title: Microarray analysis of the fiber cell maturation in the lens

status: Public on Feb 15 2005

city: Miami

first: Valery

middle: I.

last: Shestopalov

state: FL

department: Ophtalmology

institute: University of Miami

summary:
The mammalian lens consists of an aged core of quiescent cells enveloped by a layers of mature fully elongated and younger, continuously elongating and transcriptionaly active cells. In the present study, we compared global gene expression profiles in elongating and mature, non-elongating fiber cells. The microarray-based comparison of gene expression profiles was performed using Agilent 22K Mouse Oligo microarrays with two rounds of RNA amplification from samples obtained by laser capture microdissection (LCM) of paraffin-embedded lens sections.

Lenses were removed and immediately fixed using 4% paraformaldehide or methanol-based UMFIX reagent (Sakura Finetek USA, Inc.). To map the exact location of both elongating and maturing fibers on the lens slices, lenses were vibratome-sliced (Vibratome 1000, St. Louis, MO) as described previously and the GFP expression pattern was captured by confocal microscopy. For RNA extraction lens tissue was sliced into 5 micron-thick paraffin sections and microdissected using LCM (Leica Microsystems, Bannockburn, IL). The mid-saggital slices were used both for syncytium border measurements and for LCM. Control measurement (data not shown) confirmed that similar rates of shrinkage in paraformaldehyde- and UMFIX-fixed samples did not affect precision of LCM dissection.
Fiber cell samples were dissected out of 5 micron-thick slices. In one experiment we typically processed about 40 slices, which was sufficient to collect the minimum of 200 zone-specific cells pooled from P5 littermate lenses. This sample size provided a reliable representation of RNA species in experimental procedure originally designed and tested for just 1-10 cells.
Localization of the syncytium border characterized by the abrupt change of GFP labeling pattern was captured by confocal microscopy as described previously. In brief, GFP fluorescence was visualized using LSM510 instrument (Carl Zeiss, NY) equipped with an argon/krypton laser at 488 nm excitation and a 515-565 nm band pass emission filter. Physical parameters of the zones containing young and maturing fibers were measured in fixed lens slices using the morhphometric software provided by Zeiss.

Lenses were fixed in 4% paraformaldehyde/PBS and sectioned with a vibratome. Cells from elongating and maturing fiber regions were dissected out using the Leica DMLA laser capture microscope, (Leica Microsystems, Bannockburn, IL). The cut-out pieces containing captured cells were put directly into tubes containing the lysis buffer supplied in the Absolutely RNA® nanoprep kit (Stratagene, La Jolla, CA). Total RNA from the microdissected tissue sections was extracted and purified using the Absolutely RNA nanoprep kit according to the manufacturer's protocol. Caps briefly placed onto the section without laser activation were used as negative controls. Samples from several age-matched lenses were pooled together to obtain differentiation-specific samples for microarray analysis.
Target RNA amplification and labeling with Cy-3 or Cy-5 dyes from CyDye Post Labelling Reactive Dye Pack (Amersham, USA) was carried out in two rounds using the Amino Allyl MessageAmp™ aRNA Kit (Ambion, USA) as specified by the manufacturer. Quality and size distribution of the targets were determined by the Agilent 2100 Bioanalyzer (Agilent Technologies, USA). The Amino Allyl MessageAmp aRNA Kit is configured to incorporate the modified nucleotide, 5-(3-aminoallyl)-UTP (aaUTP) into the aRNA during in vitro transcription. Once purified and fragmented, the dye labeled aRNA was used for microarray hybridization.
Labeled amplified RNA was hybridized to the 22K Mouse Oligo microarrays (Agilent Technologies)) according to the manufacturer's instructions. After hybridization microarrays were washed and scanned using GenePix 4000A (Axon Instruments,Inc.).
Keywords = Lens
Keywords = differentiation
Keywords = microarray
Keywords = gene expression profiling
Keywords = laser capture microdissection
Keywords = RNA amplification
Keywords: repeat sample

first: Dmitry

last: Ivanov

first: Galina

last: Dvoriantchikova

first: Anna

last: Pestova

first: Lubov

last: Nathanson


title: Integrating phenotypic and expression profiles to map arsenic response networks

status: Public on Nov 01 2004

city: RTP

first: NIEHS

last: Microarray Core

state: NC

address: 111 T.W. Alexander Drive

department: DIR

country: USA

institute: NIEHS

summary:
The parent S. cerevisiae strain, BY4741, and specific gene deletions of the parent S.cerevisiae strain BY4741 were treated with 100uM NaAsO2 (AsIII) for 2h and compared to the untreated counterpart (yap1Δ vs. yap1Δ 2h 100uM AsIII, cad1Δ vs. cad1Δ 2h 100uMAsIII, rpn4Δ vs. rpn4Δ 2h 100uM AsIII, arr1Δ vs. arr1Δ 2h 100uM AsIII, parent vs. parent with 2h 100uM AsIII).  The untreated specific deletion strain was also compared to the untreated parent strain in order to identify differential expression of genes due to the knockout alone.
Keywords: other

