title: Transcriptional profiling of Persea americana flowers

status: Public on Jun 18 2009

city: Gainesville

first: Andre

last: Chanderbali

state: FL

address: Museum Road and Newell Drive

department: Botany

institute: University of Florida

summary: The debate on the origin and evolution of flowers has recently entered the field of developmental genetics, with focus on the design of the ancestral floral regulatory program.  Flowers can differ dramatically among angiosperm lineages, but in general, sterile perianth organs surrounding stamens (male reproductive organs) and carpels (female reproductive organs) constitute the basic floral structure. However, the basal angiosperm lineages exhibit spectacular diversity in the number, arrangement, and structure, of floral organs, while the evolutionarily derived monocot and eudicot lineages share a far more uniform floral ground plan. As such, regulatory mechanisms underlying the archetypal floral plan, for instance that of the eudicot genetic model Arabidopsis thaliana, are unlikely to apply to the original flowers. Here we show that broadly overlapping transcriptional programs characterise the floral transcriptome of the basal angiosperm Persea americana (avocado), while floral gene expression domains are typically organ-specific in Arabidopsis. Our findings extend the “fading borders” model for basal angiosperms from organ identity genes to the downstream floral transcriptome, and suggest that the combinatorial mechanism for organ identity may not operate in basal angiosperms as it does in Arabidopsis. Furthermore, fading expression of components of the stamen transcriptome in central and peripheral regions of Persea flowers resembles the developmental program of the hypothesized gymnosperm “floral progenitor”. Accordingly, in contrast to the canalized organ-specific regulatory apparatus of Arabidopsis, floral development may have been originally regulated by overlapping transcriptional cascades with fading gradients of influence from focal to bordering organs.

overall_design: Expression profiles of inflorescence buds, pre-meiotic floral buds, inner and outer tepals, stamens, carpels, initiating fruit, and leaves were assessed in an interwoven double loop design for eight samples with 16 arrays. Sample materials were collected from two individuals (biological replicates) cultivated on the University of Florida’s Gainesville campus, and RNA was isolated twice for technical replication. Thus, four RNA extractions from each of the eight tissue types listed above were individually labeled with Cy3 (twice) or Cy5 (twice) and hybridized with four other Cy3 or Cy5 labeled samples as a dual channel array system.

first: Victor

last: Albert

first: Jim

last: Leebens-mack

first: Naomi

last: Altman

first: Douglas

last: Soltis

first: Pamela


title: Detection and differentiation of Clostridium botulinum strains

status: Public on Jun 15 2009

city: Atlanta

first: Brian

middle: H

last: Raphael

state: GA

address: 1600 Clifton Road

department: Enteric Diseases Laboratory Branch

country: USA

institute: Centers for Disease Control and Prevention

summary: Strains were differentiated on the basis of hybridization to probes representing strain variable regions in C. botulinum strain ATCC 3502.  Probes for selected genes (eg. toxin genes) were also featured on the microarray to allow detection of other serotypes/subtypes.

sample_organism:
Clostridium botulinum
Clostridium butyricum
Clostridium perfringens
Clostridium sporogenes
Clostridium baratii
Clostridium argentinense
Clostridium botulinum A str. ATCC 3502

overall_design: 44 strains were evaluated for selected gene detection and/or strain differentiation.  DNA from strain ATCC 3502 was used as a control as the featured probes were based on the ATCC 3502 genome sequence.

first: Lavin

middle: A

last: Joseph

first: Loretta

middle: M

last: McCroskey

first: Carolina

last: Luquez

first: Susan

middle: E

last: Maslanka


title: Identification of the Candida albicans Cap1p regulon (Cap1-CSE-HA and Cap1-HA complete datasets)

status: Public on Jun 22 2009

city: Paris

first: Sadri

last: ZNAIDI

address: 25 rue du Docteur Roux

department: Mycology Department

country: France

institute: Institut Pasteur

type: Genome binding/occupancy profiling by genome tiling array

summary: Cap1p, a transcription factor of the basic region-leucine zipper family, regulates oxidative stress response (OSR) in Candida albicans.  Alteration of its C-terminal cysteine-rich domain (CRD) results in Cap1p nuclear retention and transcriptional activation.  To better understand the function of Cap1p in C. albicans, we used genome-wide location profiling (ChIP-on-chip) to identify its transcriptional targets in vivo.  A triple-hemagglutinin epitope was introduced at the C-terminus of wild-type Cap1p (Cap1p-HA3) or hyperactive Cap1p with an altered CRD (Cap1p-CSE-HA3).  Location profiling using whole-genome oligonucleotide tiling microarrays identified 89 targets bound by Cap1p-HA3 or Cap1p-CSE-HA3 (binding ratio ≥ 2-fold, P ≤ 0.01).  Strikingly, Cap1p binding was not only detected at the promoter region of its target genes but also at their 3'-end and within their open-reading frame, suggesting that Cap1p may associate with the transcriptional or the chromatin remodeling machinery to exert its activity.  Overrepresented functional groups of the Cap1p targets (P ≤ 0.02) included 11 genes involved in OSR (CAP1, GLR1, TRX1, SOD1, CAT1, others), 13 genes involved in response to drugs (PDR16, MDR1, FLU1, YCF1, FCR1, others), 4 genes involved in phospholipid transport (PDR16, GIT1, RTA2, orf19.932) and 3 genes involved in the regulation of nitrogen utilization (GST3, orf19.2693, orf19.3121), suggesting that Cap1p has other cellular functions in addition to OSR.  Bioinformatic analyses of the bound sequences suggest that Cap1p recognizes the DNA motif 5'- MTKASTMA.  Finally, transcriptome analyses showed that increased expression generally accompanies Cap1p binding at its targets, indicating that Cap1p functions as a transcriptional activator.

overall_design: Processing of raw data from 3 independent replicate samples for each Cap1 dataset was conducted using Tilescope (http://tilescope.gersteinlab.org:8080/mosaic/pipeline.html). Quantile normalization was applied to the data. The parameters used were as follows: window size of 400 bp, MaxGap of 60 bp and MinRun of 120 bp. The replicate data were combined and peak-finding (i.e. Cap1p-binding sites) was determined using the following criteria: pseudo-median signal threshold of ≥ 2-fold and p-value cut-off of ≤ 0.01.

last: Znaidi

first: Katherine

middle: S

last: Barker

first: Sandra

last: Weber

first: Anne-Marie

last: Alarco

first: Teresa

middle: T

last: Liu

first: Geneviève

last: Boucher

first: David

middle: P

last: Rogers

first: Martine

last: Raymond


title: Expression of survivin in lung eosinophils is associated with pathology in a mouse model of allergic asthma 1

city: Brisbane

first: Ashley

last: Connolly

address: St Lucia

country: Australia

institute: University of Queensland

summary:
Humans vary markedly in their propensity to develop asthma, despite often being exposed to similar environmental stimuli. Similarly, mouse strains vary in susceptibility to airways pathology in experimental asthma. Sensitization and aerosol challenge with ovalbumin (OVA) induces eosinophil accumulation, mucus production and airways obstruction in BALB/c and C57BL/6 mice. In contrast, CBA/Ca mice show relatively little pathology. Allergen-induced production of IL-4, IL-5, IL-10 and IFN-g was detected in all three strains, with BALB/c mice generating the highest levels of IL-4, IL-5 and IL-10. Microarray analysis was used to identify genes differentially regulated in lung tissue after OVA challenge. Differentially regulated genes in the lungs of the asthma-susceptible C57BL/6 and BALB/c strains numbered 242 and 145, respectively, whereas only 42 genes were differentially expressed in the resistant CBA/Ca strain. In C57BL/6 mice, transcripts were enriched for adhesion molecules and this was associated with high levels of eosinophil recruitment. Differentially regulated genes in the lungs of only the asthma-susceptible strains numbered 64 and several of these have not previously been associated with asthma. Many of the genes differentially regulated in the susceptible strains were enzymes involved in inflammation. Using network analysis, mRNA for the anti-apoptotic protein survivin was found to be up-regulated in the lungs following allergen challenge. Survivin mRNA and protein were also expressed at high levels in eosinophils recovered by bronchoalveolar lavage from BALB/c and C57BL/6 mice. We propose that rapid apoptosis of lung eosinophils due to low expression of survivin contributes to the limitation of pathology in CBA/Ca mice
Humans vary markedly in their propensity to develop asthma, despite often being exposed to similar environmental stimuli. Similarly, mouse strains vary in susceptibility to airways pathology in experimental asthma. Sensitization and aerosol challenge with ovalbumin (OVA) induces eosinophil accumulation, mucus production and airways obstruction in BALB/c and C57BL/6 mice. In contrast, CBA/Ca mice show relatively little pathology. Allergen-induced production of IL-4, IL-5, IL-10 and IFN-g was detected in all three strains, with BALB/c mice generating the highest levels of IL-4, IL-5 and IL-10. Microarray analysis was used to identify genes differentially regulated in lung tissue after OVA challenge. Differentially regulated genes in the lungs of the asthma-susceptible C57BL/6 and BALB/c strains numbered 242 and 145, respectively, whereas only 42 genes were differentially expressed in the resistant CBA/Ca strain. In C57BL/6 mice, transcripts were enriched for adhesion molecules and this was associated with high levels of eosinophil recruitment. Differentially regulated genes in the lungs of only the asthma-susceptible strains numbered 64 and several of these have not previously been associated with asthma. Many of the genes differentially regulated in the susceptible strains were enzymes involved in inflammation. Using network analysis, mRNA for the anti-apoptotic protein survivin was found to be up-regulated in the lungs following allergen challenge. Survivin mRNA and protein were also expressed at high levels in eosinophils recovered by bronchoalveolar lavage from BALB/c and C57BL/6 mice. We propose that rapid apoptosis of lung eosinophils due to low expression of survivin contributes to the limitation of pathology in CBA/Ca mice

overall_design: Changes in gene expression in the lungs of 4 individual of BALB/c mice challenged with OVA were monitored using the lungs of 4 individual BALB/c mice challenged with PBS as the control. The microarray analysis was performed in quadruplicate.  Changes in gene expression in the lungs of 4 individual of CBA/Ca mice challenged with OVA were monitored using the lungs of 4 individual CBA/Ca mice challenged with PBS as the control. The microarray analysis was performed in quadruplicate.  Changes in gene expression in the lungs of 4 individual of BALB/c mice challenged with OVA were monitored using the lungs of 4 individual CBA/Ca mice challenged with OVA as the control. The microarray analysis was performed in quadruplicate.  Twelve dual channel microarray slides were used in the overall design of this experiment.

first: Damon

middle: J

last: Tumes

middle: R

first: Lindsay

last: Dent


title: Small RNAs subcellular distribution in breast cancer and normal cell lines

status: Public on Jun 16 2009

city: Beijing

first: Tao

middle: Paul

last: Wu

address: Datun Road 15#

department: Systems Biology

country: China

institute: Institute of Biophysics, Chinese Academy of Sciences

summary: We detect the small RNAs subcellular distribution in breast cancer cell lines MCF-7 and MDA-MB-231, and normal cell line MCF-10A.

overall_design: Each cell line, we detected the nuclear and cytoplasmic small RNAs expression intensity; and then we could get the nuclear-cytoplasmic-ratio.

first: Runsheng

last: Chen


title: EST mining of yellow potato and gene expression profiling of its infection by P. infestans using SAGE - SOLEXA

status: Public on Jun 13 2009

city: Bogota

first: Silvia

last: Restrepo

department: Biological Science

country: Colombia

institute: Universidad de los Andes

summary:
The late blight pathogen, Phytophthora infestans has a broad host range within the Solanaceae family, including yellow potato (Solanum phureja). The disease caused by P. infestans in S. phureja is poorly understood and is a major concern in Colombia. Expressed Sequence Tag (EST) libraries obtained from a normalized library constructed from healthy plant tissue revealed high levels of sequence similarity between S. phureja and S. tuberosum. Then, utilizing Serial Analysis of Gene Expression and high-throughput sequencing (SAGE-Solexa), we characterized yellow potato gene expression during infection by P. infestans. Four-week-old yellow potato plants were inoculated with P. infestans and were collected at 12 and 72 hours post inoculation for RNA extraction. We detected differentially expressed genes by comparing inoculated to non-inoculated and resistant to susceptible plants. The discovery and characterization of the proteins mediating this host–pathogen interaction enable the understanding of the pathosystem and is the key for developing resistant plants.

