Methodology U937 Human Immune Cells Control (No treatment) (n=4) Estrogen (5 uM) (n=4) 4-nonylphenol (5 uM) (n=4) Cultured Cells, RNA Isolation, RT (to.

Slides:

Advertisements

Similar presentations

Microarray technology and analysis of gene expression data Hillevi Lindroos.

Advertisements

Microarrays Dr Peter Smooker,

Parallel human genome analysis: Microarray-based expression monitoring of 1000 genes Mark Schena, Dari Shalon, Renu Heller, Andrew Chai, Patrick O. Brown,

Figure 1: (A) A microarray may contain thousands of ‘spots’. Each spot contains many copies of the same DNA sequence that uniquely represents a gene from.

DNA Arrays …DNA systematically arrayed at high density, –virtual genomes for expression studies, RNA hybridization to DNA for expression studies, –comparative.

1 April, 2005 Chapter C4.1 and C5.1 DNA Microarrays and Cancer.

Gene_IDGene NameGene Symbol Fold change compared to control hCG retinol dehydrogenase 8 (all-trans) RDH82.09 hCG ankyrin repeat domain.

Chapter 7 Essential Concepts in Molecular Pathology Companion site for Molecular Pathology Author: William B. Coleman and Gregory J. Tsongalis.

Introduction to DNA Microarray Technology Steen Knudsen Uma Chandran.

Methodology Control (no treatment) Estrogen (5 uM) 4-nonylphenol (5 uM) Cultured Cells, Isolated RNA, RTed to cDNA Data analyzed by Spotfire software RT-PCR.

Cell culturing and treatment U937 human immune cells (ATCC, USA) were cultured in RPMI1640 medium (Invitrogen, USA). Cells were kept in a 37ºC incubator.

Microarrays and Gene Expression Analysis. 2 Gene Expression Data Microarray experiments Applications Data analysis Gene Expression Databases.

Primary Mets Node Patient 1Patient 2Patient 3 Primary Mets Node Patient 1Patient 2Patient 3 Primary Mets Node Patient 1Patient 2Patient 3 Primary Mets.

A Nanoliter-Scale Nucleic Acid Processor with Parallel Architecture Jong Wook Hong, Vincent Studer, Giao Hang, W French Andreson, Stephen R Quake presented.

Microarrays and Gene Expression Arrays

DECIPHERING THE EARLY BOVINE HOST RESPONSE AFTER Brucella melitensis INFECTION ROSSETTI CA 1 * DRAKE K 2, LAWHON S 3, NUNES J 3, GULL T 3, KHARE S 3, EVERTS.

Microarray hybridization Usually comparative – Ratio between two samples Examples – Tumor vs. normal tissue – Drug treatment vs. no treatment – Embryo.

Microarray Data Analysis The Bioinformatics side of the bench.

Genomics A Systematic Study of the Locations, Functions and Interactions of Many Genes at Once.

Genomics A Systematic Study of the Locations, Functions and Interactions of Many Genes at Once.

Statistical Analysis for Expression Experiments Heather Adams BeeSpace Doctoral Forum Thursday May 21, 2009.

NCode TM miRNA Analysis Platform Identifies Differentially Expressed Novel miRNAs in Adenocarcinoma Using Clinical Human Samples Provided By BioServe.

Human cytomegalovirus (HCMV) is a member of the herpesvirus family. About 70-90% of the population is infected with HCMV. In healthy.

Microarray: An Introduction

SSA/Ps Syndromic SSA/Ps Sporadic HPs Sporadic Genes differentially expressed ≥ 2-fold in syndromic and sporadic SSA/Ps and hyperplastic.

Figure S1 (a) (b) Fig. S1. Hydroponics culture of Arabidopsis thaliana. (a) Illustration of the hydroponics system in the growth chamber. (b) close-up.

Date of download: 6/25/2016 Copyright © 2016 American Medical Association. All rights reserved. From: Implication of SSAT by Gene Expression and Genetic.

Microarray Experiment Design and Data Interpretation

Stephen Lory, PhD, Jeffrey K. Ichikawa, PhD CHEST

Copyright © 2015 by the American Osteopathic Association.