first: Astrid

middle: C

last: Haugen

first: Ryan

last: Kelley

first: Jennifer

middle: B

last: Collins

first: Charles

middle: J

last: Tucker

first: Changchun

last: Deng

first: Cynthia

middle: A

last: Afshari

first: J

middle: M

last: Brown

first: Trey

last: Ideker

first: Bennett

last: Van Houten


title: laughter regulates postprandial blood glucose levels and gene expression

status: Public on Oct 21 2004

city: Minamidai, Kawagoe

first: Toshiro

last: Saito

state: Saitama

department: Life Science Group

country: Japan

institute: Hitachi, Ltd.

summary:
Sample tissue: peripheral blood
Disease: diabetes
Samples for gene expression analysis were obtained before the meal and 1.5 hours after the event.
Event: listening to a Japanese comic story or a monotonous academic lecture without humor.
Keywords: equivalent probe


title: Aortas of mice on high fat diet

status: Public on Aug 01 2004

city: Stanford

first: Raymond

last: Tabibiazar

state: CA

institute: Stanford

summary:
Expression profiles in aortas isolated from mouse strains C3H.2/HeJ and C57.2Bl/6. Mice were were either on a high fat diet or normal diet for 0, 4, 10, 24, or 40 weeks.
Keywords: other


title: Novel N-terminal Glucocorticoid Receptor Isoforms with Unique Transcriptional Target Genes

status: Public on Jun 20 2005

summary:
Gene expression analysis was conducted using Agilent Human1Av2 arrays (Agilent Technologies, Palo Alto, CA). Two separate biological replicates of cytoplasmic RNA samples were harvested and purified from each of the U-2 OS cell lines stably expressing hGRalpha, -A, -B, -C3, or D3 treated with 100 nM DEX or vehicle for 6 hours using RNeasy Midi kits (Invitrogen). These RNA samples were amplified using the Agilent Low RNA Input Fluorescent Linear Amplification Kit protocol. Starting with 0.5 ng of total RNA, two cRNAs, one labeled with Cy3 and one labeled with Cy5 were produced for each sample according to the manufacturer’s protocol. Consequently, four separate chips were used for each of the isoforms at each treatment condition, yielding a total of 40 chips: 5 isoforms X 2 treatments (vehicle vs. hormone) X 2 biological replicates X 2 labeling dyes. For each two-color comparison, 750 ng of each Cy3 and Cy5 labeled cRNAs were mixed and fragmented using the Agilent In Situ Hybridization Kit protocol. Hybridizations were performed for 16 hours in a rotating hybridization oven using the Agilent 60-mer oligo microarray processing protocol. Slides were washed as indicated in this protocol and then scanned with an Agilent Scanner. Data were obtained using the Agilent Feature Extraction software (v7.1), using defaults for all parameters.
Keywords: other

first: Nick

middle: Z

last: Lu

first: John

last: Cidlowski


title: Hearts_DD_Screening

status: Public on Dec 07 2004

city: Palo Alto

first: Svetlana

last: Shchegrova

institute: Agilent Technologies, Inc

summary:
Expression profiles in Adult Heart and Fetal Heart normal tissues.
Keywords: other


title: DNA methylation in Arabidopsis thaliana

status: Public on Jun 12 2004

city: Austin

first: Matthew

middle: Wayne

last: Vaughn

state: TX

address: 10100 Burnet Rd

department: Texas Advanced Computing Center

institute: University of Texas at Austin

type: Methylation profiling by genome tiling array

summary:
DNA methylation in wild type bolting plants, wild type seedlings, and ddm1 seedlings.

The purpose of the McrBC methylation microarray assay is to determine which regions of a genome are methylated versus those that are unmethylated in a single Arabidopsis thanliana genotype.

McrBC is a methylation-sensitive enzyme that restricts DNA only at purine-Cmethyl half sites when separated between 50bp and 3kb. A designated amount of DNA from a particular genotype is sheared to a size range of 1kb-10kb using nebulization. We restrict half of the nebulized DNA with McrBC, and the methylated fraction is then removed from the unmethylated fraction through gel purification of DNA fragments greater than 1kb.* The remaining nebulized DNA is subjected to the same gel purification scheme, but with no McrBC treatment.

In a single hybridization, the untreated sample is labeled with Cy5 and the McrBC-treated sample with Cy3. Thus, after labeling and microarray hybridization, the ratio of normalized Cy5 to normalized Cy3 represents the relative methylation at the sequence represented by the spot on the microarray. Dye swap analysis is carried out to take account of experimental variation by repeating the hybridization with identical samples labeled with Cy3 and Cy5, respectively.