Keywords: SAGE-Solexa, inoculation response, transcript profiling, Solanum phureja, Phytophthora infestans

overall_design: Four-week-old yellow potato (Solanum phureja) plants were inoculated with Phytophthora infestans and were collected and flash frozen in liquid nitrogen at 12 and 72 hours post inoculation, as well as mock inoculated, for RNA extraction. 2 yellow potato cultivars (resistant and susceptible) were used for each experiment. Mock inoculated plants were collected in each replicate. RNA obtained from each of the three biological replicates was pooled to obtain a single RNA sample for each timepoint X cultivar combination. A total of 6 different SAGE libraries were thus obtained. For all libraries, Illumina sequencing was performed at Canada´s Michael Smith Genome Sciences Centre.

first: Roberto

last: Sierra

first: Alejandro

last: Rojas

first: Luis

last: Rodriguez

first: Andres

last: Pinzon

first: Adriana

last: Gonzalez

first: Angela

last: Vargas

first: Sonia

last: Tinjaca

first: Carlos

last: Ñustez

last: Bernal


title: Identification of osmotic stress-responsive genes dependent on HogA and AtfA in Aspergillus nidulans

city: Sendai

first: Daisuke

last: Hagiwara

state: Miyagi

address: 6-6-10 Aoba Aramaki

department: New Industry Creation Hatchery Center

country: Japan

institute: Tohoku university

summary:
Microarray analysis was used to identify the osmotic stress-responsive genes dependent on HogA and AtfA in the filamentous fungus Aspergillus nidulans.
In order to identify such genes, we conducted the several types of experiment. One was a comparison between  wild type with and without osmotic shock (Exp.6). Others were comparison between wild type with osmotic shock and each mutant (hogA, Exp.7; atfA, Exp.8) with osmotic shock.
Compared the result of Exp.6 with other experiments, we could identify the genes whose expression was induced or repressed in response to osmotic stress in a manner dependent on HogA and AtfA.

KEY WORD; Aspergillus nidulans, osmotic stress, HogA, AtfA

overall_design:
Conidia of wild type or each mutant were cultured at 37C in 100ml CD medium containing 2% glucose for 18h, and 50 ml of 3 M sorbitol was added (final concentration; 1 M) or equivalent volume of water (as a control)  for 15 min. The mycelia were harvested and frozen in liquid nitrogen, ground to powder, and used for RNA preparation. mRNA was purified and used for hybridization experiments.

A total of 2 hybridizations were performed for each microarray experiment described in the summary. The following replicates were carried out:
1. In-slide replicates were carried out for each analysis.
2. Dye swap replicates were carried out for each experiment.

The slides were scanned with an Axon GenePix 4000B scanner (Molecular Devices). The resulting TIFF images were imported into GenePix Pro and fluorescent intensity of spots were calculated for each of the Cy3 and Cy5 channels. Global normalization was applied to all analyses. Following normalization, spots whose Cy3 or Cy5 intensity was less than 0 were removed from the data set (the exceptional case was that intensity of the other channel was more than 100). The dye-swap replicates and in-slide replicates were subjected to all analyses. Finally, gene expression ratios (channel 2/channel 1) were calculated for each replicates. Gene expression was considered to be significantly higher or lower whenever the spot intensity changed by at least 2-fold in all four replicates for each experiment.

first: Yoshihiro

last: Asano

first: Junichiro

last: Marui

first: Akira

last: Yoshimi

first: Takeshi

last: Mizuno

first: Keietsu

last: Abe


title: Identification of the Candida albicans CAP1 regulon

city: Memphis

first: Katherine

last: Barker

state: TN

address: 881 Madison Avenue, Room 305

department: Clinical Pharmacy

institute: University of Tennessee Health Science Center

summary: Cap1p, a transcription factor of the basic region-leucine zipper family, controls the oxidative stress response in Candida albicans. It was shown that alteration of the C-terminal cysteine-rich domain (CRD) of Cap1p results in nuclear retention and constitutive transcriptional activation. To further characterize the function of Cap1p in C. albicans, we used genome-wide location profiling (ChIP-on-chip), allowing the identification of Cap1p-transcriptional targets in vivo.  Location profiling using a tiled-oligonucleotide DNA microarray identified 89 targets that were bound by Cap1p-HA3 or Cap1p-CSE-HA3 (binding ratio ≥ 2-fold, P ≤ 0.01). Strikingly, Cap1p binding was not only detected at the promoter region of its target genes but also at their 3'-end and within their open-reading frame. Overrepresented functional groups of Cap1p targets (P ≤ 0.02) included notably 11 genes involved in response to oxidative stress (CAP1, GLR1, TRX1, others), 13 genes involved in response to drug (PDR16, MDR1, FLU1, others) and 3 genes involved in regulation of nitrogen utilization (orf19.2693, orf19.3121 and GST3). Bioinformatic analyses suggested that Cap1p binds to the DNA motif 5'- MTKASTMA. Transcriptome analyses showed that increased expression of most of Cap1p targets accompanies Cap1p binding at these targets, indicating that Cap1p is a transcriptional activator. We conclude that, in addition to protecting the cell against oxidative stress, Cap1p appears to have other functions including drug resistance and the regulation of nitrogen utilization. The atypical binding pattern of Cap1p suggests that this transcription factor may associate with the transcriptional or the chromatin remodeling machinery to exert its activity.

overall_design: We performed three gene expression profile comparisons: (1) benomyl-induced gene expression in a strain containing wildtype CAP1 (CJD21/PMK-CAP1 +/- benomyl), (2) benomyl-induced gene expression in a strain where CAP1 is disrupted (CJD21/PMK +/- benomyl), and (3) gene expression in a strain containing a hyperactive allele of CAP1 compared to a strain containing wildtype CAP1 (CJD21/PMK-CAP1-CSE vs. CJD21/PMK-CAP1).  There were three biological replicates (each drug-treated and untreated strain was grown/treated in three independent experiments).  Since our Affymetrix platform requires each sample is hybridized on a separate chip, there is no dye-swapping.  For each benomyl-treatment experiment (comparisons 1 and 2 above), the untreated (diluent-treated) samples are the reference samples.  For comparison 3, the CJD21-PMK-CAP1 sample is the reference sample.

first: Sadri

last: Znaidi

first: Sandra

last: Weber

first: Anne-Marie

last: Alarco

first: Teresa

last: Liu

first: Genevieve

last: Boucher

first: David

last: Rogers

first: Martine

last: Raymond


title: Tomato root culture and Al treatment

city: Nashville

first: Suping

last: Zhou

address: 3500 John A Merritt Blvd

department: IAGER

institute: Tennessee State University

summary: cDNA microarrays (Tom 1) were used to analyze changes in transcript accumulation in tomato (Solanum lycopersicum cv Money Maker) root cultures under Al-stress (20uM AlK(SO4)2). Out of the 1,362 genes that had significant changes in transcript accumulation (FDR<0.05), 569 were reduced and 793 were induced. The down-regulated genes included those found in the cell cycle (p34cdc2 protein kinase, cyclin, histone), putriscine synthesis, ABC transporter, germin-like protein. Al-induced genes included glutamate synthase, pectin modification, histidine synthesis, proline synthesis, metallothionein-like protein, multidrug efflux proteins (MATE), ethylene biosynthesis and detoxification proteins. Different isoforms in the multiple gene families of V-ATPase, glutathione transferases and others were either induced or repressed.  A wide array of genes for regulating transcription and translation activities, and signal transduction were also affected.

overall_design: Tomato (Solanum esculentum Mill cv Money Maker) seeds were surface sterilized by submerging in 50% bleach (sodium hypochloride) for 10 min followed by three washes with sterile distilled water. For seed germination, six clean seeds were placed in a  500ml flask containing 300 ml Modified Magnavaca’s solution (Magnavaca, 1982) adjusted to pH 4.5 and incubated  on a shaker (150rpm) at 25C for four days. Germinating seeds were then transferred to fresh Modified Magnavaca’s solution supplemented with 20μM AlK(SO4)2. Seedlings serving as controls were placed in Magnavaca’s solution without the Al. Cultures were maintained for 12 additional days, the solution was replaced every four days. At the termination of the experiment, root tips of ~1cm in length were dissected and soaked in pre-chilled RNA extraction buffer. Each replicate of treatment/ control has 20 flasks, and root tips (2cm in length) from the 20 flasks were pooled to make one RNA extraction. Six RNA extraction replicates were preformed for Al treated and control tissues.

first: Roger

last: Sauve

first: Braden

last: Boone

first: Shawn

last: Levy


title: Genomes of DPC5364, DPC5365, DPC5352, DPC5367, DPC5360, DPC5394, DPC5607, CNRZ32, DPC5389 vs DPC4571

status: Public on Jun 12 2009

city: Fermoy

first: Pawel

last: Kaleta

state: Co Cork

address: Moorepark

department: Biotechnology

country: Ireland

institute: TEAGASC

summary: Genomic rearrangement, often driven by insertion sequence (IS) elements, is one of the major processes in the evolution of prokaryotes. Sequence analysis of 16S rRNA of Lactobacillus  helveticus, an organism that evolved in a dairy environment and Lactobacillus acidiophilus an organism that evolved associated with the gastrointestinal tract (GIT) demonstrated 98.4% identity suggesting that they divergently evolved from a common ancestor. Moreover, complete genome sequence analysis of both organisms has demonstrated a remarkable degree of gene synteny (75% homologous genes) despite the presence of an exceptionally high number and diversity of IS elements in the Lb. helveticus genome. Array based comparative genomic hybridization (aCGH) performed on nine strains of Lb. helveticus revealed sixteen clusters of open reading frames (ORFs) flanked by IS elements. Four of these ORFs are associated with restriction/modification which may have played a role in accelerated evolution of strains in a commercially intensive ecosystem undoubtedly challenged through successive phage attack. Furthermore, analysis of the IS-flanked clusters demonstrated that the most frequently encountered IS were also those most abundant in the genome (IS1201, ISL2, ISLhe1, ISLhe2, ISLhe65 and ISLhe63). These findings contribute to the overall viewpoint on a versatile character of IS elements and the role they may play in bacterial genome plasticity.

sample_organism:
Lactobacillus helveticus
Lactobacillus helveticus CNRZ32
Lactobacillus helveticus DPC 4571

overall_design: 2164 ORFs of Lb. helvetiucs DPC4571 spotted on the array in duplicate. Two arrays per each strain.

first: John

last: O`Callaghan

first: Gerald

middle: F

last: Fitzgerald

first: Emma

last: Raftis

first: Paul

last: O`Toole

first: Tom

last: Beresford

last: Ross


title: erwinia chrysanthemi treatments-BOS1 signaling pathway in Arabidopsis thaliana / Erwinia chrysanthemi interaction

city: Evry

first: Frederique

last: Bitton

address: 2 rue Gaston Cremieux

institute: INRA

summary:
ra06-04_bos1 - erwinia chrysanthemi treatments - BOS1 signaling pathway in Arabidopsis thaliana / Erwinia chrysanthemi interaction - Comparison of 4 transcriptoms : Wt Arabidopsis ecotype infected by Wt Erwinia chrysanthemi strain, bos1 Arabidopsis mutant infected by Wt Erwinia chrysanthemi strain, Wt Arabidopsis ecotype infected by BOS1-non inducing pelBC Erwinia chrysanthemi mutant, Wt Arabidopsis ecotype buffer inoculated. Experiments were repeated 3 times.

