title: Swimming down-regulates ovarian transcriptomic response of rainbow trout Oncorhynchus mykiss

status: Public on Feb 05 2010

city: Barcelona

first: Arjan

last: Palstra

address: Av. Diagonal 645

institute: Universitat de Barcelona

summary: Metabolic processes and sexual maturation closely interact during the long-distance reproductive migration of many fish species to their spawning grounds. In the present study, we have for the first time used exercise experimentally to investigate the effects on sexual maturation in rainbow trout. Pubertal autumn-spawning seawater-raised female rainbow trout Oncorhynchus mykiss (n=26; 50-cm, 1.5-kg) were rested or swum at a near optimal speed of 0.75 body-lengths per second in a 6,000 L swim-flume under natural reproductive conditions (16 °C fresh-water, starvation, 8h-light:16h-dark photoperiod). Fish were sampled after arrival and subsequently after 10 days (resting or swimming 307 km) and 20 days (resting or swimming 636 km). Ovarian development was significantly reduced in the swimmers. Analysis of the expression of key factors in the reproductive axis included pituitary kiss1-receptor, lh and fsh and ovarian lh-receptor, fsh-receptor, aromatase and vitellogenin-receptor (vtgr). Swimmers had lower pituitary lh and ovarian vtgr expression than resters. Furthermore, the number of late vitellogenic oocytes was lower in swimmers than in resters, probably resulting from the lower vtgr expression, and vitellogenin plasma levels were higher. Therefore, swimming exercise suppresses oocyte development possibly by inhibiting vitellogenin uptake. Transcriptomic changes that occurred in the ovary of exercised fish were investigated using a salmonid cDNA microarray platform. Protein biosynthesis and energy provision were among the sixteen functional categories that were all down-regulated in the ovary. Down-regulation of the transcriptomic response in the ovary illustrates the priority of energy reallocation and will save energy to fuel exercise. A swimming-induced ovarian developmental suppression at the start of vitellogenesis during long-term reproductive migration may be a strategy to avoid precocious muscle atrophy.

overall_design:
In order to simulate the natural reproductive conditions of anadromous salmonids, experiments were performed with sea water-raised rainbow trout, Oncorhynchus mykiss. Resters and swimmers were starved during the experiment, seawater was replaced by fresh water and photoperiod was changed. These changes were expected to affect reproductive parameters but as ‘background’ effect because both resters as swimmers were experiencing these conditions at the same time in the same set-up. Any additional effects in swimming fish would thus be caused by swimming only.
Pubertal autumn spawning rainbow trout (n=26; ~50 cm, ~1.5 kg) were purchased from a Danish exporter (Frederiksvaerk Aleexport) where they had been raised for 2 years in freshwater followed by 4 months in sea water cages at 10 ‰. They were transferred by truck within 16 h to Spakenburg (Spakenburg Paling, The Netherlands) and subsequently in the same water in a similar holding tank to the swim-flume at Leiden University (The Netherlands). The next day, 5 randomly chosen fish were sampled as ‘start’-group while others were randomly divided into a ‘rest’-group (n=10) and a ‘swim’-group (n=11). During the following 4 days, the brackish 10 ‰ water was stepwise replaced by freshwater at 16 °C and photoperiod was changed from 16L:8D to 8L:16D. Fish were then acclimatized for two days to their new conditions. Subsequently, swimmers were first swum at a speed of 0.33 body-lengths per second (BL/s) which was increased the next day to the final, near optimal speed of 0.75 BL/s (0.34 m/s or 29.4 km/day). Fish were sampled after 10 days (5 resters and 5 swimmers) and 20 days (5 resters and 6 swimmers).
At sampling, fish were anesthetized using oil of cloves that was commercially purchased from a drugstore (diluted 1:10 in ethanol and used at a dosage of 1.5 ml/l). Fish were euthanized by decapitation and dissected for the ovary which was flash frozen in liquid nitrogen.
RNA was isolated from pituitary and ovary samples according to the TRIZOL reagent protocol by Invitrogen (Baro, Spain). Isolated RNA was DNAse treated using RQ1 DNAse Promega (Madison, USA) and reverse transcribed using Superscript IIITM (Baro, Spain) according to the manufacturer’s protocol.
Microarray analyses were performed on ovarian tissue for 20-days-swimmers vs. 20-days-resters. A rainbow trout cDNA microarray platform was used that was previously validated and described by Koskinen et al., 2004ab, Krasnov et al., 2005, MacKenzie et al., 2006ab, 2008 and Djordjevic et al., 2009, and deposited in GEO under accession number GPL1212, the original cDNA microarray (SFA2.0 chip), and GPL6154; the updated 1.8 K platform. This platform compromises 1818 genes representing 366 functional categories. Total RNA was extracted from flash frozen samples from experimental fish of the 20-days-swim group (n=5) and 20-days-rest group (n=5) like described before. Total RNA corresponding to pooled samples from the swim or rest groups was labeled with Cye3 and Cye5-dCTP (GE Healthcare, Barcelona, Spain) using SuperScript III reverse transcriptase (Invitrogen, Baro, Spain) and oligo(dT) primer. In our experience, reverse labeling combined with multiple replication of spots provide robust normalization and high statistical power of analyses. cDNA was purified with MicroconYM30 (Millipore). The slides were pretreated with 1% BSA(fraction V), 20 × SSC, 10% SDS (30 min at 50°C) and washed with 2 × SSC (3 min) and 0.2 × SSC (3 min) and hybridized overnight in cocktail containing 50 × Denhardt's, 20 × SSC, 10% SDS, 10μg/μl polyadenylate and 10 μg/μl yeast tRNA. All chemicals were from Sigma-Aldrich. Scanning was performed with ScanArray 5000 and images were processed with QuantArray (GSI Luminomics). The measurements in spots were filtered by criteria I/B ≥ 3 and (I-B)/(SI + SB) ≥ 0.6, where I and B are the mean signal and background intensities and SI, SB are the standard deviations. After subtraction of mean background, LOWESS normalization was performed. To assess differential expression of genes, the normalized log intensity ratios (expression ratio) were analyzed with Student's t-test (P <0.05).
Validation of the microarray was performed by Q-PCR of 6 differentially expressed genes (DEGs) as described above. The used primers were either published (Alpha-globin I-1: Donate Jimeno, 2008) or designed, again by using the Genamics software.

first: Diego

last: Crespo

first: Guido

middle: E

last: van den Thillart

first: Josep

middle: V

last: Planas


title: Mechanism of sensitivity and resistance to treatment with an TfR1 antibody fusion protein in B-cell malignancies

status: Public on Feb 09 2010

city: Los Angeles

first: Manuel

last: Penichet

state: CA

department: Department of Surgery

country: USA

institute: University of California, Los Angeles

summary:
Transferrin receptor 1 (TfR1) is an attractive target for cancer immunotherapy due to its overexpression and central role in cancer pathology. Although antibodies targeting TfR1 have shown promise against malignancies, their efficacy is variable.
The current work aims to elucidate the mechanism of sensitivity and resistance to treatment with an anti-TfR antibody fusion protein (ch128.1Av) using as models two malignant B-cell lines: the sensitive cell line IM-9 and the tolerant cell line U266.

Keywords: Time course analysis

overall_design: Duplicate samples of total mRNA from 2x10e5 IM-9 and cell line: U266 (tolerant) were collected prior treatment (time 0h) and after incubation with 10 nM ch128.1Av or buffer for 1h, 3h, 9h, and 24h

first: Gustavo

last: Helguera

first: José

middle: A

last: Rodríguez

first: Tracy

middle: R

last: Daniels

first: David

last: Casero

first: Elizabeth

last: Ortiz-Sánchez

first: Giovanni

last: Coppola

first: Matteo

last: Pellegrini

first: Otoniel

last: Martínez-Maza

middle: L


title: ArrayCGH mapping of chromosome 3 aberrations in lung Squamous Cell Carcinoma (SCC)

status: Public on Feb 10 2010

city: ILLKIRCH

first: Thomas

last: Hussenet

address: 1 rue Laurent Fries

department: Cancer Biology Department

country: France

institute: IGBMC

type: Genome variation profiling by genome tiling array

summary:
To prospectively identify new oncogenes implicated in lung Squamous Cell Carcinoma pathogenesis, we investigated chromosome 3 aberrations in advanced tumours using arrayCGH. These aberrations are indeed among the most frequent aberrations in lung SCC and correlate with SCC patient's poor prognosis.
We precisely map regions of recurrent losses at 3p and gains at 3q25-qter in a series of lung SCC. We moreover uncover 3q26.3-q27 high level amplifications in 20% of tumours.