This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series

first: M

middle: W


title: Histone methylation in wild type and ddm1 Arabidopsis thaliana seedlings

type: Genome binding/occupancy profiling by genome tiling array

summary:
Histone 3 lysine 4 and histone 3 lysine 9 methylation in wild type and ddm1 Arabidopsis thaliana seedlings. The purpose of the chromatin immunoprecipitation/microarray (ChIP/chip) experiment is to determine which regions of a genome are enriched for a particular histone modification in a single Arabidopsis thanliana genotype.

Chromatin immunoprecipitation with antibodies raised against dimethyl histone-H3 lysine-9 (H3mK9) or dimethyl histone-H3 lysine-4 (H3mK4) is performed on a selected genotype. This purified DNA from each immunoprecipiation (mH3K9, mH3K4, no antibody control) is used for random amplification to increase the quantity of DNA for microarray hybridization.

The amplified DNA from each experimental sample is then labeled with Cy5 and hybridized against total input DNA from the corresponding genotype, labeled in Cy3. In a single hybridization, the total input DNA serves as a baseline and is compared to the immunoprecipitated samples. Ratios of normalized signal intensities were calculated to identify enrichment of a particular sequence after immunoprecipitation, in comparison to the total input DNA. Dye swap analysis is carried out to take account of experimental variation by repeating the hybridization with identical samples labeled with Cy3 and Cy5, respectively.

This SuperSeries is composed of the SubSeries listed below.


type: Expression profiling by genome tiling array

summary:
mRNA levels in Wild-type versus ddm1 Arabidopsis thaliana seedlings and bolting plants.  Features found to be significantly enriched for DNA methylation were determined.

This SuperSeries is composed of the SubSeries listed below.


status: Public on Mar 10 2004

city: La Jolla

first: Jinseong

last: Cheong

institute: University of California at San Diego

summary:
The purpose of this study is to investigate the changes of global gene expression in E. coli during an oxygen shift. All cultures were grown under aerobic or anaerobic conditions in M9 minimal media supplemented with glucose. Samples were RNA-stabilized using Qiagen RNAProtect Bacterial Reagent, and total RNA was isolated from exponentially growing cells using a Qiagen RNeasy mini kit (protocols available at www1.qiagen.com). The RNA (10 µg) was then used as the template for cDNA synthesis, the product of which was fragmented, labelled, and hybridized to an Affymetrix E. coli Antisense Genome Array, which was washed and scanned to obtain an image. All of these steps were performed according to Affymetrix protocols (available at www.affymetrix.com).

This SuperSeries is composed of the SubSeries listed below.

first: Markus

last: Covert

first: Eric

last: Knight

first: Jennie

middle: L

last: Reed

last: Herrgard

first: Bernhard

middle: O

last: Palsson


title: Effect of LPS and LPS-binding protein treatment on RAW 264.4 cells

status: Public on Mar 05 2004

city: Pasadena

first: Sangdun

last: Choi

address: 1200 E. California Blvd

department: Biology

institute: California Institute of Technology

summary:
All samples were gathered from mouse RAW 264.7 cells (macrophages). Control total RNA was extracted from untreated RAW 264.7 cells cultured for either 1, 2, 4, 8, 16 or 48 hours. Test total RNA was extracted from lipopolysaccharide (100ng/ml) and lipopolysaccharide-binding protein (100pM) treated RAW 264.4 cells cultured for either 1, 2, 4, 8, 16 or 48 hours.

This SuperSeries is composed of the SubSeries listed below.

first: Yun-Anna

last: Cao

first: Mi-Sook

last: Chang

first: Sun-Young

last: Lee

first: Meghdad

last: Rahdar

first: Robert

last: Hsueh


title: Polysomal and Cytoplasmic RNA of U937 cells before and after PMA stimulation

city: Ebetsu

first: Hiroshi

last: Kitamura

address: 582 Midorimachi, Bunkyodai

department: School of Veterinary Medicine

institute: Rakuno Gakuen University

summary:
Human histomonocytic U937 cells exhibit macrophage-like properties after phorbol ester stimulation.  Accumulating evidence demonstrated that remarkable number of transcripts changed in their amount at total RNA levels.  However, post-transcriptional regulation has not been fully elucidated in this model.  Thus, we compared expression profiles between polysomal and cytoplasmic RNAs before and after phorbol 12-myristate 13-acetate stimulation (48h, 32nM).

Table 1: Detailed description of the expression data of human histomonocytic cell line U937 cells.  Data of total cytoplasmic and polysomal fractions before and after PMA stimulation is shown.  This table contains all spot data except: 1) saturated spots, 2) undetectable spots, 3) stained spots, and 4) spots only detected in less than one samples.