Keywords: normal vs disease comparison

overall_design: 9 dye-swap - CATMA arrays

first: Jean-Pierre

last: Renou

first: Frédérique

last: Van Gijsege

first: Marie-Laure

middle: L

last: Martin-Magniette


title: Using pooled earthworm RNA to test 244K probes on TA-4 test array design 2520025 antisense 2

city: Vicksburg

first: Ping

last: Gong

state: MS

address: 3909 Halls Ferry Road

department: Environmental Laboratory

institute: US Army ERDC

summary: To validate the 244K 60-mer or 40-mer probes using a custom-designed Agilent oligo array and assuming antisense orientation of all target sequences.

overall_design: A total of four test arrays were designed assuming both sense and antisense orientations. A sole pooled RNA was used for testing all 4 designs. Two arrays were hybridized for each design. This series contains test results for design 2520025.

first: Xin

last: Guan


title: Effect of deletion of cra on E. coli biofilm formation

status: Public on Jun 20 2009

city: Mumbai

first: Shamlan

last: Reshamwala

address: Powai

country: India

institute: Indian Institute of Technology Bombay

summary: Transcriptional analysis of biofilm wild-type and cra mutant E. coli cells was carried out to determine the effect of cra on biofilm formation.

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

overall_design: Samples were prepared in duplicates, with one of the replicates being dye swapped.

first: Santosh

middle: B

last: Noronha


title: Using pooled earthworm RNA to test 244K probes on TA-1 test array design 2520022 sense 1

summary: To validate the 244K 60-mer probes using a custom-designed Agilent oligo array and assuming sense orientation of all target sequences.

overall_design: A total of four test arrays were designed assuming both sense and antisense orientations. A sole pooled RNA was used for testing all 4 designs. Two arrays were hybridized for each design. This series contains test results for design 2520022.


title: Using pooled earthworm RNA to test 244K probes on TA-3 test array design 2520024 sense 2

summary: To validate the 244K 60-mer or 40-mer probes using a custom-designed Agilent oligo array and assuming sense orientation of all target sequences.

overall_design: A total of four test arrays were designed assuming both sense and antisense orientations. A sole pooled RNA was used for testing all 4 designs. Two arrays were hybridized for each design. This series contains test results for design 2520024.


title: Identification of novel candidate target genes at 3q26.2-q29 in squamous cell carcinoma of the lung

status: Public on Jun 19 2009

city: New York

first: Jiun

last: Kang

state: NY

address: 1130 St. Nicholas Avenue

department: Pathology

institute: Columbia University

type: Genome variation profiling by genome tiling array

summary: High-resolution array-CGH was performed to identify differences in the patterns of genomic imbalances between SCC and AC of non-small cell lung cancer (NSCLC). On a genome-wide profile, SCCs showed higher frequency of gains than ACs (p = 0.067). More specifically, statistically significant differences were observed across the histologic subtypes for gains at 2q14.2, 3q26.2-q29, 12p13.2-p13.33, and 19p13.3, as well as losses at 3p26.2-p26.3, 16p13.11, and 17p11.2 in SCC, and gains at 7q22.1 and losses at 15q22.2-q25.2 occurred in AC (P < 0.05).  The most striking difference between SCC and AC was gains at 3q26.2-q29, occurring in 86% (19/22) of SCCs, but in only 21% (3/14) of ACs.

overall_design: Array-CGH was performed to compare the different patterns of genetic alterations. DNA from NSCLC patients (n=36) and human reference DNA (Promega) were differentially labeled and cohybridized on the array. The Fisher exact test utilized two categories, normal and abnormal (loss and gain), with the null hypothesis that the relative proportions of each of the two imbalance categories would be expected to be the same in the groups. The counts of abnormal versus normal were summarized by subtype of NSCLC for each BAC, providing 2x2 tables for analysis. A multiple testing correction (Benjamini-Hochberg false discovery rate (FDR)) was applied to correct for the high number of false positive calls.

first: JiUn

first: SunHoe

last: Koo

first: KyeChul

last: Kwon

first: JongWoo

last: Park

first: JinMan

last: Kim


title: Gene expression profiling reveals novel molecular marker candidates of laryngeal squamous cell carcinoma

city: São Paulo

first: Eduardo

middle: Moraes

last: Reis

state: SP

address: Av. Prof. Lineu Prestes, 748

department: Biochemistry

country: Brazil

institute: University of São Paulo

summary:
Laryngeal squamous cell carcinoma is a very common in head and neck cancer, accounting for 25% of all cases, with high mortality rates and poor prognosis. In this study, we compared expression profiles of clinical samples from15 larynx tumors and 10 non-neoplastic larynx tissue using a custom-built cDNA microarray containing 332 probes for 285 genes previously identified as up- or down-regulated in head and neck tumors by the Head and Neck Annotation Consortium (Reis et al., Cancer Res 65:1693-99, 2005).
Thirty-five genes showed statistically significant differences (SNR ≥|1.0|,p-value ≤ 0.001) in expression between tumor and non-tumor larynx tissue samples. Functional annotation indicated that these genes are involved in cellular processes relevant to the cancer phenotype, such as apoptosis, cell cycle, DNA repair, proteolysis, protease inhibition, signal transduction, and transcription regulation. Six of the identified transcripts map to intronic regions of protein-coding genes and may comprise unannotated exons or yet uncharacterized long ncRNAs with a regulatory role in the gene expression program of larynx tissue.
Differential expression of 10 of these genes (ADCY6, AES, AL2SCR3, CRR9, CSTB, DUSP1, MAP3K5, PLAT, UBL1 and ZNF706) was independently confirmed by quantitative real-time RT-PCR. Among these, the CSTB gene product has cysteine protease inhibitor activity that has been associated to an antimetastatic function. Interestingly, CSTB showed low expression in all tumor samples analyzed (p-value < 0.0001).
The set of genes identified here contribute to a better understanding of the molecular basis of larynx cancer, and at the same time provide novel candidate markers for improving diagnosis, prognosis and treatment of this carcinoma.
Keywords: Gene expression profiling of larynx tumors and non-neoplastic adjacent tissue

overall_design:
Total RNA was isolated from snap-frozen tissue samples from 13 larynx tumors and 10 non-neoplastic larynx tissue.Labeled targets for hybridizations were generated from total mRNA in reverse transcription reactions using oligo-dT primers. The Cy5 (tumor or non-tumor sample) labeled cDNAs were hybridized separately with microarrays on an Automated Slide Processor (ASP, GE Healthcare) and incubated for 16 h at 42ºC. Processed slides were scanned with a 700 V PMT setting. Background-subtracted artifact-removed median intensities of Cy5 emissions were extracted for each spot from raw images, using the ArrayVision V.7.2 software (Imaging Research Inc., Ontario, Canada).
To make the experiments comparable, intensity data derived from Cy5-labeled test samples from different hybridizations were LOWESS normalized by, using as reference the intensity data from a sample with total energy comparable to the average intensity of all samples.


title: Rat liver.  Control vs. Chemical treated, 3 days

city: Osaka

first: Kayo

last: Sumida

address: 1-98, Kasugadenaka 3-chome, Konohana-ku

department: Environmental Health Science Laboratory

institute: Sumitomo Chemical Co., Ltd.

summary:
The final goal of this study is to develop a short term and highly accurate prediction method of carcinogenicity based on gene expression profile of rats administrated by carcinogens.
We conducted 3 days-, 14 days- and 28 days-repeated dose experiments in male F344 rats with 47 carcinogens and 26 non-carcinogens, and the gene expression profiles in liver were investigated by custom glass microarrays.

Supplementary file "73 Compounds for training" indicates 47 carcinogens and 26 non-carcinogens.
Supplementary file "15 Compounds for test" lists the other compounds which are for test.

overall_design: Two-condition experiment, control vs. chemical treated rat liver. Biological replicates: 4 control, 4 treated, control samples were pooled. One replicate per array.

first: Koji

last: Nakayama

first: Hideki

last: Miyaura

first: Hiroshi

last: Matsumoto

first: Fumiyo

last: Saito

first: Koichi

first: Masaru

last: Sekijima

first: Masanori

last: Otsuka

first: Yoshikuni

last: Yakabe

first: Tomoyuki

last: Shirai


title: Using pooled earthworm RNA to test 244K probes on TA-2 test array design 2520023 antisense 1

first: Ping

last: Gong

summary: To validate the 244K 60-mer probes using a custom-designed Agilent oligo array and assuming antisense orientation of all target sequences.

overall_design: A total of four test arrays were designed assuming both sense and antisense orientations. A sole pooled RNA was used for testing all 4 designs. Two arrays were hybridized for each design. This series contains test results for design 2520023.

first: Xin

last: Guan


title: Predator induced gene expression in tadpoles

city: Fujisawa

first: Tsukasa

last: Mori

state: kanagawa

address: Kameino 1866

department: Nihon University College of Bioresource Sciences

institute: Nihon University

summary: Tadpoles of the anuran species Rana pirica can undergo predator-specific morphological responses. Exposure to a predation threat by larvae of the salamander Hynobius retardatus results in formation of a bulgy body (bulgy morph) with a higher tail. The tadpoles revert to a normal phenotype upon removal of the larval salamander threat.  The objective of the present study was to use our own fabricated tadpole Rana pirica cDNA microarray to profile gene expression patterns during the predation threat.

overall_design:
Experimental design used to produce control, bulgy morph and reversion type tadpoles for the microarray analysis was as follows.  One group of tadpoles was placed with a larval salamander for 4 days to induce formation of the bulgy morph phenotype; the predator was then removed and the tadpoles were allowed to revert to the normal phenotype for 4 days.  This group is termed “8 day out tadpoles”.  A second group of tadpoles was placed with the predator for the full 8 days.  The control group was not exposed to a predator.  Tadpoles from the predator induced groups were sampled at 6 hours, 4 days and 8 days, those from the control group at 0 hour, 4 days, and 8 days.  The comparative design of the microarray analysis was performed as (Exp 6 hours VS Cont 0 hours), (Exp 4 days VS Cont 4 days), (Exp 8 days VS Cont 8 days), and (Exp 8 days-Out VS Exp 8 days), respectively.  These analyses were performed in triplicate with a dye swap experiment.
Samples 0hr~8days were as follows.
Sample plate (1~3A):Channel 1-Cy5 Experiment,Channel 2-Cy3 Control.
Sample plate (1~3B):Channel 1-Cy5 Control, Channel 2-Cy3 Experiment.
In case of 8day-out samples were as follows.
Sample plate (1~3A):Channel 1-Cy5 8day-out,Channel 2-Cy3 full 8day.
Sample plate (1~3B):Channel 1-Cy5 full 8day,Channel 2-Cy3 8day-out.

