-chloride-currents-in-rabbit-urethral-interstitial-cells-of-cajal
#6
Stephen Fedigan, Eamonn Bradley, Timothy Webb, Roddy J Large, Mark A Hollywood, Keith D Thornbury, Noel G McHale, Gerard P Sergeant
Interstitial cells of Cajal (ICC) isolated from the rabbit urethra exhibit Ca(2+)-activated Cl(-) currents (I ClCa) that are important for the development of urethral tone. Here, we examined if TMEM16A (ANO1) contributed to this activity by examining the effect of "new-generation" TMEM16A inhibitors, CACCinh-A01 and T16Ainh-A01, on I ClCa recorded from freshly isolated rabbit urethral ICC (RUICC) and on contractions of intact strips of rabbit urethra smooth muscle. Real-time quantitative PCR experiments demonstrated that TMEM16A was highly expressed in rabbit urethra smooth muscle, in comparison to TMEM16B and TMEM16F...
July 21, 2017: Pflügers Archiv: European Journal of Physiology
https://www.readbyqxmd.com/read/28732331/an-evaluation-of-dna-damage-in-human-lymphocytes-and-sperm-exposed-to-methyl-methanesulfonate-involving-the-regulation-pathways-associated-with-apoptosis
#7
Khaled Habas, Mojgan Najafzadeh, Adolf Baumgartner, Martin H Brinkworth, Diana Anderson
Exposure to DNA-damaging agents produces a range of stress-related responses. These change the expression of genes leading to mutations that cause cell cycle arrest, induction of apoptosis and cancer. We have examined the contribution of haploid and diploid DNA damage and genes involved in the regulation of the apoptotic process associated with exposure, The Comet assay was used to detect DNA damage and quantitative RT-PCR analysis (qPCR) to detect gene expression changes in lymphocytes and sperm in response to methyl methanesulfonate...
June 23, 2017: Chemosphere
https://www.readbyqxmd.com/read/28732075/comprehensive-selection-of-reference-genes-for-quantitative-rt-pcr-analysis-of-murine-extramedullary-hematopoiesis-during-development
#8
Giuliana Medrano, Peihong Guan, Amanda J Barlow-Anacker, Ankush Gosain
The purpose of this study was to perform a comprehensive evaluation and selection of reference genes for the study of extramedullary hematopoiesis during development and the early post-natal period. A total of six candidate reference genes (ACTB, GAPDH, HPRT1, PPID, TBP, TUBB3) in four organs (heart, liver, spleen, and thymus) over five perinatal time points (Embryonic days 14.5, 16.5, 18.5, Post-natal days 0, 21) were evaluated by quantitative real-time PCR. The expression stability of the candidate reference genes were analyzed using geNorm, NormFinder, Bestkeeper, Delta CT method, and RefFinder software packages...
2017: PloS One
https://www.readbyqxmd.com/read/28732013/the-kp1_4563-gene-is-regulated-by-the-camp-receptor-protein-and-controls-type-3-fimbrial-function-in-klebsiella-pneumoniae-ntuh-k2044
#9
Mei Luo, Shiya Yang, Xuan Li, Pin Liu, Jian Xue, Xipeng Zhou, Kewen Su, Xuan Xu, Ying Qing, Jingfu Qiu, Yingli Li
Klebsiella pneumoniae (K. pneumoniae) is an opportunistic pathogen that can adhere to host cells or extracellular matrix via type 1 and type 3 fimbriae. KP1_4563 is a gene encoding a hypothetical protein in K. pneumoniae NTUH-K2044. KP1_4563 is located between the type 1 and type 3 fimbrial gene clusters and is likely associated with fimbrial function given its putative conserved domains of unknown function (DUF1471). Cyclic AMP receptor protein (CRP) regulates virulence-related gene expression and is a crucial transcriptional regulator in many bacteria...
2017: PloS One
https://www.readbyqxmd.com/read/28731788/identification-of-osr2-transcriptional-target-genes-in-palate-development
#10
X Fu, J Xu, P Chaturvedi, H Liu, R Jiang, Y Lan
Previous studies have identified the odd-skipped related 2 (Osr2) transcription factor as a key intrinsic regulator of palatal shelf growth and morphogenesis. However, little is known about the molecular program acting downstream of Osr2 in the regulation of palatogenesis. In this study, we isolated palatal mesenchyme cells from embryonic day 12.5 (E12.5) and E13.5 Osr2(RFP/+) and Osr2(RFP/-) mutant mouse embryos and performed whole transcriptome RNA sequencing analyses. Differential expression analysis of the RNA sequencing datasets revealed that expression of 70 genes was upregulated and expression of 61 genes was downregulated by >1...