Column A: Gene symbol (“I_xxxxxx” is corresponding to EST clones)
Column B: Genbank accession no.
Column C: Description
Column D: Average value of expression levels in total cytoplasmic fraction in the absence of PMA
Column E: Average value of expression levels in total cytoplasmic fraction in the presence of PMA (32nM, 48h)
Column F: Average value of expression levels in polysomal fraction in the absence of PMA
Column G: Average value of expression levels in polysomal fraction in the presence of PMA
Column H: Expression ratio of polysome, PMA (-) to cytoplasm, PMA(-)
Column I: Expression ratio of polysome, PMA (+) to cytoplasm, PMA(+)
Column J: Expression ratio of cytoplasm, PMA (+) to cytoplasm, PMA(-)
Column K: Expression ratio of polysome, PMA (+) to polysome, PMA(-)
Column L: Different expression between polysome, PMA (-) and cytoplasm, PMA(-): different=1
Column M: Different expression between polysome, PMA (+) and cytoplasm, PMA(+): different=1
Column N: Different expression between cytoplasm, PMA (+) and cytoplasm, PMA(-): different=1
Column O: Different expression between polysome, PMA (+) and polysome, PMA(-): different=1

Table 2:  Candidates post-transcriptionally regulated by PMA.  We extracted 105 transcripts whose expression levels were altered only in either fraction accompanied by changes in polysome/cytoplasm expression ratio.
Column A: Gene symbol (“I_xxxxxx” is corresponding to EST clones)
Column B: Genbank accession no.
Column C: Description
Column D: Average value of expression levels in total cytoplasmic fraction in the absence of PMA
Column E Average value of expression levels in total cytoplasmic fraction in the presence of PMA
Column F: Average value of expression levels in polysomal fraction in the absence of PMA
Column G: Average value of expression levels in polysomal fraction in the presence of PMA
Column H: Expression ratio of polysome, PMA (-) to cytoplasm, PMA(-)
Column I: Expression ratio of polysome, PMA (+) to cytoplasm, PMA(+)
Column J: Expression ratio of cytoplasm, PMA (+) to cytoplasm, PMA(-)
Column K: Expression ratio of polysome, PMA (+) to polysome, PMA(-)
Keywords = phorbol ester
Keywords = macrophage
Keywords = polysome
Keywords = post-transcriptional regulation
Keywords: parallel sample

first: Tomoko

last: Nakagawa

first: Michiyo

last: Takayama

first: Osamu

last: Ohara


title: X-radiation (XR) resistant HCT116-Clone2: XR at 0 and 4 Gy

status: Public on Jun 04 2004

city: Ottawa

first: Felix

middle: M.

last: Mesak

state: ON

address: 503 Smyth Rd. 3rd Floor

department: Centre for Cancer Therapeutics

country: Canada

institute: Ottawa Regional Cancer Centre

summary:
cDNA microarray study of X-radiation (XR) resistant HCT116-Clone2 cells without XR, 10 minutes, 6 hours, or 24 hours after XR at 4Gy versus unirradiated HCT116-Clone10 cells. HCT116-Clone10 cells have similar XR response with parental HCT116 cells.

This SuperSeries is composed of the SubSeries listed below.

first: Cheng

middle: E

last: Ng

first: Wei

last: Tan


title: Mouse feeding experiment

status: Public on Feb 09 2004

city: Karlsruhe

first: Michael

middle: J.

