title: MicroRNAs altered in colon adenomas

status: Public on Aug 01 2012

city: Bethesda

first: Aaron

last: Schetter

address: 37 Convent Drive, Building 37, Room 3050

department: Laboratory of Human Carcinogenesis

institute: National Cancer Institute

summary: 95 microRNA probes were found to have altered expression in colon adenomas compared to paired non-adenoma tissue

platform_organism:
Homo sapiens
Mus musculus

overall_design: Total mRNA was extracted from colon tissue that was flash frozen immediately after surgery.  A total of 36 tissues were used for microarray analysis.  Briefly, 5 ug of RNA were biotin labeled and hybridized to OSU-CCC microRNA microarrays version 4.0.  We then analyzed associations comparing colon adenomas with paired non-adenoma tissue.

first: Curtis

last: Harris


title: Differential expression analysis of HFF cells in G0 phase compared to cells in G1 using TileShuffle

status: Public on Jun 06 2012

city: Leipzig

first: Kristin

last: Reiche

address: Perlickstr. 1

department: Diagnostics

country: Germany

institute: Fraunhofer Institute for Cell Therapy and Immunology

type: Expression profiling by genome tiling array

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Rho kinase paralog regulation of gene transcription

status: Public on Jun 01 2012

city: El Paso

first: Brad

middle: Allen

last: Bryan

state: Texas

address: 5001 El Paso Drive

department: Biomedical Sciences

country: USA

institute: Texas Tech University Health Sciences Center

summary: The serine/threonine protein kinase paralogs ROCK1 & 2 have been implicated as essential modulators of angiogenesis; however their paralog-specific roles in endothelial function are unknown.  Microarray analysis of ROCK knockdown lines revealed overlapping and unique control of global transcription by the paralogs, and a reduction in the transcriptional regulation of just under 50% of VEGF responsive genes.

overall_design: MS1 mouse endothelial cells were stably transfected with non-targeting, ROCK1 specific, or ROCK2 specific shRNA plasmids.  Cells were grown to confluence and treated with sham or 2.5 ng/ml human recombinant vascular endothelial growth factor (VEGF) for 12 hours.  RNA from six independent biological replicates from each engineered cell line were collected, pooled, and subjected to triplicate technical replicates of microarray analysis.

middle: A

first: Dianne

middle: C

last: Mitchell


title: Microarray analysis of microRNA expression in basal cell carcinoma

status: Public on May 01 2012

city: Bochum

first: Michael

last: Sand

state: NRW

address: Gudrunstr. 56

department: Depatment of Dermatology, Venereology and Allergology

institute: Ruhr-University Bochum

summary: Perturbations in the expression profiles of microRNAs (miRNAs) have been reported for a variety of different cancers. Basal cell carcinoma (BCC) of the skin has not been systematically evaluated regarding differentially expressed miRNAs. Patients with BCC (n=7) were included in the study. Punch biopsies were harvested from the center of the tumors (lesional, n=7) and from sites of adjacent non-lesional skin (intraindividual control, n=7). Microarray miRNA expression profiles were detected based on an Agilent platform and miRBase 16. For validation purposes the expression of a subset of dysregulated miRNAs were measured by means of TaqMan real-time reverse transcription polymerase chain reaction (RT-PCR).

overall_design: Seven patients with basal cell carcinoma (BCC) were enrolled in the present study. While BCCs were excised with cold steel under local anaesthesia, a 4-mm punch biopsy was taken from the center of the tumor and from adjacent non-lesional epithelial skin (control), immediately stored in RNAlater (Qiagen, HIlden, Germany) and stored at -80 °C. miRNAs were isolated with miRNeasy Mini Kit (Quiagen, Hilden, Germany) according to the manufacturer’s protocol. RNA concentration, purity and RNA integrity number (RIN) were determined with NanoDrop ND-1000 spectral photometer (Peqlab, Erlangen, Germany), Agilent 2100 Bioanalyzer and the RNA 6000 NanoLabCHip Kits (both Agilent Technologies, Santa Clara, USA). total RNA samples were spiked with the MicroRNA Spike-In Kit (Agilent Technologies, Santa Clara, USA) for assessment of labeling and hybridization efficiencies. After the spiked  total  RNA  was  treated  with  alkaline  calf  intestine  phosphatase  (CIP), a labeling reaction was started with 100 ng total-RNA per sample.  T4 RNA ligase, incorporating Cyanine 3-Cytidine biphosphate (miRNA Complete Labeling and Hyb Kit, Agilent Technologies, Santa Clara, USA) was used to label the dephosphorylated RNA. Cyanine-3-labeled miRNA samples were subsequently prepared for One-Color based hybridization (Complete miRNA Labeling and Hyb Kit, Agilent Technologies, Santa Clara, USA). Hybridization of the labeled miRNA samples with Human miRNA Microarrays Release 16.0 (Agilent Technologies, Santa Clara, USA) was done at 55°C for 20 hrs. After washing microarray slides with increasing stringency (Gene Expression Wash Buffers, Agilent Technologies, Santa Clara) they were dried with acetonitrile (Sigma-Aldich, St.Louis, USA). Agilent DNA Microarray Scanner (Agilent Technologies, Santa Clara, USA) was used detect fluorescent signal intensities. They were detected with the Scan Control A.8.4.1 Software (Agilent Technologies, Santa Clara, USA) and extracted from images by using the Feature Extraction 10.7.3.1 Software (Agilent Technologies, Santa Clara, USA). All the steps described were carried out according to the manufacturer´s instructions.

first: Marina

last: Skrygan

first: Daniel

first: Dimitrios

last: Georgas

first: Stephan

last: Hahn

first: Thilo

last: Gambichler

first: Peter

last: Altmeyer

first: Falk-Georges

last: Bechara


title: Differential expression analysis of HFF cells in G0 phase compared to cells in G1 using MAT


title: MicroRNAs expression profiling during breast cancer progression

status: Public on Jun 23 2012

city: Nottingham

first: Sarkawt

last: Khoshnaw

address: Queens Medical Centre

country: United Kingdom

institute: University of Nottingham

summary: MicroRNAs (miRNAs) are known to be deregulated in human breast cancer (BC). The purpose of the current study was to investigate the expression of miRNAs in different stages of BC to assess their biological value in BC progression. MiRNA expression was assessed in a series of BC patients (n=7) with distinct stages of tumour progression (Normal, in-situ (DCIS), primary invasive BC and nodal metastases) to evaluate miRNA differential expression. We used an Agilent miRNA microarray based platform which uses miRBase 16 to screen for 1205 Homo sapiens (hsa) and 144 human viral miRNA candidates. To validate the microarray data, the expression of two deregulated miRNAs was measured by TaqMan quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR).

overall_design: This study was conducted Formalin Fixed and Paraffin Embedded (FFPE) specimens from 7 female patients. Twenty μm thick FFPE sections were cut and mounted on PALM Membrane Slides. Under direct microscopic visualization, tissues of interest were micro-dissected using PALM non-contact Laser catapulsion instrument (P.A.L.M. Microlaser Technologies, Carl Zeiss Ltd). Total RNA, including miRNAs, was extracted with the TRIzol reagent (Invitrogen, supplied by Fisher Scientific UK Ltd) according to the manufacturer’s instructions. Total RNA was quantified with Quant-iT RiboGreen RNA Quantitation Kit (Fisher Scientific UK Ltd), which is an accurate method of measuring total RNA with a detection limit of 0.001-1ng/µl. RNA purity and RNA integrity number (RIN) were determined on an Agilent 2100 Bioanalyzer and RNA 6000 NanoLabChip Kits (both Agilent Technologies, Santa Clara, USA), and the RIN of all the samples was >2 but <3. This is expected for FFPE breast samples and is acceptable by Agilent Technologies, Santa Clara, USA, for FFPE samples to undergo miRNA microarray analysis. MiRNAs extracted from laser microdissected tissue components were profiled using Agilent microRNA microarray profiling system (Agilent Technologies, Santa Clara, USA). Total RNA samples were spiked using the MicroRNA Spike-In Kit (Agilent Technologies, Santa Clara, USA) to evaluate efficiencies of labelling and hybridisation. Total RNA was treated with Calf Intestinal Alkaline Phosphatase (CIP), and then 100 ng of total RNA was used per sample to initiate a labelling reaction. Ligation master mix for T4 RNA ligase, which includes Cyanine 3-Cytidine biphosphate (Cyanine 3-pCp) (Complete miRNA Labelling and Hyb Kit, Agilent Technologies, Santa Clara, USA) was used to label the dephosphorylated RNA. Cyanine-3-labelled miRNA samples were hybridised to human miRNA microarrays (Release 16.0, 8x60K) (Agilent Technologies, Santa Clara, USA) at 55°C for 20 hrs. Microarray slides were washed with increasing stringency (Gene Expression Wash Buffers, Agilent Technologies, Santa Clara), and subsequently dried with acetonitrile (Sigma-Aldrich, St. Louis, USA). Fluorescent signal intensities were detected on an Agilent Microarray Scanner (Agilent Technologies, Santa Clara, USA) using the Scan Control A.8.4.1 Software (Agilent Technologies, Santa Clara, USA) and extracted from the images with the Feature Extraction 10.7.3.1 Software (Agilent Technologies, Santa Clara, USA).

middle: M

first: Jorge

middle: S

last: Reis-filho

first: Maryou

middle: B

last: Lambros

first: Graham

middle: R

last: Ball

first: Emad

last: Rakha

first: Tarek

last: Abdel-Fatah

first: Christopher

last: Nolan

first: Zsolt

last: Hodi

first: R

middle: D

last: Macmillan

first: Ian

middle: O

last: Ellis

first: Andrew

last: Green


title: Transcriptome profiles of hematopoietic lineages from BAP1 WT and KO mice

status: Public on Nov 07 2012

city: South San Francisco

first: Jinfeng

last: Liu

state: CA

address: 1 DNA Way, MS93

department: Bioinformatics and Computational Biology

institute: Genentech Inc.

summary: BAP1 has been studied as a tumor suppressor. Our aim was to characterize genes that were altered in various hematopoietic cell types upon deletion of BAP1.

overall_design: BAP1 WT versus KO cells.

first: Anwesha

last: Dey


title: Unirradiated vs. irradiated wounded planarians

status: Public on Apr 30 2012

city: Cambridge

first: Danielle

last: Wenemoser

state: MA

address: 9 Cambridge Center

institute: Whitehead Institute for Biomedical Research

summary:
Transcriptional profiling of Schmidtea mediterranea planarians that have been amputated transversely and left to regenerate for different time points (UNIRR); the same experiment was performed on irradiated - neoblast depleted animals (IRR).
The purpose of this experiment was to identify transcriptional changes that are induced within the first 12h following amputation. In order to distinguish between gene expression changes in the differentiated tissue and the neoblasts, a second set of time course data was created from irradiated animals

overall_design: Two-color experiment, 3 replicates per condition (including 10 animals/replica)


title: Transcriptional profiling of Schmidtea mediterranea planarians (smed-runt-1 RNAi vs. control RNAi)

summary: Transcriptional profiling of Schmidtea mediterranea planarians that have been subjected to 2 weeks of runt-1 or control RNAi, amputated and RNA was directly collected at 5min, 9h, and 24h, or neoblasts (X1) were isolated from 9h wounded animals and subsequently RNA was extracted


title: Transcriptome profile of peripheral blood mononuclear cells in children with acute otitis media caused by Nontypeable Haemophilus influenzae

status: Public on Mar 15 2012

city: rochester

first: keyi

last: liu

address: 1425 portland ave

institute: Rochester General Research Institute

summary: Nontypeable Haemophilus influenzae (NTHi) is a common causative organism of acute otitis media (AOM) in children. A human cDNA microarray comprising 30,968 human genome probes was used to evaluate the transcriptional changes that occur in peripheral blood mononuclear cells (PBMC) at the onset of clinical AOM caused by NTHi infection in children after comparison of microarray results with the pre-infection healthy stage of the same children.