first: Hiroko

last: Kawachia

first: Youich

last: Kurata

first: Manabu

last: Sugiyama

first: Osamu

last: Kishida

first: Kinya

last: Nshimura


title: Analysis of genome rearrangements in S. cerevisiae strains with low levels of DNA polymerase alpha

city: Durham

first: Anne

last: Casper

state: NC

address: 381 CARL Building, Research Drive, Box 3054

department: Department of Molecular Genetics and Microbiology

institute: Duke University

summary: The arrays in this series were used to analyze chromosome rearrangements resulting from the repair of double-strand breaks at fragile site 2 (FS2) on S. cerevisiae chromosome III.  This site is broken under conditions of low DNA polymerase alpha, as described in Lemoine et al., Cell vol. 120, pp. 587-598 (2005).  Here, we have investigated the effect of mutations in Mre11p, Sae2p and Rad52p on the repair of these breaks.

overall_design: In all arrays, total genomic DNA from the experimental strain is labeled with Cy5 and is competitively hybridized to the array with Cy3-labeled total genomic DNA from MS71, an isogenic reference strain.


title: Human Erythroid Day 10 ChIP using GATA, PolII, H3 and H4

city: London

first: Steve

last: Best

address: 125 Coldharbour Lane

department: The James Black Centre

country: United Kingdom

institute: King's College London

summary: A large-scale analysis of histone acetylation as well as RNA polymerase II and GATA 1 interactions on chromosome 6q in human erythroid progenitor cells.

overall_design: Human erythroid progenitors from day 10 erythroid cultures were used for ChIP-chip analysis. For this purpose, the NimbleGen array system was used, following manufacturer's protocols. Samples for ChIP were taken from the same individual and processed for use with each of the following antibodies: GATA1, PolII, AcH3 and AcH4.

first: Karin

last: Wahlberg

first: Jie

last: Jiang

first: Helen

last: Rooks

first: Kiran

last: Jawaid

first: Fumihiko

last: Matsuda

first: Masao

last: Yamaguchi

first: Mark

last: Lathrop

first: Swee-Lay

last: Thein


title: Identification of the fludioxonil-responsive genes dependent on SskA, SrrA, HogA, and AtfA in Aspergillus nidulans

status: Public on Jun 10 2009

first: Daisuke

last: Hagiwara

summary:
Microarray analysis was used to identify the fludioxonil-responsive genes dependent on SskA, SrrA, HogA, and AtfA in the filamentous fungus Aspergillus nidulans.
In order to identify such genes, we conducted the several types of experiment. One was a comparison between  wild type treated with fludioxonil and without the treatment (Exp.1). Others were comparison between wild type treated with fludioxonil and each mutant (sskA, Exp.2; srrA, Exp.3; hogA, Exp.4; atfA, Exp.5) treated with fludioxonil.
Compared the result of Exp.1 with that of other experiments, we could identify the genes whose expression was induced or repressed in response to fludioxonil in a manner dependent on SskA, SrrA, HogA, or AtfA.

KEY WORD; Aspergillus nidulans, fludioxonil, SskA, SrrA, HogA, AtfA

overall_design:
Conidia of wild type or each mutant were cultured at 37C in 100ml CD medium containing 2% glucose for 18h and treated with fludioxonil (final concentration; 10 ug/ml) or DMSO (as a solvent control)  for 15 min. The mycelia were harvested and frozen in liquid nitrogen, ground to powder, and used for RNA preparation. mRNA was purified and used for hybridization experiments.

A total of 2 hybridizations were performed for each microarray experiment described in the summary. The following replicates were carried out:
1. In-slide replicates were carried out for each analysis.
2. Dye swap replicates were carried out for each experiment.

The slides were scanned with an Axon GenePix 4000B scanner (Molecular Devices). The resulting TIFF images were imported into GenePix Pro and fluorescent intensity of spots were calculated for each of the Cy3 and Cy5 channels. Global normalization was applied to all analyses. Following normalization, spots whose Cy3 or Cy5 intensity was less than 0 were removed from the data set (the exceptional case was that intensity of the other channel was more than 100). The dye-swap replicates and in-slide replicates were subjected to all analyses. Finally, gene expression ratios (test/reference) were calculated for each replicates. Gene expression was considered to be significantly higher or lower whenever the spot intensity changed by at least 3-fold in all four replicates for each experiment.

first: Yoshihiro

last: Asano

first: Junichiro

last: Marui

first: Akira

last: Yoshimi

first: Takeshi

last: Mizuno

first: Keietsu

last: Abe


title: Gene expression profiles of human mesenchymal stem cells stimulated with canonical Wnt signaling

status: Public on Jun 14 2009

city: berlin

last: li

address: Robert-Rössle-Straße 10

country: Germany

institute: MAX-DELBRÜCK-CENTRUM FÜR MOLEKULARE MEDIZIN (MDC)

summary: hMSC overexpressing activated or inactivated Wnt co-receptor LRP5 exhibit opposite phenotypes in osteogenesis and adipogenesis. To explore the underlying molecular mechanisms, we compared gene expression profiles upon Wnt3a stimulation and aimed at those genes functionally involved in determination of differentiation fate of hMSC.

overall_design:
Total RNA from hMSC overexpressing activated LRP5 (T253) or inactivated LRP5 (T244) treated with Wnt3a condition medium for 1 hour or 24 hours was compared with those treated with control condition medium. 3 replicates per condition.

The supplementary file 'GSE16186_non-normalized_data.txt' contains non-normalized data for Samples GSM406367-GSM406390.

first: Weimin

last: Qiu

first: Yuhui

last: Hu

last: Li

first: Wei

last: Chen

first: Moustapha

last: Kassem


title: GATA-2 controls megakaryocyte proliferation

city: Chicago

first: Zan

last: Huang

state: IL

address: 303 E Superior St

department: Med

institute: Northwestern University

summary: Downregulation of GATA-2 in megakaryocytic cells G1ME caused cell cycle arrest. To identify GATA-2 target genes, we transduced G1ME cells with control vector or a vector expressing shRNA specific for mouse GATA-2. The transduced cells were purified by sorting and the RNA were used for microarry.

overall_design: We performed experiments for three times. Each group had three samples.


title: Temperature-mediated inactivation of the b heterodimer during biotrophic development of U. maydis

status: Public on Jun 11 2009

city: Karlsruhe

first: Jörg

middle: Thomas

last: Kämper

address: Hertzstrasse 16, Geb. 06.40

department: Department for Applied Biosciences

institute: Karlsruhe Institute of Technology

summary: In fungi, sexual compatibility is controlled by mating type loci that prevent self-fertilization. In the plant pathogenic fungus Ustilago maydis, the b mating type locus encodes a pair of unrelated homeodomain proteins, termed bE and bW. After fusion of two compatible, haploid cells, the bE and bW proteins form a heterodimeric complex, but only if they are derived from different, compatible alleles. The active bE/bW complex is required and sufficient to initiate pathogenic development of U. maydis, which is prerequisite for sexual reproduction of the fungus. However, the role of the b heterodimer during later stages of pathogenic development was unclear. To analyze b function during in planta development, we generated a temperature-sensitive bE allele (bEts) encoding a protein with a single amino acid alteration at the border of the homeodomain. This mutation leads to a stop in pathogenic development at the restrictive temperature in planta, while bEts strains show normal development at permissive temperature. At restrictive temperature, hyphae develop enlarged, bulbous cells at their tips that contain multiple nuclei, indicating a severe defect in cell division. DNA array analysis of bEts mutant strains in planta revealed a b-dependent regulation of genes coding for secreted proteins that were shown to influence fungal virulence. Our data demonstrate that in U. maydis the b heterodimer is not only essential to establish the heterodikaryon after mating of two compatible sporidia and to initiate fungal pathogenicity, but also to sustain in planta proliferation and ensure sexual reproduction.

overall_design: Maize plants were infected with a mixture of either FB1 and FB2 (wild-type), or RAb1ts and RAb2ts (temperature-sensitive b heterodimer) and kept at 22°C (permissive conditions). Control samples of FB1/FB2 and RAb1ts and RAb2ts were taken 5 days post inoculation at 22 °C (1 replicate each), to see if strains expressing the bts heterodimer show wildtype-like biotrophic development. To measure the impact of the b heterodimer on pathogenic development, infected plants were shifted for 9 hours to resrictive conditions (31°C) 111 hours post infection. After the temperature shift FB1/FB2 (3 replicates) infections developed normal, whereas RAb1ts/RAb2ts (3 replicates) infections were not able to further proliferate in planta because of a none-functional b heterodimer.

first: Ramon

last: Wahl


title: Bluestem transcription profiles - Konza 2005

city: Fargo

first: Steve

last: Travers

state: ND

address: 218 Stevens Hall

department: Biological Sciences

institute: North Dakota State University

summary: We measured transcriptional profiles of individuals of Andropogon gerardii, a C4 grass native to North American grasslands, in a field experiment in which both temperature and precipitation have been manipulated to simulate key aspects of forecasted climate change.

overall_design: By using microarrays developed for a closely related model species, Zea mays, we were able to compare the relative influence of warming versus altered soil moisture availability on expression levels of over 7,000 genes.  The plants were located in 12 experimental plots under rainout shelters on the Konza Prairie Biological Station in Manhattan, Kansas.

first: Steven

first: Karen

last: Garrett

first: Melinda

middle: D

last: Smith


title: Tissue-specific variation in DNA methylation levels along human chromosome 1

status: Public on Jun 09 2009

city: Seattle

first: Barbara

middle: J.

last: Trask

state: WA

address: 1100 Fairview Avenue N., C3-168, P.O. Box 19024

department: Division of Human Biology

institute: Fred Hutchinson Cancer Research Center

type: Methylation profiling by genome tiling array

summary: DNA methylation is a major epigenetic modification important for regulating gene expression and suppressing spurious transcription. Most methods to scan the genome in different tissues for differentially methylated sites have focused on the methylation of CpGs in CpG islands, which are concentrations of CpGs often associated with gene promoters. Here, we use a methylation-profiling strategy that is predominantly responsive to methylation differences outside of CpG islands. The method compares the yield from two samples of size-selected fragments generated by a methylation-sensitive restriction enzyme. We profile eight different normal tissues relative to spleen for each of two human donors using a custom array of genomic clones covering the euchromatic portion of human chromosome 1 and representing 8% of the human genome. We observe gross regional differences in methylation states across chromosome 1 between tissues from the same individual, with the most striking differences detected in the comparison of cerebellum and spleen. Profiles of the same tissue from different donors are strikingly similar, as are the profiles of different lobes of the brain. Using DNMT1-deficient cells as a test case, we show that much finer-scale fluctuations in relative fragment yield can be detected when the methylation profiling is performed using a high-resolution oligomer array for chromosome 1. The varied patterns of methylation differences detected between tissues by this method reinforce the potential functional significance of regional differences in methylation levels outside of CpG islands.

overall_design: DNA digested with methylation-sensitive restriction enzyme (HpaII) and size-selected. Eleven human tissue types studied, most from two human individuals, each in triplicate, and one cell line. Two control hybrizations consisting of DNA prepared independently from the same tissue.