last: Pankratz

department: Institute of Genetics

country: Germany

institute: Forschungszentrum Karlsruhe

summary:
Mice used in this study were 129/sv males, 8-15 weeks of age, which were housed individually and received water ad libitum. Normal fed animals were fed before and during the 48h experiment with standard food ad libitum. Sugar fed animals were fed before the experiment with standard food ad libitum and during the 48h experiment with 50% (w/v) sugar solution (40% glucose, 10% sucrose). Mice were killed by CO2 asphyxiation. The livers were snap-frozen in liquid nitrogen and stored at -80 degrees C. Total RNA was prepared using the Nucleospin RNA L kit (Macherey-Nagel, Düren, Germany), with an additional step in which the final eluate was used for a subsequent elution step. PolyA RNA was extracted with the Ambion (Ambion Inc, Austin, USA) Purist kit according to the manufacturer's protocols. Equal amounts of polyA RNA were pooled to generate pools of all animals. Pool1, normal = 3 animals (N1, N2, N3), pool2, normal = 3 animals (N4, N5, N6), pool3, = 4 animals (N7, N8, N9, N10), pool1, sugar = 3 animals (Su1, Su2, Su3), pool2, sugar = 3 animals (Su4, Su5, Su6), pool3, sugar = 4 animals (Su7, Su8, Su9, Su10). Labeled cDNA was synthesized from 1.5 µg polyA RNA using the Amersham (Amersham Europe, Freiburg, Germany) direct cDNA labeling kit. Removal of the unincorporated dyes and concentration of the target were done with Microcon 30 (Millipore, Bedford, MA) spin columns according to the manufacturer's instructions. The concentrated probes were hybridized to the microarray in 1x dig easy hyb buffer (Hoffman-la Roche, Basel, CH) overnight at 42 degrees C. After hybridization the microarrays are washed 5’ in 2x SSC, 0.1% SDS at 42 degrees C; 10’ in 0.1x SSC, 0.1% SDS at RT; 4 times 1’ in 0.1x SSC at RT and shortly dipped into 0.01x SSC. Drying is performed in centrifuge on a microtiterplate holder at 130 rcf for 7’. Slides are stored in dark until they are scanned. Arrays were scanned using the dual laser scanner Axon 4000B and the corresponding software Genepix 4 (Axon Inc., Union City, CA, USA). Both channels (532 nm for Cy3 and 635 nm Cy5) were scanned in parallel and stored as 16 bit tiff files. The absolute intensity values span the range from 0 to 65,535. The scans were performed with a resolution of 10 µm. From each spot with a mean diameter of 100 µm, 100 data pixels were recorded. Individual local background area around the spots were defined, which included approximately 400 pixels and excludes neighboring spots. For each channel, the raw data was calculated as the median intensity of all foreground pixels with respect to all background pixels. Each array was scanned three times (low, medium and high scan) with different signal amplification factors (voltage settings of the photo multiplier tubes), but with the same laser power. The channels for Cy3 and Cy5 were balanced in each scan for approximately the same intensity profile. In the low scan no spot was saturated, in the high scan the signal amplification for Cy5 was set to approximately 80% of maximum and the Cy3 amplification was adjusted to this. The settings used in the medium scan lie between the low and the high scan. This method for scanning has several advantages. In the low scan where no spot is in saturation, it is possible to calculate the real ratio for genes with high expression levels; however, those with a low expression level are most likely not recognized. In order not to lose these a high scan is made; in this case the information on the saturated spots is lost so the two scans complement each other. The medium scan produces additional values for subsequent calculations. By scanning the arrays three times errors which occur while recording and which might increase the error factor in the normalization are averaged. The data processing was automated in a Visual Basic/Excel (Microsoft) program, which integrates the following steps. Raw data are derived from the result files generated by scanning and picture analysis software Genepix. The spot diameter is to lie in the range from 80 to 120 µm otherwise these spots are not included in analysis. From the pixel to pixel ratios between the foreground values of both colors, the standard deviation (SD) is calculated. Those spots which show a ratio SD of higher than 3 are excluded from analysis due to inconsistency. The foreground signal of each spot is corrected by subtracting the corresponding local background. Resulting values which are lower than the background are replaced by the background value. This defines the minimum measurement threshold as the background and avoids calculations of completely wrong ratios, e.g., if the background corrected value for one channel is close to zero. Spots are marked as saturated in one channel if more than 10% of the foreground pixels are at the highest absolute value. From the unsaturated data points of all three scans, linear regression between the low and medium, and between low and high scans, are calculated for both colors. Taking these regressions, the values of saturated spots are replaced by estimated ones which reflect the real intensity. The generated data set of each scan is normalized afterwards based on the intensity-dependent methods as described by Yang et al (Yang YH, Dudoit S, Luu P, Lin DM, Peng V, Ngai J, and Speed TP. Normalization for cDNA microarray data: robust composite method addressing single and multiple slide systematic variation. Nucleic Acids Res. 30: e15, 2002). The ratios are calculated as log2 transformed values: M=log2 (Cy5/Cy3). Number describing the spot intensity A=log2sqrt (Cy5*Cy3) is calculated according to Yang et al. A within pin-tip group loess fit to the MA-plot was made. The loess scatter plot smoother performs robust locally fits using a tricube function to weight the values relative to the median point in the intensity interval. As interval size 20% was chosen, which corresponds to 180 data points in a pin-tip group containing 30 by 30 spots. After combining the normalized data from all blocks the data sets of all scans are additionally normalized using a global approach. The normalization was performed so that the sum of all log transformed ratios (M) is 0. All statistical normalizations are based on the assumption that (1) the majority of genes are not changed in their expression and (2) that the overall up and down regulations statistically compensate each other in sum. Taking this into account, the standard deviation in the ratios of the stable genes (80% of the most unchanged genes) could be seen as the measurement noise of the array experiments. To be able to compare the different scans and different normalized microarrays, the SD of the stable genes ratios was adjusted to a medium estimated value of 0.2, which is dependent on the used array system. This value does not influence the subsequent statistical analysis because the normal distribution of the data is not changed. For each microarray the normalized and adjusted log ratios of the three scans are averaged.

This SuperSeries is composed of the SubSeries listed below.

first: Matthias

last: Bauer

first: Anne

last: Hamm

first: Melanie

last: Bonaus

first: Joerg

middle: D

last: Katzenberger

last: Gähler


title: Anti-GBM antibody effect on kidney

status: Public on Jan 15 2004

first: Sangdun

last: Choi

summary:
Control total RNA, pooled from the kidney of five normal BALB/c mice, was used as a reference against total RNA extracted from the kidney of BALB/c mice injected with anti-GBM antibody for 1, 3, 7, 11 and 16 days.