overall_design: Four to ten milliliters of heparinized peripheral venous blood was collected from children at 6 to 30 months of age when they were in acute otitis media (AOM) stage and pre-infection healthy stage. The diagnosis of AOM was based on symptoms and signs as well as Nontypeable Haemophilus influenzae culture positive in the middle ear fluid. Patients with polymicrobial infections, history of immunodeficiency, history of chronic or recurrent AOM, chronic disease, or receiving steroids or other immunomodulatory agents were excluded. Peripheral blood mononuclear cells (PBMCs) were isolated by the Ficoll gradient and total RNA was extracted from PBMCs using the QIAamp RNA blood Mini Kit (Qiagen, Maryland, USA) according to manufacturer’s instructions. Double-stranded cDNA generated from total RNA was labeled with Cyanine-5 and subsequently hybridized to Human OneArray glass slides according to the manufacturer's standard protocols (PhalanxBio Inc, CA, USA). Microarrays were scanned at 5 μm resolution using an Agilent scanner. Raw intensity signals for each microarray were captured using a Molecular Dynamics Axon 4100A scanner, measured using GenePixPro™ Software. The data from all microarrays in each experimental set was then analyzed using Omicsoft Array Studio software; control and missing features were removed, and the remaining signals were quantile normalized.

first: Keyi

first: Michael

last: Pichichero


title: MHC I-associated peptides preferentially derive from transcripts bearing miRNA response elements

status: Public on Mar 25 2012

city: Montreal

first: Diana

middle: Paola

last: Granados

state: Quebec

address: C.P. 6128, succursale Centre-ville

department: Medecine

country: Canada

institute: IRIC

summary:
MHC I-peptides (MIPs) play an essential role in normal homeostasis and diverse pathological conditions. MIPs derive mainly from defective ribosomal products (DRiPs), a subset of nascent proteins that fail to achieve a proper conformation and whose physical nature remains elusive. We used high-throughput proteomic and transcriptomic methods to unravel the structure and biogenesis of MIPs presented by HLA-A and -B molecules on human EBV-infected B lymphocytes from four subjects. We found that although HLA-different subjects present different MIPs derived from different proteins, these MIPs originate from proteins that are functionally interconnected and implicated in similar biological pathways. Secondly, the MIP repertoire of human B cells showed no bias toward conserved vs. polymorphic genomic sequences, derived preferentially from abundant transcripts and conveyed to the cell surface a cell type-specific signature. Finally, we discovered that MIPs derive preferentially from transcripts bearing miRNA response elements. Furthermore, while MIPs of HLA-disparate subjects are coded by different sets of transcripts, these transcripts are regulated by mostly similar miRNAs. Our data support an emerging model in which i) the generation of MIPs by a transcript depends on its abundance and DRiP rate, and ii) the DRiP rate is regulated to a large extent by miRNAs.

We performed genome-wide miRNA expression profiling of B cells and non-lymphoid cells to confirm that miRNAs predicted to regulate transcripts source of MHC I-associated peptides are expressed in a cell-specific manner.

overall_design: miRNA profiling was done on (i) 4 B-lymphoblastoid cell lines (B-LCLs) obtained from the peripheral blood mononuclear cells of 4 subjects, (ii) CD19+CD20- B cells and CD19+CD20+ B cells of one of the subjects, (iii) HeLa cells, and (iv) HEK293 cells.

middle: P

first: Wafaa

last: Yahyaoui

first: Céline

last: Laumont

first: Tariq

last: Daouda

first: Tara

last: Muratore-Schroeder

first: Caroline

last: Cote

first: Jean-Philippe

last: Laverdure

first: Sébastien

last: Lémieux

first: Pierre

last: Thibault

first: Claude

last: Perreault


title: Differential expression analysis of HFF cells in G0 phase compared to cells in G1 using TAS

first: Kristin

last: Reiche


title: GS003: Tight cooperation between Mot1p and NC2β in regulating genome-wide transcription, repression of transcription following heat shock induction and genetic interaction with SAGA.

city: Utrecht

first: Marian

last: Groot Koerkamp

state: Utrecht

address: Heidelberglaan 25

department: Research

country: Netherlands

institute: Princess Maxima Center for Pediatric Oncology

summary: TATA-binding protein (TBP) is central to the regulation of transcription initiation. Recruitment of TBP to target genes can be positively regulated by one of two basal transcription factor complexes: SAGA or TFIID. Negative regulation of TBP promoter association can be performed by Mot1 or the NC2 complex. Recent evidence suggest that Mot1, NC2, and TBP form a DNA-dependent protein complex. Here, we compare the functions of Mot1 and NC2beta during basal and activated transcription using the anchor-away technique for conditional nuclear depletion. Genome-wide expression analysis indicates that both proteins regulate a highly similar set of genes (r2=0.8). Upregulated genes were enriched for SAGA occupancy, while downregulated genes preferred TFIID binding. Mot1p and NC2beta depletion during heat shock resulted in a failure to downregulate gene expression after initial activation, which was accompanied by increased TBP and RNA pol II promoter occupancies. Depletion of Mot1p or NC2beta displayed preferential synthetic lethality with the TBP-interaction module of SAGA. Our results suggest that Mot1 and NC2beta cooperate in vivo to regulate TBP function, and that they are involved in maintaining basal expression levels as well as in resetting gene expression after induction by stress.

overall_design: In this study 3 S. cerevisiae strains were grown with and without addition of rapamycin. For each experimental factor two independent colonies were inoculated in Complete Synthetic Medium (CSM) with 2% glucose. Overnight cultures were diluted to 0.3 OD600 in 50 ml medium and grown to 1 OD600. Cultures were then grown for 60 minutes in the absence or presence of rapamycin at 1ug/ml (LC laboratories). Cells were harvested by centrifugation at 4000 rpm for 3 minutes and were frozen in liquid nitrogen. Amplified cRNA samples were labeled with either cy3 or cy5 and put on microarray together with an oppositely labeled common reference sample consisting of cRNA of untreated wildtype strain.

first: Gianpiero

last: Spedale

first: Claartje

last: Meddens

first: Maria

last: Koster

first: Cheuk

middle: W

last: Ko

first: Sander

last: van Hooff

first: Frank

last: Holstege

first: H

last: Timmers

first: W

last: Pijnappel


title: The evolution of low mutation rates in experimental mutator populations of Saccharomyces cerevisiae

status: Public on Feb 02 2012

city: Taipei

first: Michael

middle: Joseph

last: McDonald

address: 128 sec 2. Academia Rd

department: Molecular Biology

country: Taiwan

institute: Academia Sinica

summary:
microarray experiment to test the gene expression in long term lines of mutator and non-mutator yeast.
Here we use an experimental evolution approach to investigate the conditions required for evolution of a reduction in mutation rate and the mechanisms by which populations tolerate the accumulation of deleterious mutations. We find that after ~6700 generations four out of eight experimental mutator lines had evolved a decreased mutation rate.

overall_design:
2 condition experiment, derived experimental evolution strains compared to their ancestor strain.
We compared the expression profile of one of the mutator lines (m8) after 6700 generations with its mutator ancestor, and as a control, an evolved non mutator after 6700 generations was compared to to its non-mutator ancestor.
In order to prepare cells for expression microarray, glass tubes containing 3 ml of YPD were inoculated from overnight cultures, and grown until the OD600 was approximately 0.3.

first: Mchael

middle: J

first: Yen

middle: H

last: Yu


title: The impact of Lactobacillus plantarum sortase on target-protein sorting, gastrointestinal persistence and host immunomodulation

status: Public on Jan 12 2012

city: Ede

first: Michiel

last: Wels

address: Kernhemseweg 2

institute: NIZO food research

summary: Sortases are transpeptidase enzymes that couple surface proteins to the peptidoglycan of Gram-positive bacteria. Several sortase-dependent proteins (SDPs) have been identified that are crucial for bacterial pathogenesis and, although less frequently, for the physiology of non-pathogenic bacteria. We found that an isogenic sortase A (srtA) deletion derivative (NZ7104) of Lactobacillus plantarum WCFS1 did not express any residual SrtA activity, i.e. failed to cleave the LPQTDE SrtA recognition motif. Trypsination of intact bacterial cells, followed by mass spectrometry based peptide identification, revealed a significant decrease, but not complete loss of SDPs on the NZ7104 cell surface as compared to the wildtype. Using LiCl, we further found that several SDPs could be extracted from the cell surface of NZ7104 but not from the wildtype, demonstrating that SrtA is involved in the covalent coupling of these SDPs. Neither the gastrointestinal persistence of L. plantarum in mice, nor the cytokine secretion patterns induced in monocyte derived immature dendritic cells (iDCs) was significantly affected by the srtA deletion. However, contrary to the wild-type cells, LiCl washed NZ7104 cells induced drastically increased proinflammatory cytokine production in iDCs, indicating a role of the SDPs in attenuation of immune system stimulation.

overall_design: array were constructed in two subdesigns: one comparing all samples from logarithmic phase (WT vs mutant) and one comparing the stationary phase samples (WT vs mutant)

first: Daniela

last: Remus

first: Roger

last: Bongers

last: Kleerebezem

first: Peter

last: Bron


title: Gene expression data from isolated white winter wheat embryos at physiological maturity, environment 2 of 2.

status: Public on Feb 01 2012

city: Ithaca

first: Jesse

middle: D.

last: Munkvold

state: NY

address: 240 Emerson

department: Plant Breeding and Genetics

institute: Cornell University

summary:
Gene expression was measured in isolated wheat embryos at physiological maturity. The gene expression measurements were done across a doubled haploid white winter wheat population between the variety Cayuga and the variety Caledonia (CxC). Data can be used for mapping gene expression QTL (eQTL).
These data are from the second (Ketola 2005) of two field environments. Different target labeling was used so the environments are contained in two separate series.

overall_design: The design was a two-channel dye-swap reference design with bulked total RNA from pre-anthesis wheat spikes serving as the reference. Normalization was done using the TIGR MIDAS suite using blockwise LOWESS smoothing and variance normalization. Between slide variance normalization was also done for all slides so direct comparisons could be made. Dye-swap was submitted unmerged to facilitate flexibility in downstream processing.

first: Mark

middle: E

last: Sorrells


title: Profiling DNA copy number alterations of myxofibrosarcomas with NimbleGen array CGH chips

city: Tainan

first: Chien-Feng

last: Li

address: 901 Chunghwa Road

department: Pathology/Medical Research

institute: Chi-Mei Medical Center

type: Genome variation profiling by genome tiling array

summary: Characterized by a wide histological spectrum, myxofibrosarcoma ranges from deceptively bland-appearing lesions to frankly pleomorphic sarcomas, representing a suitable model to elucidate the molecular aberrations in multistep disease progression. To explore cancer-associated genes of myxofibrosarcoma, ultrahigh-resolution array comparative genomic hybridization (aCGH) was used to profile DNA copy number alterations in myxofibrosarcoma tumor samples and cell lines.

overall_design: Along with normal reference DNA, 12 tumor samples and 3 cell lines (NMFH-1, OH931, and NMFH2) of human myxofibrosarcoma were extracted for genomic DNAs, which were subjected to manufacturer's (NimbleGen, Inc.) outsourcing service of DNA labeling, hybridization, and normalization of data obtained from 385K or 720K oligonucleotide array CGH chips.