Example of a DNA Array (note green, yellow red colors; also note that only part of the total array is depicted)

Distinctive proliferative phase differences in gene expression in human myometrium and leiomyomata Hongbo Wang, MD, Mamatha Mahadevappa, PhD, Karen Yamamoto,

Kenneth G. Geles, Wenyan Zhong, Siobhan K

Transitory dasatinib-resistant states in KITmut t(8;21) acute myeloid leukemia cells correlate with altered KIT expression Markus D. Herrmann, Jochen.

Activated glucocorticoid and eicosanoid pathways in endometriosis

Arsenic suppresses gene expression in promyelocytic leukemia cells partly through Sp1 oxidation by Wen-Chien Chou, Hsuan-Yu Chen, Sung-Liang Yu, Linzhao.

HIS-24 regulates expression of infection-inducible genes.

Figure 1 Hierarchical clustering (HCL) outcome of all tested samples with the expression profile of the case report set as unknown Hierarchical clustering.

Learning More from Microarrays: Insights from Modules and Networks

Gene expression and genomic profiling reveal estrogen-independent ER transcriptional activity. Gene expression and genomic profiling reveal estrogen-independent.

Rose-Anne Romano, Barbara Birkaya, Satrajit Sinha

Effects of Etanercept Are Distinct from Infliximab in Modulating Proinflammatory Genes in Activated Human Leukocytes Asifa S. Haider, Irma R. Cardinale,

Volume 3, Issue 5, Pages (November 2014)

TNFα Signaling Exposes Latent Estrogen Receptor Binding Sites to Alter the Breast Cancer Cell Transcriptome Hector L. Franco, Anusha Nagari, W. Lee Kraus

Volume 12, Issue 6, Pages (December 2003)

Volume 63, Issue 4, Pages (August 2016)

Profiling Motility Signal-Specific Genes in Primary Human Keratinocytes Chieh-Fang Cheng, Jianhua Fan, Balaji Bandyopahdhay, Dennis Mock, Shengxi Guan,

Figure 1. BRCA1-associated R-Loop accumulation at a non-coding region upstream of ESR1 locus. (A) Alignment of DRIP-seq ... Figure 1. BRCA1-associated.

Characterization of the CC Chemokine Receptor 3 on Human Keratinocytes

Volume 16, Issue 4, Pages (November 2004)

Noritaka Oyama, Keiji Iwatsuki, Yoshimi Homma, Fumio Kaneko

The BRD4 bromodomain is critical for expression of SASP genes.

Volume 6, Issue 6, Pages (June 2004)

Mesenchymal Stem Cell Features of Ewing Tumors

Volume 12, Issue 9, Pages (April 2002)

ATPγS Enhances the Production of Inflammatory Mediators by a Human Dermal Endothelial Cell Line via Purinergic Receptor Signaling Kristina Seiffert,

AZA treatment induces a distinct gene-expression pattern in stromal cells. AZA treatment induces a distinct gene-expression pattern in stromal cells. (A-C)

Hierarchical clustering analysis of 7785 genes (genes with a log-ratio variation in the 25th centile and >5% missing data were excluded) (A) A heat map.

Intestinal myofibroblasts in innate immune responses of the intestine

Extracellular ATP Has Stimulatory Effects on the Expression and Release of IL-6 Via Purinergic Receptors in Normal Human Epidermal Keratinocytes Kaori.

Comparison ofMyc-induced zebrafish liver tumors with different stages of human HCC and seven mouse HCC models. Comparison ofMyc-induced zebrafish liver.

Volume 2, Issue 2, Pages (February 2014)

Multiplex Enhancer Interference Reveals Collaborative Control of Gene Regulation by Estrogen Receptor α-Bound Enhancers Julia B. Carleton, Kristofer.

Figure 1. Identification of differentially expressed messenger RNAs (mRNAs) in the The Cancer Genome Atlas (TCGA) BRCA database. (A) Heat map of the log2-fold.