This SuperSeries is composed of the SubSeries listed below.

first: Ju-Han

last: Kim

first: Il-Soo

last: Ha

first: Chang-Il

last: Hwang

first: Young-Ju

first: Ji-Hoon

first: Seung-Hee

last: Yang

first: Yon-Su

first: Dong-Sup

first: Yun-Anna

last: Cao

first: Woong-Yang

last: Park


title: Heatshock otd / OTX2

status: Public on Feb 25 2002

city: Basel

first: Thomas

last: Loop

address: Klingelbergstrasse 50

department: Institute of Zoology

country: Switzerland

institute: University of Basel

summary:
The homeobox genes of the orthodenticle (otd)/Otx family play conserved roles in embryogenesis of the head and brain.  Gene replacement experiments show that the Drosophila otd gene and orthologous mammalian Otx genes are functionally equivalent, in that overexpression of either gene in null mutants of Drosophila or mouse can restore defects in cephalic and brain development. This suggests that otd and Otx genes can control a comparable subset of downstream target genes in either organism.  Here we use quantitative transcript imaging to analyze this equivalence of Drosophila otd and human Otx gene action at a genomic level.
In this study, transgenic fly strains carrying the otd coding sequence or the human Otx2 coding sequence under control of the heat inducible Hsp70 promoter were used [Leuzinger et al.: Equivalence of the fly orthodenticle gene and the human OTX genes in embryonic brain development of Drosophila. Development 1998; 125:1703-1710.].  Stage 10-17 embryos were given a 25-min heat pulse in order to overexpress the otd or Otx2 genes and allowed to recover for 25 min.
Keywords = Drosophila melanogaster, embryo, brain, development, otd, OTX, heatshock
Keywords: ordered

first: Haiqiong

last: Montalta-He

first: Ronny

last: Leemans

first: Martin

last: Strahm

first: Ulrich

last: Certa

last: Primig

first: Dario

last: Acampora

first: Antonio

last: Simeone

first: Heinrich

last: Reichert


title: Classification of Antipsychotic Drugs by Gene Expression in the Frontal Cortex of Mice

status: Public on Sep 24 2004

city: Kagoshima

first: Shin-ichi

last: Iwata

state: Kagoshima

address: Sakuragaoka 8-35-1

department: Pharmacology

institute: Kagoshima University

summary:
Antipsychotic drugs are classified as typical and atypical based on extrapyramidal effects. However, since the frontal cortex is one of the most important regions for antipsychotic actions, this study attempted to classify antipsychotic drugs based on gene expression in the frontal cortex. Chlorpromazine and thioridazine were selected as typical antipsychotics, and olanzapine and quetiapine as atypical antipsychotics. Since these drugs have similar chemical structures, the effect of the basic structure on gene expression can be eliminated. Cluster analysis of microarray experiments showed thioridazine and olanzapine constituted a robust cluster. K-means clustering separated 4-drug-administered mice into chlorpromazine-quetiapine and thioridazine-olanzapine groups. This classification scheme is different from that which is based on criteria currently used to group the typical and atypical drugs and suggests that antipsychotic drugs can be further separated into multiple groups.
Keywords: repeat sample