first: Chien

middle: F

first: Hsuan

middle: Y

last: Huang


title: Functional profiling in yeast with zinc-limitation

status: Public on Apr 17 2012

city: Berkeley

first: Chris

last: Vulpe

state: Ca

address: 119 Morgan Hall

department: Nutritional Sciences and Toxicology

institute: UC Berkeley

summary: Zinc is an essential nutrient because it is a required cofactor for many enzymes and transcription factors. To discover genes and processes in yeast that are required for growth when zinc is limiting, we used genome-wide functional profiling. As a result, we identified over 400 different genes required for optimal growth under zinc-limiting conditions. Among these were several targets of the Zap1 zinc-responsive transcription factor. Their importance is consistent with their up-regulation by Zap1 in low zinc. We also identified genes that implicate Zap1-independent processes as important. These include endoplasmic reticulum function, oxidative stress resistance, vesicular trafficking, peroxisome biogenesis, and chromatin modification. Our studies also indicated the critical role of macroautophagy in low zinc growth. Finally, as a result of our analysis, we discovered a previously unknown role for the ICE2 gene in maintaining ER zinc homeostasis.

overall_design:
Pools of homozygous diploid deletion mutants (n = 4,607 strains) were grown in zinc defined media (LZM) at 2 concentrations for 5 and 15 generations (5g, 15g).
Each strain has a deletion in a different gene. In each strain, the gene was replaced by a deletion cassette containing a antibiotic resistance gene and two BARCODES, up and down tags (Please see Giaever et al, 2002, Nature). These tags in the DNA are specific to each strain. We pool all deletion strains and grow them under a selective condition; extract DNA; amplify barcodes using universal primers and hybridize to arrays containing complemetary sequences to the up and down tags. In this way, we can look at growth of each of the strains. Our strategy is to analyze separately the up and down tags. Therefore, for each array (CEL file), we generate two data files, one for all ups and another one for all downs. In these files, the list of genes are the same but the data come from different set of probes, up or down. The values are log2 averages of replicate probes for the same tag. These pre processed files were used to identify strains with differential growth in 1uM zinc by comparing to the 100uM zinc arrays.

first: MATTHEW

last: NORTH

first: JANET

last: STEFFEN

first: ALEX

middle: V

last: LOGUINOV

first: GINELLE

last: ZIMMERMAN

first: CHRIS

last: VULPE

first: DAVID

last: EIDE


title: Expression profiles of the NuB4/HAT-B complex and histone knockouts

status: Public on May 15 2012

first: Patrick

last: Kemmeren

address: Universiteitsweg 100

department: Department of Molecular Cancer Research

institute: UMC Utrecht

summary: Dynamic modification of histone proteins plays a key role in regulating gene expression. However, histones themselves can also be dynamic, which potentially affects the stability of histone modifications. To determine the molecular mechanisms of histone turnover we developed a parallel screening method for epigenetic regulators by analyzing chromatin states on DNA barcodes. Histone turnover was quantified by employing a genetic pulse-chase technique called RITE, which was combined with chromatin immunoprecipitation and high-throughput sequencing. In this screen, the NuB4/HAT-B complex, containing the conserved type B histone acetyltransferase Hat1, was found to promote histone turnover. Unexpectedly, the three members of this complex could be functionally separated from each other as well as from the known interacting factor and histone chaperone Asf1. Thus, systematic and direct interrogation of chromatin structure on DNA barcodes can lead to the discovery of genes and pathways involved in chromatin modification and dynamics.

overall_design: RNA isolated from a large amount of wt yeast from a single culture was used as a common reference. This common reference was used for each separate hybridization and used in the statistical analysis to obtain an average expression-profile for each deletion mutant relative to the wt. Two independent cultures were hybridized on two separate microarrays. For the first hybridization the Cy5 (red) labeled cRNA from the deletion mutant is hybridized together with the Cy3 (green) labeled cRNA from the common reference. For the replicate hybridization, the labels are swapped. Each gene is represented twice on the microarray, resulting in four measurements per mutant. Up to five deletion strains were grown on a single day. Wt cultures were grown parallel to the deletion mutants to assess day-to-day variance.

first: Kitty

last: Verzijlbergen

first: Tibor

last: van Welsem

first: Daoud

last: Sie

first: Tineke

last: Lenstra

first: Daniel

last: Turner

first: Frank

last: Holstege

first: Ron

last: Kerkhoven

first: Fred

last: van Leeuwen


title: Gene Expression Profiles of Planarians after 6,000 Rads Irradiation

status: Public on Jan 10 2012

middle: Elger

last: Wagner

institute: Whitehead Institute

summary: The goal of the study was to identify genes expressed in the proliferative cell population (neoblasts) of adult planarians.  Proliferating cells are rapidly and specifically eliminated following lethal irradiation.  Genes expressed by neoblasts, therefore, were expected to show decreased level of expression following 6,000 rads, relative to control animals.

overall_design: Total RNA was collected from control (unirradiated) animals and from animals 6, 12, 24, and 48 hours following 6,000 rads irradiation.  Between three and five replicates were used for each sample

first: Peter

last: Reddien


title: Gene expression data from isolated white winter wheat embryos at physiological maturity, environment 1 of 2.

first: Jesse

last: Munkvold

summary:
Gene expression was measured in isolated wheat embryos at physiological maturity. The gene expression measurements were done across a doubled haploid white winter wheat population between the variety Cayuga and the variety Caledonia (CxC). Data can be used for mapping gene expression QTL (eQTL).
These data are from the first (Helfer 2004) of two field environments. Different target labeling was used so the environments are contained in two separate series.

first: Mark

last: Sorrells


title: Transgenic mice carrying the human PLN gene

status: Public on Dec 15 2011

city: Athens

first: Despina

last: Sanoudou

address: Soranou Efesiou 4

country: Greece

institute: Foundation for Biomedical Research, Academy of Athens

summary: These mice have been genetically modified so that the mouse PLN gene has been replace with the human PLN gene. Phopspholamban (PLN), the reversible inhibitor of the sarcoplasmic reticulum (SR) Ca2+-ATPase (SERCA2a), is a key regulator of myocyte Ca2+-cycling with a significant role in heart failure. We previously showed that the single amino acid difference between human and mouse PLN results in increased inhibition of Ca2+-cycling, cardiac remodeling and attenuated stress responses, in transgenic mice expressing the human PLN in the null background (hPLN). This set of microarray experiments aimed at deciphering the molecular mechanisms implicated in this process.

overall_design: 6 wild type, 6 transgenic mice. All at 11 weeks of age. FVB/N background. Whole hearts isolated, snap frozen and used for total RNA extraction.


title: Profiles of gene expression and alternative splicing in K562 cells exposed to  2 mM sodium valproate for 12 hours assessed using exon array

status: Public on Nov 11 2011

city: Beijing

first: wang

last: liang

address: 7A North Yongchang Road BDA

country: China

institute: BioChainBJ

summary: To investigate the effect of valproate (VPA) treatment on K562 cell line, the growth and survival of the K562 cell line were investigated using the Annexin V/PI dual staining method, and global profiles of gene expression and alternative splicing in K562 cells were assessed using exon microarray. A significant increase in cell apoptosis was observed in valproate exposed K562 cells using flow cytometry. A total of 628 transcripts were identified as being significantly differentially expressed. The number of genes demonstrating increased expression levels was greater than the number of genes demonstrating decreased expression levels (445 genes vs. 183 genes, respectively). The significant enrichment analysis of GO terms for the differentially expressed genes revealed that these genes are involved in many important biological processes such as apoptosis. Six of observed differentially expressed genes that might be involved in apoptosis were selected to undergo qRT-PCR validation. In total, 198 candidates of alternative splicing variants were identified. Among them, three alternative splicing events were selected for validation, and CBLC and TBX1 were confirmed as alternative splicing by semi-nested PCR. In conclusion, valproate exposure facilitated cell apoptosis, altered mRNA expression and alternative splicing events in the K562 cell line.

overall_design: Gene expression/alternative splicing in K562 cell line were measured after exposure to  2 mM valproate for 12 hours or left untreated as a control using exon array.

first: Xiang-Zhong

last: Zhang

first: Ai-Hua

last: Yin

first: Xiao-Yu

last: Zhu

first: Qian

last: Ding

first: Chun-Huai

last: Wang

first: Yun-Xian

last: Chen


title: Beyond KRAS mutation status: influence of KRAS copy number status and microRNAs on response to cetuximab in metastatic colorectal cancer patients

status: Public on Jan 01 2012

city: Nijmegen

first: Roland

middle: P.

last: Kuiper

address: Geert Grooteplein 10

department: Human Genetics

institute: Radboud University Nijmegen Medical Centre

summary:
KRAS mutation is a negative predictive factor for treatment with anti-epidermal growth factor receptor (EGFR) antibodies in metastatic colorectal cancer (mCRC). Novel predictive markers are required to further improve the selection of patients for this treatment. Here, we assessed the influence of modification of KRAS by gene copy number aberration (CNA) and microRNAs (miRNAs) in correlation to clinical outcome in mCRC patients treated with cetuximab in combination with chemotherapy and bevacizumab. Formalin-fixed paraffin-embedded primary tumour tissue was used from 34 mCRC patients in a phase III trial, who were selected based upon their good (n=17) or poor (n=17) progression-free survival (PFS) upon treatment with cetuximab in combination with capecitabine, oxaliplatin, and bevacizumab. Gene copy number at the KRAS locus was assessed using high resolution genome-wide array CGH and the expression levels of 17 miRNAs targeting KRAS were determined by real-time PCR. Good response was associated with 12p12.1 copy number loss, even in patients with a KRAS mutation, while copy number gain in wild-type KRAS patients was correlated with a poor response. In KRAS mutated tumours increased miR-200b and decreased miR-143 expression were associated with a good response. In wild-type KRAS patients, miRNA expression did not predict response in a multivariate model. Thus, assessment of KRAS CNA and miRNAs targeting KRAS might further optimize the selection of patients eligible for anti-EGFR therapy.
Copy number detection was performed using NimbleScan and Nexus software

overall_design: Formalin-fixed paraffin-embedded primary tumour tissue was used from 34 metastisized colorectal cancer patients in a phase III trial (CKTO 2005-02; ClinTrials.gov NCT00208546) of the Dutch Colorectal Cancer Group (DCCG), who were selected based upon their good (n=17) or poor (n=17) progression-free survival (PFS) upon treatment with cetuximab in combination with capecitabine, oxaliplatin, and bevacizumab.

first: Leonie

last: Mekenkamp

first: Jolien

last: Tol

first: Jeroen

last: Dijkstra

first: Inge

last: de Krijger

first: M

last: Vink-Börger

first: Steven

last: Teerenstra

first: E

last: Kamping

first: Eugène

middle: T

last: Verwiel

first: Miriam

last: Koopman

first: Gerrit

middle: A

last: Meijer

first: J

last: van Krieken

first: Cornelis

last: Punt

first: Iris

last: Nagtegaal


title: Transcription profiling of Rab25 transfected ovarian A2780 cancer cell

status: Public on Dec 30 2011

city: Houston

first: Kwai Wa

last: Cheng

address: 1515 Holcombe Blvd.

department: Systems Biology

institute: UT-MD Anderson Cancer Center

summary: To elucidate potential mechanisms by which Rab25 contributes to tumor aggressiveness, we determined the transcription profile induced by expression of Rab25.

overall_design: Twenty-four independent samples, 12 from pcDNA transfected and 12 from Rab25 expressing A2780 ovarian cancer cells were profiled on Affymetrix GeneChip HT-HG_U133A Early Access Array.

first: Kwai

first: John

last: Lahad

first: Joe

last: Gray

first: Gordon

middle: B

last: Millls


title: Comparative expression profile of Porphyromonas gingivalis parent strain W83 and the HU protein (PG0121) mutant during exponential growth in rich medium.