first: Cecilia

last: De Bustos

first: Edward

last: Ramos

first: Janet

last: Young

first: Robert

middle: K

last: Tran

first: Uwe

last: Menzel

first: Cordelia

last: Langford

first: Evan

last: Eichler

first: Li

last: Hsu

last: Henikoff

first: Jan

last: Dumanski


title: Disease-specific expression of host genes during downy mildew infection of Arabidopsis

city: Amsterdam

first: Mark

middle: de

last: Jong

address: Kruislaan 318

country: Netherlands

institute: University of Amsterdam

summary: Here we report on the identification of Arabidopsis genes that are induced during compatible but not during incompatible interactions with the downy mildew pathogen Hyaloperonospora arabidopsidis. This set of so-called compatible specific (CS) genes contrasts the large group of defence-associated genes that is differentially expressed during both compatible and incompatible interactions. From the 17 identified CS-genes, 6 belong to the ERF family of transcription factor genes, suggesting that these ERFs have a role during compatibility. The majority of CS-genes are differentially regulated in response to various forms of abiotic stress. In silico analysis of the CS-genes revealed an over-representation of DREB1A/CBF3 binding sites and EveningElement motifs in their promoter regions. The CS-ERFs are closely related to the CBF transcription factors and could potentially bind the DREB1A/CBF3 promoter elements in the CS-genes. Transcript levels of CS-genes peak at 2-3 days post inoculation, when pathogen growth is highest, and decline at later stages of infection. The induction of several CS-genes was found to be isolate-dependant. This suggests that the identified CS-genes could be the direct or indirect targets of downy mildew effector proteins that promote disease susceptibility.

overall_design:
Compatible interaction - Arabidopsis Ler inoculated with Cala2 compared to Ler with mock inoculation with 3 biological replicates. Each biological replicate is analysed by 4 technical dyeswapped replicates.
Incompatible interaction - Arabidopsis Ler inoculated with Waco9 compared to Ler with mock inoculation with 3 biological replicates. Each biological replicate is analysed by 4 technical dyeswapped replicates.

first: Robin

last: Huibers

last: de Jong

first: René

last: Dekter

first: Guido

last: Van den Ackerveken


title: Preliminary exploration of Oryza sativa small RNAs - Run 1 and 2

status: Public on Jun 08 2009

city: Davis

first: Venkatesan

last: Sundaresan

state: CA

address: One Shields Ave

department: Plant Biology

institute: University of California Davis

type: Non-coding RNA profiling by high throughput sequencing

summary:
The small RNAs presented here were produced as a preliminary exploration of small RNAs in rice, and as such, various tissues and stress conditions were sampled.  Small RNAs present in these samples were all mapped to the rice genome TIGR version 5.  The total number of distinct mapped sequences are 12879 for Run 1 and 88508 for Run 2.  The total number of sequence reads were respectively 70406 and 191682.

The datasets contain Oryza sativa var Nipponbar endogenous small RNA sequences in the size range 18 to 34 nt. Plants were grown in a Conviron Environmental Chamber at high light intensity using both high pressure sodium and metal halide lamps for 10.5 hr at 28 degrees C and for 13.5 hr at 26 degrees C in the dark. RNA was extracted from rice tissues at various stages of development and under different abiotic and biotic stresses. The small RNAs presented here were all mapped to the rice genome TIGR version 5. The total number of distinct mapped sequences are 12879 for Run 1 and 88508 for Run 2.  The total number of sequence reads were respectively 70406 and 191682.

overall_design:
Run 1: A small RNA library was generated from whole cell RNA isolated from inflorescence, 30 and 60 day leaves, 10 and 25 day seedlings and from seedling polysomes using Bartel primers (5' ATCGTAGGCACCTGAAA - smRNA - CTGTAGGCACCATCAAT 3').  This was combined with a library derived from whole cell RNA isolated from 30 and 60 day leaves and 10 and 25 day seedlings using Bartel A primers (5' AGCTATCGTAGGCACCTGAAA - smRNA - CTGTAGGCACCATCAATACTG 3').
Run 2: Mixed tissues from unstressed plants. Library prepared from equal amounts of adult root, adult stem and inflorescence from unstressed plants. Amplified with Standard primers (5' ATCGTAGGCACCTGAAA - smRNA - CTGTAGGCACCATCAAT 3').     Seedlings stressed by cold, salt desiccation, heat and dark. Tissues were mixed in a 2:2:1:1:1:1 ratio.  Amplified with A primers (5' AGCTATCGTAGGCACCTGAAA - smRNA - CTGTAGGCACCATCAATACTG 3').     Kinase library. RNA was isolated from tissues used in 'Adult whole plant' (~30%), 'Stressed' (~25%), from unstressed seedlings of various ages (~15%) from 30 and 60 day leaves (20%) and from immature inflorescence (~10%).  RNA isolated from tissues from unstressed plants were treated with polynucleotide kinase before second ligation.  This eliminates the dependence of a 5` phosphate as for all the other libraries.  Amplified with B primers (5' CAGTATCGTAGGCACCTGAAA - smRNA - CTGTAGGCACCATCAATAGCT 3').

first: Cameron

last: Johnson

first: Alex

last: Adai

first: Anna

last: Kasprzewska

first: Kristin

last: Tennessen

first: John

last: John

first: Guo-Ling

first: Virginia

last: Walbot

first: Vicki

last: Vance

first: Lewis

middle: H

last: Lewis


title: VLA lyssavirus genotyping dataset

city: Addlestone, Surrey

first: Akbar

last: Dastjerdi

address: Woodham Lane

department: Virology

institute: Animal and Plant Health Agency

summary:
Data from the VLA lyssavirus genotyping microarray. The array platform for this data is GEO accession GPL8066, and consists of 624 oligos representing two viral families. The data set itself consists of 14 arrays, 7 hybridised with RNA from mice brains infected with 7 genotypes of lyssaviruses, 1 hybridised with RNA from normal mouse brain, and 6 hybridised with RNA from coded samples consisting of infected mouse brains or control mouse brains.
Keywords: Lyssavirus genotyping microarray

sample_organism:
Lyssavirus rabies
Lyssavirus mokola
unidentified
Lyssavirus lagos
Lyssavirus duvenhage
European bat lyssavirus 1
European bat lyssavirus 2
Lyssavirus australis

overall_design: Data from the VLA lyssavirus genotyping microarray. The array platform for this data is GEO accession GPL8066, and consists of 624 oligos representing two viral families. The data set itself consists of 14 arrays, 7 hybridised with RNA from mice brains infected with 7 genotypes of lyssaviruses, 1 hybridised with RNA from normal mouse brain, and 6 hybridised with RNA from coded samples consisting of infected mouse brains or control mouse brains.   Statistical analysis of the data was done with DetectiV software (Watson et al., 2007). The median and array methods of normalization were used in the statistical analysis of the results.  In the median method, DetectiV software calculates the mean fluorescence for each set of probes and normalised against background fluorescence of all probes, assuming that most probes are not hybridized.  The array method utilizes an entire control array, e.g. RNA from a known uninfected animal, as the negative control and all probe values are divided by their respective elements from the control array.

first: R

last: Gurrala

first: A

first: N

first: J

last: Nunez-Garciac

first: S

last: Grierson

first: F

last: Steinbach

first: M

last: Banks


title: H4K12 acetylation in Human Spermatozoa

city: Leeds

first: David

last: Miller

address: Clarendon Way

department: Leeds Institute for Genetics and Health Therapeutics

institute: Leeds University

summary: H4K12 acetylation mapped by chromatin Chromatin immunoprecipitation (ChIP) in human sperm.

overall_design: ChIP mapping of chromatin modification. Three biological replicates hybridized to each of two chips.

first: Ali

last: Arpanahi

first: Martin

last: Brinkworth

last: Iles

first: Stephen

middle: A

last: Krawetz

first: Adrian

middle: E

last: Platts

first: Agnieszka

last: Paradowska

first: Klaus

last: Steger

first: Philip

last: Tedder


title: RNA expression at chromosome 8q24 in human colon

city: Boston

first: Mark

last: Pomerantz

state: MA

address: 44 Binney St.

department: Medical Oncology

institute: Dana-Farber Cancer Institute

type: Expression profiling by genome tiling array

summary: An inherited variant on chromosome 8q24, rs6983267, is significantly associated with cancer pathogenesis. We present evidence that this region is a transcriptional enhancer, that the risk region physically interacts with the MYC proto-oncogene, and that the alleles of rs6983267 differentially bind transcription factor 7-like 2 (TCF7L2). These data provide strong support for a biological mechanism underlying this non-protein coding risk variant.

overall_design: Total RNA and genomic DNA were isolated from colon tissue in 5 individuals.  cDNA and DNA were analyzed on separate tracks, with the genomic DNA used to identify sprurious cross-hybridization

first: Matthew

last: Freedman

first: John

last: Manak


title: Acid- and bile-stress induced trnscriptomes of epidemic and environmental strains of Listeria monocytogenes

city: Pullman

first: Devendra

last: Shah

address: 100 Dairy Rd

department: Veterinary Microbiology and Pathology

institute: Washington State University

summary:
Listeria monocytogenes is a gram-positive, food-borne pathogen responsible for invasive infections with high overall mortality. Early host defenses encountered by L. monocytogenes following ingestion include low pH of the stomach and bile present in the small intestinal lumen. We hypothesized that “epidemic” strains are better able to withstand exposure to low pH and bile encountered in the gastrointestinal tract as compared to most “environmental” strains. Furthermore, we hypothesized that epidemic and environmental strains would have distinct transcriptional programs upon exposure to these conditions. Our treatments included 1 hr exposure to acid (pH 5.5 and 3.5) and bile (0.3%) stress. Strains were pre-exposed to pH 5.5 (1 hr) before being treated with pH 3.5. We used a collection of 12 previously characterized epidemic and environmental strains and each strainXtreatment combination included 3 biological replicates for each microarray experiment. All microarray experiments were two color competitive hybridizations that paired experimental conditions with the same strain at neutral pH for acid stress and pH 5.5 for bile stress. Transcriptomes of environmental strains exposed to acid and bile stress showed remarkably greater number of genes with differences of ≥2-fold expression levels as compared to epidemic strains (5 and 7, respectively). Environmental strains were characterized by up-regulation of several stress related genes and down-regulation of several cell envelope biosynthesis and virulence related genes, suggesting that complex regulatory networks orchestrate the cellular changes in the environmental strains to overcome stressful environments. The transcriptome of epidemic strains, in contrast, showed muted responses to these stress conditions implying their pre-adaptability to acid and bile stress encountered during natural infection that may enable epidemic strains to survive and become “primed” for subsequent colonization and infection in the lower gastrointestinal tract.
Keywords: stress response, comparative transcriptomics, acid-adaptation, differential virulence, acid-stress response, bile-stress response

overall_design: The treatments included 1 hr exposure to acid (pH 5.5 and 3.5) and bile (0.3%) stress. Strains were pre-exposed to pH 5.5 (1 hr) before being treated with pH 3.5. We used a collection of 12 previously characterized epidemic and environmental strains and each strainXtreatment combination included 3 biological replicates for each microarray experiment. All microarray experiments were two color competitive hybridizations that paired experimental conditions with the same strain at neutral pH for acid stress and pH 5.5 for bile stress. Technical replicates were performed for both pH 3.5 acid stress and bile stress.

