title: Transcriptome of soleus muscle of 129/SV mice

city: Berne 9

first: Martin

last: Flück

state: Bern

address: Bühlstrase 26

department: Department of Anatomy

country: Switzerland

institute: University of Berne

summary:
1 year-old male 129/SV mice.
Animals were acclimatized to housing in single cages for one week. Soleus muscle was harvested, total RNA isolated and subjected to transcript profiling with custom microarrays with custom-designed ATLASTM cDNA nylon filter as described (Dapp, C., Schmutz, S., Hoppeler, H. & Fluck, M. (2004) Physiol Genomics 20, 97-107).
Keywords: Transcriptome

overall_design: Seven replicas

last: Flueck


title: Transcriptome of soleus muscle of Tenascin-C deficient 129/SV mice

status: Public on Aug 08 2008

summary:
1 year-old male Tenascin-C deficient 129/SV mice with the targeted insertion of a beta-lactamase cassette in the NcoI site of exon 2 of the TNC gene (Forsberg, E., Hirsch, E., Frohlich, L., Meyer, M., Ekblom, P., Aszodi, A., Werner, S. & Fassler, R. (1996) Proc. Natl. Acad. Sci U. S. A 93, 6594-6599) were used for the study. Animals were derived from the original strain and back-crossed with wildtype 129/SV mice (Institut für Labortierkunde, University of Zurich).
Animals were acclimatized to housing in single cages for one week. Soleus muscle was harvested, total RNA isolated and subjected to transcript profiling with custom microarrays with custom-designed ATLASTM cDNA nylon filter as described (Dapp, C., Schmutz, S., Hoppeler, H. & Fluck, M. (2004) Physiol Genomics 20, 97-107).
Keywords: Transcriptome

overall_design: Seven biological replicas


title: Transcriptome of reloaded soleus muscle of Tenascin-C deficient 129/SV mice

summary:
1 year-old male Tenascin-C deficient 129/SV mice with the targeted insertion of a beta-lactamase cassette in the NcoI site of exon 2 of the TNC gene (Forsberg, E., Hirsch, E., Frohlich, L., Meyer, M., Ekblom, P., Aszodi, A., Werner, S. & Fassler, R. (1996) Proc. Natl. Acad. Sci U. S. A 93, 6594-6599) were used for the study. Animals were derived from the original strain and back-crossed with wildtype 129/SV mice (Institut für Labortierkunde, University of Zurich).
Animals were acclimatized to housing in single cages for one week followed by 7 days of hindlimb suspension and one day of reloading.
Soleus muscle was harvested, total RNA isolated and subjected to transcript profiling with custom microarrays with custom-designed ATLASTM cDNA nylon filter as described (Dapp, C., Schmutz, S., Hoppeler, H. & Fluck, M. (2004) Physiol Genomics 20, 97-107).

Keywords: Transcriptome

overall_design: Six biological replicas


title: Genomic location data for ectopic p73 in H1299 lung carcinoma cells

status: Public on Aug 06 2008

city: Nashville

first: Jennifer

last: Pietenpol

state: TN

address: 23rd Ave South at Pierce

department: Biochemistry

country: USA

institute: Vanderbilt University Medical Center

type: Genome binding/occupancy profiling by high throughput sequencing

summary:
The integral role of p53 in tumor suppression has promted many laboratories to perform extensive analyses of signaling pathways downstream of the p53 family of sequence-specific DNA binding transcription factors (p53 and its homologs p63 and p73).  Despite the ability of p73 to regulate many p53 family target genes, little is known about the specific pathways that modulate p73 during development, tumorigenesis and tumor therapy.  In this study we present a gene signature-based approach for connecting signaling pathways to transcription factors, as exemplified by p73.  We generated a p73 gene signature by integrating whole-genome chromatin immunoprecipitation and expression profiling.

Keywords: Transcription factor analysis

overall_design: H1299 lung carcinoma cells were transduced with TAp73beta or GFP expressing adenoviruses.  Microarray analysis (on the GFP and TAp73beta samples) and ChIPSeq analysis (on the TAp73beta sample) were performed to identify candidate p73 target genes.

middle: M

last: Rosenbluth

middle: A


title: Global proteomic characterization of photosystem II complexes from Synechocystis 6803

status: Public on Aug 18 2008

city: Saint Louis

first: Eric

middle: Agnew

last: Welsh

state: MO

address: One Brookings Drive; Campus Box 1137

department: Biology

institute: Washington University in Saint Louis

summary:
In this study, we undertook a global proteomics analysis of isolated PSII complexes, comparing protein profiles of HT3 (C-terminal hexahistidine tagged CP47 protein) to those of deltaPsbV HT3, deltaPsbP HT3, and deltaPsbQ HT3. The sensitivity of these techniques allowed for identification of not only stoichiometric components of active PSII complexes, but also for the identification of proteins that are transiently associated with PSII throughout its lifecycle, such as assembly, repair, or degradation partners. From the results, we identified an operon of unknown function which contains binding domains for photosynthetic cofactors and which we have named the cofactor integration operon  (cio). Upon deletion of the operon, photosynthetic capacity is decreased, indicating a function in PSII-mediated activity.
Keywords: proteomic, photosystem II, cyanobacteria

overall_design: Comparison of wild type (HT3) to 3 mutants (deltaPsbV HT3, delataPsbP HT3, and deltaPsbQ HT3)


title: Transcriptome of reloaded soleus muscle of 129/SV mice

summary:
1 year-old male 129/SV mice.
Animals were acclimatized to housing in single cages for one week followed by 7 days of hindlimb suspension and one day of reloading.
Soleus muscle was harvested, total RNA isolated and subjected to transcript profiling with custom microarrays with custom-designed ATLASTM cDNA nylon filter as described (Dapp, C., Schmutz, S., Hoppeler, H. & Fluck, M. (2004) Physiol Genomics 20, 97-107).
Keywords: transcriptome


title: Frequent switching of Polycomb repressive marks and DNA hypermethylation in the PC3 prostate cancer cell line

status: Public on Aug 07 2008

city: Rehovot

first: Amos

last: Tanay

address: The Weizmann Insititute

department: Computer Science and Applied Math

country: Israel

institute: The Weizmann Institute

type: Methylation profiling by genome tiling array

type: Genome binding/occupancy profiling by genome tiling array

summary:
Epigenetic reprogramming is commonly observed in cancer, and is hypothesized to involve multiple mechanisms, including DNA methylation and Polycomb repressive complexes (PRCs). Here we devise a new experimental and analytical strategy using customized high density
tiling arrays to investigate coordinated patterns of gene expression, DNA methylation and Polycomb marks which differentiate prostate cancer cells from their normal counterparts. Three major changes in the epigenomic landscape distinguish the two cell types. Developmentally significant genes containing CpG islands which are silenced by PRCs in the normal cells acquire DNA methylation silencing and lose their PRC marks (epigenetic switching). Since these genes are normally silent this switch does not cause de novo repression but might significantly reduce epigenetic plasticity. Two other groups of genes are silenced by either de novo DNA methylation without PRC occupancy (5mC reprogramming) or by de novo PRC occupancy without DNA methylation (PRC reprogramming). Our data suggest that the two silencing mechanisms act in parallel to reprogram the cancer epigenome, and that DNA hypermethylation may replace Polycomb based repression near key regulatory genes, possibly reducing their regulatory plasticity.

overall_design: Profiling 5mC and H3K27m3/Suz12 occupancy in the PC3 cancer line and the PrEC system. Hybridization over a custom array including a representative set of promoters (-2.5 to +1 kb) with variable CpG content levels and variable expression levels in the two cell systems.

first: Einav

last: Nili Gal-Yam

first: Gerda

last: Egger

first: Peter

last: Jones


title: Genomic profiling of microRNA and messenger RNA reveals deregulated microRNA expression in prostate cancer.

city: Bethesda

first: Robert

middle: Scott

last: Hudson

state: MD

address: National Cancer Institute Room Blgd.37/3044

department: NCI

institute: NIH

summary:
MicroRNAs are small non-coding RNAs that regulate mRNA function. Recent studies have shown that microRNA expression is altered in tumors. We studied the expression of both microRNAs and mRNAs in 60 primary prostate tumors and 16 non-tumor prostate tissues to evaluate the involvement of microRNAs in prostate cancer. Global microRNA expression was determined in RNA isolated from fresh-frozen human tissues with a custom oligonucleotide microarray chip. Expression analysis of mRNAs using Affymetrix gene chips revealed that Dicer, a key component of microRNA processing, and two microRNA host genes, MCM7 and C9orf5, were significantly up-regulated in prostate tumors. Consistent with the findings, tumors expressed at higher levels the miR-25 cluster (miR-25/miR-93/miR-106b), which maps to intron 13 of MCM7, and miR-32, which maps to intron 14 of C9orf5, than non-tumor prostate tissues. Other microRNAs that were overexpressed included miR-26a, miR-31, miR-182, miR-196a, and miR-200c, among others, and homologues of the miR-25 cluster, such as miR-92 and miR-106a. Among the down-regulated microRNAs in tumors were the miR-1/miR-133a cluster, miR-490, miR-494 and miR-520h. Differences in microRNA expression were also observed between high and low Gleason score and between tumors that either showed or did not show extraprostatic extension. A 37-probeset signature, representing 23 different mature microRNAs, correctly classified all non-tumor tissues and 80% of the tumors. In summary, our data indicate that alterations in microRNA expression occur in the development and progression of human prostate cancer. Such changes may prove useful in the development of novel diagnostic and prognostic markers.
Keywords: Marcodissected tissues

platform_organism:
Homo sapiens
Mus musculus

overall_design: Sixty fresh-frozen prostate tumors were obtained from the NCI Cooperative Prostate Cancer Tissue Resource (CPCTR) and the Department of Pathology at the University of Maryland (UMD). All tumors were resected adenocarcinomas that had not received any therapy prior to prostatectomy. The macro-dissected CPCTR tumor specimens were reviewed by a CPCTR-associated pathologist, who confirmed the presence of tumor in the frozen specimens. Surrounding non-tumor prostate tissue was collected from 16 patients with prostate cancer. All tissues were collected between 2002 and 2004. Information on race/ethnicity was either extracted from medical records (CPCTR) or obtained through an epidemiological questionnaire (UMD). Clinicopathological characteristics of the patients, including age at prostatectomy, histology, Gleason score, pathological stage, PSA at diagnosis, tumor size, extraprostatic extension, margin involvement, and seminal vesicle invasion were obtained from CPCTR.  For UMD cases, this information was extracted from the medical and pathology records, if available. The study was approved by the institutional review boards of the participating institutions.                                                                                Total RNA was isolated using the TRIZOL reagent according to the manufacturer’s instructions (Invitrogen, Carlsbad, CA). RNA integrity for each sample was confirmed with the Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA). Each RNA was then split into two pools that were either processed for the microRNA microarray or the mRNA microarray.