status: Public on Nov 30 2011

first: Mary

middle: Ellen

last: Davey

address: 245 First Street

department: Molecular Genetics

institute: The Forsyth Institute

summary: HU protein regulates expression of genes involved in cell-division, iron acquisition, and transcriptional regulation in Porphyromonas gingivalis

overall_design: Analysis involved RNA extracted from the parent strain W83 and the PG0121 (HU protein) mutant during exponential growth in rich medium. Each of the four experiments used a different preparation of RNA. Comparative expression profiles were determined by hybridizing the microarray slide with a mixture containing equal amount of cDNA generated from each strain. Dye swap experiments were performed.

first: Carla

last: Cugini

first: Richa

last: Priyadarshini

first: Annette

last: Arndt


title: Gene expression profiles of Six1/2-1(RNAi) animals and POU2/3(RNAi) animals

status: Public on Nov 28 2011

first: Mansi

last: Srivastava

institute: Whitehead Institutue

summary: The goal of this study was to identify genes that were expressed in the protonephridial system. Because Six1/2(RNAi) and POU2/3(RNAi) animals lose the tubules, flame cells and accessory cells, these animals were expected to show decreased levels of the genes expressed in the nephridial system relative to control RNAi animals.

overall_design: RNA from Six1/2(RNAi) and POU2/3(RNAi) animals was harvested with Trizol at days 8 and 15 after RNAi initiation for the former and days 15 and 21 for the latter. Three biological replicates were used. RNA from the four samples were competetively hybridized on a microarray with RNA from control RNAi animals of the corresponding days.

first: M

middle: L

last: Scimone

first: George

last: Bell

first: Peter

last: Reddien


title: Overexpression of Ssz1 S295F and Zuo1 amino acids 365-433 S427G in S. cerevisiae

status: Public on Aug 30 2011

city: Madison

first: Elizabeth

last: Craig

state: Wisconsin

address: 433 Babcock Dr.

institute: University of Wisconsin

summary:
WT strains were transformed with a control TAP plasmid, plasmid containing the Hsp70 chaperones Ssz1 containing a mutation in S295F, or J-domain containing protein Zuo1 amino acids 365-433 with the S427G mutation.
These mutations are known to enhance the induction of the pleiotropic drug resistance pathway.

overall_design: RNA from three biological replicates were pooled and analyzed by Nimblegen array.

first: Amy

last: Prunuske


title: Comparison of plant responses upon Aphanomyces euteiches infection in wild type (A17) and NFP allelic mutants

status: Public on Sep 02 2011

city: Beaucouze

first: sandrine

last: balzergue

address: Rue G. Morel

country: France

institute: INRA

summary:
affy_aphanomyces_medicago - NFP is one of the putative Nod factor receptor and plays therefore a key role in the establishment of symbiosis. nfp allelic mutants are more susceptible to Aphanomyces euteiches (Ae), a root pathogen, than the A17 wild type (WT) line.  In this study we want to compare the early plant responses, 1 day post inoculation (dpi) between WT and the nfp2 mutant, in order to identify NFP-dependent gene networks during infection.-Fifteen-day-old A17 or nfp-2 plants were grown in vitro on M medium. The entire roots were then inoculated (or not = controls) with a A. euteiches zoospores (105/ml) and harvested one day after inoculation. Three independent repeats were performed.

Keywords: normal vs ems mutant comparison,near isogenic lines (hif)

platform_organism:
Sinorhizobium meliloti
Medicago sativa
Medicago truncatula

overall_design: 12 arrays - Medicago

first: Sandrine

last: Balzergue

first: Christophe

last: Jacquet

first: Marie-Laure

last: Martin-Magniette


title: MiRNA expression profiles of trophoblastic cells

status: Public on Sep 23 2011

city: Jena

first: Udo

middle: R.

last: Markert

address: Bachstraße 18

department: Department of Obstetrics

institute: Universitätsklinikum Jena

summary:
To gain more insight in the biological expression and role of microRNAs in trophoblast cells, we then set up to determinate the miRNA expression pattern of four trophoblastic cell lines and compare them with the miRNA signature of isolated trophoblast
Total RNA was reverse-transcribed, pre-amplified with miRNA-specific primers, and loaded into TLDA cards. Quantitative real-time PCR was performed for 768 miRNAs. For each of two cards per sample (A and B), 9 ul of pre-amplified diluted sample was mixed with TaqMan reaction mix (Applied Biosystems) and loaded into miRNA TLDA cards (Applied Biosystems) by centrifugation. Cards were sealed, and qRT-PCR was performed with a real-time thermocycler (ABI-7900, Applied Biosystems) per manufacturer's recommendations.

overall_design: MiRNA signatures of isolated trophoblasts were compared with those in four trophoblastic cell lines

first: Diana

middle: M

last: Morales-Prieto

first: Wittaya

last: Chaiwangyen

first: Stephanie

last: Ospina-Prieto

first: Uwe

last: Schneider

first: Jörg

last: Herrmann

first: Bernd

last: Gruhn

middle: R


title: AURATUS goldfish environmental genomics projects: SKF 38393 in the hypothalamus

status: Public on Aug 22 2011

city: Ottawa

first: Jason

middle: T.

last: Popesku

state: ON

address: 20 Marie Curie

department: Biology

institute: University of Ottawa

summary:
Sexually mature female goldfish were administered 2 i.p. injections:
1)  0.1% DMSO in 0.6% saline or  SKF 38393 (40 µg/g) dissolved in 0.1% DMSO in 0.6% saline, and;
2) 0.6% saline.

platform_organism:
Carassius auratus
Cyprinus carpio

overall_design:
A common reference design was used:
4 independent groups of hypothalami from sexually mature female goldfish treated with 40 ug/g SKF 38393 hybridized against a common vehicle control.
Slides 9, 19, 33, 44.

first: Chris

middle: J

last: Martyniuk

first: Susanna

middle: C

last: Wiens

first: Vance

last: Trudeau


title: AURATUS goldfish environmental genomics projects: quinpirole in the hypothalamus

summary:
Sexually mature female goldfish were administered 2 i.p. injections:
1) 0.1% DMSO in 0.6% saline, and;
2) 0.6% saline OR quinpirole (2 ug/g) dissolved in 0.6% saline.

overall_design:
A common reference design was used:
4 independent groups of hypothalami from sexually mature female goldfish treated with 40 ug/g SKF 38393 hybridized against a common vehicle control.
Slides 5, 11, 14, 26.


title: Hybridization of human oocyte cDNA library on the multi-species oocyte array to assess for gene conservation using a temperature stringency criteria

status: Public on Dec 15 2010

city: Quebec

first: Eve-Lyne

last: Sylvestre

address: 2440 Boul Hochelaga

institute: Université Laval

summary:
Human oocyte cDNA library was hybridized on a multi-species oocyte array (Bovine, Mouse, Frog)
Temperature stringency criteria was used to evaluate the conservation degree of oocyte genes among vertebrates (Bovine, Mouse, Frog)

platform_organism:
Xenopus laevis
Bos taurus
Mus musculus

overall_design: 2 technical replicates on distinct array were made at each pre-determined hybridization temperature (45°C, 50°C, 55°C), overall, the experiment includes 6 arrays

first: Claude

last: Robert

first: Sophie

last: Pennetier

first: Rémi

last: Labrecque

first: Isabelle

last: Gilbert

last: Dufort

first: Marie-Claude

last: Léveillé

first: Marc-André

last: Sirard


title: Peritoneal and testicular macrophage mrna expression after exposure to UPEC CFT073

status: Public on Oct 20 2010

city: Giessen

first: Andreas Christian

last: Geisler

address: Frankfurter Str. 107

department: Med. Mikrobiologie

institute: Justus Liebig Universität Giessen

summary: Ascending bacterial infections of the male genitourinary tract by Escherichia coli are an important cause of male infertility. Thus understanding mechanisms by which immunocompetent cells such as testicular macrophages (TM) respond to infection and how bacterial pathogens manipulate defense pathways is of great relevance. Following infection with uropathogenic E. coli (UPEC) whole genome expression profiling revealed induction of the calcium-dependent nuclear factor of activated T cells (NFAT) signaling pathway in TM as well as in peritoneal macrophages (PM) that were used in comparison. UPEC-dependent NFAT activation was confirmed in cultured TM and in TM using an in vivo infection model. Elevated expression of NFATC2-regulated anti-inflammatory cytokines was found in TM (IL-4, IL-13) and PM (IL-3, IL-4, IL-13). NFATC2 is activated by a rapid influx of calcium caused by the UPEC pore forming toxin alpha-hemolysin. Mutant analysis identified alpha-hemolysin as the main virulent factor responsible for UPEC-dependant suppression of IL-6 and TNF-a cytokine release from PM. Alpha-hemolysin caused differential activation of MAP kinase and AP-1 signaling pathways in TM and PM leading to differential expression profiles of key pro-inflammatory cytokines in PM (IL-1a, IL-1b and IL-6 downregulated) and TM (IL-1b and IL-6 upregulated). In contrast to PM, treatment with lipopolysaccharide did not result in activation of the NFkB pathway in TM as shown by lack of degradation of IkBa and lack of pro-inflammatory cytokine secretion (IL-6, TNF-a). In conclusion, these results propose a mechanism how TM can defend against microbes, while at the same time they are able to maintain the immune privileged status of the testis.

overall_design: 2 Macrophage populations ( peritoneal, testicular)  infected with UPEC CFT073; timepoints of mrna harvest at 0 min (control), 30 min and 60 min; of each point 2 biological replicates; of each repliccate 2 technical replicates: summa summarum: 2*((1+1+1)*2*2)=24 samples

first: Sudhanshu

last: Bhushan

first: Hamid

last: Hossain

first: Yongning

last: Lu

first: Andreas

last: Klug

first: Svetlin

last: Tchatalbachev

first: Trinad

last: Chakraborty

last: Meinhardt

first: Zbigniew

last: Mikulski

first: Gerhard

last: Schuler


title: Computational analysis of genome-wide DNA-methylation during the differentiation of human embryonic stem cells along the endodermal lineage

status: Public on Jul 27 2010

city: Heidelberg

first: Lukas

last: Chavez

address: Im Neuenheimer Feld 280

institute: German Cancer Research Center (DKFZ)

summary: The generation of genome-wide data derived from methylated DNA immunoprecipitation followed by sequencing (MeDIP-Seq) has become a major tool for epigenetic studies in health and disease. The computational analysis of such data, however, still falls short on accuracy, sensitivity and speed. We propose a statistical method that is able to cope with the inherent complexity of MeDIP-Seq data and outperforms computation time of existing methods by orders of magnitude with similar performance. In order to demonstrate the computational approach, we have analysed alterations in DNA methylation during the differentiation of hESCs to definitive endoderm. We show improved correlation of normalized MeDIP-Seq data in comparison to available whole-genome bisulphite sequencing data and investigated the effect of differential methylation on gene expression. Furthermore, we analyzed the interplay between DNA-methylation, histone modifications, transcription factor binding, and show that in contrast to de-novo methylation, de-methylation is mainly associated with regions of low CpG densities.