Keywords: ArrayCGH, Lung Squamous Cell Carcinoma, 3q26.3, SOX2

overall_design:
Profiling of 26 advanced (stage III) lung SCC.
Replicates : each tumor sample is hybridized together with a normal dna sample to one microarray. Each microarray contain 3 replicates per BAC clone.

first: Stanislas

last: du Manoir


title: IMPACT OF FUNGAL SECONDARY METABOLITES ON TRANSCRIPT REGULATION OF FOLSOMIA CANDIDA

status: Public on Feb 03 2010

city: Amsterdam

first: Thierry

last: Janssens

address: de Boelelaan 1085

department: Ecological Sciences

country: Netherlands

institute: VU  University Amsterdam

summary: Genetic and molecular evidence to support the hypothesis that fungal secondary metabolites play a significant role in protecting the fungi against fungivory is scarce. We investigated the impact of fungal secondary metabolites on transcript regulation of stress related expressed sequence tags (ESTs) of the Collembola Folsomia candida feeding on mixed vs. single diets. Aspergillus nidulans wildtype (WT; Ascomycota) able to produce secondary metabolites including sterigmatocystin (ST) and a knockout mutant with reduced secondary metabolism (A. nidulans ΔLaeA) were combined with the high quality fungus Cladosporium cladosporioides as mixed diets or offered as single diets. We hypothesized that (i) A. nidulans WT triggers more genes associated with stress responses compared to the A. nidulans ΔlaeA strain with suppressed secondary metabolism, (ii) C. cladosporioides causes significantly different transcript regulation than the A. nidulans strains ΔlaeA and WT, and (iii) mixed diets will cause significantly different transcript expression levels than single diets. All three hypotheses are generally supported despite the fact that many functions of the affected ESTs are unknown. The results bring molecular evidence for the existence of a link between fungal secondary metabolites and responses in springtails supporting the hypothesis that fungal secondary metabolites act as a shield against fungivory.

overall_design: Twenty-three day old Folsomia candida were fed ad libitum for five days to fungal cuts respectively Cladosporium cladosporoides, Aspergillus nidulans WT, Aspergillus nidulans ΔLaeA and two mixed diets of C.cladosporoides/A. nidulans WT (mix 1) and C. cladosporoides/A. nudlans ΔLaeA (mix2) respectively. Four biological replicates were used for every treatment and a dye swap was used with the Cy3/Cy5 labels. This resulted in 20 samples which were analysed in 10 hybridisations executed in an interwoven loop design. The C. cladosporoides diet was used as the reference in the data analysis.

middle: K

first: Swantje

last: Staaden

first: Bauke

last: Ylstra

first: Janine

last: Mariën

first: Dick

last: Roelofs

first: Nico

middle: M

last: van Straalen


title: The microstructural, mineralogical and transcriptional developments of shell biomineralization of Pinctada maxima

city: Pacific Grove

first: Luke

last: Gardner

state: California

address: 120 Ocean View Blvd, Hopkins Marine Station

institute: Stanford University

summary:
Molluscan larval ontogeny is a highly conserved process typical of 3 principal developmental stages. A characteristic unique to each of these stages is shell design, termed prodissoconch I, prodissoconch II and dissoconch. These shells vary in morphology, mineralogy and microstructure. The discrete temporal transitions in shell biomineralization between these larval stages are utilized in this study to investigate transcriptional involvement in several distinct biomineralization events. Scanning electron microscopy and X-ray diffraction analysis of P. maxima larvae and juveniles collected throughout post-embryonic ontogenesis, document the mineralogy and microstructure of each shelled stage as well as establishing a timeline for transitions in biomineralization. P. maxima larval samples most representative of these biomineralization distinctions and transitions were analyzed for differential gene expression on the microarray platform PmaxArray 1.0. A number of transcripts are reported as differentially expressed in correlation to the mineralization events of P. maxima larval ontogeny. Some of those isolated are known shell matrix genes while others are novel, these are discussed in relation to potential shell formation roles. This interdisciplinary investigation has married the shell developments of P. maxima larval ontogeny with corresponding gene expression profiles, furthering the elucidation of shell biomineralization.

Keywords: Temporal expression profiling by array

overall_design: Microarray is used to examine the temporal differential expression of transcripts from several bivalve larval development stages including 24hrs post fertilization, 3 days, 17 days, 20 days, 23 days, 26 days, 30 days, 35 days, 40 days. Differential expression profiles for transcripts of all the temporal samples was determined based on comparison to a common reference of unfertilized eggs. Each temporal larval sample included in the study has at least 3 replicate hybridizations. Dye flips have been incorporated in the replicates. A total of 46 microarray hybridizations were performed in this investigation for differential expression analysis.

middle: D

first: Abigail

last: Elizur


title: Classifier analysis of gene expression profiles reveals a discriminatory gene set for soil containing different metals

first: Benjamin

last: Nota

state: Noord Holland

address: Plesmanlaan 125

department: Research Facilities

institute: Sanquin

summary: Environmental pollution is a worldwide problem, and metals are the largest group of contaminants in soil. Microarray toxicogenomic studies with ecologically relevant organisms such as springtails, supplement traditional ecotoxicological research, but are presently rather descriptive. Classifier analysis, a more analytical application of the microarray technique, is able to predict biological classes of unknown samples. We used the uncorrelated shrunken centroid (USC) method to classify gene expression profiles of the springtail Folsomia candida exposed to soil spiked with six different metals (barium, cadmium, cobalt, chromium, lead, and zinc). We identified a gene set (classifier) of 188 genes that can discriminate between six different metals present in soil, which allowed us to predict the correct classes for samples of an independent test set with an accuracy of 83% (error rate = 0.17). This study shows further that in order to apply classifier analysis to actual contaminated field soil samples, more insight and information is needed on the transcriptional responses of soil organisms to different soil types (properties) and mixtures of contaminants.

overall_design: Gene expression was measured in springtails after exposure of 2 days to soil containing either EC10 or EC50 of 6 different metals. The exposure experiment was performed in two separate series (1 and 2), both containing a separate non-spiked (LUFA 2.2) soil control. Also, two field soil samples were tested. The samples were divided into a separate training set and a validation set for USC classifier analysis.

first: Rudo

last: Verweij

first: Douwe

last: Molenaar


title: Transcriptional responses of Folsomia candida exposed to polar and apolar narcotizing compounds.

summary: Narcosis or baseline toxicity is the inert toxicity of hydrophobic compounds, supposed to take place at the level of the cellular membranes. Based on the linear relationship between the  toxicity (logEC50) and the hydrophobicity (logKow), class I and II narcotizing compounds are  recognized, in which the latter group is assumed to exert additional toxic mechanisms by  hydrogen bond donor acidity by their polar groups. Chlorinated anilines, which occur  often in the environment as degradation products of certain pesticides , are considered to be  narcotizing compounds. In this study the transcriptional responses of the soil arthropod Folsomia candida are investigated upon exposure to a series of anilines with increasing chlorination. A discrimination between class1 and class2 narcotizing compound is being made.

overall_design: Twenty-three day old Folsomia candida were exposed in LUFA 2.2 standard soil for two days to the EC50 concentrations of the following compounds: aniline, 4-chloroaniline, 3.5-dichloroaniline, 2,3,4-trichloroaniline, 2,3,5,6-tetrachloroaniline, pentachloroaniline and 1,2,3,4-tetrachlorobenzene.1,2,3,4-tetrachlrobenzene was included in the experimental design as a posiitive control of a narcotic class 1 compound. Four biological replicates were used for every treatment and a dye swap was used with the Cy3/Cy5 labels. This resulted in 32 samples which were analysed in 16 hybridisations executed in an interwoven loop design. The solvent (for spiking the soil) control was used as the reference treatment in the data analysis.

first: Daniel

last: Giesen

first: Kees

last: van Gestel


title: Expression profiling of the three clonotypic lineages of Toxoplasma gondii

status: Public on Feb 02 2010

city: Philadelphia

first: David

last: Roos

state: PA

address: 304B Carolyn Lynch Laboratories

department: Department of Biology

institute: University of Pennsylvania

summary: Expression profiling of the three clonotypic lineages dominating T. gondii populations in North America and Europe provides a first comprehensive view of the parasite transcriptome.

overall_design: Prugniaud, RH, and VEG strain parasites were cultured in human foreskin fibroblast (HFF) cells as previously described (Roos, Donald et al. 1994).  Prior to host cell rupture, cells were scraped from the flask and spun at 300g for 9 min.  The resultant pellet was lysed with Buffer RLT from the Qiagen RNeasy Mini Kit (Valencia, CA) and RNA was extracted according to the manufacturer’s instructions.  Labeled cRNA was created using the One-Cycle Labeling protocol in the Affymetrix GeneChip® IVT Labeling Kit (Santa Clara, CA).  Hybridization, washing, and scanning of arrays was performed using standard Affymetrix instrumentation and protocols for 11 micron, 169 format arrays.  Biological triplicates were generated for each strain and expression values computed using the RMA implementation (default parameters) in the affy package from Bioconductor (Gentleman, Carey et al. 2004).

first: Amit

last: Bahl

first: Paul

middle: H

last: Davis

first: Michael

last: Behnke

first: Florence

last: Dzierszinski

first: Manjunatha

last: Jagalur

first: Feng

last: Chen

first: Dhanasekaran

last: Shanmugam

middle: W

last: White

last: Kulp

middle: S


title: Analysis of the global gene expression profile for pearl oyster, Pinctada maxima, exposed to organotin (tributyltin)

last: Gardner

summary:
Bivalves are well known sentinel organism in the detection of environmental pollutants. Bioaccumulation of these contaminants in bivalves often manifests as specific alterations of their biological processes, which are used as biomarkers for environmental pollution. Tributyltin (TBT) is one such pollutant previously used as a biocide in marine antifouling paints, it now causes a number deleterious effects in bivalves leaching out of sediments in marine ecosystems. One effect extensively documented is shell abnormalities, including shell thickening and chambering. Changes in amino acid compositions of the shell matrix are associated with these deformations suggesting that TBT mode of action influences the biological control of shell biomineralization. This environmental toxicants effect on shell biomineralization was analyzed in this investigation at a transcriptional level in order to elucidate the normal shell biomineralization process. P. maxima animals were exposed to TBT in laboratory conditions and a concentration range for chronic and acute toxicity has been established. Animals exposed to chronic concentrations were analyzed for differential gene expression using PmaxArray 1.0 microarray platform and compared against control animals. Genes indentified as differentially expressed in association with TBT exposure included up-regulation of immunity and detoxification related genes and down-regulation of several shell matrix genes. A number of novel transcripts were additionally identified. The potential actions of these genes are discussed with reference to TBT toxicity and shell biomineralization. This investigation has used a microarray to determine transcriptional effects of TBT on P. maxima and proposed the involvement of novel components in shell formation, aiding the elucidation of the process.