first: Lisa

last: Orfe

first: Cristina

middle: W

last: Cunha

first: Nairanjana

last: Dasgupta

first: Douglas

last: Call


title: Clara cell regulation of the inflammatory response

status: Public on Jun 04 2009

first: Barry

last: Stripp

address: 106 Research Drive, DUMC Box 103000

department: Medicine

summary: Chronic lung diseases are marked by excessive inflammation and epithelial remodeling. Reduced Clara cell secretory function and corresponding decreases in the abundance of the major Clara cell secreted protein, CCSP, are characteristically seen in these disease states. We sought to define the impact of Clara cell and CCSP depletion on regulation of the lung inflammatory response. We used chemical and genetic mouse models of Clara cell and CCSP deficiency (CCSP-/-) coupled with P. aeruginosa lipopolysaccharide (LPS) elicited inflammation. Exposure of Clara cell depleted or CCSP-/- mice to LPS resulted in augmented inflammation as assessed by polymorphonuclear leukocyte recruitment to the airspace. Gene expression analysis and pathway modeling of the CCSP-/- inflammatory response implicated increased TNF-alpha signaling. Consistent with this model was the demonstration of significantly elevated TNF-alpha in airway fluid of LPS-stimulated CCSP-/- mice compared to similarly exposed wild type mice. Increased LPS-elicited TNF-alpha production was also observed in cultured lung macrophages from CCSP-/- mice compared to wild type mice. We demonstrate that macrophages from Clara cell depleted and CCSP-/- mice displayed increased TLR4 surface expression. Our results provide evidence that Clara cells can attenuate inflammation through regulation of macrophage behavior and suggest that epithelial remodeling leading to reduced Clara cell secretory function is an important factor that increases the intensity of lung inflammation in chronic lung disease.

overall_design: 24 samples analyzed (total lung RNA from Mus musculus), 12 wild-type and 12 CCSP knock-out, samples were from control or LPS exposed (1.5, 6, and 12 hours post exposure). N = 3 per group, for a total of 12 per genotype.

first: Joshua

middle: C

last: Snyder

first: Susan

last: Reynolds

last: Hollingsworth

first: Zhuowei

last: Li

first: Naftali

last: Kaminski


title: Novel and highly recurrent chromosomal alterations in Sézary syndrome

city: Leiden

first: Marloes

last: van Kester

address: Einthovenweg 20

department: Dermatology

institute: Leiden University Medical Center

summary:
This study was designed to identify highly recurrent genetic alterations typical of Sézary syndrome (Sz), an aggressive cutaneous T-cell lymphoma/leukemia, possibly revealing pathogenetic mechanisms and novel therapeutic targets. High-resolution array-based comparative genomic hybridization was done on malignant T cells from 20 patients. Expression levels of selected biologically relevant genes residing within loci with frequent copy number alteration were measured using quantitative PCR. Combined binary ratio labeling-fluorescence in situ hybridization karyotyping was done on malignant cells from five patients. Minimal common regions with copy number alteration occurring in at least 35% of patients harbored 15 bona fide oncogenes and 3 tumor suppressor genes. Based on the function of the identified oncogenes and tumor suppressor genes, at least three molecular mechanisms are relevant in the pathogenesis of Sz. First, gain of cMYC and loss of cMYC antagonists (MXI1 and MNT) were observed in 75% and 40% to 55% of patients, respectively, which were frequently associated with deregulated gene expression. The presence of cMYC/MAX protein heterodimers in Sézary cells was confirmed using a proximity ligation assay. Second, a region containing TP53 and genome maintenance genes (RPA1/HIC1) was lost in the majority of patients. Third, the interleukin 2 (IL-2) pathway was affected by gain of STAT3/STAT5 and IL-2 (receptor) genes in 75% and 30%, respectively, and loss of TCF8 and DUSP5 in at least 45% of patients. In sum, the Sz genome is characterized by gross chromosomal instability with highly recurrent gains and losses. Prominent among deregulated genes are those encoding cMYC, cMYC-regulating proteins, mediators of MYC-induced apoptosis, and IL-2 signaling pathway components. [Cancer Res 2008;68(8):2689–98]
Keywords: aCGH, COBRA, FISH, PLA

overall_design: In this multicenter study, we aimed to obtain a detailed catalogue of numerical and structural chromosomal abnormalities in a group of 20 well-defined Sz patients using high-resolution array-based CGH in combination with combined binary ratio labeling (COBRA), fluorescence in situ hybridization (FISH), and in situ proximity ligation assays (PLA).

first: Maarten

last: Vermeer

first: Remco

last: van Doorn

last: Dijkman

first: Xin

last: Mao

first: Sean

last: Whittaker

first: Pieter

last: van Voorst Vader

first: Marie-Jeanne

middle: P

last: Gerritsen

first: Marie-Louise

last: Geerts

first: Sylke

last: Gellrich

first: Ola

last: Söderberg

first: Karl-Johan

last: Leuchowius

first: Ulf

last: Landegren

first: Jacoba

middle: J

last: Out-Luiting

first: Jeroen

last: Knijnenburg

first: Marije

last: IJszenga

first: Karoly

last: Szuhai

first: Rein

last: Willemze

first: Cornelis

last: Tensen


title: Hybrid Poplar H11-11 Systemic Herbivory Response Dataset 2

status: Public on Jun 03 2009

city: Vancouver

first: Joerg

last: Bohlmann

state: BC

department: Michael Smith Laboratories

country: Canada

institute: University of British Columbia

summary: We investigated the transcriptional response of hybrid poplar (Populus trichocarpa x deltoides) source and sink leaves to simulated herbivory (mechanical wounding plus the application of Malacosoma disstria oral secretions) over a 24 hour time course. Experiments were conducted using clonal trees under greenhouse conditions at the University of British Columbia. We used the 15.5K poplar cDNA microarray platform previously described by Ralph et al. (Molecular Ecology 2006, 15:1275-1297). Differentially expressed genes were determined using three criteria: fold-change between treated and untreated control leaves > 1.5-fold, P value < 0.05 and Q value < 0.05. This study identified > 3,000 differentially expressed genes in response to simulated insect feeding damage, which possess distinct source/sink and treated/systemic patterns.

overall_design: For simulated herbivory analysis, we compared RNA from systemic sink leaves of four individual treated trees versus four individual control trees, with dye balance. We also compared pooled RNA from systemic sink leaves of five treated or five untreated trees, for a total of 8 slides. Based on an earlier study using pooled biological replicates (90 trees) to examine a large combination of leaf types and time points, we selected the peak response group for the simulated herbivory treatment for further investigation using individual tree replicates (systemic sink leaves, 2hrs).

first: Ryan

middle: N

last: Philippe

first: Steven

middle: G

last: Ralph


title: Ovarian Cancer CGH

city: Stanford

first: Jonathan

last: Pollack

address: 269 Campus Drive, CCSR3245A

department: Pathology

institute: Stanford University

summary:
OBJECTIVES: Amplification of the 11q13 locus is commonly observed in a number of human cancers including both breast and ovarian cancer. Cyclin D1 and EMS1 have been implicated as candidate oncogenes involved in the emergence of amplification at this locus. Detailed analysis of the 11q13 amplicon in breast cancer led to the discovery of four regions of amplification suggesting the involvement of other genes. Here, we investigate the role of EMSY, a recently described BRCA2 interacting protein, as a key element of the 11q13 amplicon in ovarian cancer. EMSY maps to 11q13.5 and is amplified in 13% of breast and 17% of ovarian carcinomas. METHODS: EMSY amplification was assessed by fluorescent in-situ hybridization (FISH) in 674 ovarian cancers in a tissue microarray and correlated with histopathological subtype and tumor grade. A detailed map of the 11q13 amplicon in 51 cases of ovarian cancer was obtained using cDNA-array-based comparative genomic hybridization (aCGH). To further characterize the role of EMSY within this amplicon, we evaluated both the amplification profiles and RNA expression levels of EMSY and two other genes from the 11q13 amplicon in an additional series of 22 ovarian carcinomas. : EMSY amplification was seen in 52/285 (18%) high grade papillary serous carcinomas, 4/27 (15%) high grade endometrioid carcinomas, 3/38 (8%) clear cell carcinomas, and 3/10 (30%) undifferentiated carcinomas. aCGH mapping of 11q13 in ovarian cancer showed that EMSY localized to the region with the highest frequency of copy number gain. Cyclin D1 and EMS1 showed a lower frequency of copy number gain. A highly significant correlation between EMSY gene amplification and RNA expression was also observed (P = 0.0001). This was a stronger correlation than for other genes at 11q13 including Cyclin D1 and PAK1. CONCLUSIONS: These findings support the role of EMSY as a key oncogene within the 11q13 amplicon in ovarian cancer.
Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc.

Keywords: Logical Set

overall_design: Using regression correlation

first: Hanna

last: Kim


title: Expression Analysis of Geobacter sulfurreducens During Growth on a Flow-through Potentiostat

city: Amherst

first: Derek

last: Lovley

address: University of Massachusetts at Amherst, Dept. of Microbiology, Morrill IV north room 202

department: Microbiology

institute: University of Massachusetts Amherst

summary:
G. sulfurreducens can generate electricity from the oxidation of organic compounds. This is because it can take electrons from organic compounds and ship them out to the outer surface of the cell where they can then be deposited on various insoluble electron acceptors including electrodes. Cells attatched to the surface of an electrode oxidize acetate and and deposit the electrons derived from acetate onto the surface of the electrode after which they can travel through an electrical circuit, producing a current. Microbial fuel cells powered by acetate oxidation by Geobacter species are called Geobatteries.

In this experiment we compared gene expression in a biofilm of the wild type strain growing on the surface of an electrode within a current-producing Geobattery to gene expression in a wild type biofilm that is not producing current, but is growing on the surface of an electrode.

In both cases, the cells were growing in a flow-through two chambered H-cell Geobattery setup. This consists of two glass chambers, an anoxic anode chamber containing G. sulfurreducens, a graphite electrode, a reference electrode and growth medium and an oxic chamber containing the counter electrode. The two chambers are connected by a cation selective membrane and a wire connected to a potentionstat. A potentiostat is an instrument which maintains the redox potential of the anode at a fixed value relative to a reference electrode. Media continuously flowed through the anoxic anode chamber at a dilution rate of 0.15.

In the experimental condition, the Geobattery was operational. The circuit was closed and G. sulfurreducens attached to the electrode generated current as it oxidized acetate. The redox potential at the anode was poised at 300 mV by the potentiostat. In the control condition, everything was the same, except that the medium in the anode chamber contained fumarate as electron acceptor, and the anode was not hooked up to the potentiostat i.e. the circuit was open. This prevented the anode from serving as an electron acceptor. Nevertheless a thick G. sulfurreducens biofilm grew on the surface of the electrode. The control and experimental geobatteries were harvested when current in the operational/experimental Geobatteries reached 10 mA.
Keywords: two condition comparison

overall_design: Three biological replicates were hybridized in duplicate on 12K Combimatrix sense-detecting arrays, the experiment (10 mA potentiostat) was labeled Cy5, and the control (fumarate biofilm) was labeled with Cy3.

first: Kelly

last: Nevin

first: Sean

middle: F

last: Covalla

first: Jessica

last: Johnson

first: Barbara

last: Methe

first: Trevor

middle: L

last: Woodard

first: Raymond

last: DiDonato

middle: R


title: Gene expression patterns under conditions of oxidative stress in Geobacter uraniumreducens

summary:
Previous analysis of gene transcript levels of Geobacter species in groundwater during in situ
bioremediation of a uranium-contaminated aquifer detected expression of genes encoding
superoxide dismutase (sodA) and cytochrome d ubiquinol oxidase (cydA), proteins known to be involved in the response to oxidative stress in other microorganisms.  In order to further elucidate gene expression patterns that could be attributed to oxygen
exposure, G. uraniumreducens was grown with acetate
as the electron donor and fumarate as the electron acceptor in the presence of oxygen and compared to non-oxygen treated cultures.

overall_design: Three biological replicates were hybridized in duplicate. Experimental (oxygen treated) was labeled with cy5, control (no oxygen) was labeled with cy3.

first: Paula

last: Mouser

first: Dawn

last: Holmes

first: Lorrie

last: Perpetua


title: RNAi knockdown of NMD in D Melanogaster

city: Berkeley

first: Kasper

middle: Daniel

last: Hansen

address: 461 Koshland Hall

department: PMB

institute: UC Berkeley

summary:
Two core factors of the NMD machinery in D. melanogaster, Upf1 and Upf2, were knocked down using RNAi, as described in Rehwinkel et al. (2005) RNA, PMID: 16199763. Each of the two knockdowns were compared to mock RNAi knockdowns as described in Blanchette et al. (2005) Genes Dev, PMID: 15937219 in a dual channel experiment, using a custom splice-junction microarray design, see Blanchette et al. (2005), PMID: 15937219. The aim of the experiment was to identify which isoforms of alternatively spliced genes were affected by NMD knockdown and thereby gain insight into the NMD mechanism in Drosophila.