first: Stefan

last: Ambs

first: Ming

last: Yi


title: Analysis of HSF2 target genes using a global promoter ChIP-chip strategy on mouse wild-type testis

city: Turku

first: Lea

last: Sistonen

address: Tykistokatu 6 B

country: Finland

institute: Åbo Akademi University

type: Genome variation profiling by genome tiling array

summary:
Mapping of in vivo targets for HSF2 by using mammalian tissue material in a global ChIP-chip promoter screen. Chromatin was isolated from three wild-type (WT) mouse testes, crosslinked and sonicated into fragments of 100-500 bp, and the quality of the DNA was controlled before the immunoprecipitation, showing no signs of degradation.
The DNA amplified from the HSF2 immunoprecipitation samples was labeled and hybridized against the total input DNA samples, on a 1.5 kb promoter tiling array (NimbleGen Systems Inc.), covering ~26000 promoters of the mouse genome. After hybridization and scanning, HSF2 hybridization signals were Lowess normalized to produce log2 ratios  (HSF2 ChIP/Input DNA) for all  replicate arrays.
The promoter values were calculated as average log2 ratios over all probes for each promoter. To identify the HSF2 target population, we used a non-arbitrary analysis method called RankProd described in Breitling et al. 2004. RankProd has been published as an R/Bioconductor package.
Keywords: 1.5 kb mouse promoter ChIP-chip (MM5, NimbleGen Systems Inc.), mouse testis

overall_design: HSF2 ChIP sample vs. Input sample (reference),  biological replicates: 3, one replicate per array.

first: Malin

last: Åkerfelt

first: Eva

last: Henriksson

first: Asta

last: Laiho


title: Gene expression after transfection of IGF-II specific siRNAs

city: Mannheim

first: Xiaolei

last: Yu

address: Theodor-Kutzer-Ufer

department: ZMF

country: Germany

institute: University Heidelberg

summary:
Total RNA was isolated from HuH-7 cells after transfection of IGF-II specific siRNAs. Gene expression profiling was performed using the Affymetrix Human Genome U133A 2.0 Arrays. The raw data were analysed using mixed model ANOVA.
Keywords: transfection with siRNA

overall_design: Cells transfected with two independent siRNAs (ds1 and ds2) as well as untransfected cells were examined. For each condition biological duplicates were used for gene expression profiling


title: Effects of diet and age on canine cerebral cortex gene expression

status: Public on Aug 05 2008

city: Urbana

first: Kelly

last: Swanson

state: IL

address: 1207 W. Gregory Dr.

institute: University of Illinois

summary:
Mechanisms contributing to age-related cognitive decline are poorly defined.  Thus, we used canine microarrays to compare gene expression profiles of brain tissue from geriatric and young adult dogs.  Cerebral cortex samples were collected from 6 geriatric (12 yr-old) and 6 young adult (1 yr-old) female beagles after being fed one of two diets (animal protein-based versus plant-protein based) for 12 months.  RNA samples were hybridized to Affymetrix GeneChip Canine Genome Arrays.  Statistical analyses indicated that the age had the greatest impact on gene expression, with 963 transcripts differentially expressed in geriatric dogs.  Although not as robust as age, diet affected mRNA abundance of 140 transcripts.  As demonstrated in aged rodents and humans, geriatric dogs had increased expression of genes associated with inflammation, stress response, and calcium homeostasis and decreased expression of genes associated with neuropeptide signaling and synaptic transmission.  In addition to its existing strengths, availability of gene sequence information and commercial microarrays make the canine a powerful model for studying the effects of aging on cognitive function.

Keywords: age; diet

overall_design: Six geriatric (11.1 yr old) and 6 weanling (8 wk old) female beagles were used.  Three dogs of each age were assigned to one of two dietary treatments and fed for 12 months.  Diets tested in this experiment were previously shown to manipulate energy metabolism.  One diet was an animal-protein based diet (APB) and was composed primarily of highly digestible ingredients and animal-derived protein and fat sources (brewer’s rice, poultry by-product meal, poultry fat) and was formulated to contain 28% protein, 23% fat, and 5% dietary fiber.  The other diet was a plant-protein based diet (PPB) and was composed primarily of moderately digestible plant-derived ingredients (corn, soybean meal, wheat middlings, and meat and bone meal) and was formulated to contain 26% protein, 11% fat, and 15% dietary fiber.  Although the two diets were very different in terms of ingredient and chemical composition, both were formulated to meet or exceed all nutrient requirements for canine growth according to the Association of American Feed Control Officials.  Young dogs were fed ad libitum to allow for adequate growth, while geriatric dogs were fed to maintain baseline BW throughout the experiment.  To produce the desired metabolic effects, the PPB diet was formulated to contain a lower caloric density (APB = 5.38 kcal/g; PPB = 4.75 kcal/g) and have a lower nutrient digestibility than the APB diet.  Thus, dogs fed the PPB diet needed to consume a greater (P<0.05) quantity of food (237 g/d; 1123 kcal/d) than dogs fed the APB diet (166 g/d; 893 kcal/d) to grow (young) or maintain BW (geriatrics).  Even though metabolic indices were altered, mean BW among dietary treatments was not different at any time over the course of the study for young or geriatric dogs.  After 12 months on experiment, animals were fasted for 12 hr and then given a lethal dose (130 mg/kg BW) of sodium pentobarbital (Euthasol, Virbac Corp., Fort Worth, TX) intravenously into the left forearm.  Death was confirmed by lack of respiration and a corneal reflex, and absence of a heartbeat detected with a stethoscope placed under the left elbow.   Cerebral cortex samples were collected immediately after death was confirmed, flash frozen using liquid nitrogen, and stored at -80oC until further analysis.

middle: S

first: Lawrence

middle: B

last: Schook


title: Lifestyle of lactobacillus plantarum in the mouse cecum

status: Public on Aug 01 2008

city: Amsterdam

first: Douwe

last: Molenaar

address: De Boelelaan 1085

department: Systems Bioinformatics

country: Netherlands

institute: Vrije Universiteit Amsterdam

summary:
Lactobacillus plantarum is a common inhabitant of mammalian gastrointestinal tracts and specific strains belonging to this species are marketed as probiotics intended to confer beneficial health effects. To assist in determining the physiological status and host-microbe interactions of L. plantarum in the digestive tract we assessed changes in the transcriptome of L. plantarum WCFS1 during colonization of the cecum of germ-free mice. According to the transcript profiles L. plantarum WCFS1 was metabolically active and not under severe stress in this intestinal compartment. Carbohydrate metabolism was the most strongly affected functional gene category whereby many genes encoding diverse sugar transport and degradation pathways were induced in mice even compared to L. plantarum grown in a mouse chow-derived laboratory medium. This suggests that the ability of L. plantarum WCFS1 to consume diverse energy sources including plant-associated and host-derived carbohydrates was increased during its residence in the digestive tract. Many of these genes were also induced in L. plantarum colonizing germ-free mice fed a humanized Western-style diet. Similarly a core set of genes encoding cell surface-related properties were differentially expressed in mice. This set includes genes required for the D-alanylation and glycosylation of lipoteichoic acids that were strongly down-regulated in mice. In total L. plantarum exhibits a distinct in vivo transcriptome directed towards adaptation to the mouse intestinal environment.
Keywords: cell type comparison

overall_design: Six-week old germ-free C57 Black-6 male mice were inoculated with a single dose of 109 CFU of exponential-phase L. plantarum WCFS1 cells. The mice were sacrificed 15 days later, after sufficient time had passed for several turnovers of the intestinal epithelium and its overlying mucosal layer. Four mice were fed on Chow diet and two mice were fed on western style diet. RNA was isolated from the cecum of these mice. The transcriptome of L. plantarum in these mice was compared to that of L. plantarum grown on MRS broth, Chow broth, or on chemically defined media with either glucose or lactose as carbon- and energy source.

first: Maria

middle: L

last: Marco

first: Theodorus

middle: H

last: Peters

first: Roger

last: Bongers

first: Saskia

last: vanHemert

first: Jeffrey

middle: I

last: Gordon

first: Michiel

last: Kleerebezem


title: Global transcriptional profiling of Saccharomyces cerevisiae treated with thymol

city: Changchun

first: Xuming

last: Deng

address: Xi'an Street 5333#

country: China

institute: College of Animal Science and Veterinary Medicine, Jilin University

summary:
We employed CapitalBio Corporation to investigate the global transcriptional profiling of Saccharomyces cerevisiae treated with thymol.

Keywords: gene expression array-based, count

overall_design: S. cerevisiae strain L1190 was incubated in a rotary shaker. Subsequently, thymol was added to the culture and DMSO to the control. Then, the yeast cells were further incubated for a certain time and total RNA was extracted by a hot acidic phenol method. The experimental sample and control sample were incorporated and dissolved in hybridization solution after labelled. Arrays hybridization was preformed in a CapitalBio BioMixerTM II Hybridization Station overnight and washed with two consecutive solutions. Arrays were scanned with a confocal LuxScanTM scanner and the images obtained were then analyzed using LuxScanTM 3.0 software. A space- and intensity-dependent normalization based on a LOWESS program was employed.


title: Evaluation of genes involved in watermelon fruit ripening and development

city: Charleston

first: Patrick

last: Wechter

state: SC

address: 2700 Savannah Highway

department: U.S. Vegetable Laboratory

institute: USDA-ARS

summary:
A microarray and quantitative Real-Time PCR-based study was conducted in watermelon [Citrullus lanatus (Thunb.) Matsum. & Nakai var. lanatus] in order to elucidate the flow of events associated with fruit development and ripening in this species.  RNA from three different maturation stages of watermelon fruit, as well as leaf, were collected from field grown plants during three consecutive years, and hybridized to high-density, photolithography microarrays.
Keywords: developmental time course, gene expression

overall_design: This experiment contained a single biological replicate, two tissue types (leaf, fruit flesh), and three time points (12 days post-pollination, 24 days post-pollination, and 36 days post-pollination.  One hundred and twenty-seven genes were chosen from this experiments and used in conjunction with quantitative-PCR to examine two additional biological replications of the experiment.

first: W

middle: P

first: Amnon

last: Levi

first: Karen

middle: R

last: Harris

first: Angela

last: Davis

first: Zangjun

last: Fei

first: Nurit

last: Katzir

first: James

middle: J

last: Giovannoni

first: Ayelet

last: Salman-minkov

first: Alvaro

last: Hernandez

first: Jyothi

last: Thimmapuram

first: Yaakov

last: Tadmor

first: Vitaly

last: Portnoy

first: Tova

last: Trebitsh


title: Comparative expression analysis between SdrP (TTHA1359) deletion mutant and wild-type of T. thermophilus HB8

status: Public on Aug 03 2008

city: Hyogo

first: Akeo

last: Shinkai

address: 1-1-1, Kouto, Sayo

department: SPring-8 Center

country: Japan

institute: RIKEN Harima Institute

summary:
We observed the expression profile of the total mRNA of TTHA1359 deletion mutant of Thermus thermophilus HB8 strain grown in a rich (TR) medium at 70°C compared with that of wild type.
Keywords: cell type comparison

overall_design: Four wild-type and mutant T. thermophilus HB8 strains were each pre-cultured at 70°C for 16 h in 3 ml of TR medium containing 0.8% polypeptone, 0.4% yeast extract, 0.2% NaCl, 0.4 mM CaCl2, and 0.4 mM MgCl2, which was adjusted to pH 7.2 with NaOH. The cells (2 ml) were inoculated into 1 liter of the same medium and then cultivated at 70°C. Cells were collected after 680 min, and then crude RNA was extracted. In order to compare mRNA expression in the TTHA1359 deletion mutant with that of wild type, the expression of each mRNA was analyzed on a GeneChip as described under Sample Description Sheet of each sample in this website.