Keywords: Expression profiling by array, stress response

overall_design: In order to determine to differential expression profiles for transcripts relevant to TBT exposure, 9 animals treated with TBT 50 ng1-1 were compared to 9 control animals untreated on a dual channel (Cy3 and Cy5) cDNA microarrays. The RNA for the 9 control animals was pooled together for a common reference while the RNA from the 9 treated animals was separated into 3 pooled replicates, each containing RNA from 3 individual animals. Each of the pooled treatment replicates were labeled (Cy3 or Cy5) as was the controls (opposing treatment label) and hybridized to a separate microarray chip, totaling 3 chips. Each chip had duplicate spot grids printed on the left and right of the chip providing technical replication. In total 6 microarrays were challenged and analyzed comprising 3 biological replicates each with 2 technical replicates.


title: Molecular signatures of mucosal immunity that distinguish HIV controllers from non-controllers

city: San Francisco

first: Sajeev

last: Batra

institute: UCSF

summary:
HIV “controllers” are individuals infected with the human immunodeficiency virus, type I (HIV) who maintain long-term control of viremia without antiviral therapy and who usually do not develop the acquired immune deficiency syndrome (AIDS). In this study, we have used oligonucleotide expression arrays to characterize the mucosal immune responses of these subjects in relation to untreated HIV+ individuals with high viral loads and progressive disease (“non-controllers”). Recto-sigmoid biopsies were analyzed from 9 controllers and 11 non-controllers. All of the genes identified to be significantly different were more highly expressed in the non-controllers. Many of these genes are involved in immunity and defence. These results underscore the importance of the sustained inflammatory response that attends progressive HIV disease.

Keywords: Recto-sigmoid biopsy profiles from HIV infected individuals

overall_design: We analyzed a series of 20 HEEBO arrays on which were hybed RNA amplified from the recto-sigmoid biopsies of HIV infected individuals that either have progressive disease or can maintain long term control of viremia.

first: Png

last: Loke

last: Favre

first: Peter

last: Hunt

first: Jacqueline

last: Leung

first: Bittoo

last: Kanwar

first: Jeffrey

middle: N

last: Martin

first: Steven

middle: G

last: Deeks

first: Joseph

last: McCune


title: Gene expression profiles after infection of zebrafish with Viral haemorrhagic septicemia rhabdovirus (VHSV)

status: Public on Feb 01 2010

city: Madrid

first: Julio

last: Coll

state: Madrid

address: crt Coruna km7

department: Biotechnology

country: Spain

institute: INIA

summary: We describe for the first time transcripts induced after infection of zebrafish with VHSV. Two days after infection, differentially expressed transcript levels from selected immune-related zebrafish genes were studied  using qRT-PCR in both fins and internal organs (pooled spleen, head kidney and liver).

overall_design: VHSV-induced gene expression in zebrafish fins and organs was measured by qRT-PCR at 2 days after infection.  Six (fins) and three (organs) independent experiments were performed for each treatment and their corresponding uninfected controls.


title: Gene expression in whole-roots of a Populus trichocarpa and Populus deltoides pseudo-backcross population

city: Gainesville

first: Matias

last: Kirst

state: FL

institute: University of Florida

summary: A microarray analysis of whole-genome gene expression in roots was carried out  in a (Populus trichocarpa X Populus deltoides) X Populus deltoides pseudo-backcross pedigree. Genetic variation in gene expression was quantified for 55,793 predicted gene models based on a single probe per gene. Resultant data contributed to the analysis of the genetic architecture of gene expression in roots of  Populus.

overall_design: Data include one biological replicate of 163 individuals segregating from a pseudo-backcross pedigree of (Populus trichocarpa X Populus deltoides) X Populus deltoides analyzed for gene expression (GE) in roots using one probe per gene for 55793 independent gene models (probes E_POPLARSxxxxxPxxxxx) and single feature sequence polymorphism (SFP) using one probe per gene for 12084 independent gene models (probes G_POPLARSxxxxxPxxxxx). GE and SFP probes were selected from 6-7 probes per gene previously tested in a pilot study of the two parent trees of the cross (Populus deltoides X Populus trichocarpa)


title: TSKMLO microarray validation experiment

city: East Lansing

first: Kelly

last: Sporer

state: MI

address: 3365 Anthony Hall

department: Food Science and Human Nutrition

institute: Michigan State University

summary: A 6K oligonucleotide microarray (TSKMLO) was designed as a community resource for researchers interested in domestic turkey muscle biology as it pertains to growth and development and their association with downstream meat quality. A series of quality control experiments performed to assess the functionality of the newly constructed array, which included a set of validation hybridizations performed in order to initially assess the functionality and repeatability of the newly designed TSKMLO array.  Strong correlations between these test comparisons confirmed that the array was able to hybridize fluorescently-labeled aRNA samples from turkey skeletal muscle at different developmental stages, that there was little dye bias, and that results were repeatable.

overall_design: Three experiments were designed.  1) A “Self-Self” array was hybridized using the same RNA sample (a randomly chosen RBC2, 18de) divided into two aliquots: one labeled with Cy3 and the other with Cy5 to ensure equal hybridization of each fluorescent dye-labeled sample.  2) Two “Dye Swap” arrays were hybridized using randomly chosen F, 18de and F, 1d samples.  The fluorescent dye assignments were reversed in the two arrays.  The objectives of this experiment were to observe expected differential expression between embryonic (hyperplasia) and neonatal (hypertrophy) muscle samples as well as to ensure similar hybridization regardless of dye assignment.  3) A “Repeat” array was hybridized on a different day using the same samples and dye assignments as the “Dye Swap 2” array to ensure reproducibility of hybridization results.

first: Wen

last: Chiang

first: Robert

middle: J

last: Tempelman

first: Catherine

last: Ernst

first: Kent

last: Reed

first: Sandra

last: Velleman

first: Gale

last: Strasburg


title: Analysis of the turkey skeletal muscle transcriptome between genetic lines within a developmental stage (Experiment 2)

summary: The purpose of Experiment 2 was two-fold: a) to evaluate a unique set of negative, distance, and mismatch control probes on the newly designed 6K TSKMLO microarray, and b) to identify differences in gene expression between two genetic lines of turkeys that phenotypically display differences in growth. Negative and mismatch controls confirmed specific hybridization using this array, and genes identified as differentially expressed between developmental stages by microarray analysis were confirmed by qRT-PCR. Novel candidate genes and pathways were identified as playing potentially crucial roles in skeletal muscle development between 2 distinct lines of turkeys with implications for improving turkey meat quality defects and growth-induced myopathies long-term.

overall_design: This experiment utilized turkeys from two genetic lines: RBC2, a randomly bred control line representative of a 1,967 commercial turkey, and F, a subline genetically selected from the RBC2 line for increased 16-week body weight. Skeletal muscle was collected at three critical developmental stages: 18-day embryo (18de; peak of hyperplasia), 1-day post-hatch (1d; shift from myoblast-mediated growth to satellite cell modulated growth by hypertrophy), and 16-week-old birds (16wk; completion of muscle formation and approximate age of commercial slaughter). Experiment 2 was designed to directly compare the two genetic lines, RBC2 vs. F, for each of the three stages of skeletal muscle development: 18de, 1d, and 16wk. Experiment 2 contained 10 arrays per comparison, therefore, it contained 30 arrays total. Birds were randomly assigned to an array, and hybridizations were performed in random order.


title: Identification and function of non-coding small RNAs in Streptococcus pneumoniae

city: Bloomington

first: Ho-Ching

middle: Tiffany

last: Tsui

state: IN

address: 1001 E. Third St.