July 1, 2017: Journal of Dental Research
https://www.readbyqxmd.com/read/28731464/trophoblast-survival-signaling-during-human-placentation-requires-hsp70-activation-of-mmp2-mediated-hbegf-shedding
#11
Chandni V Jain, Philip Jessmon, Charbel T Barrak, Alan D Bolnick, Brian A Kilburn, Michael Hertz, D Randall Armant
Survival of trophoblast cells in the low oxygen environment of human placentation requires metalloproteinase-mediated shedding of HBEGF and downstream signaling. A matrix metalloproteinase (MMP) antibody array and quantitative RT-PCR revealed upregulation of MMP2 post-transcriptionally in human first trimester HTR-8/SVneo trophoblast cells and placental villous explants exposed to 2% O2. Specific MMP inhibitors established the requirement for MMP2 in HBEGF shedding and upregulation. Because α-amanitin inhibited the upregulation of HBEGF, differentially expressed genes were identified by next-generation sequencing of RNA from trophoblast cells cultured at 2% O2 for 0, 1, 2 and 4 h...
July 21, 2017: Cell Death and Differentiation
https://www.readbyqxmd.com/read/28731269/biofilm-formation-and-antibiotic-resistance-in-klebsiella-pneumoniae-urinary-strains
#12
Claudia Vuotto, Francesca Longo, Chiara Pascolini, Gianfranco Donelli, Maria Pia Balice, Maria Francesca Libori, Valentina Tiracchia, Antonino Salvia, Pietro Emanuele Varaldo
AIMS: Multi-drug resistant Klebsiella pneumoniae has become a relevant healthcare-associated pathogen. Capsule, type 1 and 3 fimbriae (mrkA gene), type 2 quorum-sensing system (luxS), synthesis of D-galactan I (wbbM), LPS transport (wzm) and poly-beta-1,6-N-acetyl-D-glucosamine (pgaA) seem involved in K. pneumoniae biofilm. Non-enzymatic antibiotic resistance is related to non-expression or mutation of porins (OmpK35 and OmpK36), and efflux pump (acrB) over-expression. The aim of this study was to analyse some virulence factors of K...
July 21, 2017: Journal of Applied Microbiology
https://www.readbyqxmd.com/read/28730955/itraq-based-proteomic-analysis-of-appsw-ind-mice-provides-insights-into-the-early-changes-in-alzheimer-s-disease
#13
Nan Li, Pinghong Hu, Tiantian Xu, Huan Chen, Xiaoying Chen, Jianwen Hu, Xifei Yang, Lei Shi, Jian-Hong Luo, Junyu Xu
BACKGROUND: Several proteins have been identified as potential diagnostic biomarkers in imaging, genetic, or proteomic studies in Alzheimer disease (AD) patients and mouse models. However, biomarkers for presymptom diagnosis of AD are still under investigation, as are the presymptom molecular changes in AD pathogenesis. OBJECTIVE: In this study, we aim to analyzed the early proteomic changes in APPSw,Ind mice and to conduct further functional studies on interesting proteins...
July 19, 2017: Current Alzheimer Research
https://www.readbyqxmd.com/read/28730333/identification-of-a-novel-leukemic-specific-splice-variant-of-dnmt3b-and-its-stability
#14
Prachi Singh, Sarvagalla Sailu, Elango Palchamy, Mohane Selvaraj Coumar, Sudhakar Baluchamy
DNA methyltransferases make use of alternative splicing mechanism to generate various splice variants, but their role(s) in modulating DNA methylation patterns in the cells is unclear. Notably, DNMT3B alone contains nearly 40 different splice variants. In this study, we have identified a novel splice variant of DNMT3B, which lacks exon 7 and 10 from leukemic cell lines which we termed as DNMT3B9. The exon 7 codes for the major part of PWWP domain, and exon 10 inclusion serves as a pluripotent marker. By quantitative RT-PCR using exon-exon junction-specific primers, we showed higher level of DNMT3B9 transcripts in several leukemic cell lines...
August 2017: Medical Oncology
https://www.readbyqxmd.com/read/28730102/anti-scarring-effects-of-butaprost-on-human-subconjunctival-tenon-s-fibroblasts
#15
Jong Hoon Shin, Je Hyun Seo, Jae Ho Jung, Tae Woo Kim
AIM: To investigate the toxicity of the E-prostanoid 2 (EP2) receptor agonist, butaprost against human subconjunctival (Tenon's capsule) fibroblasts, and to determine the underlying mechanism. METHODS: We isolated Tenon's fibroblasts from the subconjunctival area of healthy subjects and evaluated the types of EP receptors expressed using quantitative real-time reverse transcription polymerase chain reaction (RT-PCR). The toxicity of butaprost against the fibroblasts was evaluated using methyl thiazolyl tetrazolium and lactic dehydrogenase assays...