overall_design:
Male 13-week-old ddY mice (Japan SLC Co., Hamamatsu, Japan) weighing 38-43 g were used. Animals were housed with free access to standard food in an air-conditioned room under a constant dark-and-light cycle (light: 7:00 a.m. to 7:00 p.m.) at a temperature of 22 to 24°C and 60 to 70% relative humidity. Ether was used for anesthesia during decapitation. All efforts were made to minimize animal suffering and to reduce the number of animals used. The present experiments were carried out after obtaining permission from the Committee of Animal Experimentation of the Graduate School of Medical and Dental Sciences at Kagoshima University.
Doses of 25 mg/kg chlorpromazine HCl (Sigma-Aldrich Co., St. Louis, MO), 25 mg/kg thioridazine HCl (Sigma-Aldrich Co.), 1.25 mg/kg olanzapine (gift from Eli Lilly and Company, Indianapolis, IN) and 18.75 mg/kg quetiapine fumarate (gift from AstraZeneca, Macclesfield, UK) were used in the study. The dosages for these drugs were determined from therapeutically equivalent doses previously reported (Lehman et al, 2003; Woods, 2003). The drugs were dissolved in acetic anhydride and diluted with 0.9% saline, resulting in a final concentration of acetic acid of 0.5%. The drugs were injected intraperitoneally once daily for 28 consecutive days in a volume of 0.1 ml/10 g body weight.
Microarray experiments were performed using an Agilent G4121A Mouse Oligo Microarray Kit (Agilent Technologies, Palo Alto, CA) as per the manufacturer’s instructions. The frontal cortex was immediately removed and the RNA was stabilized in RNAlater RNA Stabilization Reagent (Qiagen, Valencia, CA) and stored at -80°C until use. Total RNA was extracted using the RNeasy Mini Kit (Qiagen). The RNA was amplified and labeled by the Low RNA Input Fluorescent Linear Amplification Kit (Agilent). To synthesize cDNA, 200 ng total RNA was used. Vehicle-injected controls were labeled by cyanine 3 (PerkinElmer Life Sciences, Inc., Boston, MA) and drug-injected mice were labeled by cyanine 5 (PerkinElmer Life Sciences, Inc.). Hybridizations to the microarray were performed using the In situ Hybridization Kit Plus (Agilent). Doses of 750 ng cyanine 3-labeled cRNA, 750 ng cyanine 5-labeled cRNA, and control targets were mixed and fragmented in the kit’s fragmentation buffer, and then hybridized to the microarrays for 17 hours at 60°C in a hybridization rotator (Agilent) set to rotate at 4 rpm. Microarrays were washed in 6×SSC with 0.005% Triton X-102 at RT for 10 min, followed by 0.1×SSC with 0.005% Triton X-102 at 4°C for 5 min. The slides were dried, and then scanned by the Agilent G2565BA Microarray Scanner System. Data were analyzed using the Agilent Feature Extraction Software version 7.1. A rank consistency filter and LOWESS were used for dye normalization. Control mice and drug-injected mice were processed in parallel. The data discussed in this publication are presented in accordance with the MIAME guidelines (http://www.mged.org/Workgroups/MIAME/miame.html) and were deposited in NCBIs Gene Expression Omnibus (GEO, http://www.ncbi.nlm.nih.gov/geo/). They can be accessed using the GEO series accession number GSE1501.
Cluster analysis was performed using free software written by M. Eisen (http://rana.lbl.gov/EisenSoftware.htm). The cclust package in the “R” statistical software system (www.cran.r-project.org) was used to find the most appropriate number of clusters (i.e. “k” in the k-means clustering).

first: Hirofumi

last: Morioka


title: angiogenesis course

status: Public on Nov 25 2003

city: Adelaide

first: Anna

last: Tsykin

state: SA

address: Frome Road

department: Hanson Institute

country: Australia

institute: SA Pathology

summary:
This is a time course experiment with 4 - 6 slides per time point where all samples were hybridised to time zero.  The microarray slides were scanned using a GenePix 4000B Scanner (Axon Instruments, Foster City, CA, USA). The “SPOT” software package (CSIRO) was used to identify spots and subtract backgrounds. The extracted information was normalised using the “Statistics in Microarray Analysis" (SMA) open source R package.  Scaled print tip group lowes normalisation was performed for each slide and replicate slides were scaled to each other.
The cDNAs used in this microarray were identified as part of collaborative project between the IMVS and Bionomics Limited. The project aims were to identify genes up-regulated during in vitro capillary tube formation as targets for angiogenesis-based therapeutics.

Any requests for further information regarding the data generated from this collaboration should be directed to Bionomics Limited (www.bionomics.com.au) at the
following address:
Bionomics Limited
31 Dalgleish Street
Thebarton, South Australia
Australia, 5031
Phone: 618 8354 6104
Fax: 618 8354 6199
Email: busdev@bionomics.com.au

This SuperSeries is composed of the SubSeries listed below.

first: C

last: Hahn

first: J

middle: R

last: Gamble

first: M

last: Vadas

middle: N

last: Drogemuller

first: S

last: Waterman

first: A

first: G

last: Goodall


title: Analysis of murine liver tissue

status: Public on Jul 27 2004

city: Raleigh

first: Jack

last: Odle

department: Functional Genomics/Animal Science

institute: North Carolina State University

summary:
Functional genomic analysis of liver tissue extracted from polygenic obese mice fed either a control (1% linoleic acid) or treatment (1% trans 10, cis 12 conjugated linoleic acid) diet after fourteen days.
Keywords = Obesity
Keywords = conjugated linoleic acid
Keywords = polygenic obese mice
Keywords = lipid metabolism
Keywords: repeat sample

first: Ralph

last: House

first: Joseph

middle: P

last: Cassady

first: Eugene

last: Eisen

first: Thomas

last: Eling


title: Cancer Genome Anatomy Project (CGAP) SAGE Libraries

status: Public on Nov 29 2001

city: Bethesda

first: Cancer Genome Anatomy Project

last: CGAP

state: MD

department: Cancer Genome Anatomy Project

institute: National Cancer Institute

summary:
This series represents the Cancer Genome Anatomy Project SAGE library collection.  Libraries contained herein were either produced through CGAP funding, or donated to CGAP.
The Cancer Genome Anatomy Project (CGAP: http://cgap.nci.nih.gov) is an interdisciplinary program established and administered by the National Cancer Institute (NCI: http://www.nci.nih.gov) to generate the information and technological tools needed to decipher the molecular anatomy of the cancer cell.