department: Biology

institute: Indiana University

summary: We report here an experimental search for small non-coding RNAs in a low GC, gram-positive, human pathogenic bacteria Streptococcus pneumoniae.  Based on the bioinformatic analyses by Livny et al. (2006, Nucleic Acids Res. 34:3484-93), we tested 36 candidates by Northern analyses and confirmed the expression of 8 novel and one previously-reported sRNAs (CcnA) in the genome of D39 serotype 2 strain.  Some of the sRNAs showed differential expression in stationary phase or after treatment with competence stimulatory peptide compared to untreated exponential cells. One sRNA we detected is CcnA, one of five CiaR-controlled non-coding RNA (CcnA to E) reported previously.  By ectopically expressing this sRNA in a ΔciaR strain, we provide evidence that CcnA plays important roles in the regulation of growth and chain length of S. pneumoniae.  Microarray analysis comparing global gene expression of CcnA-expressing and CcnA-nonexpressing ΔciaR strains showed significant differences in expression of competence related genes and five putative transcriptional regulators. QPCR and transformation assays further confirmed that CcnA sRNA mediates a strong negative effect on competence development.  Our findings suggested many of the CiaRH function may be mediated via Ccn sRNAs and that sRNAs play regulatory roles in S. pneumoniae, an organism that lacks Hfq RNA chaperone.

overall_design: Three independent hybridizations using independent RNA preparations from the strain IU2678 (a ciaR knockout strain with no ccnA expression, reference strain) and strain IU2841 (a ciaR knowckout strain with ccnA expression at an ectopic site) were performed. Dye swap was performed with the reference strain labeled with Cy3 in two hybridizations and Cy5 in the one hybridizations.

middle: T

first: Valerie

last: Ray

first: Dhriti

last: Mukherjee

first: Lok-To

last: Sham

first: Kyle

last: Wayne

first: Andrew

last: Feig

first: Malcolm

last: Winkler


title: Comparison of C. cinerea vegetative dikaryotic to vegetative monokaryotic mycelia

first: Claire

last: Burns

address: 1001 E 3rd St

summary: Coprinopsis cinerea vegetative dikaryotic mycelia compared to vegetative monokaryotic mycelia

overall_design: Vegetative dikaryon cDNA was comparitively hybridized with monokaryotic vegetative cDNA. Labelled cDNA was hybridized to arrays in a 2 channel reaction.


title: Gene expression in leaves of a Populus trichocarpa and Populus deltoides pseudo-backcross population

summary: A microarray analysis of whole-genome gene expression in leaves was carried out  in a (Populus trichocarpa X Populus deltoides) X Populus deltoides pseudo-backcross pedigree. Genetic variation in gene expression was quantified for 55,793 predicted gene models based on a single probe per gene. Resultant data contributed to the analysis of the genetic architecture of gene expression in leaves of  Populus.

overall_design: Data include one biological replicate of 183 individuals segregating from a pseudo-backcross pedigree of (Populus trichocarpa X Populus deltoides) X Populus deltoides analyzed for gene expression (GE) in roots using one probe per gene for 55793 independent gene models (probes E_POPLARSxxxxxPxxxxx) and single feature sequence polymorphism (SFP) using one probe per gene for 12084 independent gene models (probes G_POPLARSxxxxxPxxxxx). GE and SFP probes were selected from 6-7 probes per gene previously tested in a pilot study of the two parent trees of the cross (Populus deltoides X Populus trichocarpa)


title: Cyst formation in the PKD2 (1-703) transgenic rat precedes deregulation of proliferation-related pathways

city: Mannheim

first: Carsten

last: Sticht

address: Theodor-Kutzer-Ufer

department: ZMF

country: Germany

institute: University Heidelberg

summary: Polycystic Kidney Disease is characterized by the formation of large fluid-filled cysts that eventually destroy the renal parenchyma leading to end-stage renal failure. Although remarkable progress has been made in understanding the pathologic mechanism of the disease, the precise orchestration of the early events leading to cyst formation is still unclear. Abnormal cellular proliferation was traditionally considered to be one of the primary irregularities leading to cyst initiation and growth. Consequently, many therapeutic interventions have focused on targeting this abnormal proliferation, and some have even progressed to clinical trials. However, the role of proliferation in cyst development was primarily examined at stages where cysts are already visible in the kidneys and therefore at later stages of disease development. In this study we focused on the cystic phenotype since birth in an attempt to clarify the temporal contribution of cellular proliferation in cyst development. Using a PKD2 transgenic rat model (PKD2 (1-703)) of different ages (0-60 days after birth) we performed gene expression profiling and phenotype analysis by measuring various kidney parameters. Phenotype analysis demonstrated that renal cysts appear immediately after birth in the PKD2 transgenic rat model (PKD2 (1-703)). On the other hand, abnormal proliferation occurs at later stages of the disease as identified by gene expression profiling. Interestingly, other pathways appear to be deregulated at early stages of the disease in this PKD model.  Our data suggest that cystogenesis precedes deregulation of proliferation-related pathways, suggesting that proliferation abnormalities may contribute in cyst growth rather than cyst formation.

overall_design: In this study we focused on the cystic phenotype since birth in an attempt to clarify the temporal contribution of cellular proliferation in cyst development. Using a PKD2 transgenic rat model (PKD2 (1-703)) of different ages (0,6 and 24 days after birth) we performed gene expression profiling of kidney cells.

first: Panayiota

last: Koupepidou

first: Kyriacos

last: Felekkis

first: Bettina

last: Kränzlin

first: Norbert

last: Gretz

first: Constantinos

last: Deltas


title: Gene expression of Streptomyces turgidiscabies pathogenicity island

city: Helsinki

first: Panu

last: Somervuo

address: Viikinkaari 4

country: Finland

institute: University of Helsinki

summary:
Gene expression in Streptomyces turgidiscabies pathogenicity island was studied after transferring cells into thaxtomin A inducing medium OBB using Agilent 60mer oligonucleotide array with probes designed for S.scabies and S.turgidiscabies.
Gene expression study was used to confirm results of gene prediction.

platform_organism:
Streptomyces scabiei
Streptomyces turgidiscabies

overall_design: Time course gene expression experiment with two replicates

first: Marja

last: Aittamaa

first: Jari

last: Valkonen


title: Early response of bovine alveolar macrophages to infection with live and heat-killed Mycobacterium bovis

status: Public on Jan 31 2010

city: Roslin

first: Michael

middle: Bryan

last: Watson

state: Midlothian, EH25 9PS

address: Roslin Biocentre

department: ARK-Genomics

country: United Kingdom

institute: The Roslin Institute

summary: Bovine tuberculosis, caused by Mycobacterium bovis, is a disease of considerable economic importance yet comparatively little is known about the bovine immune response to the disease. Alveolar macrophages are one of the first cells to encounter mycobacteria following infection. In this experiment we investigated the early transcriptional response of bovine alveolar macrophages following infection with M. bovis. The transcriptional response to heat-killed M. bovis was also investigated to look for genes that are only differentially transcribed in response to the live organism.

overall_design: Five-condition experiment, uninfected, live and heat-killed M. bovis-infected bovine alveolar macrophages from five cattle infected for two and four hours. Comparisons were within animal. Dye swaps were incorporated into the design.

first: Stephanie

last: Widdison

first: Tracey

last: Coffey


title: Winter and fall gene expression in rainbow smelt liver

city: Halifax

first: Kathryn

middle: Vanya

last: Ewart

state: NS

address: 1411 Oxford St.

department: Marine Bioactives Group

country: Canada

institute: NRC Institute for Marine Biosciences

summary:
Rainbow smelt (Osmerus mordax) are freeze-resistant fish that accumulate glycerol and produce an antifreeze protein during winter.  Quantitative reverse transcription-PCR (qPCR) and subtractive hybridization studies have revealed a few genes in smelt liver to be differentially regulated in winter in comparison with the fall when water temperatures are warmer.  In order to further define the suite of processes that are regulated seasonally, we undertook a large-scale analysis of gene expression by hybridization of smelt cDNA to the salmonid 16K cGRASP cDNA microarray.
These microarray experiments were conducted as a focused sieving exercise, which identified informative genes for further study in the microarray samples and over a seasonal sampling series using quantitative reverse-transcription PCR.

overall_design: Total RNA was obtained from livers of 3 male fish sampled on October 20th and aliquots were pooled to contain equimolar concentrations of each RNA preparation.  Total RNA was prepared from 3 male fish on January 3rd and processed in the same manner.  Fluorescently labeled cDNA preparations were made using the fall (October) and winter (January) pools and mixed before hybridization to the microarray.  The cDNAs were hybridized to three separate microarrays in order to generate technical replicates.  Differential expression was assessed in order to select genes for further study.