overall_design: Total RNA obtained from three biological replicates of hESCs (passage 53, Control) and from three biological replicates of Activin-A induced differentiated hESCs (definitive endoderm (DE), Treatment)


title: Induced overexpression of Pdx1 and Ngn3 in a mouse ES cell-derived endoderm population induces pancreatic differentiation

status: Public on Aug 24 2010

city: South San Francisco

first: Robert

last: Stull

address: 384 Oyster Point Blvd, Suite #8

department: Genomics

institute: VistaGen Therapeutics

summary:
Induced overexpression of Pdx1 in activin-induced endoderm population resulted in the upregulation of pancreas-related genes such as insulin 1 and 2 at day 20.  To enhance the developmental progression from the pancreatic bud to the formation of the endocrine lineages, we next expressed neurogenin3 (Ngn3) together with Pdx-1.  Induced overexpression of Pdx1 together with Ngn3 dramatically increased Insulin 1 mRNA by day 9 differentiation.  The levels of insulin 1 mRNA present in the induced EBs represented approximately 100 % of that found in insulinoma cell line, betaTC6.
We also confirmed insulin and C-peptide staining by immunohistochemistry.  These cells process and secrete insulin and respond to various insulin secretagogues.  These inductive effects were restricted to c-kit+ endoderm enriched EB-derived populations suggesting that Pdx1/Ngn3 functions at the level of pancreatic specification of endoderm in this model.  Microarray analysis showed that Pdx1/Ngn3 regulated the expression of a broad spectrum of pancreatic endocrine cell-related genes.

overall_design: On day 4 of a multiday differentiation protocol, EBs were dissociated and cultured in serum-free growth medium supplemented with differentiation facots, with or without Dox (1 ug/ml).  On day 6, EBs were replated on gelatin in 12-well low-cluster dishes (Nunc) to obtain non-adherent floating EBs with or without Dox (1 ug/ml) and cultured out to day 13, at which time RNA was harvested for microarray analysis.

first: Atsushi

last: Kubo

first: Mitsuaki

last: Takeuchi

first: Kristina

last: Bonham

first: Valerie

last: Gouon-Evans

first: Masayuki

last: Iwano

last: Sho

first: Yoshihiko

last: Saito

first: Gordon

last: Keller

first: Ralph

last: Snodgrass


title: Genome-wide gene expression analysis in mouse embryonic stem cells

status: Public on Nov 17 2010

city: Granada

first: Angel

last: Concha

address: 2 Fuerzas Armadas

country: Spain

institute: Servicio Andaluz de Salud

summary: Embryonic stem cells studies have generated a great interest due to their ability to form a wide variety of matured cells. However, there remains a poor understanding of mechanisms regulating the cell state of ESCs and of the genes they express during early differentiation. Gene expression analysis may be a valuable tool to elucidate either the molecular pathways involved in self-renewal and pluripotency or early differentiation and to identify potential molecular therapy targets. The aim of this study was to characterize at molecular level the undifferentiated mouse ESCs state and the early development towards embryoid bodies. To attemp this issue we performed CodeLink Mouse Uniset I 20K bioarrays in a well-characterized mouse ESC line, MES3, 3- and 7 day-old embryoid bodies and we compared our findings with those in adult tissue cells. Gene expression results were subsequently validated in a commercial stem cell line, CGR8 (ATCC). SAM analysis was used to identify statistically significant changes in microarray data. We identified 3664 genes expressed at significantly greater levels in MES3 stem cells than in adult tissue cells, which included 611 with 3-fold higher gene expression level versus the adult cells. We also investigated the gene expression profile during early embryoid body formation, identifying 2040 and 2243 genes that were up-regulated in 3- and 7-day-old EBs, respectively. Our gene expression results in MES3 cells were partially confirmed in CGR8 cells, showing numerous genes that are expressed in both mouse stem cells. In conclusion, our results suggest that commonly expressed genes may be strong candidates for involvement in maintenance of a pluripotent and undifferentiated phenotype and in early development.

overall_design: This study characterizes at molecular level the undifferentiated mouse ESCs state and the early development towards embryoid bodies. We identified numerous genes not been previously reported, which may offer novel insights into the behaviour of mouse ESC, the maintenance of their undifferentiated state and the mechanisms underlying their pluripotency, self-renewal and early differentiation.

first: Juan

last: Sainz

first: Fernando

last: Garcia-Alcalde

first: Armando

last: Blanco


title: Analysis of gene expression profiles in primary embryonic fibroblasts deficient for the Ing1 locus

status: Public on Oct 01 2010

city: MAJADAHONDA

first: Javier

last: Alonso

state: MADRID

address: CTRA. MAJADAHONDA-POZUELO KM 2

department: AREA DE GENÉTICA HUMANA

institute: INSTITUTO DE SALUD CARLOS III - IIER

summary: The ING familiy of tumor suppressor proteins control several cellular functions relevant to anti-tumor protection, like cell-cycle control, apoptosis, senescence or migration. ING proteins are functionally linked to the p53 pathway, and they participate in transcriptional control via the recognition of histone marks and recruitment of protein complexes with chromatin-modifying activity to specific promoters. Here, we have investigated the global impact of ING1 in gene regulation through genome-wide analysis of expression profiles in primary embryonic fibroblasts deficient for the Ing1 locus. We find that Ing1 has a predominant role as transcriptional repressor in this setting, affecting the expression of genes involved in a variety of cellular functions. Within the subset of genes showing differential expression, we have identified DGCR8, a protein involved in the early steps of microRNA biogenesis.

overall_design: In this study we have taken an unbiased approach to identify Ing1 targets, consisting in the comparison of expression profiles by genome-wide microarray analysis (Codelink mouse whole-genome expression microarrays), of mouse embryonic fibroblasts from embryos with the Ing1 locus inactivated by the insertion of a gene trap cassette, and their wild-type littermates. Three biological replicates were done for each phenotype.

first: Daniel

last: Gómez-Cabello

first: Ignacio

last: Palmero


title: The landscape of promoter DNA hypomethylation in liver cancer (MeDIP-chip data)

status: Public on Jun 04 2011

city: Montreal

first: Moshe

last: Szyf

address: McIntyre Medical Building, 3655 Promenade Sir William Osler, Room 1309

department: Pharmacology

institute: McGill University

type: Methylation profiling by genome tiling array

summary: Extensive loss of DNA methylation is a hallmark of cancer. The role of hypomethylation in altering gene expression in cancer cells has been poorly understood. Hepatic cellular carcinoma (HCC) is one of the most common human cancers. We use HCC as a model to investigate hypomethylation in cancer by a combination of methylated DNA immunoprecipitation and hybridization with comprehensive promoter arrays. We identify approximately 2,800 promoters that are hypomethylated in tumor samples. The hypomethylated promoters appear in clusters across the genome suggesting a high-level organization behind the epigenomic changes in cancer. The genes whose promoters are demethylated are mainly involved in cell growth, cell adhesion and communication, signal transduction, mobility and invasion; functions that are essential for cancer progression and metastasis. Previous studies suggested that MBD2 was involved in demethylation of uPA and MMP2 genes in human breast and prostate cancer cell lines. We extend these results here showing that whereas MBD2 depletion in normal liver cells has little or no effect, its depletion in the human hepatocellular carcinoma cell line HepG2 and the adenocarcinoma cell line SkHep1 results in suppression of cell growth, anchorage-independent growth and invasiveness, as well as an increase in promoter methylation and silencing of several of the genes that are hypomethylated in tumors. Our studies establish for the first time the rules governing hypomethylation of promoters in liver cancer and define the potential functional role of hypomethylation in cancer.

overall_design: Cancerous and normal adjacent tissue samples were obtained from 11 patients with HCC from the Chinese National Human Genome Center at Shanghai, China (Dr. Ze-Guang Han). For three patients, the cancer samples were dissected using laser capture microdissection technique. All patients provided written informed consent, and the Ethics Committee from the Chinese National Human Genome Center at Shanghai approved all aspects of this study.  Human HCC HepG2 cells and adenocarcinoma SkHep1 cells were purchased from ATCC (HB8065 and HTB52, respectively, USA), whereas human untransformed hepatocytes (normal hepatocytes, NorHep) were obtained from Celprogen (33003-02, USA).  Purified DNA from cancerous samples and normal adjacent samples, as well as HepG2 and NorHep cells, was enriched for methylated DNA using the MeDIP protocol developed by Cedar's group (PMID 16444255). The labeled input and bound DNA samples were hybridized to a custom-designed 244K promoter tiling array that contained probes covering all transcription start sites at intervals from 800 bp upstream to 200 bp downstream of all genes described in Ensembl (version 44) and within 250 bp of the ~400 microRNAs from miRBase, all at 100 bp-spacing.

first: Barbara

last: Stefanska

first: Bishnu

last: Bhattacharyya

first: Jian

last: Huang

first: Ze-Guang

last: Han

first: Michael

last: Hallett

first: Matthew

last: Suderman


title: Comparison of gene expression induced in hearts excised from mice 24 or 48 hours after DMA or DMSO (vehicle) treatment.

status: Public on Mar 01 2010

city: Campinas

first: Ana

middle: Carolina

last: Deckmann

state: Sao Paulo

address: Cidade Universitaria Zeferino Vaz

department: Medical Sciences

country: Brazil

institute: State University of Campinas

summary:
In isolated heart preparations, we observed that previous treatment with quinazoline DMA evoked both acute and late phases of cardioprotection. In this study, we adopted whole genome microarrays approach to investigate cardiac transcriptional responses induced in hearts from DMA treated mice in comparison to vehicle-only treatment.

Deckmann AC, Rocco SA, Marin RM, Rolim L, Carazzolle MF, Barau JG, Pereira GAG and Franchini KG. DELAYED CARDIOPROTECTION INDUCED BY ADENOSINE KINASE INHIBITION STIMULATES ADIPOCYTOKINES AND ACTIVATORS OF PRO-SURVIVAL KINASES OF RISK PATHWAY. Unpublished yet.

Keywords: Mus musculus, ischemic heart diseases, quinazoline compound, late cardioprotection

overall_design: Balb/c mice were randomly assigned to two groups, in which they were gavaged with 30 mg DMA/kg body wt (treatment group, T) or DMSO (control group, C). After 24 hours, half of the animals of each group had their hearts excised and the total RNA was isolated as a pool. Same procedures were performed with the remaining animals 48 hours after treatment.

first: Silvana

last: Rocco

first: Rodrigo

last: Marin

first: Luis

last: Rolim

first: Marcelo

last: Carazzolle

first: Joan

middle: G

last: Barau

first: Gon�alo

last: Pereira

first: Kleber

last: Franchini


title: Comparison of infrarenal and suprarenal sections of mouse ApoE-/- abdominal aorta

status: Public on May 12 2010

city: Townsville

first: Jonathan

last: Golledge

state: Queensland

address: School of Medicine

department: School of Medicine

country: Australia

institute: James Cook University

summary:
The ApoE -/- mice model of abdominal aortic aneurysm (AAA) involves introducing Angiotensin II subcutaneously to 14 week old male mice for 4 weeks by osmotic pump. A significant number of mice will develop aneurysm-like dilations in the suprarenal section of the abdominal aorta (SRA) that have a number of similarities to the human condition and make this a useful model of AAA. The mouse infrarenal aorta is very resistant to aneurysm formation while in humans AAA predominately occurs in the infrarenal section of the aorta (IRA).
There have been a number of theories proposed to explain the site selctivity of aneurysm formation in AAA and this mice model. This study was designed to ascertain differences between SRA and IRA that may explain this site selectivity.
Keywords: tissue type comparison

overall_design: The aortas of five groups of 2 male (14 weeks old) ApoE-/- mice  were harvested and divided into infrarenal and suprarenal sections. RNA and proteins were extracted from each group of pooled segments to allow comparison between individual gene expression levels in IRA and SRA. The gene expression ratios of IRA to SRA were determined for each group and compared between groups.

first: Moses

last: Nyara

first: Catherine

last: Rush

first: Mirko

last: Karan


title: Cu2+ response in Arabidopsis

status: Public on Feb 28 2010

city: Valencia

first: Miguel

last: Perez-Amador

state: Valencia

address: Avda. de los Naranjos s/n

institute: Inst. Biologia Molecular y Celular de Plantas (IBMCP-CSIC)

summary:
Transcription profiling of Arabidopsis plants grown in high versus low Cu2+ concentration.