SAGE libraries are named according to the following convention:
* SAGE_Organ_histology_code_unique identifier, e.g., SAGE_Colon_adenocarcinoma_CL_Caco2
* Codes: B = bulk; CL = cell line; CS = short-term cell culture; MD = micro-dissected; AP = antibody purified.

These libraries were downloaded from the CGAP Web site on March 17, 2009

This SuperSeries is composed of the SubSeries listed below.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

first: R

last: Strausberg

first: D

last: Gerhard

first: Gregory

last: Riggins


title: Analysis of murine epididymal adipose tissue

status: Public on Jul 23 2004

summary:
Functional genomic analysis of epididymal adipose tissue extracted from polygenic obese mice fed either a control (1% linoleic acid) or treatment (1% trans 10, cis 12 conjugated linoleic acid) diet after five or fourteen days.
Keywords = Obesity
Keywords = conjugated linoleic acid
Keywords = polygenic obese mice
Keywords = adiposity
Keywords = lipid metabolism
Keywords = apoptosis
Keywords: repeat sample

first: Jennifer

last: Collins

first: Sherry

middle: F

last: Grissom


title: Endothelial in vivo vs. in vitro responses to LPS

status: Public on Feb 26 2004

city: Dallas

first: John

middle: G

last: Maresh

address: 5323 Harry Hines

department: Internal Medicine

institute: U.T. Southwestern Medical Center

summary:
Endothelial responses to LPS using endothelial cells freshly isolated from Tie2 GFP animals vs. cultured cell line.

This SuperSeries is composed of the SubSeries listed below.


title: B cell response to CD40, Anti-IgM, ELC, IL4, terbutaline and lipopolysaccharide treatment

status: Public on Feb 16 2004

first: Sangdun

last: Choi

summary:
Temporal analysis (60, 180, 360 min) of B cells treated with either:
CD40
Anti-IgM
ELC
IL4
Lipopolysaccharide
Terbutaline
CD40 and IL4
CD40 and Lipopolysaccharide
CD40 and Anti-IgM
Anti-IgM and ELC
Anti-IgM and Terbutaline
ELC and Lipopolysaccharide

This SuperSeries is composed of the SubSeries listed below.

first: Yun-Anna

last: Cao

first: Mi-Sook

last: Chang

first: Sun-Young

last: Lee

first: Meghdad

last: Rahdar

first: Robert

last: Hsueh


title: Gliogenesis in Drosophila

status: Public on Feb 04 2003

first: Boris

last: Egger

state: BS

department: University of Basel

institute: Institute of Zoology

summary:
In Drosophila, the glial cells missing (gcm) gene encodes a transcription factor that controls the determination of glial versus neuronal fate. In gcm mutants, presumptive glial cells are transformed into neurons and, conversely, when gcm is ectopically misexpressed, presumptive neurons become glia. Although gcm is thought to initiate glial cell development through its action on downstream genes that execute the glial differentiation program, little is known about the identity of these genes. To identify gcm downstream genes in a comprehensive manner, genome-wide oligonucleotide arrays were used to analyze differential gene expression in wild-type embryos versus embryos in which gcm is misexpressed throughout the neuroectoderm.  Transcripts were analyzed at two defined temporal windows during embryogenesis.  This first genome-wide analysis of gene expression events downstream of a key developmental transcription factor presents a novel level of insight into the repertoire of genes that initiate and maintain cell fate choices in CNS development.
Keywords: other

first: B

last: Leemans

first: T

last: Loop

first: L

last: Kammermeier

first: Y

last: Fan

last: Radimerski

first: M

middle: C

last: Strahm

first: U

last: Certa

first: H

last: Reichert


title: Global transcriptional response to transient cell wall damage

city: Madrid

first: Javier

last: Arroyo

state: Madrid

address: Plaza Ramon y Cajal S/N

department: Microbiologia II

country: Spain

institute: Facultad de Farmacia (UCM)

summary:
Data processing included:
1. Filtering of those bad quality image spots (these includes alterations on the shape or hybridization)
2. Filtering of low intensity which was done flagging with -50 those spots whose feature intensity was lower in any of both channels than the average of the local background in the corresponding channel for all spots. However, those genes whose feature intensity in one channel was under the limit but in the other channel was 5 times over limit were considered as on-off genes and were not filtered.
3. Filtering of low reproducibility intrachip replicated spots. Normal distribution was created with the value obtained from the formula: log2(Rax1/Rb) which means logarithm (two-based) of one ratio multiplied by the inverse of its replicate. This value for each gene shoud keep between the range of average ± 3SD. Those genes out of these range are flagged as -25
4. Normalization of ratios was done by Lowess mathematical method. Correction factor was 0.33

This SuperSeries is composed of the SubSeries listed below.

first: Raúl

last: García

first: Clara

last: Bermejo

first: Cecilia

last: Grau

first: Rosa

last: Pérez

first: Jose

last: Rodríguez-Peña

first: Jean

last: Francois

first: César

last: Nombela