first: Robert

middle: C

last: Richards

first: Connie

middle: E

last: Short

first: William

last: Driedzic

first: K


title: SAGE libraries from cell bodies and axons of NGF-differentiated sympathetic neurons

city: London

first: Catia

last: Andreassi

address: Gower street, 1.03

department: MRC LMCB and Dept. of Neuroscience, Physiology and Pharmacology

institute: University College London

summary: The subcellular localization of specific mRNAs is an evolutionary conserved mechanism that underlies the establishment of cellular polarity and specialized cell functions.  In neurons, mRNA trafficking and local protein translation in dendrites provides an important mechanism that mediates synaptic development and plasticity.  The significance of mRNA targeting and protein synthesis in axons, however, is still unclear.  Only a small number of transcripts have been identified in axons to date, and their contribution to axon growth and neuronal survival remains largely unknown. Here, we report the results of a novel screen that allowed the separate identification of mRNAs localized in cell bodies and in axons of developing neurons. Using compartmentalized cultures of sympathetic neurons and Sequential Analysis of Gene Expression (SAGE), the screen identified more than 200 axonal mRNAs, including ones that encode cytoskeletal proteins and proteins that function in neural development and signal transduction.  Importantly, several classes of transcripts were selectively enriched in axons, indicating that an active process drives the targeting of specific mRNAs from the cell bodies to the axons.  This study is the first comprehensive and unbiased analysis of mRNA localization in subcellular domains of any neuronal cell type.

overall_design: We used compartmentalized chambers to culture neonatal rat sympathetic neurons (Campenot, 1977).   In these cultures, the cell bodies are separated from the distal axons by a 1 mm wide Teflon divider, which maintains the cell bodies and axon terminals in separate fluid compartments.  Primary rat sympathetic neurons are especially suitable for compartmentalized culture because they can be grown as a highly homogeneous population without glial cells.  Neurons were seeded in the central compartment with nerve growth factor (NGF), and after a few days, the NGF was lowered in this compartment and supplied only to the peripheral compartment to stimulate axon growth.  The anti-mitotic agent cytosine arabinoside C (Ara-C) was added to both compartments to remove non-neuronal cells. mRNA was then isolated after 12 days in culture (DIV) from cell body or axon compartments. As the initial mRNA content in axons was not sufficient to perform the SAGE analysis, both axon and cell body mRNAs were subjected to two rounds of linear amplification to obtain antisense RNA (aRNA).  The amplified aRNA was then reverse transcribed and second strand synthesis was performed to proceed with the SAGE assay using the LongSAGE kit (Invitrogen) according to the manufacturer’s protocol.

first: Richard

last: Mitter

first: Antonella

last: Riccio


title: LPS-induced gene expression in wildtype and MAL knockout mouse macrophages

status: Public on Jan 29 2010

city: Dublin

first: Stephen

middle: Francis

last: Madden

address: Glasnevin

department: National Institute for Cellular Biotechnology

country: Ireland

institute: Dublin City University

summary: Comparison of gene expression in WT and MAL knockout (MALKO) mouse macrophages treated with 10ng/ml lipopolysaccharide (LPS) with that of mock-treated cells incubated for the same time (10 days). Cells from 4 mice of each genotype were used and each individual served as its own control. Hybridizations of treated and control samples were dye swapped.

overall_design: Two experiments: WT vs. LPS-treated WT, and MALKO vs. LPS-treated MALKO.  4 individual biological replicates for each genotype, and each mouse served as its own mock control.

middle: F

first: Susan

middle: B

last: Carpenter

first: Katherine

middle: A

last: Fitzgerald

first: Harry

last: Bjorkbacka


title: Profiling of copy-number variation between BN and SHR rats

status: Public on Jan 30 2010

city: Utrecht

first: Victor

last: Guryev

address: Uppsalalaan 8

institute: Hubrecht Institute

summary: We used Nimblegen HD aCGH to detect copy-number variants between genomes of BN and SHR rats

overall_design: Comparison of single BN against single SHR individual in a dye-swap experiment

first: Edwin

last: Cuppen


title: Striatum of Huntington's disease model mice [Illumina data]

city: Vancouver

first: Paul

last: Pavlidis

state: British Columbia

address: 177 Michael Smith Laboratories 2185 East Mall

department: Centre for High-Throughput Biology / Psychiatry

institute: University of British Columbia

summary:
Evaluation of transcriptional changes in the striatum may be an effective approach to understanding the natural history of changes in expression contributing to the pathogenesis of Huntington disease (HD). We have performed genome-wide expression profiling of the YAC128 transgenic mouse model of HD at 12 and 24 months of age using two platforms in parallel; Affymetrix and Illumina. We performed gene expression profiling on the same striatal mRNA across both platforms.

Keywords: time course,disease state analysis

overall_design: Transgenic mice expressing human HD huntingtin with 120 CAG repeats (YAC128) and wildtype littermates were used for the described experiments. The mice were group housed in polystyrene cages under a normal light-dark cycle (6 am to 8 pm) in a clean facility and with free access to water and standard rodent chow. All experiments were performed in accordance with the University of British Columbia animal care committee. Striatal tissue was collected from 12 and 24 month old YAC128 mice and wildtype littermates.

first: Kristina

last: Becanovic


title: HNF-1alpha Triggers Initiation of the Human Intestinal Cell Differentiation Program in Cooperation with Cdx-2 and GATA-4

city: Sherbrooke

first: Jean-François

last: Beaulieu

state: Québec

address: 3001, 12ème avenue nord

department: Anatomie & biologie cellulaire

institute: Université de Sherbrooke

summary:
Transcriptional profiling of undifferentiated HIEC cells stably infected with a retroviral inducible expression vector (RevTet system) comparing DOX-induced HIEC pTetON with DOX-induced HIEC cells expressing either HNF1 and CDX2 or HNF1, CDX2 plus GATA4.
Keywords: Differential gene expression

overall_design: Three-condition experiment, DOX-induced HIEC expressing HNF1-CDX2 vs DOX-induced HIEC pTetON, DOX-induced HIEC expressing HNF1-CDX2 plus GATA4 vs DOX-induced HIEC pTetON.

first: Fabrice

last: Escaffit

first: Yannick

last: Benoît

first: Frédéric

last: Paré

first: Caroline

last: Francoeur

first: Dminique

last: Jean

first: Eric

last: Tremblay

first: François

last: Boudreau


title: Chromatin signature of embryonic pluripotency is established during zygotic genome activation [Danio rerio]

status: Public on Jan 26 2010

city: Shanghai

first: Yong

last: Zhang

address: 1239 Siping Road

country: China

institute: Tongji University

type: Genome binding/occupancy profiling by genome tiling array

summary:
[PROJECT] After fertilization the embryonic genome is inactive until transcription is initiated during the maternal-zygotic transition (MZT). This universal process coincides with the formation of pluripotent cells, which in mammals can be used to generate embryonic stem (ES) cells. To study the changes in chromatin structure that accompany zygotic genome activation and pluripotency, we mapped the genomic locations of histone H3 modifications before and after MZT in zebrafish embryos. Repressive H3 lysine 27 trimethylation (H3K27me3) and activating H3 lysine 4 trimethylation (H3K4me3) are only detected after MZT. H3K4me3 marks more than 80% of genes, including many developmental regulatory genes that are also occupied by H3K27me3. Sequential chromatin immunoprecipitation demonstrates that both methylation marks occupy the same promoter regions, revealing that the bivalent chromatin domains found in cultured ES cells also exist in embryos. In addition, we find a large group of genes that are monovalently marked by H3K4me3 but not H3K27me3. These H3K4me3 monovalent genes are neither expressed nor stably bound by RNA polymerase II. Closer inspection of in vitro data sets reveals similar monovalent H3K4me3 domains in ES cells. The analysis of an inducible transgene indicates that H3K4me3 domains can form in the absence of sequence-specific transcriptional activators or stable association with RNA pol II. These results suggest that bivalent and monovalent domains might poise embryonic genes for activation and that the chromatin profile associated with pluripotency is established during MZT.
[SAMPLES] ChIPchip analysis of histone modifications (H3K4me3, H3K27me3, H3K36me3) and RNA polymerase II in pre MZT (256-cell) and post MZT (4hpf; dome/30% epiboly) wt zebrafish embryos.

overall_design: H3K4me3, H3K27me3, H3K36me3 and PolII ChIP-chip at 256 cell stage (one replicate) and 4hpf (dome/30% epiboly) (two replicates)

first: Nadine

middle: L

last: Vastenhouw

first: Ian

last: Woods

first: Farhad

last: Imam

first: Aviv

last: Regev

first: X

last: Liu

first: John

last: Rinn

first: Alexander

last: Schier


title: Comparison of rat thoracic duct to vena cava

status: Public on Jan 27 2010

city: Temple

first: David

middle: C.

last: Zawieja

state: TX

address: 702 SW HK Dodgen Loop

department: Dept. of Systems Biology and Translational Medicine

institute: Texas A&M Health Science Center

summary:
While blood vessels have muscular walls that undergo tonic contractions to alter vascular resistance and, thus, control blood flow, lymphatics at the level of the collecting vessels and higher have muscular walls capable of rapid phasic contractions that generate lymph flow in addition to tonic contractions that regulate lymph flow resistance.  While the ability of lymphatics to undergo rapid phasic contractions has been known for several centuries, the biological elements governing this phenomenon remain unknown.  In an attempt to gain insight into the structural and regulatory elements that give lymphatic vessels their unique contractile capabilities, we utilized two-color microarray analysis to compare the thoracic duct of the rat to the vena cava of the same donor animal.  Total cellular RNA was isolated immediately following vessel isolation and amplified in the presence of amino allyl dUTP.  The resulting modified aRNA was conjugated to either Cy3 or Cy5 dye prior to hybridization to a rat 5.7K oligonucleotide array.  Analysis and filtering of the data obtained from the microarray image yielded several contractile and regulatory genes with altered expression in the thoracic duct relative to the vena cava.  Further evaluation of the data obtained in this study may aid in illustrating the unique properties of the lymphatic vessel and its muscular wall.
Keywords: Thoracic duct, lymphatics, microarray

overall_design: Four unique thoracic duct/vena cava sample pairs were individually analyzed via two-color microarray analysis yielding 4 biological replicates.  To minimize dye bias, a dye balance design was utilized in which the orientation of dye assignment was alternated between vessel pairs (i.e. two thoracic duct samples were labeled with Cy3 and two were labeled with Cy5).  Prior to analysis, the data from 2 of the replicates was transformed to accomodate the dye balance such that all thoracic duct data is interpreted as Cy5 and all vena cava data is interpreted as Cy3.