2017: International Journal of Ophthalmology
https://www.readbyqxmd.com/read/28728902/role-of-mirnas-in-development-and-disease-lessons-learnt-from-small-organisms
#16
REVIEW
Swati Chandra, Divya Vimal, Divya Sharma, Vipin Rai, Subash Chandra Gupta, D Kar Chowdhuri
MicroRNAs (miRNAs) constitute a class of small (21-24 nucleotides) non-coding RNAs that regulate gene expression at the post-transcriptional level. Caenorhabditis elegans, Drosophila melanogaster, and many other small organisms have been instrumental in deciphering the biological functions of miRNAs. While some miRNAs from small organisms are highly conserved across the taxa, others are organism specific. The miRNAs are known to play a crucial role during development and in various cellular functions such as cell survival, cell proliferation, and differentiation...
July 17, 2017: Life Sciences
https://www.readbyqxmd.com/read/28728562/low-oxygen-tension-increases-mitochondrial-membrane-potential-and-enhances-expression-of-antioxidant-genes-and-implantation-protein-of-mouse-blastocyst-cultured-in-vitro
#17
Yun-Yi Ma, Huei-Wen Chen, Chii-Ruey Tzeng
BACKGROUND: In human IVF, embryos cultured with a lower O2 tension (5%) can give rise to higher success rates when compared with normoxic conditions (20%). However, the mechanisms behind the beneficial effects of reduced oxygen tension in embryogenesis remain unclear. The aim of this study was to evaluate the expression of oxygen related and antioxidant genes and mitochondrial function in mouse embryo cultured under hypoxic and normoxic conditions, to investigate the beneficial effect of low oxygen tension in preimplantation embryogenesis...
July 20, 2017: Journal of Ovarian Research
https://www.readbyqxmd.com/read/28727318/appropriate-cyclic-tensile-strain-promotes-biological-changes-of-cranial-base-synchondrosis-chondrocytes
#18
F Chu, Q Feng, Z Hu, G Shen
OBJECTIVES: This study was designed to clarify biological changes of cranial base synchondrosis chondrocytes (CBSCs) upon cyclic tensile strain (CTS) loading which simulated orthopaedic mechanical protraction on cranial base synchondroses (CBS). MATERIAL AND METHODS: A two-step digestion method was used to isolate CBSCs obtained from 1-week-old Sprague Dawley rats. Immunohistochemical staining of type II collagen and Sox9 was conducted to identify chondrocytes. A CTS of 1 Hz and 10% elongation was applied to the second passage of CBSCs by FX-5000™ Tension System for 24 hours...
August 2017: Orthodontics & Craniofacial Research
https://www.readbyqxmd.com/read/28725241/mir-378-suppresses-the-proliferation-migration-and-invasion-of-colon-cancer-cells-by-inhibiting-sdad1
#19
Mingxi Zeng, Linlin Zhu, Liangping Li, Changming Kang
BACKGROUND: MicroRNAs (miRNAs) play important roles in the growth and metastasis of colon cancer. It is known that one set of miRNAs are dysregulated in colon cancer cells, but the mechanism of their role in cancer development is still largely unknown. Our study focuses on the role of miR-378 in colon cancer cells. METHODS: Human colon cancer tissues and adjacent non-tumor tissues were collected from patients diagnosed in pathological examinations. In addition, human colon cancer cell lines LoVo, CaCo2, SW1116, SW480 and HCT-116, and a normal colonic mucosa cell line NCM460 were included...
2017: Cellular & Molecular Biology Letters
https://www.readbyqxmd.com/read/28725105/porcine-reproductive-and-respiratory-syndrome-virus-prrsv-in-pig-meat
#20
Philippe Raymond, Christian Bellehumeur, Malliga Nagarajan, Diane Longtin, Alexandra Ferland, Peter Müller, Rachel Bissonnette, Carole Simard
Porcine reproductive and respiratory syndrome, caused by the porcine reproductive and respiratory syndrome virus (PRRSV), is an economically important disease in the swine industry. Previous studies demonstrated the presence of the virus in pig meat and its transmissibility by oral consumption. This study further analyzed the infectivity of PRRSV in commercial pig meat. Fresh bottom meat pieces (n = 1500) randomly selected over a period of 2 y from a pork ham boning plant located in Quebec, Canada, were tested by reverse transcriptase polymerase chain reaction (RT-PCR)...
July 2017: Canadian Journal of Veterinary Research, Revue Canadienne de Recherche Vétérinaire
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