Keywords: Stress response

overall_design: Three replicates for each sample category (Arabidopsis Col0 grown either in high or low Cu2+) were generated, and compared directly to each other, in a non-paired direct comparisson. Two technical replicas from each biological replica were carried out.

first: Nuria

last: Andres-Colas

first: Eavan

last: Dorcey

first: Lola

last: Peñarrubia


title: Altered ovarian gene expression in the lethal yellow mouse

status: Public on Feb 18 2010

city: Vermillion

first: Kathleen

last: Eyster

state: SD

address: 414 E. Clark St.

department: Basic Biomedical Sciences

institute: University of South Dakota

overall_design:
Experiment Design:
Goal of the experiment:  To examine differential gene expression the ovaries of obese Lethal Yellow mice (C57BL/6 Ay/a) mice at 90 and 180 days of age compared to their lean black littermates (C57BL6 a/a).

Brief description of the experiment:  Mice possessing the lethal yellow (LY) agouti mutation (C57BL/6J Ay/a) exhibit adult-onset obesity, altered metabolic regulation, and early reproductive senescence.  The present study was designed to test the hypothesis that aging obese mice possess differences in expression of ovarian genes involved in reproductive function relative to age-matched lean mice.  To synchronize ovarian state and exclude gonadotropin-mediated effects, 90- and 180-day old diestrous LY and lean black (C57BL/6J a/a) mice were suppressed with GnRH antagonist (Antide®) for five days, then injected with 5 IU equine chorionic gonadotropin (eCG).  Complementary RNA derived from RNA extracts of whole ovarian homogenates collected 36h post eCG were run individually on CodeLink Mouse Whole Genome Bioarrays (Applied Microarrays).  Unidentified genes, EST, genes whose expression was below sensitivity limits, and those with less than a 2-fold difference in expression between groups were excluded from analysis.  After exclusions, 52 genes had a significant (p<0.05) difference in expression between 180-day-old obese LY and lean black mice.  LY exhibited elevated ovarian expression of agouti (350x), leptin (6.5x), and numerous genes involved in cholesterol/lipid transport and metabolism, e.g., lanosterol synthase, CYP51, and steroidogenic acute regulatory protein (StAR).  Fewer genes showed lower expression in LY mice, e.g. angiotensinogen.  In contrast, none of these genes showed differential expression in 90-day-old LY and black mice, which are of similar body weight.  Interestingly, 180-day-old LY mice had a 2-fold greater expression of 11β-hydroxysteroid dehydrogenase (HSD) type 1 and a 2-fold lesser expression of 11β-HSD type 2, differences not seen in 90-day-old mice.  Consistent with altered 11β-HSD expression, ovarian concentrations of corticosterone (C) were elevated in aging (180-day) LY mice relative to black mice, but C levels were similar in young (90-day) LY and black mice.  The data suggest that reproductive dysfunction in aging obese mice is related to modified intraovarian gene expression that is directly related to acquired obesity, but independent of gonadotropic state.

Keywords:  obesity, ovary, aging

Experimental factors:  age, presence vs absence of LY mutation

Experimental design:    The ovarian cycles of 90- and 180-day old mice with the lethal yellow mutation in the agouti gene (C57BL/6J Ay/a) and their non-mutated black littermates (C57BL6 a/a) (n=3 per group) were synchronized with the GnRH antagonist, Antide.  At 36 hours after the Antide injections, the mice were stimulated to ovulate with an injection of eCG.  At 36 hours after eCG injection the mice were euthanized and the ovaries collected.  One ovary from each animal was preserved in RNAlater for DNA microarray analysis.

Quality control steps:  The cRNA that was synthesized from each ovary was used for hybridization to a single CodeLink (Applied Microarrays) Whole Mouse Genome microarray.  Only one sample was hybridized with each slide and only one dye (Alexa 647) was used so no dye swaps were necessary.  Bacterial control spikes were used as per manufacturer's instructions.

Samples used, extract preparation and labelling:
The origin of each biological sample:    Ovaries of mice with the lethal yellow mutation in the agouti gene (C57BL/6J Ay/a) and their non-mutated black littermates (C57BL6 a/a) (n=3 per group).

Manipulations of biological samples and protocols used:  The ovarian cycles of 90- and 180-day old mice with the lethal yellow mutation in the agouti gene (C57BL/6J Ay/a) and their non-mutated black littermates (C57BL6 a/a) (n=3 per group) were synchronized with the GnRH antagonist, Antide.  At 36 hours after the Antide injections, the mice were stimulated to ovulate with an injection of eCG.  At 36 hours after eCG injection the mice were euthanized and the ovaries collected.  One ovary from each animal was preserved in RNAlater for DNA microarray analysis.

Experimental factor:  age, presence vs absence of LY mutation

Technical protocols:  The ovaries were collected into RNAlater (Ambion) and stored at -80C until processed.  The RNAlater was blotted from the ovaries and each was individually homogenized in TRI reagent (MRC).  Bromochloropropane and sodium acetate were added, and the samples were centrifuged to separate the phases.  The RNA-containing layer was removed and the RNA purified on an RNeasy extraction column (Qiagen).  The sample was treated with an on-column DNase treatment (RNase-free DNase, Qiagen).  The purity and quantity were evaluated by an Agilent Bioanalyzer using the RNA 6000 Nanoassay LabChip.

Labelled cRNA was prepared using the manufacturer's Instruction Protocol (www1.amershambiosciences.com, CodeLink Gene Expression System: Manual Labelled cRNA Target Preparation).  The source of reagents was the CodeLink Expression Assay Reagent Kit, Manual Prep, except where specified otherwise.  2.0 micrograms of total endometrial cell RNA was mixed with bacterial control RNA spikes and primed with T7 oligo(dT) primer for 10 min at 70C.  (The bacterial control spikes included araB, entF, fixB, gnd, hisB, and leuB.)  The first strand of cDNA was synthesized with first strand buffer, dNTP mix, RNase inhibitor, and reverse transcriptase for 2 h at 42C.  The second strand cDNA synthesis reaction was prepared using second strand buffer, dNTP mix, DNA polymerase mix, and RNase H; the reaction was carried out for 2h at 16C.  The double-stranded cDNA was purified on QIAquick columns (Qiagen) and the eluent was dried down in a SpeedVac concentrator.  The double-stranded cDNA was resuspended in a mixture containing T7 reaction buffer, T7 ATP, T7 GTP, T7 UTP, T7 CTP, biotin-11-UTP, and T7 enzyme mix for the synthesis of cRNA.  The cRNA synthesis reaction was terminated after 14h at 37C by purifying the cRNA on RNeasy columns (Qiagen).  The concentration of cRNA was determined by spectrophotometry.

Hybridization procedures and parameters:  10 micrograms of cRNA was mixed with fragmentation buffer and heated to 94C for 20 min.  The fragmented cRNA was mixed with CodeLink hybridization buffer, loaded on the microarray slides, and hybridized for 18 hours at 37C.
The slides were washed in 0.75x TNT (Tris-HCl, NaCl, Tween-20) at 46C for 1h then incubated with streptavidin-Alexa 647 fluorescent dye for 30 min at room temperature.  The Alexa fluor was prepared in TNB blocking buffer (0.1M Tris-HCl, 0.15M NaCL, 0.5% NEN Blocking Reagent-PerkinElmer)  The slides were then washed 4 times for 5 min each in 1x TNT and twice in 0.05% Tween 20 for 5 sec each.  The slides were dried by centrifugation and scanned in an Axon GenePix 4000B scanner.

first: John

last: Brannian

first: Maureen

last: Diggins


title: Sip1 regulates sequential fate decisions through feedback signalling from postmitotic neurons to progenitor cells.

status: Public on Oct 30 2009

city: Leuven

first: Rekin's

last: Janky

address: Herestraat 49 Box 816

department: Nucleomics Core

country: Belgium

institute: VIB

summary: In the developing cerebral cortex different types of neurons and glial cells are born through a precisely controlled sequence of events. The fate of cortical progenitors, in turn, is determined by an elusive conundrum of temporally and spatially regulated signalling mechanisms. We found the DNA-binding transcription factor Sip1 (also known as Zfhx1b) to be produced at high levels in postmitotic neurons of the cerebral cortex. Conditional deletion of Sip1 in young neocortical neurons was found to induce premature and increased production of upper layer neurons at the expense of deep layer neurons. Furthermore, it caused precocious and increased generation of glial precursors during late corticogenesis, leading subsequently to enhanced astrocytogenesis at early postnatal stages.  Expression profiling analysis indicated that the temporal shift in upper layer production coincides with overexpression of the neurotrophin-3 (NT3) gene and altered growth factor signalling in progenitors, while the premature gliogenesis is preceded by upregulation of fibroblast growth factor-9 (Fgf9) gene expression. Chromatin immunoprecipitation and in situ hybridization validates NT3 as a direct target of Sip1 in the cortex and confines the transcriptional repression by Sip1 to postmitotic neurons. Moreover, we show that exogenous application of Fgf9 in solution or via coated beads to wild-type cortical slices induces premature and excessive generation of glial precursors in the germinal zone. In conclusion, our data suggest that throughout corticogenesis Sip1 acts to restrain the level of production of secreted signalling factors in postmitotic neurons. These factors feed back to progenitor cells in order to regulate the timing of cell fate switch and the numbers of neurons and glial cells produced in the developing cerebral cortex.

overall_design: In a first series of arrays, we examined differential gene expression in neocortex and hippocampal tissue from 2 control (WT|Nestin) and 2 mutant (Sip1|Nestin) E18.5 littermate mice, each taken from 2 different litters (x|Nestin1 and x|Nestin2). In a separate experiment, we tested differential gene expression in E14.5 neocortex and hippocampal tissue from 2 control (WT|NEX) and 2 mutant (Sip1|NEX) littermate mice. RNA from mouse neocortex and hippocampus tissues was isolated using RNeasy kit according to the manufacturer’s protocol (QIAGEN).  Total RNA was controlled for integrity and purity using an Agilent Bioanalyzer and a NanoDrop spectrophotometer, respectively. All samples were of similar RNA quality. Starting with 1ug of total RNA, the RNA amplification was performed by in vitro transcription (IVT) with a biotin labeling reaction during the IVT, according to the recommendations of the manufacturer (Amersham Biosciences). The probes were purified and analyzed again for yield (> 20 ug) and purity (260:280 nm and 260:230 nm >1.8). Ten micrograms of the resulting antisense RNA was fragmented according to the recommendations of the manufacturer (Amersham Biosciences) and resuspended in 260 ml of hybridization buffer. Codelink Mouse Whole Genome array is a single array representing 35,000 transcripts. The gene array chips were hybridized in a shaker-incubator at 37°C at 300 rpm for 18 hours and washed and stained with Cy5-Streptavidin according to the recommendations of the manufacturer (Amersham Biosciences). The DNA Microarray scanner of Agilent was used for scanning and image analysis was performed with the Codelink Expression Analysis 4.1 software

first: Eve

last: Seuntjens

first: A

last: Nityanandam

last: Miquelajauregui

first: J

last: Debruyn

last: Stryjewska

first: S

last: Goebbels

first: K

last: Nave

first: D

last: Huylebroeck

first: V

last: Tarabykin


title: Impact of Developmental Lead Exposure on Splenic Factors

status: Public on Dec 21 2009

city: Rensselaer

first: Sridar

last: Chittur

address: One Discovery Drive, CRC 342G

department: Center for Functional Genomics

institute: SUNY-University at Albany

summary:
This study focuses on the impact of developmental Pb exposure on splenic gene expression. Pb exposure from gestational day 8 (gd8) to post-natal day 21 (pnd21) was used to obtain insight into how Pb induces anemia and negatively influences the immune response
Keywords: exposure analysis

overall_design: This study uses Codelink Mouse whole genome microarrays. Mouse spleen from 3 control and 3 experimental (treated with 0.1mM Pb since gd8) BALB/c female mice on pnd21 were used to study gene expression.