last: Bridenbaugh


title: M.ruminantium B.proteoclasticus coculture vs monocultures

status: Public on Jan 28 2010

city: Urbana

first: Carl

middle: James

last: Yeoman

state: IL

address: 1207 W Gregory ave

department: Institute for Genomic Biology

institute: University of Illinois

summary: This study was undertaken with two major goals in mind: i) To investigate the potential for interspecies hydrogen transfer between a hemicellulytic rumen bacterium (B. proteoclasticus) and a methanogenic achaea (M. ruminantium) Microscopic examination had previously shown B. proteoclasticus in co-culture with M. ruminantium rapidly formed cell to cell co-aggregates. ii) To examine the expression of genes involved in methanogenesis under more rumen-like conditions.

sample_organism:
Butyrivibrio proteoclasticus B316
Methanobrevibacter ruminantium M1

overall_design:
Cells of M.ruminantium and B. proteoclasticus were grown in triplicate as mono- or co-cultures until a mid-exponential growth phase was reached. RNAs were extracted and purified. The relative quantities of RNAs contributed by each organism to the co-culture samples were determined by rt qPCR using genes previously shown to be constitutively expressed. The mono-culture RNAs were then combined in the determined proportions to normalise mRNA abundance with their co-culture replicates. cDNAs were synthesized and labelled and hybridised to the arrays. 3 biological replicates were used and the cDNA from each was divided in two to allow each to undergo a dyeswap (total of six arrays) giving technical replicates. Each gene was probed in triplicate adding further technical replication. Blanks and negative controls were also included.
The log ratios for each probes replicates (biological and technical) were averaged prior to further analysis.


title: Comparative analysis of gene expression profiles between Nodal-overexpressing embryos with control embryos

city: Kochi

first: Kaoru

last: Mita

address: 2-5-1, Akebono-cho

country: Japan

institute: Kochi University

summary:
A comprehensive search for downstream genes is necessary for understanding of functions of Nodal signaling in embryos of the ascidian Ciona intestinalis.  The level of mRNA expression in Nodal-overexpressing embryos was compared with that in normal embryos at the late gastrula stage by a microarray analysis.  Overexpression of Ci-nodal elevated the level of expression of 115 genes more than two-fold.  These candidate target genes include those encoding transcription factors, proteins involved in cell-cell communication, and extracellular matrix components.  Many of these genes were expressed in the neural plate at the late gastrula stage.  Overexpression of nodal also affected the expression of genes involved in the planar cell polarity pathway and Wnt/Ca2+ pathway (prickle, Rho-like2, and PLC2), and delta1-protocadherin-like.
Keywords: ascidian, Ciona intestinalis, Nodal, embryogenesis, gene expression profile

overall_design:
two samples
Dye Swap design with 2 arrays

first: Ryo

last: Koyanagi

last: Azumi

first: Shigeki

last: Fujiwara


title: Gene expression after SPVD infection in resistant and susceptible sweet potato cultivars

city: Athens

first: Cecilia

last: McGregor

state: GA

address: Miller Plant Sciences

department: Horticulture

institute: University of Georgia

summary: Sweet potato virus disease (SPVD) is one of the most devastating diseases affecting sweetpotato (Ipomoea batatas), an important food crop in developing countries. SPVD develops when sweetpotato plants are dually infected with sweet potato feathery mottle virus (SPFMV) and sweet potato chlorotic stunt virus (SPCSV).  In the current study, global gene expression between SPVD affected plants and virus-tested control plants (VT) were compared in the susceptible ‘Beauregard’ and resistant ‘NASPOT 1’ (Nas) sweetpotato cultivars at 5, 9, 13 and 17 days post inoculation (DPI).

overall_design:
Two-color (Cy3 and Cy5) microarrays.
2 cultivars: Beauregard (Bx) and Naspot 1 (Nas); 4 time points: 5dpi, 9dpi, 13 dpi, 17 dpi; Treatments : SPVD and Virus Tested Control; 4 replicates (plants) for each cultivar x treatment combination.
Please note that the loop design used for this study necessitated the use of 2 "samples" for each array. So there are 64 sample entries and 32 arrays. Use the title of the supplementary file to determine which samples were hybridized on the same array. For example GSM467650_BNHS1_1.gpr.gz and GSM467651_BNHS1_2.gpr.gz  indicated that GSM467650 and GSM467651 were hybridized on the same array (Lab array # BNHS1).

first: Don

last: LaBonte

first: Douglas

last: Miano

first: Mary

last: Hoy

first: Chris

last: Clark

first: Guilherme

last: Rosa


title: Vinyl Chloride Degradation by Mixed Microbial Community KB1

city: Toronto

first: Elizabeth

last: Edwards

address: 200 College st.

department: Department of Chemical Engineering and Applied Chemistry

institute: University of toronto

summary: A shotgun metagenome microarray was created and used to investigate gene transcription during vinyl chloride (VC) dechlorination by a microbial enrichment culture called KB1.  The array was constructed by spotting genomic fragments amplified from short-insert libraries of KB1 metagenomic DNA.  Subsequently, the microarrays were interrogated with RNA extracted from KB1 during VC dechlorination (VC+methanol), and in the absence of VC (methanol-only).  The most differentially expressed spots, and spots with the highest intensities, were then chosen to be sequenced.  Sequencing revealed that Dehalococcoides (Dhc) genes involved in transcription, translation and energy generation were up-regulated during VC degradation.  Furthermore, the results indicated that the reductive dehalogenase homologous (RDH) gene KB1rdhA14 is the only RDH gene up-regulated upon VC degradation, and that multiple RDH genes were more highly transcribed in the absence of VC.  Numerous hypothetical genes from Dehalococcoides were also more highly transcribed in methanol only treatments and indicate that many uncharacterized proteins are involved in cell maintenance in the absence of chlorinated substrates.  Spots with genes from Spirochaetes, Chloroflexi, Geobacter, Methanogens and phage organisms were differentially expressed and sequencing provided information from these uncultivated organisms that can be used to design primers for more targeted studies.  This array format is powerful, as it does not require a priori sequence knowledge.  This study provides the first report of such arrays being used to investigate transcription in a mixed community, and shows that this array format can be used to screen metagenomic libraries for functionally important genes.

overall_design: 2 Biological replicate experimens conducted 1 month apart.  In the first there were 2 dye-swapped duplicates (total 4) of VC+MeOH versus MeOH only.  In the second experiment there was one set of dye swapped arrays.  Thus 6 arrays were performed including biological replicates, dye swapped replicates and technical duplicates.

first: Alison

middle: S

last: Waller


title: Arp2/3 complex mutants show a pronounced lack of hyphal specific gene expression in Candida albicans

status: Public on Jan 23 2010

city: Montréal

first: Elias

last: Epp

address: 1205 Docteur Penfield

institute: McGill University

summary: Candida albicans is a diploid fungal pathogen lacking a defined complete sexual cycle, and thus has been refractory to standard forward genetic analysis. Instead, transcription profiling and reverse genetic strategies based on Saccharomyces cerevisiae have typically been used to link genes to functions. To overcome restrictions inherent in such indirect approaches, we have investigated a forward genetic mutagenesis strategy based on the UAU1 technology. We screened 4700 random insertion mutants for defects in hyphal development, and linked two new genes (ARP2 and VPS52) to hyphal growth. Deleting ARP2 abolished hyphal formation, generated round and swollen yeast phase cells, disrupted cortical actin patches and blocked virulence in mice. The mutants also showed a global lack of induction of hyphae-specific genes upon the yeast-to-hyphae switch. Surprisingly, both arp2Δ/Δ and arp2Δ/Δarp3Δ/Δ mutants were still able to endocytose FM4-64 and Lucifer Yellow, although as shown by time-lapse movies internalization of FM4-64 was somewhat delayed in mutant cells. Thus the non-essential role of the Arp2/3 complex discovered by forward genetic screening in C. albicans showed that uptake of membrane components from the plasma membrane to vacuolar structures is not dependent on this actin nucleating machinery.