The samples were hybridized in a dual channel setup, to custom designed Splice-Junction arrays manufactured by Agilent.

overall_design: A total of 6 independent knockdowns (3 Upf1 and 3 Upf2) were generated and hybridized to 6 different arrays. On each array a control sample was hybridized as well. The control sample is a pool of 3 independent mock RNAi knockdowns.

first: Macro

last: Blanchette

first: Ed

last: Green


status: Public on Jun 02 2009

city: Muenster

first: Carsten

last: Müller-Tidow

address: Domagkstr. 3

department: Department of Medicine, Hematology and Oncology

institute: University of Muenster

type: Genome binding/occupancy profiling by array

summary:
Chromatin modifications are increasingly recognized as a key mechanism in cancer. The histone methyl-transferase Enhancer of Zeste, Drosophila, Homolog 2 (EZH2) is the enzymatic subunit of the polycomb PRC2 complex and methylates histone H3K27, thereby, mediating gene silencing. Down-regulation of EZH2 by RNA interference in ET suppressed oncogenic transformation, tumor development and metastasis in a respective mouse model. Further microarray analysis of EZH2 knock down, or functionally linked HDAC-inhibitor treatment revealed an undifferentiated reversible phenotype in ET maintained by EZH2. EZH2 suppression resulted in a generalized loss of H3K27me3 as well as an increase in H3 acetylation levels. In addition, ChIP-Chip assays for H3K27me3 identified genes that had specifically lost H3K27me3 upon EZH2 silencing. These findings suggested that stemness features are preserved via epigenetic mechanisms. Taken together, the genetic EWS-Fli1 translocation is intimately linked to global and gene specific epigenetic alterations in ET biology. EZH2 mediates neuroectodermal and endothelial embryonal tumor stem cell growth and metastasic spread induced by a translocation derived chimeric transcription factor.

Keywords: PNET, Ewing Tumor Epigenetic regulation

overall_design: A673 transiently transfected with siRNA-EZH2 and siRNA-scrambled were harvested and Chromatin-IPs of H3K27me3 or H3 followed by microarray hybridisation were carried out. For detailed procedures see Burdach et al. Epigenetic Maintenance of Stemness and Malignancy in Peripheral Neuroectodermal Tumors by EZH2.

first: Stefan

last: Burdach

first: Stephanie

last: Plehm

first: Rebekka

last: Unland

first: Arndt

last: Borckhardt

first: Martin

middle: S

last: Steage

first: Günther

last: Richter


title: Hybrid Poplar H11-11 Systemic Herbivory Response Dataset 1

first: Joerg

last: Bohlmann

overall_design: A factorial hybridization design was chosen to assess gene expression among the untreated control leaves, and leaves subjected to mechanical wounding plus the application of forest tent caterpillar (Malacosoma disstria) oral secretions. For each tree, the lowest five mature leaves were caged in nylon mesh bags, treated or left untreated as a control. Treated leaves, acropetally adjacent source leaves, and sink leaves at crown were harvested 2, 6 or 24 hours after the initiation of each treatment and total RNA was individually isolated from each leaf sample. For each treatment and time point, equal amounts of total RNA were combined from each of the five biological replicate trees prior to cDNA microarray analysis. Total RNA from treated source and untreated source and sink leaves was compared using a balanced loop consisting of direct and indirect comparisons across treatments and time points, with dye balance, using a total of 54 hybridizations.

first: Ryan

last: Philippe

first: Steven

last: Ralph


title: Characterization of Zur-dependent genes and direct Zur targets in Yersinia pestis

status: Public on Jun 01 2009

city: Beijing

first: Ruifu

last: Yang

address: No.20, DOngdajie, Fengtai

institute: Institute of Microbiology and Epidemiology

summary:
  The zinc uptake regulator Zur is a Zn2+-sensing metalloregulatory protein involved in the maintenance of bacterial zinc homeostasis. Up to now, regulation of zinc homeostasis by Zur is poorly understood in Y. pestis.
  We constructed a zur null mutant of Y. pestis biovar microtus strain 201. Microarray expression analysis disclosed a set of 154 Zur-dependent genes of Y. pestis under zinc rich condition. Real-time reverse transcription (RT)-PCR was subsequently used to validate the microarray data. Based on the 154 Zur-dependent genes, predicted regulatory Zur motifs were used to screen for potential direct Zur targets including three putative operons znuA, znuCB and ykgM-RpmJ2. The LacZ reporter fusion analysis verified that Zur greatly repressed the promoter activity of the above three operons. The subsequent electrophoretic mobility shift assay (EMSA) demonstrated that a purified Zur protein was able to bind to the promoter regions of the above three operons. The DNase I footprinting was used to identify the Zur binding sites for the above three operons, verifying the Zur box sequence as predicted previously in γ-Proteobacteria. The primer extension assay was further used to determine the transcription start sites for the above three operons and to localize the -10 and -35 elements. Zur binding sites overlapped the -10 sequence of its target promoters, which was consistent with the previous observation that Zur binding would block the entry of the RNA polymerase to repress the transcription of its target genes.
  Zur as a repressor directly controls the transcription of znuA, znuCB and ykgM-RpmJ2 in Y. pestis by employing a conserved mechanism of Zur-promoter DNA association as observed in γ-Proteobacteria. Zur contributes to zinc homeostasis in Y. pestis likely through transcriptional repression of the high-affinity zinc uptake system ZnuACB and two alternative ribosomal proteins YkgM and RpmJ2.

overall_design:
The wild-type (WT) Y. pestis strain 201 belongs to a newly established Y. pestis biovar, Microtus, which was thought to be avirulent to humans, but highly virulent to mice. An in-frame deletion of the zur gene was constructed by using one step inactivation method based on the lambda phage recombination in which PCR primers provide the homology to the target gene, as described previously by Datsenko and Wanner. The entire coding region of zur was replaced by a kanamycin resistance (KnR) cassette, which was verified by PCR and DNA sequencing. The resulting mutant strain was referred to as Δzur.
Both the WT strain and the Zur mutant were pre-cultivated at 26 ºC to the middle exponential growth phase (OD620 about 1.0) in TMH medium. The cell cultures were then diluted 1:20 in fresh TMH medium and grown at 26°C until an OD620 of about 1.0. Finally, 5mM ZnCl2 was added into each cell culture to ensure zinc rich conditions. Growth was continued for 30 min at 26°C before harvested for total RNA isolation.
Gene expression profiles were compared between WT and Δzur. RNA samples were isolated from four individual bacterial cultures, as biological replicates, for each strain. The dual-fluorescently (Cy3 or Cy5 dye) labeled cDNA probes, for which the incorporated dye was reversed, were synthesized from the RNA samples, and then hybridized to four separated microarray slides, respectively.


title: Adaptation of Sulfolobus solfataricus P2 to fluctuating O2 concentrations

city: marseille cedex 9

first: Gwenola

last: Simon

address: 163 avenue de Luminy

department: ESIL case 925

institute: IRD

summary: Sulfolobus solfataricus P2 was grown aerobically, with O2 concentrations ranging from 1.5 to 26 % (v/v; gas phase). To gain some insight in control of the respiratory system, transcriptomes of the strain cultivated in different O2 concentrations (1.5 % vs 21 %, 1.5 % vs 26 %) were compared using a DNA microarray approach.

overall_design: Two-condition experiments: 1.5% O2 vs two other O2 concentrations (21 or 26% O2). Biological replicates independently grown. One replicate per array. Dye swaps.

first: Jasper

last: Walther

first: Nathalie

last: Zabeti

first: Yannick

last: Combet-Blanc

first: Richard

last: Auria

first: John

last: van der Oost


title: Clostridium beijerinckii NCIMB 8052 wild-type fermentation time course

city: Urbana

first: Zhen

last: Shi

address: 1206 West Gregory Drive

department: Institute for Genomic Biology

institute: University of Illinois

summary: The Clostridium beijerinckii NCIMB 8052 wild-type culture was monitored from exponential growth to stationary phase. During this period the culture underwent a shift from acidogenesis to solventogenesis. Acetone and butanol production was initiated with the onset of the solventogenic phase. Using DNA microarray changes in gene expression were examined during the transitional period.

overall_design: RNA samples were taken from Clostridium beijerinckii NCIMB 8052 wild-type fermentation culture at individual time points during the acidogenic phase and the solventogenic phase. The samples were used for microarray hybridization.

first: Hans

last: Blaschek


title: Chilling requirement fulfillment in Vitis riparia overwintering buds

city: Brookings

first: Kathy

last: Mathiason

state: SD

address: NPB 201 Box 2140A

department: Horticulture, Forestry, Landscape, Parks

institute: South Dakota State University

summary:
This study investigated changes in gene expression of controlled environment chilled (4C) grape overwintering buds as they accumulated from 0 to 2000 chilling hours.
Keywords: time course, chilling, endodormancy release, axillary bud, grape

platform_organism:
Vitis vinifera
Vitis riparia

overall_design: A loop design with 3 biological replicates (RNA from buds collected at 0, 500, 1000, 1500, and 2000 hr of chilling in 2002, 2004, and 2005).

first: Anne

middle: Y

last: Fennell


title: Transcriptional responses of Mycobacterium tuberculosis to lung surfactant

city: Ithaca

first: Ute

middle: Elisabeth

last: Schwab

department: Microbiology-Immunology

institute: Cornell University Veterinary School

summary: This study uses microarray analyses to examine gene expression profiles for Mycobacterium tuberculosis (Mtb) induced by exposure in vitro to bovine lung surfactant preparations that vary in apoprotein content: (i) whole lung surfactant (WLS) containing the complete mix of endogenous lipids and surfactant proteins (SP)-A, -B, -C, and -D; (ii) extracted lung surfactant (CLSE) containing lipids plus SP-B and -C; (iii) column-purified surfactant lipids (PPL) containing no apoproteins, and (iv) purified human SP-A. Exposure to WLS evoked a multitude of transcriptional responses in Mtb, with 52 genes up-regulated and 23 genes down-regulated at 30 min exposure, plus 146 genes up-regulated and 27 genes down-regulated at 2 h.  Notably, WLS rapidly induced several membrane-associated lipases that presumptively act on surfactant lipids as substrates, and a large number of genes involved in the synthesis of phthiocerol dimycocerosate (PDIM), a cell wall component known to be important in macrophage interactions and Mtb virulence. Exposure of Mtb to CLSE, PPL, or purified SP-A caused a substantially weaker transcriptional response (≤ 20 genes were induced) suggesting that interactions among multiple lipid-protein components of WLS may contribute to its effects on Mtb transcription.