first: Jane

last: Kasten-Jolly

first: David

last: Lawrence


title: Gene expression profiling of the ethanol tolerant strain HG228 in comparison to its parental strain Escherichia coli MG1655

status: Public on Jun 15 2010

city: San Francisco

first: Hani

last: Goodarzi

department: Biochemistry and Biophysics

institute: UCSF

summary: The goal of this experiment is to identify the pathways that are deregulated as part of an adaptive response to high levels of ethanol in the media.

sample_organism: Escherichia coli str. K-12 substr. MG1655

overall_design: Ethanol tolerant cells (HG228) grown in minimal media plus 1.5% (v/v) ethanol were hybridized against E. coli MG1655 grown under similar conditions. The experiment was done in duplicates using biological replicates.

first: Bryson

middle: D

last: Bennett

first: Sasan

last: Amini

first: Marshall

last: Reaves

first: Alison

middle: K

last: Hottes

first: Joshua

last: Rabinowitz

first: Saeed

last: Tavazoie


title: Whole mouse genome expression analysis of host responses in Citrobacter rodentium infection

status: Public on Jul 01 2009

city: La Jolla

first: Martina

last: Spehlmann

address: 9500 Gilman Drive

institute: UCSD

summary: Profiling of a total of 34,790 genes revealed a wide range of expression changes during the course of C. rodentium infection in murine colon. The majority of changes were observed during weeks 1 and 2, while relatively fewer changes were seen at week 3. Interestingly, chemokines made up 20% of the top twenty upregulated genes.

overall_design: Mouse colon from pooled control or C. rodentium-infected mice.

first: M

middle: E

last: Dann

first: P

last: Hruz

first: E

last: Hanson

middle: F

last: McCole

first: L

last: Eckmann


title: Ozone Exposure of Populus

status: Public on Apr 01 2009

city: Umeå

first: Nathaniel

middle: Robert

last: Street

address: Umeå University

department: Plant Physiology

country: Sweden

institute: Umeå Plant Science Centre

summary: This SuperSeries is composed of the SubSeries listed below.

sample_organism:
Populus trichocarpa
Populus trichocarpa x Populus deltoides
Populus deltoides

platform_organism:
Populus trichocarpa x Populus deltoides
Populus euphratica
Populus tremula x Populus alba

overall_design: Refer to individual Series


title: Src kinase Induces Tumor Formation in the cSRC C57BL/6 Mouse

status: Public on May 08 2009

city: Hershey

first: Rosalyn

middle: B.

last: Irby

state: PA

address: 500 University Drive

department: Medicine

institute: Pennsylvania State University Hershey Medical Center

summary: We developed the c-SRC transgenic mouse in the C57BL/6 strain to address the issue of carcinogenesis in cells with high levels of Src expression. The transgene was constructed with the human c-SRC gene downstream of the mouse metallothionein promoter to create zinc inducible gene expression. In these C57BL/6 mice, Src protein was increased in a number of tissues both with and without zinc induction, but most highly in the liver.

overall_design:
Src expression was inducible with zinc, but typically zinc induction was unnecessary. No additional carcinogenic agent was administered. After 20 months, mice were assessed for tumor development in the liver and GI tract, as well as other organs. 15% of c-Src transgenic mice developed liver tumors while no wild type mice developed tumors. We performed microarray analysis on the livers of wild type and transgenic mice to determine the possible genes downstream of Src that might be involved in carcinogenesis.

Microarray runs of Wild Type liver sample #3 and c-SRC transgenic liver sample #2 both gave very low signals, and were not included in the final analysis.

first: Christina

last: Kline

first: Rosalind

last: Jackson

first: Robert

last: Engelman

first: Warren

last: Pledger

first: Timothy

last: Yeatman


title: Ozone exposure of Populus deltoides and Populus trichocarpa

summary:
Populus deltoides and Populus trichocarpa were exposed to either ambient air or an acute ozone exposure of 200 ppb for 9 hrs and ozone response was profiled for each genotype by hybridising control against ozone-exposed samples per genotype.
Keywords: stress response, genotype comparrison, ozone exposure

sample_organism:
Populus trichocarpa
Populus deltoides

overall_design: RNA was extracted from the fifth leaf below the first fully unfurled leaf for each plant. Control and ozone-exposed plants were then randomly paired for hybridisation.

first: James

last: Tucker

first: Gail

last: Taylor


title: Hyperthermia and p53 effects on global gene expressions

status: Public on May 01 2009

city: College Station

first: Jennifer

last: Goldsby

state: TX

address: 112 Cater-Mattil

department: Nutrition and Food Sciences

institute: Texas A&M University

summary: Neural tube defects (NTDs) are one of the most common human birth defects, with a prevalence of approximately 1 in 1000 live births in the United States. In animal studies, deletion of p53 and hyperthermia leads to a significant increase in embryos that exhibit exencephaly. To evaluate the effects of hyperthermia on p53-deficient mouse embryos we used microarray approach to find genes modified by deletion of p53 and hyperthermia treatment in day 8.5 mouse embryos to identify genes that may be involved in the mechanisms underlining NTDs and begin to define the developmental role of p53 in the etiology of NTDs.

overall_design: After day 8.5 mouse embryos from p53 heterozygous crosses were treated in either control (38C) water bath or hot (43C) water bath for 10 minutes, they were collected, genotyped, and embryos of similar genotype (+/+; +/-; -/-) and treatment were pooled. Total RNA of biological quarduplicate samples were isolated each from p53 +/+, +/-, -/- embryos. Total RNA of each sample was obtained from 3-7 embryos. Gene expression differences between the three genotypes were examined.

first: Hiromi

last: Hosako

middle: S

last: Martin

first: Marrianne

last: Barrier

first: Philip

last: Mirkes


title: Notch targets in DmD8 and KC Drosophila cells

status: Public on Feb 17 2009

city: Cambridge

first: Sarah

last: Bray

address: Downing Street

department: PDN

institute: University  of Cambridge

summary:
To identify genes upregulated in response to Notch signalling in DmD8 cells.

To identify genes upregulated in response to Notch signalling in KC cells.

Keywords: Expression analysis at a single timepoint (30' after Notch activation)

overall_design:
DmD8 cells were obtained from the Drosophila Genomics Resource Center (http://dgrc.cgb.indiana.edu). In the first experiment, 'control cells versus Notch activated cells' mRNA was extracted from DmD8 cells incubated in the absence or presence of EDTA to activate Notch (cells were harvested 30 minutes after addition of EDTA). In a second experiment, 'Notch activated in presenilin inhibited cells versus Notch activated cells', DmD8 cells were split and one 1/2 pretreated with DFK-167 overnight.  Untreated and DFK-treated cells were then exposed to EDTA and harvested after 30minutes. RNA was isolated using Trizol (Sigma) and reverse transcription was performed with Superscript III Reverse Transcriptase (Invitrogen) and oligo-dT primers (Sigma). Control and experimental samples were labeled with Cy3 or Cy5, mixed together and hybridized on Drosophila transcriptome long-oligonucleotide microarrays (FlyChip, FL002;  http://www.flychip.org.uk/services/core/FL002/). Three biological replicates and dye swaps were performed for each experiment (5 arrays in total/experiment). Slides were scanned by Genepix 400B dual laser scanner (Axon) and spots were found and quantified by Dapple software.

KC cells were obtained from Dr Martin Zeidler. Control versus Notch activated KC cells: mRNA was extracted from KC cells incubated in the absence or presence of EDTA to activate Notch (cells were harvested 30 minutes after addition of EDTA). RNA was isolated using TRIzol (Sigma) and reverse transcription was performed with Superscript III Reverse Transcriptase (Invitrogen) and oligo-dT primers (Sigma). Control and experimental samples were labeled with Cy3 or Cy5, mixed together and hybridized on Drosophila transcriptome long-oligonucleotide microarrays (FlyChip, FL002;  http://www.flychip.org.uk/services/core/FL002/). Three biological replicates and dye swaps were performed (6 arrays in total/experiment). Slides were scanned by Genepix 400B dual laser scanner (Axon) and spots were found and quantified by Dapple software.

first: Alena

last: Krejci


title: Gene Expression Differences in p53-Deficient Day 8.5 Mouse Embryos

summary:
Neural tube defects (NTDs) are one of the most common human birth defects, with a prevalence of approximately 1 in 1000 live births in the United States.  In animal studies, deletion of p53 leads to a significant increase in embryos that exhibit exencephaly.  Whereas several studies have closely investigated the morphological changes of p53-deficient embryos, there is no study that has reported the molecular-level alternations in p53-deficient embryos.  Here we use microarray approach to find genes modified by deletion of p53 in day 8.5 mouse embryos to identify genes that may be involved in the mechanisms underlining NTDs and begin to define the developmental role of p53 in the etiology of NTDs.
Keywords: genetic modification

overall_design: Day 8.5 mouse embryos from p53 heterozygous crosses were collected, genotyped, and embryos of similar genotype (+/+; +/-; -/-) were pooled.  Total RNA of biological quarduplicate samples were isolated each from p53 +/+, +/-, -/- embryos.  Total RNA of each sample was obtained from 3-7 embryos. Gene expression differences between the three genotypes were examined.

first: Marianne


title: Control of gene expression by Ott1/RBM15 protein

status: Public on Jun 18 2008

city: Frederick

first: Andrei

last: Zolotukhin

state: MD

address: P.O. Box B

institute: NCI-Frederick

summary: The conserved mRNA nuclear export receptor NXF1 (Mex67 in yeast) assembles with messenger ribonucleoproteins (mRNP) in the nucleus and guides them through the nuclear pore complex (NPC) into the cytoplasm. The DEAD-box RNA helicase Dbp5 is essential for nuclear export of mRNA and is thought to dissociate Mex67 from mRNP upon translocation, thereby generating directional passage. However, the molecular mechanism by which Dbp5 recognizes Mex67-containing mRNP is not clear. Here we report that the human NXF1-binding protein RBM15 binds to DBP5 specifically and enables its direct contact with mRNA. We found that RBM15 is enriched at the nuclear envelope and colocalizes with DBP5 at the NPC. Knockout of RBM15’s orthologue in mouse (Ott1) leads to global changes in mRNA expression and excessive mRNA accumulation in the cytoplasm, indicating an essential role of RBM15 in mRNA export regulation. We propose that RBM15 acts locally at the NPC, by transiently bridging the NXF1-mRNP complexes to DBP5, thereby contributing to the substrate specificity of mRNA export.

overall_design: Mice harboring a low expression Ott1 allele were mated and embryos at day E11.5 were dissected and genotyped. Total RNA was isolated from whole embryos and subject to microarray analysis. The study included biological replicates of all genotypes: wt, n=2, embryos##11, 13; heterozygous, n=3, embryos##3, 10, 15 and homozygous, n=3, embryos##2, 7, 16.

first: Hiroaki

last: Uranishi

first: Susan

last: Lindtner

first: Soren

last: Warming

first: Irina

last: Tretyakova

first: Gen-Mu

last: Zhang

first: Jenifer

last: Bear

first: Neal

last: Copeland

first: Nancy

middle: A

last: Jenkins

first: George

middle: N

last: Pavlakis

first: Barbara

last: Felber


title: Pioglitazone treatment of Lethal Yellow mice

status: Public on Jul 08 2008

first: Kathleen

last: Eyster

summary:
Goal of the experiment:  To examine differential gene expression in the ovaries of heterozygous Lethal Yellow (C57BL/6J Ay/a) mice treated with vehicle or pioglitazone.