overall_design: By forward genetic screening, we have identified genes that are essential for hyphal formation. One of the hits is the Arp2/3 complex, which is essential for hyphal formation, but not for viability in Candida albicans. To gain insights into cellular processes affected by disrupting Arp2/3 complex functions, we performed transcriptional profiling under yeast growth conditions (YPD at 30°C for three hours) or hyphal induction (YPD + 10% FBS at 37°C for three hours) and compared transcriptional consequences of deleting ARP2 to MYO5 and SLA2 microarray data sets (Oberholzer et al., 2006).

first: Andrea

last: Walther

first: Guylaine

last: Lépine

first: Zully

last: Leon

first: Alaka

last: Mullick

first: Martine

last: Raymond

first: Jürgen

last: Wendland

first: Malcolm

last: Whiteway


title: ChIP-chip from the follicular lymphoma cell line RL with H3K4me3, H3K37me3 and Suz12

status: Public on Jan 24 2010

city: Augusta

first: Huidong

last: Shi

department: Georgia Cancer Center

institute: Augusta University

summary: We conducted genome-wide bisulfite sequencing analysis of the follicular lymphoma cell line RL and found that a large amount of methylated genes are polycomb target genes in ES cells. We therefore conducted a ChIP-chip experiment to determine the methylated genes that are bound by the polycomb protein Suz12. Although 28% of MRIs are PRC2 target genes in ES cells, our ChIP-on-Chip analysis showed that only 13% of MRIs are associated with H3K27Me3 marks and only 5% of the MRIs are bound by Suz12 in RL cells in vivo.

overall_design: Comparison of DNA methylation with histone modifications in RL cells.


title: Analyses of Human Brain Metastases of Breast Cancer Reveal the Association between HK2 Up-Regulation and Poor Prognosis

status: Public on Jan 22 2010

city: Bethesda

first: Sean

last: Davis

state: MD

address: 37 Convent Drive, Room 6138

institute: National Cancer Institute

summary:
Analyses of Resected Human Brain Metastases of Breast Cancer Reveal the Association between Up-Regulation of Hexokinase 2 and Poor Prognosis.
Brain metastases of breast cancer seem to be increasing in incidence as systemic therapy improves. Metastatic disease in the brain is associated with high morbidity and mortality. We present the first gene expression analysis of laser-captured epithelial cells from resected human brain metastases of breast cancer compared with unlinked primary breast tumors. The tumors were matched for histology, tumor-node-metastasis (TNM) stage, and hormone receptor status. Most differentially expressed genes were down-regulated in the brain metastases, which included, surprisingly, many genes associated with metastasis. Quantitative real-time PCR analysis confirmed statistically significant differences or strong trends in the expression of six genes: BMP1, PEDF, LAMγ3, SIAH, STHMN3, and TSPD2. Hexokinase 2 (HK2) was also of interest because of its increased expression in brain metastases. HK2 is important in glucose metabolism and apoptosis. In agreement with our microarray results, HK2 levels (both mRNA and protein) were elevated in a brain metastatic derivative (231-BR) of the human breast carcinoma cell line MDA-MB-231 relative to the parental cell line (231-P) in vitro. Knockdown of HK2 expression in 231-BR cells using short hairpin RNA reduced cell proliferation when cultures were maintained in glucose-limiting conditions. Finally, HK2 expression was analyzed in a cohort of 123 resected brain metastases of breast cancer. High HK2 expression was significantly associated with poor patient survival after craniotomy (P = 0.028). The data suggest that HK2 overexpression is associated with metastasis to the brain in breast cancer and it may be a therapeutic target.

overall_design: Common reference design, disease state design.

first: Diane

last: Palmieri

first: Paul

last: Meltzer

first: Patricia

last: Steeg


title: Stromal gene expression in human esophageal cancer

city: North Wales

first: James

last: Hardwick

address: 351 N. Sumneytown Pike

department: Molecular Profiling

institute: Merck & Co., Inc.

summary: The incidence of esophageal and junctional adenocarcinoma has increased 6 fold in the last 30 years and 5 year survival remains less than 14%. Once considered an inert scaffold, the stromal compartment is now recognized to play a central role in invasive cancer. However, whether the stroma plays a role in the transition from pre-invasive to invasive disease is unknown. In this study we hypothesized that the altered expression of stromal genes influences the progression from pre-invasive to invasive disease. Using Barrett's esophagus as a model system, we performed an unbiased, genome-wide analysis of gene expression in micro-dissected stroma from different disease progression stages.

overall_design: Total RNA isolated from laser-capture microdissected stromal cells from human tissue sections was used to make fluorescently labeled cRNA that was hybridized to DNA oligonucleotide microarrays. Briefly, 4 µg of total RNA was used to synthesize dsDNA through reverse transcription. cRNA was produced by in vitro transcription and labeled postsynthetically with Cy3 or Cy5. Two populations of labeled cRNA, a reference population and an experimental population, were compared with each other by competitive hybridization to microarrays. Two hybridizations were done with each cRNA sample pair using a fluorescent dye reversal strategy. Human microarrays contained oligonucleotide probes corresponding to approximately 21,000 genes. All oligonucleotide probes on the microarrays were synthesized in situ with inkjet technology (Agilent Technologies, Palo Alto, CA). After hybridization, arrays were scanned and fluorescence intensities for each probe were recorded. Ratios of transcript abundance (experimental to control) were obtained following normalization and correction of the array intensity data. Gene expression data analysis was done with the Rosetta Resolver gene expression analysis software (version 7.0, Rosetta Biosoftware, Seattle, WA).

first: A

last: Saadi

first: N

last: Shannon

first: P

last: Lao-Sirieix

first: M

last: O'Donovan

first: E

last: Walker

last: Clemons

first: J

first: C

last: Zhang

last: Das

first: V

last: Save

last: Novelli

first: F

last: Balkwill

first: R

last: Fitzgerald


title: Effects of IGF 1 on CCL-171 fibroblasts

status: Public on Jan 21 2010

city: Zurich

first: Michal

last: Rajski

address: Winterthurerstrasse 190

department: Institute of Physiology

country: Switzerland

institute: University of Zurich

summary: To characterize the effects of IGF 1 on CCL171 cells, we cultured pre-starved CCL 171 fibroblasts with or without 50ng/ml IGF 1 for 24 h. We then profiled gene expression changes using Human Exonic Evidence Based Oligonucleotide (HEEBO) microarrays. After stimulation, total RNA was extracted and amplified using a modified Eberwine procedure. The amplified RNA was labeled with the fluorescent dye Cy5 and pooled with Cy3 labeled reference RNA, and then the pooled RNA was hybridized onto HEEBO microarrays. After hybridization and washing, arrays were scanned on a fluorescent microscope scanner.

overall_design: An all pairs experiment design type is where all labeled extracts are compared to every other labeled extract.


title: Effects of IGF 1 on primary breast fibroblasts (normal and carcinoma associated)

summary: To characterize the effects of IGF 1 on primary breast fibroblasts, we cultured pre-starved primary breast fibroblasts (normal and carcinoma associated) from three patients with or without 50ng/ml IGF 1 for 24 h. We then profiled gene expression changes using Human Exonic Evidence Based Oligonucleotide (HEEBO) microarrays. After stimulation, total RNA was extracted and amplified using a modified Eberwine procedure. The amplified RNA was labeled with the fluorescent dye Cy5 and pooled with Cy3 labeled reference RNA, and then the pooled RNA was hybridized onto HEEBO microarrays. After hybridization and washing, arrays were scanned on a fluorescent microscope scanner.


title: Effects of IGF 1 on MCF-7 breast cancer cells

summary: To characterize the effects of IGF 1 on MCF 7 breast cancer cells, we cultured pre-starved MCF 7 cells with or without 50ng/ml IGF 1 for 24 h. We then profiled gene expression changes using Human Exonic Evidence Based Oligonucleotide (HEEBO) microarrays. After stimulation, total RNA was extracted and amplified using a modified Eberwine procedure. The amplified RNA was labeled with the fluorescent dye Cy5 and pooled with Cy3 labeled reference RNA, and then the pooled RNA was hybridized onto HEEBO microarrays. After hybridization and washing, arrays were scanned on a fluorescent microscope scanner.


title: Transcriptional responses of allotetraploid cotton species to salt stress

first: Alan

middle: R

last: Gingle

address: 111 Riverbend Rd.

department: Center for Applied Genetics Technologies

summary:
Five allotetraploid cotton species have adapted, through their transcriptional responses, to unique environments with distinct levels of inherent abiotic stresses. The transcriptional responses of leaf and root tissue in five allotetraploid cotton species (Gossypium hirsutum, G. barbadense, G. tomentosum, G. mustelinum, and G. darwinii) under salt stress have been investigated in this study using cotton long oligonucleotide microarrays. Physiological responses to salinity such as stomatal conductance, ion and osmoprotectant contents were also measured as indicators of imposed stress. Accessions from these five cotton species were hydroponically grown and gradually introduced to a NaCl treatment (15 dS m-1). The microarray results identified 2721 and 2460 differentially expressed genes under salt stress that were significant in leaf and root tissue, respectively. Many of these genes were classified under gene ontology (GO) categories that suggest abiotic stress. These allotetraploid cottons shared transcriptional responses to salinity, but also showed responses that were species-specific. No consistent differences in transcriptional response among the previously estimated phylogenetic branches were found. Stomatal conductance, ion accumulation, and betaine, trigonelline, and trehalose contents also indicated salt stress. This global assessment of transcriptional and physiological responses to salt stress of these cotton species may identify possible gene targets for crop improvement and evolutionary studies of cotton.