first: Yoshihiro

last: Agari

first: Aiko

last: Kashihara

first: Emi

last: Nakai

first: Noriko

last: Nakagawa

first: Sigeyuki

last: Yokoyama

first: Seiki

last: Kuramitsu


title: Iron regulation in Vibrio vulnificus CMCP6

city: Portland

first: Jorge

last: Crosa

state: OR

address: 3181 SW Sam Jackson Park Road

department: Molecular Microbiology and Immunology

institute: Oregon Health and Science University

summary:
Vibrio vulnificus multiply rapidly in host tissues under iron overloaded conditions. To understand the effects of iron in the physiology of this pathogen we performed a genome-wide transcriptional analysis of this bacterium growing under three different iron concentrations. V.vulnificus CMCP6 cells were grown under three different iron concentrations (TSBS + EDDA 50uM, TSBS and TSBS + FAC 250 ug/ml) and samples taken at log phase.
Keywords: Response to the iron concentration of the media

overall_design: Strains were grown to an OD600nm of 0.6 to 0.8 in TSBS, TSBS with the addition of 250 μg/ml FAC or TSBS with the addition of 50 μM EDDA. Three independent cultures of the V. vulnificus cells grown in each media, were combined and treated as a single sample for the RNA extraction to minimize culture variation. Two samples per condition were used for the microarray analysis. Cells were centrifuged and the pellets resuspended in RNAWiz reagent (Ambion®, Austin, TX). Total RNA was extracted from each strain according to the manufacturer’s instructions

first: Alejandro

middle: F

last: Alice

first: Hiroaki

last: Naka


title: Largemouth Bass E2 exposure

city: Ocean SPrings

first: Joe

last: Griffitt

state: MS

address: 703 East BEach Drive

department: Coastal Sciences

institute: University of Southern Mississippi

summary:
A novel custom microarray for largemouth bass (Micropterus salmoides) was designed from sequences obtained from a normalized cDNA library using the 454 Life Sciences GS-20 pyrosequencer.  The GS-20 yielded in excess of 58 million bases of high-quality sequence.  The sequence information was combined with 2,616 reads obtained by traditional suppressive subtractive hybridizations to derive a total of 31,391 unique sequences. Annotation and coding sequences were predicted for these transcripts where possible. 16,350 annotated transcripts were selected as target sequences for the design of the custom largemouth bass oligonucleotide microarray.  The microarray was validated by examining the transcriptomic response in male largemouth bass exposed to 17 -oestradiol.  Transcriptomic responses were assessed in liver and gonad, and indicated gene expression profiles typical of exposure to oestradiol.  The results demonstrate the potential to rapidly create the tools necessary to assess large scale transcriptional responses in non-model species, paving the way for expanded impact of toxicogenomics in ecotoxicology.
Keywords: E2 exposure, array validation

overall_design: This experiment tested two organs - liver and gonad from either E2-exposed or control fish, 4 fish (biological replicates) per treament (control and E2).

first: Natalia

last: Garcia-Reyero

first: William

middle: G

last: Farmerie

first: Robert

first: David

last: Barber

first: Li

last: Liu

first: Nancy

middle: D

last: Denslow

first: Kevin

last: Kroll


title: Transcriptional analysis of Leishmania major methotrexate resistant strains using full-genome DNA microarrays

city: Québec

first: Marc

last: Ouellette

state: Québec

address: 2705 bd Laurier

department: Département médecine

country: Canada

institute: Université Laval

summary:
Transcriptional analysis of the drug resistant strain L. major MTX60.4 compared to the wild-type strain LV39
The full-genome DNA microarrays includes one 70-oligonucleotides probe for each gene of L.major LV39
Keywords: drug resistance

platform_organism:
Leishmania major
Leishmania infantum

overall_design: Two-condition experiment, resistant strain MTX60.4 vs wild-type LV39. Four biological replicates for each strain, independently grown and harvested. One replicate per array

first: J

last: Ubeda

first: D

last: Légaré

first: F

last: Raymond

first: A

last: Ouameur

first: S

last: Boisvert

first: P

last: Rigault

last: Corbeil

first: M

last: Tremblay

last: Olivier

first: B

last: Papadopoulou


title: Global gene transcription analysis of Anaplasma phagocytophilum in tick and human cells using tiling microarrays

status: Public on Aug 04 2008

city: St. Paull

first: Michael

last: Herron

state: MN

address: 1980 Folwell Avenue, Room 219 Hodson Hall

department: Entomology

institute: Michael Herron

type: Expression profiling by genome tiling array

summary:
A whole genome tiling microarray based on the published sequence of the Anaplasma phagocytophilum (Ap) strain Hz genome was used to measure transcriptional activity of Ap grown to late stage in three cell lines representing the diversity of host cells naturally infected by Ap: two human cell lines, HL-60 (promyelocytic leukocyte) and HMEC-1 (microvascular endothelial), and the tick (Ixodes scapularis) cell line ISE6.

Keywords: cell type comparison

sample_organism:
Ixodes scapularis
Homo sapiens
Anaplasma phagocytophilum str. HZ

overall_design:
Total RNA was isolated from Ap (HZ strain) infected cells, used to make biotinylated cDNA fragments, and hybridized to Ap whole genome tiling arrays to measure relative abundance of mRNA transcripts.
.CEL files generated by the University of Minnesota’s microarray facility were joined to Affymetrix BPMAP files specific to the tiling array using Affymetrix® Tiling Analysis Software (TAS).  TAS generated a list of signal intensities and arranged them in order of genomic location and DNA strand. The intensity values were normalized via quantiles, and imported into the IgorPro data analysis program (WaveMetrics Lake Oswego OR, USA) along with the ORF and structural RNA annotations available from NCBI (http://www.ncbi.nlm.nih.gov). A script was written to index a trapezoidal integration algorithm of the intensity list with the start and end genomic positions indicated on the annotation. This script operation generated a list of 1411 “areas under the curve” (see Supplementary file at the foot of this record).

first: Curtis

last: Nelson

first: Roderick

last: Felsheim

first: Timothy

last: Kurtti

first: Ulrike

last: Munderloh


title: Comparative transcriptomics in Yersinia pestis: a global view of environmental modulation of gene expression

city: Beijing

first: Ruifu

last: Yang

address: No.20, DOngdajie, Fengtai

institute: Institute of Microbiology and Epidemiology

summary:
Background: Environmental modulation of gene expression in Yersinia pestis is critical for its life style and pathogenesis. Using cDNA microarray technology, we have analyzed the global gene expression of this deadly pathogen when grown under different stress conditions in vitro.
Results: To provide us with a comprehensive view of environmental modulation of global gene expression in Y. pestis, we have analyzed the gene expression profiles of 25 different stress conditions. Almost all known virulence genes of Y. pestis were differentially regulated under multiple environmental perturbations. Clustering enabled us to functionally classify co-expressed genes, including some uncharacterized genes. Collections of operons were predicted from the microarray data, and some of these were confirmed by reverse-transcription polymerase chain reaction (RT-PCR). Several regulatory DNA motifs, probably recognized by the regulatory protein Fur, PurR, or Fnr, were predicted from the clustered genes, and a Fur binding site in the corresponding promoter regions was verified by electrophoretic mobility shift assay (EMSA).
Conclusion: The comparative transcriptomics analysis we present here not only benefits our understanding of the molecular determinants of pathogenesis and cellular regulatory circuits in Y. pestis, it also serves as a basis for integrating increasing volumes of microarray data using existing methods.
Keywords: stress responses

overall_design:
The genome-wide transcriptional changes upon the environmental perturbations were monitored by the standard dual-fluorescent hybridization method with a Y. pestis whole-genome DNA microarray spotted with PCR-amplified ORFs. For the stimulon analysis, the gene expression pattern of WT strain grown under a stimulating condition was compared with that of an unperturbed control. For the regulon analysis, we compared the expression profiles of deletion mutant strains of the target regulator to Yersinia pestis strain 201 (wild-type).
For microarray hybridization, two independent bacterial cultures for each growth condition were prepared as biological replicates for RNA isolation. Three separate labeled probes were made for each RNA preparation as technical replicates. Pairwise comparisons were made using dye swaps to avoid labeling bias.
For microarray hybridization, two or three independent bacterial cultures for each growth condition were prepared as biological replicates for RNA isolation. Three or four separate labeled probes were made for each RNA preparation as technical replicates. Pairwise comparisons were made using dye swaps to avoid labeling bias.


title: Gene expression induced in Escherichia coli O157:H7 upon exposure to model apple juice

city: East Lansing

first: Thomas

last: Whittam

state: MI

address: 165 Food Safety & Tox

department: Microbiology

institute: Michigan State University

summary:
Escherichia coli O157:H7 has caused serious outbreaks of foodborne illness via transmission in a variety of food vehicles, including unpasteurized apple juice, dried salami, and spinach.  To understand how this pathogen responds to the multiple stresses of the food environment, we compared global transcription patterns after exposure to apple juice.  Transcriptomes of mid-exponential and stationary phase cells were evaluated after 10 minutes in model apple juice (pH3.5) using microarrays probing 4,886 ORFs.  Significant changes in gene expression were determined using R/MAANOVA and the Fs test. A total of 331 ORFs were significantly induced upon exposure of cells to model apple juice and included genes involved in the acid and osmotic stress responses as well as the oxidative stress response and envelope stress.  Genes involved in the acid and osmotic stress responses, including asr, osmC, osmB, and osmY were significantly induced in response to model apple juice.   Genes involved in the envelope stress response, known to be controlled by CpxR (cpxP, degP, and htpX), were significantly induced 2 to 15 fold upon exposure to apple juice, independent of growth phase.  Inactivation of CpxRA resulted in a significant decrease in survival of O157:H7 in model apple juice compared to the isogenic parent strain.  Of the 331 ORFs induced in model apple juice, 104 are O157-specific ORFs, including those encoding type three secretion effectors espJ, espB, espM2, espL3 and espZ.  By elucidating the response of O157:H7 to acidic foods, we hope to gain insights into how this pathogen is able to survive in food matrices and how exposure to foods affects subsequent transmission and virulence.

Keywords: stress

overall_design: The results are based on O157:H7 Sakai exponential and stationary phase cultures grown in MOPS minimal medium and then exposed to model apple juice (pH 3.5, 37C) for 10 minutes.  Differences in transcript levels were determined using a mixed model ANOVA in R/MAANOVA which tested for significant differences due to growth phase (exponential or stationary), treatment (MOPS or MAJ) and the interaction of these two factors using the following linear model: array+dye+sample (biological replicate)+ phase+treatment+phase*treatment.   We incorporated the dye-swaps among the biological replicates.

first: Teresa

last: Bergholz

first: Sivapriya

last: Kailasan Vanaja


title: Comparative expression analysis between logarithmic and stationary phase of T. thermophilus HB8 on a synthetic medium

first: Akeo

last: Shinkai

summary: We observed the expression profile of the total mRNA of Thermus thermophilus HB8 wild-type strain grown in a synthetic medium at 70°C.

overall_design: Three wild-type T. thermophilus HB8 strains were pre-cultured at 70°C for 16 h in 3 ml of synthetic medium. The cells (2 ml) were inoculated into 500 ml of the same medium and then cultivated at 70°C. Cells were collected after 10 h and 20 h, and then crude RNA was extracted. The expression of each mRNA at each time point was analyzed on a GeneChip as described under Sample Description Sheet of each sample in this website.