overall_design:
Whole lung surfactant (WLS) was obtained by centrifugation of lavage fluid (0.15M NaCl) from the intact lungs of freshly killed calves (ONY, Inc, Amherst, NY). Centrifugation was done at 12,000 x g at room temperature to pellet the active large aggregate surfactant fraction. CLSE was prepared by extraction of WLS into chloroform-methanol by the method of Bligh and Dyer (1959), as described in our prior work (Notter et al. 2002). CLSE is conceptually identical to the substance of the clinical surfactant Infasurf®. The composition of CLSE by weight was approximately 94% phospholipid, 4.5% cholesterol, and 1.5% protein (a mixture of SP-B and SP-C) as defined previously (Notter et al. 2002). Mixed lung surfactant phospholipids (PPL) were isolated from CLSE by column chromatography with an elution solvent of 1:1 (v/v) chloroform-methanol plus 5% 0.1 N HCl. Two passes through a Sephadex LH-20 column (Pharmacia-LKB Biotechnology, Piscataway, NJ) separated phospholipids away from hydrophobic surfactant apoproteins and neutral lipids. Acid remaining in PPL fractions was removed by a second chloroform-methanol extraction. Final PPL had a protein content below the limits of detection of both the Folin-phenol assay of Lowry et al. (1951) and the amido black assay of Kaplin and Pedersen (1989) modified by the addition of 15% sodium dodecyl sulfate (SDS) to allow accurate quantitation in the presence of lipid.  Human SP-A was a gift from Dr. J. R. Wright at Duke University, and was isolated from bronchoalveolar lavage from patients with alveolar proteinosis as described previously (Wright et al. 1987).
For exposure studies with Mtb, amounts of WLS, CLSE, and PPL used per experiment were 3.3 mg in a total volume of 4 ml. In preliminary studies, this concentration was shown to induce a transcriptional response in Mtb without affecting viability after a 2 h treatment as determined by serial dilutions compared to untreated Mtb.   SP-A was studied at a level of 0.165 mg based on its putative concentration of approximately 5% in endogenous surfactant.
To determine the effects of lung surfactant on the transcriptional response of Mtb, preparations of WLS, CLSE, or PPL (in 650 µl PBS) were initially coated onto the bottom surface of a 25 cm2 cell culture flask for 30 min at 37oC to mimic a surfactant layer. A minimum of two independent biological replicates were studied for each surfactant preparation. Purified human SP-A was also examined for its effects on Mtb transcription, and phosphate buffered saline (PBS) was used as a “no treatment” control. In experiments with SP-A (in 650 µl PBS), the pure apoprotein was either pre-coated on the flask as done with the surfactant preparations, or was directly added and incubated with Mtb during study. Mtb was added to the flask in 3.35 ml of DMEM (supplemented with 2% FCS and 5 mM HEPES) and exposed to coated surfactant/SP-A or PBS control for 30 min or 2h at 37oC, followed by RNA isolation as detailed by Rohde et al. (2008). GTC lysis/stabilization buffer was added, and pelleted Mtb were washed and then digested with lysozyme. TRIzol (Invitrogen) was added, and organisms were disrupted using glass beads in a Mini-Bead Beater (BioSpec Products, Inc. Bartlesville, OK). After extraction of Mtb lysates with chloroform, ethanol was added to the aqueous phase before loading samples onto RNeasy mini columns (Qiagen, Valencia, CA). Subsequent steps were then performed according to the manufacturer’s instructions. Following elution with RNase-free water, the RNA samples were treated with DNase (Turbo DNAfree, Applied Biosystems, Foster City, CA), and stored at -70oC until analysis.
The MessageAmpTMII-Bacteria system (Applied Biosystems) was used to amplify mycobacterial RNA for all samples [16]. Total RNA was reverse transcribed to yield double-stranded amino-allyl modified aRNA, which was labeled with Alexa Fluor 555 and Alexa Fluor 647 (Invitrogen). Excess reactive dyes were removed with a Mega Clear purification kit (Applied Biosystems).  Next, Alexa-labeled aRNA from paired samples was dried using a Speedvac, and re-suspended in hybridization buffer (5x SSC, 25% formamide, 0.15% SDS, and 25 μg salmon sperm DNA). The mixed samples were denatured by being heated to 95oC for and then cooled to 50oC prior to loading them onto microarrays.  The arrays were hybridized for approximately 15 h at 45oC, washed and then dried by centrifugation.  Fluorescence intensity data from each array were collected with a GenePix 4000B scanner (Axon Instruments, Union City, CA) using GenePixPro3.0 software (Axon Instruments).  Image analysis, i.e. spot identification, background measurements, quality assessment and flagging were conducted with Imagene software (Version 6.0, BioDiscovery, Inc., El Segundo, CA).  Poor quality spots were filtered and removed from the analysis.  Subsequent normalization, statistical analysis (i.e. t-test), and visualization of array data was conducted using Genespring (Version 7, Agilent Technologies, Palo Alto, CA).

first: Kyle

last: Rohde

first: Zhengdong

last: Wang

first: Patricia

last: Chess

first: Robert

last: Notter

first: David

last: Russell


title: Transcriptomic computational analysis of mastic oil-treated Lewis lung carcinomas

city: Athens

first: Fragiskos

last: Kolisis

address: 48 Vasileos Constantinou ave.

department: Institute of Biological Research and Biotechnology

country: Greece

institute: National Hellenic Research Foundation

summary: Mastic oil from Pistacia lentiscus variation chia, a blend of bioactive terpenes with recognized medicinal properties, has been recently shown to exert anti-tumor activity. Lewis lung carcinoma (LLC) cells are mastic oil-susceptible cells and were used in this work to study the effects of mastic oil at the transcriptomic level.

overall_design:
Expression profiling by array.
LLC cells were treated with mastic oil or DMSO (control) for 3h, 6h, 12h, 24h, or 48h. Two replicates per treatment/time point. Data were normalized using rank invariant and quantile normalization.

first: Olga

last: Papadodima

first: Panagiotis

last: Moulos

first: Aristotelis

last: Chatziioannou

first: Heleni

last: Loutrari

first: Charis

last: Roussos


title: Expression of lignin-degrading enzymes in soils using targeted microarrays

city: Richland

first: Katrina

middle: M

last: Waters

address: 902 Battelle Blvd; MSIN J4-18

department: Biological Sciences Division

institute: Pacific Northwest National Laboratory

summary:
Microarrays have become established tools for describing microbial systems, however the assessment of expression profiles for environmental microbial communities still presents unique challenges.  Notably, the concentration of particular transcripts are likely very dilute relative to the pool of total RNA, and PCR-based amplification strategies are vulnerable to amplification biases and the appropriate primer selection.  Thus, we apply a signal amplification approach, rather than template amplification, to analyze the expression of selected lignin-degrading enzymes in soil.  Controls in the form of known amplicons and cDNA from Phanerochaete chrysosporium were included and mixed with the soil cDNA both before and after the signal amplification in order to assess the dynamic range of the microarray.  We demonstrate that restored prairie soil expresses a diverse range of lignin-degrading enzymes following incubation with lignin substrate, while farmed agricultural soil does not.  The mixed additions of control cDNA with soil cDNA indicate that the mixed biomass in the soil does interfere with low abundance transcript changes, nevertheless our microarray approach consistently reports the most robust signals.

Keywords: comparative analysis, microbial ecology, soil microbial communities

sample_organism:
synthetic construct
unidentified
Phanerodontia chrysosporium

platform_organism:
Cryphonectria parasitica
Neurospora crassa
Phlebia radiata
Pleurotus ostreatus
Pleurotus eryngii
Trametes versicolor
Schizophyllum commune
Agaricus bisporus
Lentinula edodes
Fusarium oxysporum
Trametes cinnabarina
Heterobasidion annosum
synthetic construct
Botrytis cinerea
Gelatoporia subvermispora
Coriolopsis gallica
Dichomitus squalens
Trametes coccinea
Fusarium proliferatum
Phanerodontia chrysosporium

overall_design: We used lignin degradation as a model process to demonstrate the use of an oligonucleotide microarray for directly detecting gene expression in soil communities using signal amplification instead of template amplification to avoid the introduction of PCR bias. In the current study, we analyzed mRNA isolated from two distinct soil microbial communities and demonstrate our ability to detect the expression of a small subset of lignin degrading genes following exposure to a lignitic substrate.  We also included purified control amplicons and mixed target experiments with pure P. chrysosporium genomic cDNA to determine the level of interference from soil biomass on target hybridization.

first: Vanessa

last: Bailey

first: Sarah

last: Fansler


title: Gene expression patterns in the head horns, thoracic horns, and legs of the horned beetles Onthophagus taurus

city: Bloomington

first: Justen

last: Andrews

state: IN

address: 915 East Third Street

department: Biology

institute: Indiana University

summary: This series examines gene expression patterns in the head horns, thoracic horns, and legs of the horned beetles Onthophagus taurus.  Expression in each of these tissues was compared to that in common non-appendage reference - abdominal epithelium.

overall_design: The series consists of three pair-wise comparisons: head horn versus abdominal epithelium, thoracic horn versus abdominal epithelium and legs versus abdominal epithelium.  Each tissue sample was obtained by pooling tissue dissected from four pupae.  Samples compared on the same array were derived from tissues dissected from the same four animals.  Five independent biological replicates were performed for each comparison with dye flips (three in one direction and two in the opposite direction).

first: Teiya

last: Kijimoto

first: Armin

last: Moczek


title: RNAcompete: a method for rapid and systematic analysis of the RNA recognition specificities of RNA-binding proteins

status: Public on Jun 07 2009

city: San Francisco

first: Esther

middle: T

last: Chan

state: California

address: 1500 Owens Street, Suite 500

institute: Celgene Corporation

summary: Metazoan genomes encode hundreds of RNA binding proteins (RBPs) but relatively few have well-defined RNA-binding preferences. Current techniques for determining RNA targets, including those involving in vitro selection and RNA co-immunoprecipitation, require significant time and labour investment. Here we introduce RNAcompete, a new method for the systematic analysis of RNA-binding specificities that employs a single binding reaction to determine the relative preferences of RBPs for short RNAs that containing a complete range of k-mers in structured and unstructured RNA contexts. We tested RNAcompete by analyzing nine diverse RBPs (HuR, Vts1, FUSIP1, PTB, U1A, SF2/ASF, SLM2, RBM4, and YB1). RNAcompete identified both expected and previously unknown RNA binding preferences. Using in vitro and in vivo binding data, we demonstrate that preferences for individual 7-mers identified by RNAcompete are a more accurate representation of binding activity than conventional motif models. We anticipate that RNAcompete will be a valuable tool for the large-scale study of RNA-protein interactions.

overall_design: The bound RNA from each RNA binding protein pulldown assay is analyzed on a custom Agilent microarray using a pool RNA control as a reference.

first: Debashish

last: Ray

first: Hilal

last: Kazan

first: Lourdes

last: Pena-Castillo

first: Sidharth

last: Chaudhry

first: Shaheynoor

last: Talukder

first: Benjamin

last: Blencowe

first: Quaid

middle: D

last: Morris

first: Timothy

last: Hughes