A, unsupervised hierarchical clustering of the expression of probe sets differentially expressed in the oral mucosa of smokers versus never smokers. A,

A Light-Independent Allele of Phytochrome B Faithfully Recapitulates Photomorphogenic Transcriptional Networks Wei Hu, Yi-Shin Su, J. Clark Lagarias

Heat map of genes for which CR significantly altered expression versus AL. Cluster analysis of genes significantly changed by the CR intervention compared.

Genome-wide Functional Analysis Reveals Factors Needed at the Transition Steps of Induced Reprogramming Chao-Shun Yang, Kung-Yen Chang, Tariq M. Rana

Gene expression profiles of T cells.

Presentation transcript:

Methodology U937 Human Immune Cells Control (No treatment) (n=4) Estrogen (5 uM) (n=4) 4-nonylphenol (5 uM) (n=4) Cultured Cells, RNA Isolation, RT (to cDNA) Applied cDNA to microarray chips Scanned chips for detection of gene expression by chemiluminescence Data analyzed by Spotfire software RT-PCR (using ER-beta) Reference gene: B-actin 2% agarose gel electrophoresis A) Scatterplots of gene expression analyzed by DNA microarray. The probe ID’s for each gene is represented by the x-axis and the fold changes in gene expression for the experimental samples based on the fluorescence-based detection signals of the mRNA levels in the control are represented by the y-axis. (B) Remaining genes after filtering by modulation and normalization with the b-actin reference gene. Genes that exhibited 2-fold regulation A B Genes altered with E2 treatment after 48 hours Genes altered with 4-NP treatment after 48 hours This figure indicates that E2 and NP have the potential to down regulate the estrogen receptor beta (ESR1) and the estrogen receptor related beta (ESRR-β) and up regulate repressor estrogen receptor activity (REA) (at their respective concentrations). ESR1 does not exhibit a significant pattern. These data appear to demonstrate that NP as well as enhanced estrogen levels diminishes estrogen beta activity negatively. Probe_IDGene SymbolGene NameControlE2 5µM4NP 5µM ESR1estrogen receptor ESR2estrogen receptor 2 (ER beta) ESRRBestrogen-related receptor beta REA repressor of estrogen receptor activity RT-PCR Analysis of Intensity of ESR2 gene expression normalized with the internal control gene beta-actin. This data serves as a validation for the microarray analysis. RT-PCR Gene Expression Profile ESR2 β-actin These gels exhibit expression of the ESR2 gene the β-actin reference gene. Lane 1=control, Lane 2 =E2 5uM, Lane 3=control and Lane 4 = 4-NP 5uM. Gene profiles of BRCA1 and BRCA2. The E2 treament induced a fold repression of more than two fold while the NP treatment had no significant effect. This phenomenon was observed for 23/45 genes in the portfolio involved in the onset of breast cancer. Gene Expressions Profiles of BRCA1 and BRCA2 Relative Gene Expression (Signal of treatment chip/signal of chip treatment) Relative Gene Expression (Signal of treatment chip/signal of chip treatment) # Of Genes Induced By E2 and 4-NP by a > or = 2-Fold Change from the Control E24-NP 14,913 (45.0%) 414 (1.2%) 54 (60.0%) 7 (7.8%) 8 (8.9%) 633 (1.9%) E24-NP All genes from the whole- genome nanochips (33,155) Genes from customized portfolio (90) A (A) Venn Diagram which shows the number of genes that exhibited significant changes in gene expression after the application of E2 and concentrations of NP. (B) A normalized hierarchical clustering heat map performed in order to similarities of genes and to what extent they were affected. Genes that were downregulated significantly are illustrated by the green while red denotes genes that were upregulated by at least 2-fold. Comparison of Induced Genes/Hierarchical Clustering Heat Map About 45.0% (14,913) exhibited significant changes in gene expression (> 2-fold) by E2 while approximately 1.91% (633) B E2 (5uM) 4-NP (5uM) Hierarchical Clustering Normalized Heat Map B Graphical Representation of Significantly Altered Genes Signal-to-Noise Values of Four Genes Involved in Estrogen Signaling