first: Yoshihiro

last: Agari

first: Miwa

last: Ohmori

first: Seiki

last: Kuramitsu


title: Pig SISP jejunal segments: Lactobacillus plantarum and msa/srt mutant strains vs control, perfusion 4 or 8 hours

city: Lelystad

first: Gabriele

last: Gross

address: P.O. Box 65

institute: Animal Sciences Group of Wageningen UR

summary:
Mannose-specific interactions of Lactobacillus plantarum 299v with jejunal epithelium were investigated using an in situ pig small intestinal segment perfusion (SISP) model. L. plantarum 299v wildtype strain was compared to two isogenic mutant strains either lacking the gene encoding for the mannose-specific adhesin (msa) or sortase (srtA; responsible for anchoring of cell surface proteins like Msa to the cell wall). Salmonella typhimurium served as a positive control for gene expression analysis. Scrapings from jejunal segments were collected after perfusion with bacterial suspensions or PBS (control) for 4 or 8 hours, and host gene expression was assessed using a home-made cDNA porcine microarray.
Keywords: host-microbe interaction, Lactobacillus plantarum, mannose-specific adhesion

overall_design: A Small Intestinal Segment Perfusion (SISP) test was performed using 4 pigs. 10 segments were prepared in the jejunum of each pig and perfused with Lactobacillus plantarum 299v wildtype, Lactobacillus plantarum 299v msa mutant strain, Lactobacillus plantarum 299v srtA mutant strain, Salmonella typhimurium or PBS (control) for 4 or 8 hours. Pooled samples from each treatment at each timepoint were used for microarray analysis. 8 comparisons were done: L. plantarum wildtype vs control (4 hours), L. plantarum wildtype vs control (8 hours), L. plantarum msa mutant vs control (4 hours), L. plantarum msa mutant vs control (8 hours), L. plantarum srt mutant vs control (4 hours), L. plantarum srt mutant vs control (8 hours), S. typhimurium vs control (8 hours), samples taken at the beginning of the experiment vs control (8 hours). Dye-swaps were performed for each comparison.

first: Jan

last: van der Meulen

first: Johannes

last: Snel

first: Roelof

last: van der Meer

first: Theo

last: Niewold

first: Marcel

last: Hulst

first: Mari

last: Smits


title: LNCap cells treated with 3-diol; No_Tmt and 1, 3 and 6hrs after treatment

city: Oklahoma City

first: Hsueh-Kung

last: Lin

state: OK

address: Room 462, 800 Research Parkway

department: Urology

institute: University of Oklahoma Health Sciences Center

summary:
5α-androstane-3α,17β-diol (3α-diol) is reduced from the potent androgen, 5α-dihydrotestosterone (5α-DHT), by reductive 3α-hydroxysteroid dehydrogenases (3α-HSDs).  3α-diol is believed to be a weak androgen, and has to be oxidized to 5α-DHT before it can exert its androgenic activity on androgen target tissues including the prostate.  However, we repeatedly demonstrated that 3α-diol can be a potent androgen, activates cytoplasmic signaling pathway, and may be responsible for androgen-independent prostate cancer growth.  A cancer-specific, cDNA-based membrane array was used to determine 3α-diol-mediated gene expressions in prostate cancer progression.  Several canonical pathways appeared to be affected by 3α-diol-regulated signaling in LNCaP cells; among them are apoptosis signaling, PI3K/Akt signaling, and death receptor signaling pathways.  Biological analysis of 3α-diol-activated signaling confirmed that 3α-diol augmented PI3K/Akt activation can be independent from the classical androgen receptor (AR) signaling.  These observations sustained our previous report that 3α-diol continues to supported prostate cell survival and proliferation regardless the status of the AR.  We provided the first global analysis of 3α-diol-activated gene expressions and identified cytoplasmic signaling pathways as important components of this response in human prostate cells.  3α-diol may play, therefore, a significant role for transition from androgen-dependent to androgen-independent prostate cancer progression in the presence of androgen blockade.
Keywords: array, time course, androgen, prostate, cancer

overall_design:
Human prostate cancer cell line LNCaP were maintained in the complete medium consisting of RPMI 1640 supplemented with 10% FBS, 5 units/ml penicillin, and 5 µg/ml streptomycin at 37C and 5% CO2.  Cells were passaged every 3-5 days or whenever cells reach 70-80% confluence.  To perform androgen stimulation followed by total RNA and protein extraction, 1 x 106 LNCaP cells were first seeded in 60 mm tissue culture plates, subjected to serum starvation by culturing the cells with OPTI-MEM supplemented with 1% CD FBS for 24 hours followed by 3α-diol stimulation.

To determine the involvement of AR in 3α-diol regulated cytoplasmic signaling, AR was suppressed by transfecting LNCaP cells with the plasmid vector-based siRNA constructs.  Briefly, 2×106 LNCaP cells were first mixed with 3 µg AR specific or control siRNA plasmid in 100 µl of Nucleofector solution R; and transfection was accomplished using the Nucleofector transfection device (Amaxa Biosystems; Gaithersburg, MD).  Cells were then returned to pre-warmed completed medium for 24 hours followed by serum deprivation for another 24 hours and 3α-diol stimulation.

To identify transcription profiling in LNCaP cells following 3α-diol stimulation, cDNA-based membrane arrays were used.  There were a total of 1,186 genes present in the Atlas Human Cancer 1.2 Expression Array. Following the washing steps, array membranes were exposed to phosphor-imaging screens (Packard BioScience; Meriden, CT); and images were captured by a Cyclone storage phosphorimager system (Packard BioScience).  Results from the phosphor-imaging system were presented as digital light units (DLU) and interpreted using OptiQuant image analysis software (Packard BioScience).

The background of the array membranes was determined by the average and standard deviation (SD) of all 276 background spots for each array consisting of spots between probes areas and spots on the rims indicated in the array membranes.  Total system noise around the true zero point was normally distributed with right-tail positively skewed.  Points exceeding 3 SD above the mean were eliminated until the true normal distribution was successively observed; usually this procedure involved the removal of no more than 5-10 points.  The mean of the remaining background measurements was subtracted from all expression measurements.  Genes whose expression levels exceeded above 2 SD of the background levels were considered to be regulated genes.  In contrast, genes whose expression levels did not exceed the 2 SD thresholds were marked as below noise threshold (BNT).  3α-diol-regulated genes were identified if their expression levels changed from BNT to exceed the 2 SD thresholds (off-on genes) or the opposite (on-off genes) following 3α-diol stimulation.

The collective responsive genes were used to identify underlying pathways regulated by 3α-diol in prostate cells.  The responsive genes were analyzed as one set by Ingenuity Pathway Analysis, a web-based bioinformatics application, to identify canonical pathways over-represented in the data set.  The identified pathways were then evaluated against the full expression data to identify each gene’s potential participation in 3α-diol-regulated cellular responses.  The responsive genes were not ranked by expression levels.
Human prostate cancer cell line LNCaP was obtained from ATCC (CRL-1740; Manassas, VA).  3α-diol and mouse anti-β-actin antibody were purchased from Sigma-Aldrich (St. Louis, MO).  [α-32P]dATP (3,000 Ci/mmol, 10 µCi/µl) was acquired from Amersham Pharmacia (Piscataway, NJ).  RPMI 1640 medium, OPTI-MEM, penicillin-streptomycin, fetal bovine serum (FBS), and Lipofectamine 2000 were purchased from Invitrogen (Grand Island, NY).  Charcoal-dextran treated (CD) FBS was obtained from HyClone (Logan, UT).  Atlas nylon Human Cancer 1.2 cDNA Expression Array was obtained from Clontech Laboratories (Palo Alto, CA).  Rabbit anti-phospho Akt Ser(473) antibody was purchased from Cell Signaling Technology (Danvers, MA).

For Western blot analysis total cellular proteins were harvested with RIPA buffer supplemented with proteinase inhibitors.  Protein concentration was determined by bicinchoninic acid (BCA) protein assay kit (Pierce; Rockford, IL).  Total proteins separation and transfer were also performed.  Phospho-Akt and b-actin expressions were detected by incubating protein membranes with appropriate primary antibodies against these molecules followed by peroxidase-conjugated secondary antibodies incubation.  Immunoreactive proteins were detected using the enhanced chemiluminescent (ECL) reagent (Pierce).

first: Mikhail

last: Dozmorov

first: Qing

first: Adam

last: Matwalli

first: Robert

middle: E

last: Hurst

first: Daniel

last: Culkin

first: Bradley

last: Kropp


title: Pair wise comparison between STD-MO and klf4-MO embryos

city: St Lucia

first: Darrin

last: Taylor

state: Qld

address: Research Rd

department: Genomics and Bioinformatics

country: Australia

institute: Institute for Molecular Bioscience, Uni of Qld

summary:
Direct pairwise comparison of the differential expression patterns between embryos injected with Standard olignucleotide morpholinoes and those injected with an anti-klf4 morpholino.
Keywords: Direct pairwise comparison

overall_design: Each "arm" of the comparison is a pairwise comparison containing a dye swap to eliminate dye bias.


title: Time course of the mRNA expression in wild-type Thermus thermophilus HB8 strain grown on a rich medium.

first: Akeo

last: Shinkai

summary:
We observed the expression profile of the total mRNA of wild-type Thermus thermophilus HB8 strain grown in a rich (TR) medium at 70°C.
Keywords: time course

overall_design: Three wild-type T. thermophilus HB8 strains were pre-cultured at 70°C for 16 h in 3 ml of TR medium containing 0.8% polypeptone, 0.4% yeast extract, 0.2% NaCl, 0.4 mM CaCl2, and 0.4 mM MgCl2, which was adjusted to pH 7.2 with NaOH. The cells (2 ml) were inoculated into 1 liter of the same medium and then cultivated at 70°C. Cells were collected after 180, 240, 300, 360, 420, 480, 540, 600, 680, and 760 min, and then crude RNA was extracted. The expression of each mRNA at each time point was analyzed on a GeneChip as described under Sample Description Sheet of each sample in this website.

first: Aiko

last: Kashihara

first: Emi

last: Nakai

first: Noriko

last: Nakagawa

first: Yoshihiro

last: Agari

first: Sigeyuki

last: Yokoyama


title: Transcription profile of an azole resistant mutant of Mycobacterium truberculosis

status: Public on Jul 31 2008

city: Padua

first: Roberta

last: Provvedi

address: Via Gabelli 63

department: Biologia

country: Italy

institute: University of Padua

summary: We used DNA microarrays to detect differentially expresses genes in a spontaneous mutant of Mycobacterium tuberculosis H37Rv resistant to azoles. We identified three adjacent upregulated genes encoding a transcription regulatory protein and two membrane proteins belonging to the family of RND efflux transporters.

overall_design: Hybridization were performed using RNA extracted from three different biological samples coming from cultures grown in mid-exponential phase.

first: Riccardo

last: Manganelli

first: Giovanna

last: Riccardi


title: Structure and Function of a Transcriptional Network Activated by Hog1

status: Public on Jul 29 2008

city: Cambridge

first: Andrew

last: Capaldi

state: MA

address: Bauer 307, 7 Divinity Ave

department: MCB

institute: Harvard University

summary:
Cells regulate gene expression using a complex network of signaling pathways, transcription factors and promoters.  To gain insight into the structure and function of these networks we analyzed gene expression in single and multiple mutant strains to build a quantitative model of the Hog1 MAPK-dependent osmotic stress response in budding yeast. Our model reveals that the Hog1 and general stress (Msn2/4) pathways interact, at both the signaling and promoter level, to integrate information and create a context-dependent response.