Brief description of the experiment:  The objective of this study was to use whole mouse genome DNA microarrays (Codelink)  to examine gene expression the ovaries of heterozygous Lethal Yellow (C57BL/6J Ay/a) mice.
Women with polycystic ovary syndrome (PCOS) treated with thiazolidinediones (TZD), such as pioglitazone, show reduced androgen levels and improved ovulatory function.  Recent evidence suggests that TZD, acting via peroxisome proliferator-activated receptor gamma (PPAR gamma), alter the expression of ovarian genes involved in insulin/IGF signaling that may account for some of the observed effects.  Since TZD can alter the expression of a large variety of genes, there may be other mechanisms involved in the improvement of ovarian function with TZD treatment.  Lethal yellow (LY) mice exhibit progressive obesity, reproductive dysfunction, and altered metabolic regulation (e.g. insulin and leptin resistance) similar to women with PCOS.  This  study was designed to test the hypothesis that prolonged treatment of aging LY mice with pioglitazone would alter the expression of ovarian genes involved in reproduction.
Beginning at 120-days and continuing until 180-days of age, female LY mice received daily oral doses of either 0.1 mg pioglitazone (n = 4) or an equal volume of vehicle (0.1 ml 100% DMSO; n = 4).  At the end of the treatment regimen, ovaries were removed and RNA extracted.  Total RNA extracts were run individually on Codelink Mouse Whole Genome Bioarrays (GE).  Relative gene expression was compared between treatments by t-test using GeneSpring software.

Keywords:  mouse, drug treatment

Experimental factors:  drug treatment

Keywords: drug treatment

overall_design:
Experimental design:  Female heterozygous Lethal Yellow (C57BL/6J Ay/a) mice were randomized to vehicle treatment group (0.1 ml 100% DMSO, n=4) or pioglitazone treatment group (0.1 mg pioglitazone in 0.1 mg 100% DMSO).  The drug/vehicle was administered by gavage daily from 120 days to 180 days of age.  The mice were euthanized on day 60 of treatment (180 days of age) and the ovaries were removed and placed in RNAlater (Ambion).

Quality control steps:  The cRNA that was synthesized from each mouse ovary was used for hybridization to a single CodeLink (Amersham/GE) whole mouse microarray.  Only one sample was hybridized with each slide and only one dye (Alexa 647) was used so no dye swaps were necessary.  Bacterial control spikes were used as per manufacturer's instructions.

Samples used, extract preparation and labelling:
The origin of each biological sample:  The samples were ovarian tissue obtained from heterozygous Lethal Yellow (C57BL/6J Ay/a) mice that had been treated by gavage with vehicle (DMSO) or pioglitazone (0.1 mg/day) for 60 days.

Manipulations of biological samples and protocols used:  The ovaries were dissected free from the animals, cleaned of adhering fat, placed in RNAlater, and frozen at -80C until RNA was extracted.

Experimental factor:  drug treatment

Technical protocols:  The ovaries were homogenized in TRI reagent (MRC) using a Polytron (Brinkmann).  The RNA-containing layer was removed and the RNA purified on an RNeasy extraction column (Qiagen).  The sample was treated with an on-column DNase treatment (RNase-free DNase, Qiagen).  The purity and quantity were evaluated by an Agilent Bioanalyzer using the RNA 6000 Nanoassay LabChip.

Labelled cRNA was prepared using the manufacturer's Instruction Protocol (www1.amershambiosciences.com, CodeLink Gene Expression System: Manual Labelled cRNA Target Preparation).  The source of reagents was the CodeLink Expression Assay Reagent Kit, Manual Prep, except where specified otherwise.  2.0 micrograms of total ovarian RNA was mixed with bacterial control RNA spikes and primed with T7 oligo(dT) primer for 10 min at 70C.  (The bacterial control spikes included araB, entF, fixB, gnd, hisB, and leuB.)  The first strand of cDNA was synthesized with first strand buffer, dNTP mix, RNase inhibitor, and reverse transcriptase for 2 h at 42C.  The second strand cDNA synthesis reaction was prepared using second strand buffer, dNTP mix, DNA polymerase mix, and RNase H; the reaction was carried out for 2h at 16C.  The double-stranded cDNA was purified on QIAquick columns (Qiagen) and the eluent was dried down in a SpeedVac concentrator.  The double-stranded cDNA was resuspended in a mixture containing T7 reaction buffer, T7 ATP, T7 GTP, T7 UTP, T7 CTP, biotin-11-UTP, and T7 enzyme mix for the synthesis of cRNA.  The cRNA synthesis reaction was terminated after 14h at 37C by purifying the cRNA on RNeasy columns (Qiagen).  The concentration of cRNA was determined by spectrophotometry.

Hybridization procedures and parameters:  10 micrograms of cRNA was mixed with fragmentation buffer and heated to 94C for 20 min.  The fragmented cRNA was mixed with CodeLink hybridization buffer, loaded on the microarray slides, and hybridized for 18 hours at 37C.
The slides were washed in 0.75x TNT (Tris-HCl, NaCl, Tween-20) at 46C for 1h then incubated with streptavidin-Alexa 647 fluorescent dye for 30 min at room temperature.  The Alexa fluor was prepared in TNB blocking buffer (0.1M Tris-HCl, 0.15M NaCL, 0.5% NEN Blocking Reagent-PerkinElmer)  The slides were then washed 4 times for 5 min each in 1x TNT and twice in 0.05% Tween 20 for 5 sec each.  The slides were dried by centrifugation and scanned in an Axon GenePix 4000B scanner.

first: John

last: Brannian

first: Maureen

last: Diggins


title: Gene expression profiles of four murine tumors after carbon-ion (C-ion) or gamma irradiation.

status: Public on Jun 09 2008

city: Chiba

first: Kazunori

last: Nojiri

address: 4-9-1, Anagawa, Inage-ku

country: Japan

institute: National Institute of Radiological Sciences

summary:
This study uses microarray analysis to examine the fate of multiple C-ion-irradiated tumors in vivo to gain a comprehensive overview of the changes in gene expression induced by C-ion irradiation. Four murine tumors were irradiated in vivo with C-ions at a single dose, and gamma-rays were used as a reference beam.

Keywords: mouse squamous cell carcinoma and fibrosarcoma,  carbon ion-irradiation, gamma-irradiation, resected sample, transplanted tissues

overall_design: We examined gene expression changes after carbon-ion (C-ion) irradiation (290 MeV/m, SOBP 6 cm middle, 50 kev/mm) with a single dose of 30 Gy in four mouse tumors (NR-S1, SCCVII, NFSa and #8520) transplanted into the hind legs of C3H/HeNrs mice, using 44K single-color oligomicroarrays at 6 hours, 1 day and 3 days after irradiation. Gamma rays of 30 Gy and 50 Gy were used as a reference beam.

first: Kaori

last: Imadome

first: Mayumi

last: Iwakawa


title: Radiation-induced genes expressed in two murine tumors.

status: Public on Mar 20 2009

summary:
Gene expression profiling  was used to identify genes that display radiation-induced transcriptional change over tumor histopathology.
Keywords: mouse squamous cell carcinoma, fibrosarcoma and mammary carcinoma, gamma-irradiation, carbon ion irradiation,  resected sample, transplanted tissues

overall_design:
C3H/HeMs male mice were irradiated by gamma-rays (30Gy, 50Gy, 70Gy) or carbon ions (30Gy) as local irradiation in single doses to hind legs where each tumor was transplanted. Animals were sacrificed either
before irradiation (pre) or 1 day after irradiation (day 1)
for expression analysis.

first: Takashi

last: Imai


title: Histone deacetylase 4 represses dystrophin-glycoprotein (DGC) complex expression

status: Public on Mar 25 2008

city: Durham

first: Zachary

middle: Conrad

last: Hartman

state: NC

address: Research Drive MSRB rm 414

department: Surgery

institute: Duke University

summary:
Dysfunction of the dystrophin-glycoprotein complex (DGC) is a frequent cause of hereditary forms of muscular dystrophy. Although DGC function in maintaining skeletal muscle integrity has been well characterized, little is known about how the DGC complex is coordinately regulated at the transcriptional level.  To test this hypothesis, we engineered HDAC4 stably overexpressing and control myotubes in an in vitro model of muscle differentiation.  Here we present evidence that HDAC4, a neural activity-responsive histone deacetylase, is a critical transcriptional regulator of the DGC complex. We show that HDAC4 can repress multiple components of the DGC complex, including dystrophin and sarcoglycan family members in both cultured myotubes.  To confirm this finding, the protein levels of core DGC complex members including dystrophin, sarcoglycan complex members, and additional dystrophin-associated proteins were evaluated in differentiated myotubes by western analysis.
Keywords: genetic modification and cell type comparison of muscle cells

overall_design: C2C12 mouse myotubes were infected with either a HDAC4 expressing or control (Neo) retroviruses.  After stable selection, myotubes were differentiated at 90% confluency in 2% horse serum (Hyclone) for 4 days.  At this time point, RNA was extracted and the two types of cells compared in a microarray analysis.

first: Todd

last: Cohen

first: Tomasa

last: Barrientos

first: Sean

last: Garvey

first: Gregory

last: Cox

first: Richard

last: Bedlack

first: Tso-Pang

last: Yao


title: Effect of forced uncoupling protein-1 expression on 3T3-L1 adipocyte gene expression

status: Public on Mar 24 2008

first: Arul

last: Jayaraman

address: 3122 TAMU

department: Artie McFerrin Department of Chemical Engineering

summary:
The aim of this study was to investigate the molecular basis underlying the response of 3T3-L1 adipocytes to forced expression of UCP1.
Keywords: adipocyte, Uncoupling protein 1, microarray

overall_design: 3T3L1 adipocytes expressing control and Uncoupling protein 1 were grown to confluence.  Total RNA was collected and hybridized to Codelink mouse whole  genome Bioarrays.  All the samples were done in triplicate.

first: Fatih

last: Senocak

first: Yaguang

last: Si

first: Kyongbum

last: Lee


title: Ozone exposure of tolerant and sensitive F2 genotypes of Populus

first: Nathaniel

last: Street

summary:
Three tolerant and three sensitive genotypes of the inbred QTL mapping population Family 331 were exposed to either ambient air or an acute ozone exposure of 200 ppb for 9 hrs and ozone response was profiled for each genotype by hybridising control against ozone-exposed samples per genotype.
Keywords: stress response, genotype comparrison, ozone exposure

overall_design: Three replicates were grown per genotypes used and RNA was extracted from the fifth leaf below the first fully unfurled leaf. After RNA extraction, equal quantities of RNA from each replicate of a genotype per treatment were pooled to provide a single pool per treatment per genotype. Control and ozone-exposed sample per genotype were then hybridised. Hald of all arrays were hybridised with the control sample in the Cy3 channel and half with control in the Cy5 channel.

first: James

last: Tucker

first: Gail

last: Taylor


title: Prostate cancer cell line with slienceing of PDEF or control (GFP) gene using siRNA.

status: Public on Dec 20 2007

city: Boston

first: Towia

last: Libermann

address: 99 Brookline Ave.

institute: Beth Israel Deaconess Medical Center

summary:
PDEF(Prostate Derived Ets factor) is a member of Ets transcription factor with specific expression in Prostate. We identified target genes for PDEF in prostate cancer cell line PC-3 using Affymetrix Genechip Microarray technology.
Keywords: target gene identification for transcription factor

overall_design: Stable cell lines were generated by infecting with Lentivirus contained silence RNA for PDEF and control GFP in PC-3 cells. 48 h after infection, total RNA was isolated from those samples.