Keywords: Stress response network analysis using genetically modified cells

overall_design: 85 samples were used to dissect the structure and function of the Hog1 network (Critical Samples measured in triplicate).  The overall strategy was to double mutant or epistasis analysis to break down the influence that genes in the Hog1 network have on each other and the genome-wide stress response. This was done by comparing the expression in strains with different combinations of genes deleted and fitting the data to quantitative models.  See Capaldi et. al. Nature Genetics 2008 for details.

first: Tommy

last: Kaplan

first: Ying

last: Liu

first: Naomi

last: Habib

first: Aviv

last: Regev

first: Nir

last: Friedman

first: Erin

middle: K

last: O'Shea


title: Genome-associated RNA Polymerase II includes the dissociable Rpb4/7 subcomplex

status: Public on Jul 30 2008

city: Munich

first: Anna

middle: J.

last: Jasiak

address: Feodor-Lynen-Str. 25

department: Department of Chemistry and Biochemistry

institute: Gene Center, Ludwig-Maximilians-University of Munich

summary:
Yeast RNA polymerase (Pol) II consists of a ten-subunit core enzyme and the Rpb4/7 subcomplex, which is dispensable for catalytic activity and dissociates in vitro. To investigate whether Rpb4/7 is an integral part of DNA-associated Pol II in vivo, we used chromatin immuno-precipitation coupled to high-resolution tiling microarray analysis. We show that the genome-wide occupancy profiles for Rpb7 and the core subunit Rpb3 are essentially identical. Thus, the complete Pol II associates with DNA in vivo, consistent with functional roles of Rpb4/7 throughout the transcription cycle.

Keywords: ChIP-chip

overall_design: Comparison of Rpb3 vs. Rpb4/7 genome-wide occupancy profiles. Data obtained from independent ChIP-chip experiments on two yeast strains: S288C Rpb3TAP vs. Rpb7-TAP strain and W303 Rpb3-TAP vs. wild type strain (IP with the monoclonal antibody for Rpb4/7). Biological replicates W303 strains: 1 x Rpb3-TAP, 1 x wt with Rpb4/7 antibody, independently grown and harvested. Biological replicates S288C strains: 3 x Rpb3-TAP, 2 x Rpb7-TAP, independently grown and harvested. One replicate per array.

first: Holger

last: Hartmann

first: Elena

last: Karakasili

first: Marian

last: Klaocsay

last: Flatley

first: Elisabeth

last: Kremmer

first: Katja

last: Sträßer

first: Dietmar

last: Martin

first: Johannes

last: Söding

first: Patrick

last: Cramer


title: Inflammatory markers in ABCG1-/- lungs

city: Los Angeles

first: Angel

last: Baldan

state: CA

address: 675 Charles Young Dr.

institute: UCLA

summary:
Transcriptional profiling of cytokines and its receptors in lungs from 8 month old chow-fed wild-type and ABCG1-/- mice
Keywords: genotype comparison; inflammatory mediators

overall_design: 4 individual mice per group; RNA from inferior left lobe; WT vs. ABCG1-/-

first: Peter

middle: A

last: Edwards


title: Transcriptional Response of Candida albicans Cells Following Phagocytosis by Mouse Macrophages

status: Public on Jul 28 2008

city: Montreal

first: Andre

last: Nantel

state: QC

address: 6100 Royalmount

department: Biotechnology Research Institute

institute: National Research Council of Canada

summary:
Transcriptional profiling of Candida albicans that had been phagocytosed for 1, 2, or 4 hours by either bone marrow-derived mouse macrophaged (BMDM) or immortalized RAW264.7 cells
Keywords: Time course

overall_design: Two-condition experiment, phagocytosed cells vs cells maintained in D-10 media. Time course: 1, 2, and 4 hours. Biological replicates: 4 independent experiments at each cell-time combinations.

first: Anne

last: Marcil

first: Christine

last: Gadoury

first: Josee

last: Ash

first: Jianchun

last: Zhang

first: Malcolm

last: Whiteway


title: p53 status and AAI gene expression response

city: Sutton

first: Ian

last: Giddings

state: Surrey

address: 15 Cotswold Road

department: Section of Molecular Carcinogenesis

country: United Kingdom

institute: Institute of Cancer Research

summary:
Human colon carcinoma cells (HCT116) differing in TP53 status were exposed to aristolochic acid I (AAI) (50 and 100 uM for up to 48 h), and their gene expression responses compared by cDNA microarray technology.
Keywords: other

overall_design: 36 samples  - cells exposed to AAI dissolved in water were compared to cells treated to water alone


title: Experimental analysis of oligonucleotide microarray design criteria to detect deletions by CGH

status: Public on Jul 26 2008

city: Vancouver

first: Stephane

last: Flibotte

state: BC

address: 2370-6270 University Blvd.

department: Faculty of Science

institute: University of British Columbia

summary:
BACKGROUND:
Microarray comparative genomic hybridization (CGH) is currently one of the most powerful techniques to measure DNA copy number in large genomes. In humans, microarray CGH is widely used to assess copy number variants in healthy individuals and copy number aberrations associated with various diseases, syndromes and disease susceptibility. In model organisms such as Caenorhabditis elegans (C. elegans) the technique has been applied to detect mutations, primarily deletions, in strains of interest. Although various constraints on oligonucleotide properties have been suggested to minimize non-specific hybridization and improve the data quality, there have been few experimental validations for CGH experiments. For genomic regions where strict design filters would limit the coverage it would also be useful to quantify the expected loss in data quality associated with relaxed design criteria.
RESULTS:
We have quantified the effects of filtering various oligonucleotide properties by measuring the resolving power for detecting deletions in the human and C. elegans genomes using NimbleGen microarrays. Approximately twice as many oligonucleotides are typically required to be affected by a deletion in human DNA samples in order to achieve the same statistical confidence as one would observe for a deletion in C. elegans. Surprisingly, the ability to detect deletions strongly depends on the oligonucleotide 15-mer count, which is defined as the sum of the genomic frequency of all the constituent 15-mers within the oligonucleotide. A similarity level above 80% to non-target sequences over the length of the probe produces significant cross-hybridization. We recommend the use of a fairly large melting temperature window of up to 10 C, the elimination of repeat sequences, the elimination of homopolymers longer than 5 nucleotides, and a threshold of -1 kcal/mol on the oligonucleotide self-folding energy. We observed very little difference in data quality when varying the oligonucleotide length between 50 and 70, and even when using an isothermal design strategy.
CONCLUSIONS:
We have determined experimentally the effects of varying several key oligonucleotide microarray design criteria for detection of deletions in C. elegans and humans with NimbleGen's CGH technology. Our oligonucleotide design recommendations should be applicable for CGH analysis in most species.

sample_organism:
Caenorhabditis elegans
Homo sapiens

platform_organism:
Caenorhabditis elegans
Homo sapiens

overall_design: One-copy deletions were studied in both the C. elegans (2 samples) and human (1 sample) genomes. There were no replicate or dye-swap hybridizations.

first: Donald

last: Moerman


title: TJ411(fer-15(b26)II; age-1(hx542)II) vs. BA713(fer-15(b26)II)

city: Stanford

first: Stuart

last: Kim

department: Department of Developmental Biology

institute: Stanford University Medical Center

summary:
To identify changes in gene expression in age-1 mutants, we compared expression in fer-15(b26) animals to expression in fer-15(b26); age-1(hx542) mutants.  We prepared RNA from young adult fer-15(b26) mutants grown at 25?C (five samples prepared at Stanford University and six samples prepared at the University of Colorado) and young adult fer-15(b26); age-1(hx542) mutants (four samples at Stanford University and six samples at the University of Colorado).
Set of arrays that are part of repeated experiments
Place of growth: Stanford vs Colorado
Genotype: mutant strains vs wt (N2) mixed stage; some expts are reverse dye

overall_design:
Biological Replicate
Computed

first: Min

last: Jiang

first: Pat

last: Tedesco


title: Transcriptional analysis of Leishmania infantum methotrexate resistant strains using full-genome DNA microarrays

first: Marc

last: Ouellette

summary:
Transcriptional analysis of the drug resistant strain L. infantum MTX20.5 compared to the wild-type strain
The full-genome DNA microarrays includes one 70-oligonucleotides probe for each gene of L. infantum
Keywords: drug resistance

overall_design: Two-condition experiment, resistant strain MTX20.5 vs wild-type. Four biological replicates for each strain, independently grown and harvested. One replicate per array

first: J

middle: M

last: Ubeda

first: D

last: Légaré

first: F

last: Raymond

first: A

last: Ouameur

first: S

last: Boisvert

first: P

last: Rigault

last: Corbeil

first: M

last: Tremblay

last: Olivier

first: B

last: Papadopoulou


title: Transcriptome analysis of infection of the archaeon Sulfolobus solfataricus with STIV

status: Public on Jul 25 2008

city: Bozeman

first: Kate

last: McInnerney

state: MT

address: 109 Lewis Hall

institute: Montana State University

summary: Time series of infection of Sulfolobus solfataricus strain 2-2-12 cells infected with STIV are compared with uninfected cells over 32 h.

overall_design: Two-condition experiment, Control vs. Infected cells sampled at 5 time points (0, 8, 16, 24, 32 h post infection). Biological replicates: 3 control, 3 infected, paired samples independently grown. One replicate per array. Dye swaps.

first: Alice

middle: C

last: Ortmann

first: Susan

last: Brumfield

first: Jasper

last: Walther

first: Kathleen

first: Stan

last: Broun

first: Harmen

last: vandeWerken

first: Brian

last: Bothner

first: Trevor

last: Douglas

first: John

last: vanderOost

first: Mark

last: Young


title: A New Class of Homoserine Lactone Quorum Sensing Signals

city: Seattle

first: Yasuhiro

last: Oda

state: WA

address: Box 357242, 1705 N.E. Pacific Street, HSB K-354

department: Department of Microbiology

institute: University of Washington

summary:
Quorum sensing is a term used to describe cell-to-cell communication that allows cell density-dependent gene expression. Many Gram-negative bacteria use acyl-homoserine lactone (acyl-HSL) synthases to generate fatty acyl-HSL quorum sensing signals, which function with signal receptors to control expression of specific genes. The fatty acyl group is derived from fatty acid biosynthesis and provides signal specificity, but the variety of signals is limited. We have discovered that the photosynthetic bacterium Rhodopseudomonas palustris uses an acyl-HSL synthase to produce p-coumaroyl-HSL by using environmental p-coumaric acid rather than fatty acids from cellular pools. The bacterium has a signal receptor with homology to fatty acyl-HSL receptors that responds to p-coumaroyl-HSL to regulate global gene expression. We also found that p-coumaroyl-HSL is made by other bacteria including Bradyrhizobium BTAi1 and Silicibacter pomeroyi DSS-3. This discovery extends the range of possibilities for acyl-HSL quorum sensing and raises fundamental questions about quorum sensing within the context of environmental signaling.
Keywords: Comparison of transcriptome profiles

overall_design: Transcriptome profiles between Rhodopseudomonas palustris cells grown in the in the presence or absence of pC-HSL were compared.

first: Amy

last: Schaefer

first: Everett

last: Greenberg

first: Colin

last: Oliver

first: Jean

last: Huang

first: Gili

last: Bittan-Banin

first: Caroline

last: Peres

first: Silke

last: Schmidt

first: Katarina

last: Juhaszova

first: Janice

last: Sufrin

middle: S

last: Harwood


title: TJ1081)fer-15(b26)II; daf-16(m26)I) vs. BA713(fer-15(b26)II)

first: Stuart

last: Kim

summary:
To identify changes in gene expression in daf-16 mutants, we compared expression in fer-15 young adults to expression in fer-15(b26); daf-16(m26) young adults grown at 25?C.  At Stanford University, four samples of fer-15 and five samples of fer-15; daf-16 animals were prepared.  At the University of Colorado, seven samples of fer-15 and six samples of fer-15; daf-16 was prepared.
Set of arrays that are part of repeated experiments
Place of growth: At Stanford vs Colorado
Genotype: mutant strains vs wt (N2) mixed stage; some expts are reverse dye (wt in channel 2)

first: Pat

last: Tedesco

first: Min

last: Jiang


title: Cellular Aging of Mesenchymal Stem Cells (miRNA)

city: Aachen

first: Wolfgang

last: Wagner

address: Pauwelsstrasse 20

department: Helmholtz Institute for Biomedical Engineering

institute: RWTH Aachen University

summary:
To determine miRNA expression changes during in vitro senescence of mesenchymal stem cells (MSC) we have analyzed differential expression of the corresponding early passage (P2) and senescent passage (PX).
Keywords: miRNA, senescence, mesenchymal stromal cells, mesenchymal stem cells, MSC

overall_design:
Mesenchymal Stem Cells (MSC) were isolated from human bone marrow (BM) as described before  (Wagner et al., 2005, Exp Hematol, 33, 1402-1416; Wagner et al., Exp Hematol, 34, 536-548). Cells were always replated when grown to about 70% confluency. A sample for RNA isolation was taken at every passage until the cells finally became senescent: they became much larger with irregular and flat shape. The nucleus became more circumscribed in phase contrast microscopy. The cytoplasm began to be granular with many inclusions and there was more cell debris. Global gene expression profiles were analyzed to determine molecular changes between corresponding early passage (P2) and senescent passage (PX) in three donor samples. In addition we have analyzed different passages of donor 1 (P2, P3, P4, P5, P6, P7, P8, P9, P10, and P11) to determine changes in the course of cellular aging.
The medians of four background-corrected replicas for each miRNA capture probe were used for further analysis. Data of each hybridization was median normalized (each value was divided by the median of values). Differential expression was compared between P2 and later passages of the corresponding donor sample.

first: Mirco

last: Castoldi

first: Patrick

last: Horn

first: Vladimir

last: Benes

first: Jonathon

last: Blake

first: Anthony

last: Ho


title: Genetic Structure of Duplicated Sequences Revealed by Genotyping Single Sperm

city: Piscataway

first: Minjie

last: Luo

state: NJ

address: 675 Hoes Lane

department: Microbiology & Molecular Genetics

institute: University of Medicine and Dentistry of New Jersey

type:
Genome variation profiling by SNP array
SNP genotyping by SNP array

summary:
Three different experimental approaches were evaluated for discrimination of genomic variance in and between duplicated sequences using 48 markers in duplicon regions and 17 SNPs in unique sequences previously characterized in another study. We found only the method high-throughput single sperm typing could conclusively resolve the alleles of all markers. Resulting data from single sperm analysis were also used to examine the genetic structure of duplicon markers in the human population. Single sperm typing can be a rapid, efficient and accurate method for initial screening and assessment of genetic variation and for detailed genetic analysis of duplicon markers.

Keywords: Genotyping

overall_design: Sixty-five markers including 17 MSVs, 12 PSVs, 19 SIDs and 17 SNPs in unique sequences described in Fredman et al. were selected for study. The samples include 40 genomic DNA samples from four ethnic groups, semen samples from 11 donors, and 10 to 20 sperm from each donor except one, AB012, for whom 65 sperm were analyzed. Both genomic and sperm DNA samples were subject to multiplex amplification followed by microarray analysis. Genotypes were determined by using the Accutyping software. Semen samples were genotyped on both strands. Allele status in these samples were compared and analyzed. The single sperm typing method allowed us to identify markers residing in non-unique sequence, to analyze the detailed genetic structure of the duplicons and to learn whether different alleles are present for the duplicon sequences in the human population.

first: David

last: Fredman

first: Xiangfeng

last: Cui

last: Brookes

first: Marco

last: Azaro

first: Danielle

last: Greenawalt

first: Guohong

last: Hu

first: Hui

middle: Y

last: Wang

first: Irina

middle: V

last: Tereshchenko

first: Yue

last: Shentu

first: Richeng

last: Gao

first: Li

last: Shen

first: Honghua

last: Li


title: Saccharomyces pastorianus strains comparison

status: Public on Jul 23 2008

first: Barbara

last: Dunn

department: Genetics

institute: Stanford University

summary:
Saccharomyces pastorianus is the yeast used to make lager beer; it is known to be an interspecific hybrid formed by the fusion between S. cerevisiae and S. bayanus genomes.  This data set queries 17 S. pastorianus strains, collected at various times over the last 125 years from various breweries located in different geographical locations, which were obtained from CBS and DBVPG culture collections.  The data in this set represent array-CGH experiments performed with these strains, using "2-species" custom Agilent arrays (the "2-species" arrays contain probes spaced every ~2 kb across the whole genomes of both S. cerevisiae and S. bayanus; the probes are unique and specific for each genome).  The data set also contains 3 self-self hybridizations (S. cerevisiae + S. bayanus DNA mixed together in equimolar amounts, then labeled green or red in separate reactions, then hybridized to the "2-species" arrays) used for normalization in CGH-Miner analysis.
A strain or line experiment design type assays differences between multiple strains, cultivars, serovars, isolates, lines from organisms of a single species.

overall_design: Keywords: strain_or_line_design


title: Gene Expression and evolution of antifungal drug resistance

city: Mississauga

first: James

middle: B

last: Anderson

state: Ontario

address: 3359 Mississauga Road N.

department: Biology

institute: University of Toronto

summary:
We examined the gene expression
changes resulting from the evolution of resistance in
experimental populations of the yeast Saccharomyces
cerevisiae subjected to two antifungal drugs, fluconazole
(FLC) and amphotericin B (AmB). Fluconazole resistance may involve increased efflux or changes in sterol metabolism, while AmB resistance
generally involves changes in sterol metabolism; for all of
these types of resistance, the gene expression changes are
extensive. The goal of these experiments was to test
whether failure of gene expression changes all downstream
of the original mutation for drug resistance would affect
the ability of a mutant cell to evolve and/or to support a
drug-resistant phenotype.

overall_design:
In this study, replicate populations of Saccharomyces
cerevisiae were evolved in increasing concentrations of AmB
over hundreds of generations to yield resistant types among
which there were permanent changes in gene-expression.
Evolution experiments were started with 100 μL of an
overnight culture that was derived from a single colony
inoculated into 10 ml of YPD containing 0.25 μg/ml AmB. Five
different populations were evolved in the presence of AmB
in two different regimens, long and short. The rational
for choosing different evolution regimens was to increase
the chance of recruiting different mechanisms of AmB
resistance among all of the replicates. In the long
regimen, three populations started at 0.25 μg/ml AmB and
this concentration was doubled every 100 generations for a
total of 1100 generations and a final AmB concentration of
125 μg/ml (sce2468, sce2469, sce2470).
In the short regimen, two populations were also
started at 0.25 μg/ml AmB, but were given as much time as
necessary to grow to high density, usually two to four
days. The populations were then transferred a second time
into the same AmB concentration. At the next transfer, the
AmB concentration was doubled for another two growth
cycles, and so on, until the populations had been through
two transfers in 128 μg/ml AmB (sce2539 and sce2542).

first: Caroline

last: Sirjusingh

first: Nazia

last: Syed

first: Shantelle

last: LaFayette


title: Sister chromatid cohesion establishment by acetylation

status: Public on Jul 24 2008

city: London

first: Tamar Tom

last: Rolef Ben-Shahar

address: 44 Lincoln's Inn Fields

institute: Cancer Research UK London Research Institute

summary: Cohesion between sister chromatids depends on the chromosomal cohesin complex and allows the spindle apparatus in mitosis to recognize replicated chromosomes for segregation into daughter cells. Sister chromatid cohesion is established concomitant with DNA replication, and requires the essential Eco1 protein, a replication fork-associated acetyl transferase. The mechanism by which Eco1 establishes sister chromatid cohesion is not known. Here, we show that the cohesin subunit Smc3 is acetylated in an Eco1-dependent manner during S phase to establish sister chromatid cohesion. We isolated spontaneous suppressors of the thermosensitive eco1-1 allele in budding yeast, and identified the suppressor mutations from the hybridization pattern of genomic DNA on oligonucleotide tiling arrays. An acetylation mimicking mutation of a conserved lysine in Smc3 to asparagine (K113N) makes Eco1 dispensable for cell growth, indicating that Smc3 acetylation is Eco1’s only essential function. We identified a second set of eco1-1 suppressor mutations in the budding yeast ortholog of the cohesin regulator Wapl (Wpl1/Rad61). Wapl destabilizes cohesin on chromosomes, and Eco1-dependent Smc3 acetylation during S-phase might render cohesin resistant to Wapl. Our results clarify the role of Eco1 in the establishment of sister chromatid cohesion, and suggest that Eco1 modifies cohesin to stabilize an Eco1-independent cohesion establishment reaction.

overall_design: High density oligonucleotide tiling arrays were used to hybridize genomic DNA from the Saccharomyces cerevisiae eco1-1 mutant and from spontaneous suppressor mutants that rescue eco1-1 thermosensitive growth. Hybridization intensities at each oligonucleotide probe were compared using the algorithm discribed in [D. Gresham et al. Science 311, 1932 (2006)] to locate the nucleotide changes causing the suppression. One such mutation was in the SMC3 gene, introducing an acetylation mimicking asparagine mutation to an evolutionarily conserved lysine that serves as a substrate to Eco1 acetyltransferase activity. A separate complementation group had mutations in the RAD61 gene (here renamed WPL1), the ortholog of human and fission yeast Wapl. WPL1 inactivation allowed cell growth in the absence of Eco1. Tiling arrays were used again to analyze Wpl1 and cohesin chromatin immunoprecipitates, to demonstrate their colocalization along chromosomes in G2.

first: Tom

middle: T

first: Sebastian

last: Heeger

first: Chris

last: Lehane

first: Philip

last: East

first: Helen

last: Flynn

first: Mark

last: Skehel

first: Frank

last: Uhlmann


title: Gene expression patterns of sulfur starvation in Synechocystis sp. PCC 6803

city: St. Paul

first: Zhigang

last: Zhang

address: 1479 Gortner Avenue, Suite 140

department: BioTechnology Institute

institute: University of Minnesota

summary:
The unicellular cyanobacterium Synechocystis sp. PCC 6803 is a model system for studying biochemistry, genetics and molecular biology of photobiological processes. Despite its importance in basic and applied research, the genome-wide picture of transcriptional regulation in this bacterium is limited. Characteristic transcriptional responses to changes in the growth environment are expected to provide a scaffold for describing the Synechocystis transcriptional regulatory network as well as efficient means for functional annotation of genes in the genome. We designed, validated and used Synechocystis genome-wide oligonucleotide (70-mer) microarray (representing 96.7% of all chromosomal ORFs) to study transcriptional activity of the cyanobacterial genome in response to S deprivation. The microarray data were verified by quantitative RT-PCR. We made five main observations: 1) Transcriptional changes upon sulfate withdrawal were relatively moderate, but significant and consistent with growth kinetics; 2) S acquisition genes encoding for a high-affinity sulfate transporter were significantly induced, while decreased transcription of genes for phycobilisome, photosystems I and II, cytochrome b6/f, and ATP synthase indicated reduced light-harvesting and photosynthetic activity; 3) S deprivation elicited transcriptional responses associated with general growth arrest and stress; 4) A large number of genes regulated by S availability encode hypothetical proteins or proteins of unknown function; 5) Hydrogenase structural and maturation accessory genes were not identified as differentially expressed, even though increased hydrogen evolution was observed. The expression profiles recorded by using this oligonucleotide-based microarray platform revealed that during transition from the condition of plentiful sulfur to no sulfur, Synechocystis undergoes coordinated transcriptional changes, including genes whose products are involved in sensing nutrient limitations and tuning bacterial metabolism. The transcriptional profile of the nutrient limitation was dominated by decrease in abundances of many transcripts. However, these changes were unlikely due to the across-the-board, non-specific shut down of transcription in a condition of growth arrest. Down-regulation of transcripts encoding proteins whose function depends on a cellular sulfur status indicated that the observed repression has a specific regulatory component. The repression of certain sulfur-related genes was paralleled by activation of genes involved in internal and external S scavenging.

Keywords: stress response, time course

overall_design: Synechocystis sp. PCC 6803 was grown photoautotrophically in BG-11 medium supplemented with 8mM NaHCO3 and buffered with 10mM HEPES (pH 7.4). The cells were grown in 250ml flasks at 32oC under a light intensity of 25µmol photons m-2 s-1. Cultures were bubbled with sterile air containing 1% (v/v) CO2. Log phase cells (OD730nm=0.6) were harvested by centrifugation (2000×g for 12 min) washed once and then re-suspended in sulfate-free media (MgSO4 replaced by the same molarity of MgCl2). In addition, all S-containing trace metals in BG-11 were replaced by non-S containing metals. Cells were harvested and fixed for microarray analysis by adding 10% (v/v) ice-cold 5% phenol in ethanol stop solution at the following time points: before S-depravation (time 0, control), 1, 3, 6, 12, 24, 48 and 72 hr after S-depravation. S-deprivation with HEPES buffering control experiment was performed as described above, except that HEPES buffer was used upon sulfate removal. Bacterial samples for a time course were taken at time 0, 1, 12 and 24 hrs after sulfate withdrawal. Growth stage control experiment was done in parallel with S deprivation experiments. Samples were taken at 0, 1, 2.5, 4, 7, 11 and 48 hr after OD730nm reached 0.60. All the experiments were done in biological replicates.

first: Ninad

last: Pendse

first: Arkady

last: Khodursky


title: Glucocorticoid Signaling Defines a Novel Commitment State during Adipogenesis In Vitro

city: Berkeley

first: Jason

last: Huff

address: 211 Koshland Hall

department: Plant & Microbial Biology

institute: University of California, Berkeley

summary: Differentiation of 3T3-L1 preadipocytes can be induced by a 2-d treatment with a factor "cocktail" (DIM) containing the synthetic glucocorticoid dexamethasone (dex), insulin, the phosphodiesterase inhibitor methylisobutylxanthine (IBMX) and fetal bovine serum (FBS). We temporally uncoupled the activities of the four DIM components and found that treatment with dex for 48 h followed by IBMX treatment for 48 h was sufficient for adipogenesis, whereas treatment with IBMX followed by dex failed to induce significant differentiation. Similar results were obtained with C3H10T1/2 and primary mesenchymal stem cells. The 3T3-L1 adipocytes differentiated by sequential treatment with dex and IBMX displayed insulin sensitivity equivalent to DIM adipocytes, but had lower sensitivity to isoproterenol-stimulated lipolysis and reduced triglyceride content. The nondifferentiating IBMX–then-dex treatment produced transient expression of adipogenic transcriptional regulatory factors C/EBP{beta} and C/EBP{delta}, and little induction of terminal differentiation factors C/EBP{alpha} and PPAR{gamma}. Moreover, the adipogenesis inhibitor preadipocyte factor-1 (Pref-1) was repressed by DIM or by dex-then-IBMX, but not by IBMX-then-dex treatment. We conclude that glucocorticoids drive preadipocytes to a novel intermediate cellular state, the dex-primed preadipocyte, during adipogenesis in cell culture, and that Pref-1 repression may be a cell fate determinant in preadipocytes.

overall_design: We sought to define the genes differentially expressed in response to dexamethasone (dex) in 3T3L1 mouse preadipocytes.  We treated 3T3L1 cells with dex or DMSO vehicle for 2 or 36 hrs and then prepared total RNA.  We hybridized each sample (Cy5) to a pool of the samples, which served as the reference (Cy3) channel. All linear models for differential gene expression were constructed with the "limma" package (Smyth, 2004) in BioConductor (Gentleman et al., 2004). A linear model for dex was constructed to compare the 4 biological replicates of DMSO vehicle treatment with dex treatment at either 2 or 36hr. Genes were considered differentially expressed if at least one probe had log-odds greater than 50:50 in the linear model.

first: Carlos

last: Pantoja

first: Keith

last: Yamamoto


title: Target genes associated to siRNA mediated repression of Pea3 and Erm transcription factors in mammary cancerous cells

city: Lille

first: Anne

last: Chotteau

address: 1 rue du professeur calmette

country: France

institute: CNRS UMR 8161

summary:
Pea3 and Erm are transcription factors belonging to the ETS family which are involved in tumorigenesis, particularly mammary oncogenesis. To elucidate the mechanism of Pea3/Erm induced tumorigenesis in mouse mammary cancerous MMT cells (ATCC CCL-51), we have down-regulated Pea3 or Erm expression in this cell line by using small interfering RNA and performed a microarray analysis using Applied Biosystems AB1700 technology to define target associated genes.
Keywords: Comparative transcriptome hybridization, small interfering RNA, mouse mammary cancerous cells

overall_design:
Mouse mammary cancerous MMT cells (ATCC CCL-51) have been transfected with a siRNA control, an erm-directed siRNA or an pea3-directed siRNA. MMT cells were also used to determine the modifications due to the transfection protocol.
These conditions correspond to four sample.
Two biological replicate were used for the MMT cells (control sample), three biological replicates were used for the siRNA control (control sample) and erm-directed siRNA (reference sample) MMT cells and four biological replicates were used for the pea3-directed siRNA (reference sample) MMT cells. Transcriptome profiles (28.218 validated mouse genes) were acquired using Applied Biosystems AB1700 technology, which make use of chemiluminescence-based detection chemistry.

first: Virginie

last: Firlej

first: Franck

last: Ladam

first: Guillaume

last: Brysbaert

first: Yvan

last: de Launoit

first: Arndt

last: Benecke

last: Chotteau-Lelievre


title: A DNA polymerase α accessory protein, Mcl1, is required for propagation of centromere structures in fission yeast

status: Public on Jul 22 2008

city: Bunkyo-ku

first: Katsuhiko

last: Shirahige

state: Tokyo

address: 1-1-1 Yayoi

department: Research Center for Epigenetic Disease

institute: The University of Tokyo

summary:
Specialized chromatin exists at centromeres and must be precisely transmitted during DNA replication. The mechanisms involved in the propagation of these structures remain elusive. Fission yeast centromeres are composed of two chromatin domains: the central CENP-ACnp1 kinetochore domain and flanking heterochromatin domains. Here we show that fission yeast Mcl1, a DNA polymerase α (Polα) accessory protein, is critical for maintenance of centromeric chromatin. In a screen for mutants that alleviate both central domain and outer repeat silencing, we isolated several cos mutants, of which cos1 is allelic to mcl1. The mcl1-101 mutation causes reduced CENP-ACnp1 in the central domain and an aberrant increase in histone acetylation in both domains. These phenotypes are also observed in a mutant of swi7+, which encodes a catalytic subunit of Polα. Mcl1 forms S-phase-specific nuclear foci, which colocalize with those of PCNA and Polα. These results suggest that Mcl1 and Polα are required for propagation of centromere chromatin structures during DNA replication.
Keywords: ChIP-chip analysis, S. pombe

overall_design: Antibodies used were α-acetyl-histone H4 antiserum (Upstate) and α-histone H4 pan antibody (Upstate). ChIP-on-chip was carried out using IVT amplification method. ChIP-chip data were comfirmed by real-time PCR.

first: Toyoaki

last: Natsume

first: Yasuhiro

last: Tsutsui

first: Takashi

last: Sutani

first: Elaine

last: Dunleavy

first: Alison

middle: L

last: Pidoux

first: Hiroshi

last: Iwasaki

first: Robin

last: Allshire

first: Fumiaki

last: Yamano


title: Human cholangiocarcinoma cells treated with Clonorchis sinensis excretory-secretory products.

city: Seoul

first: Dong-Wook

last: Kim

address: 221, Heukseok-Dong, Dongjak-Ku

department: Lifescience

country: South Korea

institute: Chung-Ang University

summary:
Clonorchiasis is associated with bile duct malignancy and the subsequent development of cholangiocarcinoma. Although this is likely caused by adult Clonorchis sinensis and its excretory-secretory products (ESP), the precise molecular mechanisms remain obscure. To evaluate the effect of C. sinensis infection on differential gene expression in host hepatocytes, we used cDNA microarrays in human cholangiocarcinoma cells through the mimicking of C. sinensis infestation, and analyzed differential mRNA expression patterns of host cells.

Keywords: Time course

overall_design: Metacercariae of C. sinensis were obtained from P. parva and then it infected into albino rabbits. Adult worms were collected from infected rabbits, and incubated in PBS. The supernatant was centrifuged and filtered with syringe-driven filter. HuCCT1 cell were treated with prepared ESP in order to profile the differentially expressed genes at five time points: 0 (T01), 1(T02), 3(T03), 9(T04), and 24(T05) hours.

first: Jhang

middle: H

last: Pak

first: Ju

last: Moon

first: Joo-Hyun

last: Nam

first: Jong-Hyeok

first: Jung

middle: W

last: Ju

first: Tong-Soo

first: Sang-Beom

last: Seo


title: Keratinocyte specific Fos-deletion in K5-Sos-F mouse tumor model

status: Public on Sep 01 2008

first: Daniel

last: Haag

state: Santa Clara

address: 265 Campus Drive

department: ISCBRM

institute: Stanford School of Medicine

summary:
Expression and function of the oncogenic transcription factor AP-1 (mainly composed of Jun and Fos proteins) is required for neoplastic transformation of mouse and human keratinocytes in vitro and tumor promotion as well as malignant progression in vivo. Here, we describe the identification of novel Fos target genes using global gene expression profiling with samples from a tumor model of mouse skin (K5-SOS-F). We could identify 366 differentially expressed genes comparing expression profiles from tumor samples of control animals with samples derived form mice with a specific deletion of fos in keratinocytes.
Keywords: Fos-deletion, Fos-floxed, K5-SOS-F mouse tumor model, skin papilloma, global gene expression, microarray, Fos target in skin carcinogenesis

overall_design: In the present Experiment two different genotypes in combination with an established mouse tumor model were compared. An active form of the Sos (son of sevenless) is expressed under a keratin5 promoter leading to a constitutive activation of the Ras-Raf pathway, which results in skin papilloma formation. In the context of this tumor model an epidermis specific homozygote Fos-deletion was compared to homozygous floxed Fos-alleles. Both genotypes (Fos-deleted and Fos-floxed) were treated equally. The specimens were taken from skin papilloma at the mouse tail. A total number of six specimens from six different individuals (three biological replicates per genotype) were taken. Each sample was prepared and hybridized against Universal Mouse Reference RNA (Stratagene) in a dye-swap experiment, resulting in two technical replicates for each specimen, respectively.

first: Moritz

last: Durchdewald

first: Juan

last: Viniegra


title: Bj_Heterotrophy vs. Chemoautotrophy

status: Public on Jul 19 2008

city: Arlington

first: Woo-Suk

last: Chang

state: TX

address: 501 S. Nedderman Dr. , Life Science Building 216

institute: University of Texas at Arlington

summary:
Transcriptional profiling of chemoautotrophically grown cells
Keywords: Comparison of different lifestyles

overall_design: Two independent biological materials were prepared for chemoautotrophically cultured cells and heterotrophically cultured cells. Total 4 arrays including dye swap were analyzed.

first: William

last: Franck

first: Jing

last: Qiu

first: Stephanie

last: Smith

first: Gary

last: Stacey


title: Chip-chip of condensin subunits Brn1 and Smc4 and Scc2/4 and cohesin and Tfc3 (TFIIIC) on budding yeast chromosomes

status: Public on Jul 21 2008

first: claudio

last: d'ambrosio

institute: Cancer Research UK

summary:
Eukaryotic chromosomes reach their stable rod-shaped appearance in mitosis in a reaction dependent on the evolutionarily conserved condensin complex. Little is known about how and where condensin associates with chromosomes. Here, we analyse condensin binding to budding yeast chromosomes using high resolution oligonucleotide tiling arrays. Condensin binding sites coincide with those of the loading factor Scc2/4 of the related cohesin complex. The sites map to tRNA genes, ribosomal protein genes, and other places characterised by the RNA polymerase III transcription factor TFIIIC. An ectopic B-Box element, recognised by TFIIIC, constitutes a minimal condensin binding site, and TFIIIC and the Scc2/4 complex promote productive condensin association with chromosomes. A similar pattern of condensin binding is conserved along fission yeast chromosomes. This reveals that TFIIIC binding sites, including tRNA genes, constitute a hitherto unknown chromosomal feature with important implications for chromosome architecture during both interphase and mitosis.

Keywords: Chip-chip, cell type comparison

sample_organism:
Schizosaccharomyces pombe
Saccharomyces cerevisiae

overall_design: Pattern of binding of condensin Brn1, cohesin Scc1, Scc2 and Tfc3 on budding yeast chromosomes as well as condensin Cnd2 binding on fission yeast chromosomes.

first: Claudio

last: D'Ambrosio

first: Christine

last: Schmidt

first: Yuki

last: Katou

first: Gavin

last: Kelly

first: Takehiko

last: Itoh

first: Katsuhige

last: Shirahige

first: Frank

last: Uhlmann


title: Influence of nalidixic acid on gene expression of Escherichia coli

status: Public on Jul 20 2008

city: Bremen

last: Klockow

address: Celsiusstr. 1

institute: Max Planck Institute for Marine Microbiology

summary:
The intention of this study is to analyse the effect of antibiotics on the gene expression of Escherichia coli. Shaking-flask cultivations of Escherichia coli K12GFP-UTL2 were carried out with a medium containing nalidixic acid. Cultures with antibiotic-free medium, which were run in an identical way, served as reference. Samples were taken at different times during the cultivations, the RNA was isolated and hybridised on whole genome yeast microarrays.
Keywords: Influence of toxins on gene expression in E. coli

overall_design: A timeserial experiment of the influence of nalidixic acid on the gene expression in Escherichia coli was performed. Effects of the growth curve were eliminated by bionformatic methods.

first: Ken

last: Reardon

last: Stahl

first: Bernd

last: Hitzmann

first: Thomas

last: Scheper


title: Bj_Chemoautotrophy vs. Arabinose supplemented chemoautotrophy

last: Chang

summary:
Transcriptional profiling of chemoautotrophically and arabinose supplemented chemoautotrophically grown cells
Keywords: Comparison of different lifestyles

overall_design: Three independent biological materials were prepared for arabinose supplemented chemoautotrophically cultured cells and chemoautotrophically cultured cells. Total 6 arrays including dye swap were analyzed.

last: Franck


title: Comparison of ES cells with relevant compartments of early embryos

city: Cardiff

first: Fiona

middle: Catherine

last: Mansergh

state: Wales

address: Museum Avenue

department: Biosciences

institute: Cardiff University

summary:
Embryonic stem cells (ES cells) are pluripotent stem cells that can contribute to all lineages of the embryo proper and were first isolated in 1981 in this laboratory (Evans and Kaufmann, 1981). They have contributed immeasurably to biology, both in terms of studying pluripotent stem cell maintenance in culture and in vitro differentiation, and because they can be easily genetically manipulated in culture, allowing for the production of mice with designed specific mutations (targeted mice), via homologous recombination. The recent isolation of human ES cell lines is promising in terms of stem cell therapy development.
Mouse ES cell lines are isolated from the inner cell masses of mouse blastocysts between days 3.5 and 5.0 post coitum, or from the inner cell masses of delayed blastocysts. However, it has been suggested that some aspects of their biology may be more similar to embryonic ectoderm from early post implantation embryos, than to pre-implantation blastocysts. It has also been suggested that human embryonic stem cells are more similar to mouse stem cells derived from early embryonic ectoderm, than to mouse ES cells. Until recently, however, it has been difficult to answer these questions definitively, as the quantity of tissue available from early embryos is extremely limiting. However, we have used two rounds of RNA amplification on early embryonic samples and individual colonies picked from ES cell culture to compare the transcriptomic profiles of mouse ES cells with the embryonic tissues from with these cells are commonly derived, and with slightly later embryonic stages. We have found that ES cells are notably most similar in profile to embryonic ectoderm at day 5.5  (EE5.5) . ES cells cannot be derived from this post implantation embryonic tissue, so this result is in some ways surprising. However, ES cells show key differences from EE5.5, including the expression of markers of pluripotency such as Oct4 and Nanog (see gene lists acompanying this record), which explain how their ability to contribute to all tissues of the embryo proper is retained.
Keywords: cell type comparison

overall_design: We used the NIA 15K array set, which is perfect for this study as the cDNA clones that comprise this set are derived from germ line tissues, germ cells and various embryonic stages. The pooled control used in this study was assembled using all of the samples used originally when isolating clones for the NIA15K set. This reference sample should therefore hybridize to most of the clones in the NIA set, providing a good background signal with which to compare samples from specific embryonic stages. This pooled control was always labelled with Cy5, allowing all of the experimental samples (IMT11 ES cells, ICM88, ICM105, DICM136, DICM180, EE5.5 and EE6.5) to be labelled with Cy3. Since these samples were always labelled with cy3, they could be compared with each other without risk of dye bias, since they were all labelled with the same dye. 6 repetitions were carried out for all samples except DICM180 (which was extremely difficult to isolate), for which there are 4 repetitions. 3 biological replicates were carried out for each stage (2 slides per biological replicate gives 6 reps total). RNA was isolated using the Zymo Mini RNA isolation kit, and then amplified twice using the Arcturus RiboAmp kit according to the manufacturer's protocol. This yielded enough amplified RNA for target labelling and array hybridization. The Atlas™ PowerScript™ Fluorescent Labelling Kit and 5μg of amplified RNA were used for labelling according to the manufacturer’s protocol (Promega). This labelling method is indirect (amino-allyl), reducing the chances of direct cyanine dye incorporation affecting the efficiency of hybridization.

first: Susan

last: Hunter

first: Martin

middle: J

last: Evans


title: Profiling protease and protease inhibitor gene expression in tumors and their microenvironment

city: Detroit

first: Kami

last: Moin

address: 540 E. Canfield

department: Pharmacology

institute: Wayne State University

summary:
To assist in the identification of proteases, their endogenous inhibitors, and proteins that interact with proteases or proteolytic pathways in biological tissues, a dual species oligonucleotide microarray has been developed in conjunction with Affymetrix, Inc. To investigate the performance of the array, gene expression profiles were analyzed in pure mouse and human samples and orthotopically implanted xenografts of human A549 lung and MDA-MB-231 breast carcinomas. The Hu/Mu ProtIn array is a valuable discovery tool for the identification of components of human and murine proteolytic pathways in health and disease, and has particular utility in the determination of cellular origins of proteases and protease inhibitors in xenograft models of human cancer.
Keywords: gene expression profiles were analyzed in pure mouse and human samples and orthotopically implanted xenografts of human A549 lung and MDA-MB-231 breast carcinomas

sample_organism:
Homo sapiens
Mus musculus

overall_design: Commercial pan human and mouse total RNA replicate samples (n=3) were profiled. In addition, human ductal stage II or III breast carcinoma biopsies (n=18) were compared to normal human breast biopsies (n=13). Spontaneous spontaneous murine mammary carcinomas from MMTV-PyMT+(FVB/n) (n=8) and MMTV-PyMT+/uPARAP-/-(FVB/n) (n=2) transgenic mice were compared to normal mouse mammary fat pads (n=4; female Rag-1 tm1Mom retired breeders at >6 weeks post lactation). In vitro cultures of MDA-MB-231 human breast carcinoma cells (n=3) and MDA-MB-231 orthotopic xenografts (n=3) were profiled. The xenografts were compared to normal mammary fat pads. A549 human lung adenocarcinoma cells (n=2), A549 orthotopic xenografts (n=3) and normal mouse lung tissue (n = 3) were profiled.

first: Donald

middle: R

last: Schwartz

first: Kamiar

first: Bin

last: Yao

first: Lynn

last: Matrisian

first: Lisa

last: Coussens

last: Bugge

first: Bonnie

middle: F

last: Sloane

first: Brian

last: Fayz


title: Bj_Heterotrophy vs. Arabinose supplemented chemoautotrophy

first: Woo-Suk

summary:
Transcriptional profiling of arabinose supplemented chemoautotrophically grown cells
Keywords: Comparison of different lifestyles

overall_design: Three independent biological materials were prepared for arabinose supplemented chemoautotrophically cultured cells and heterotrophically cultured cells. Total 6 arrays including dye swap were analyzed.

first: William

first: Jing

last: Qiu

first: Stephanie

last: Smith

first: Gary

last: Stacey


title: Genomic profiling identifies GATA6 as a candidate oncogene amplified in pancreatobiliary cancer

status: Public on Jul 18 2008

first: Kevin

last: Kwei

address: 269 Campus Dr.

institute: Stanford

summary:
Pancreatobiliary cancers have among the highest mortality rates of any cancer type.  Discovering the full spectrum of molecular genetic alterations may suggest new avenues for therapy.  To catalogue genomic alterations, we carried out array-based genomic profiling of 31 exocrine pancreatic cancers and 6 distal bile duct cancers, expanded as xenografts to enrich the tumor cell fraction.  We identified numerous focal DNA amplifications and deletions, including in 19% of pancreatobiliary cases gain at cytoband 18q11.2, a locus uncommonly amplified in other tumor types.  The smallest shared amplification at 18q11.2 included GATA6, a transcriptional regulator previously linked to normal pancreas development.  When amplified, GATA6 was overexpressed at both the mRNA and protein level, and strong immunostaining was observed in 25 of 54 (46%) primary pancreatic cancers compared to 0 of 33 normal pancreas specimens surveyed.  GATA6 expression in xenografts was associated with specific microarray gene-expression patterns, enriched for GATA binding sites and mitochondrial oxidative phosphorylation activity.  siRNA mediated knockdown of GATA6 in pancreatic cancer cell lines with amplification led to reduced cell proliferation, cell cycle progression, and colony formation.  Our findings indicate that GATA6 amplification and overexpression contribute to the oncogenic phenotypes of pancreatic cancer cells, and implicate GATA6 as a lineage-specific oncogene in pancreatobiliary cancer, with implications for novel treatment strategies.
Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc.
Tissue type: cancer xenograft from diff tissues
Keywords: Logical Set

overall_design:
cDNA microarrays from the Stanford Functional Genomics Facility were used to perform gene expression profiling on 28 out of 31 of the exocrine pancreatic cancer and all 6 distal bile duct cancer xenografts.
Using regression correlation

first: Murali

last: Bashyam