Keywords: CEGC Cotton oligo salt stress

sample_organism:
Gossypium barbadense
Gossypium hirsutum
Gossypium mustelinum
Gossypium darwinii
Gossypium tomentosum

platform_organism:
Gossypium barbadense
Gossypium hirsutum
Gossypium arboreum
Gossypium raimondii

first: Rozaura

middle: V

last: Hall

first: Jonathan

last: Shortt

first: Michelle

last: Withers

first: Von

middle: D

last: Jolley

first: W

last: Johnson

first: Joshua

last: Udall


title: The meiotic transcriptome of the basidomycete fungus Coprinopsis cinerea (Coprinus cinereus)

city: Bloomington

first: Claire

last: Burns

address: 1001 E 3rd St

institute: Indiana University

summary:
Coprinopsis cinerea exhibits synchronised meiosis  in the gill tissue of the fungus, which is 75% meiotic.  The mushroom develops from a dikaryon, which contains two separate nuclei.  These nucleifuse in the basidia (karyogamy).  After karyogamy, the nuclei enter an extended meiotic prophase, in which pahcytene occurs at 6 hours post karyogamy (K+6).  The tetrads produced by the second meiotic division are present 12 hours after karyogamy (K+12).  To examine a comprehensive timecourse of meiosis in this organism, we took samples over a 15-hour period, 3 hours apart:  K-3, K, K+3, K+6, K+9, K+12.

Keywords: time course

overall_design: Four biological replicate samples  of each timepoint were taken.  Labelled cDNA was hybridized to arrays in a 2 channel reaction.  The reference sample was a mixture of timepoint samples.


title: Gene expression response of Populus tremuloides cell suspension cultures to salicyl alcohol feeding

first: Chung-Jui

last: Tsai

address: 120 Green Street

department: School of Forestry and Natural Resources

summary: Aspen cell cultures were fed 5 mM salicyl alcohol at the early exponential phase (5 days after subculture). Cells were harvested from fed (SL) or unfed (C) cells after 48 hours.  Gene expression changes were analyzed using a custom 7K aspen EST array.

overall_design: RNA was extracted from 3 independent control (C) and 3 salicyl alcohol-fed (SL) samples, and one control and one treated sample were randomly paired for hybridization (3 biological replicates).  Dye-swap hybridizations were carried out for each pair of samples.

first: Raja

last: Payyavula


status: Public on Jan 20 2010

first: Beau

last: Fenner

address: 123 Saint Stephen's Green

institute: Royal College of Surgeons in Ireland

summary: Signal transduction by the NF-kappaB pathway is a key regulator of a host of cellular responses to extracellular and intracellular messages. The NEMO adaptor protein lies at the top of this pathway and serves as a molecular conduit, connecting signals transmitted from upstream sensors to the downstream NF-kappaB transcription factor and subsequent gene activation. The position of NEMO within this pathway makes it an attractive target from which to search for new proteins that link NF-kappaB signaling to additional pathways and upstream effectors. In this work, we have used protein microarrays to identify novel NEMO interactors. A total of 112 protein interactors were identified, with the most statistically significant hit being the canonical NEMO interactor IKKbeta, with IKKalpha also being identified. Of the novel interactors, more than 30% were kinases, while at least 25% were involved in signal transduction. Binding of NEMO to several interactors, including CALB1, CDK2, SAG, SENP2 and SYT1, was confirmed using GST pulldown assays and coimmunoprecipitation, validating the initial screening approach. Overexpression of CALB1, CDK2 and SAG was found to stimulate transcriptional activation by NF-kappaB, while SYT1 overexpression repressed TNFalpha-dependent NF-kappaB transcriptional activation in human embryonic kidney cells. Corresponding with this finding, RNA silencing of CDK2, SAG and SENP2 reduced NF-kappaB transcriptional activation, supporting a positive role for these proteins in the NF-kappaB pathway. The identification of a host of new NEMO interactors opens up new research opportunities to improve understanding of this essential cell signaling pathway.

sample_organism:
Escherichia coli
Homo sapiens

overall_design: For microarray screening, Invitrogen Protoarray v4.0 protein microarrays were used. Human NEMO expressed as a C-terminal GST fusion was purified from E. coli lysates and labelled with biotin. NEMO or biotinylated GST were applied to the microarrays and binding partners detected using streptavidin-Alexa Fluor 647. Significant interactors on both arrays were detected using Invitrogen Protoarray Prospector software and a Z-score cutoff of 3.0. Following subtraction of interactors present on the GST control array, a final set of significant NEMO interactors was derived. Full experimental details are supplied in Fenner, B. J., Scannell, M. & Prehn, J. H. M. (2010). Expanding the substantial interactome of NEMO using protein microarrays. PLoS ONE (in press).

middle: J

first: Michael

last: Scannell

first: Jochen

middle: H

last: Prehn


title: Copy Number Abnormalities in Sporadic Canine Colorectal Cancers

status: Public on Jan 19 2010

first: Jie

last: Tang

address: 120 Green St

department: Institute of Bioinformatics

summary: Human colorectal cancer (CRC) is one of the better-understood systems for studying the genetics of cancer initiation and progression. To develop a cross-species comparison strategy for identifying CRC causative gene or genomic alterations, we performed array comparative genomic hybridization (aCGH) to investigate copy number abnormalities (CNAs), one of the most prominent lesion types reported for human CRCs, in 10 spontaneously occurring canine CRCs. The results revealed for the first time a strong degree of genetic homology between sporadic canine and human CRCs. First, we saw that between 5 and 22% of the canine genome was amplified/deleted in these tumors, and that, reminiscent of human CRCs, the total altered sequences directly correlated to the tumor’s progression stage, origin, and likely microsatellite instability status. Second, when mapping the identified CNAs onto syntenic regions of the human genome, we noted that the canine orthologs of genes participating in known human CRC pathways were recurrently disrupted, indicating that these pathways might be altered in the canine CRCs as well. Lastly, we observed a significant overlapping of CNAs between human and canine tumors, and tumors from the two species were clustered according to the tumor subtypes but not the species. Significantly, compared with the shared CNAs, we found that species-specific (especially human-specific) CNAs localize to evolutionarily unstable regions that harbor more segmental duplications and interspecies genomic rearrangement breakpoints. These findings indicate that CNAs recurrent between human and dog CRCs may have a higher probability of being cancer-causative, compared with CNAs found in one species only

sample_organism:
Homo sapiens
Canis lupus familiaris

platform_organism:
Homo sapiens
Canis lupus familiaris

overall_design: Comparison of human and dog colon cancer CNAs

first: Shoshona

last: Le

first: Mucheng

last: Zhang

last: Zwick

first: Shaying

last: Zhao


title: Gene expression microarray analysis of heat stress in the soil invertebrate Folsomia candida

status: Public on Jan 16 2010

city: Amsterdam

first: Benjamin

last: Nota

address: Plesmanlaan 125

department: Research Facilities

institute: Sanquin

summary: Temperature is an important ecological condition, and sudden temperature changes in soil can induce stress in soil-dwelling invertebrates. Soil animals can move to more favorable habitats and/or adapt physiologically to a stressful environment. Hyperthermic conditions will impact gene expression as one of the first steps. We use a transcriptomics approach to identify the transcripts of which expression changed in response to heat stress in the springtail Folsomia candida using a 5,131 probe microarray. A temperature shift from 20°C to 30°C for 30 minutes significantly altered the expression of 142 genes, of which 116 were upregulated, and 26 downregulated. Many upregulated genes encoded heat shock proteins (Hsps) or enzymes involved in the synthesis of ATP, such as members of the electron transport chain. Furthermore, genes involved in oxidative stress and anion-transporting ATPases were upregulated. Downregulated were glycoside hydrolases, involved in catalysis of certain disaccharides, which indicate an accumulation of stress-protective disaccharides. The microarray results from this study, which were validated using quantitative RT PCR, reveal a mild response to heat shock in this soil invertebrate, relative to other organisms. This may be due to specific ecological factors during evolution of soil invertebrates, such as the relatively stable temperatures in the soil habitat. This study presents potential candidate genes for future functional studies concerning thermal stress in soil-dwelling invertebrates, like e.g., the investigation of the heat hardening process.

overall_design: Folsomia candida was first acclimated to LUFA 2.2 soil at 20 C for two days. Then animals were either exposed to 30 C for 30 minutes (heat shock treated), or were not heat shocked (reference). For each treatment 4 biological replicates were used, replicate samples consisted of total RNA extracted from ~30 animals exposed in the same jar to either reference or heat shock. Each unique heat shock treated sample was hybridized to a unique reference sample. In total in 4 hybridizations of 8 samples, was performed. The dyes were evenly distributed, which means that for each treatment two samples were labeled with cy3 and the other two with cy5.

first: Nico

middle: M

last: van Straalen

first: Bauke

last: Ylstra

first: Dick

last: Roelofs


title: Transcriptional profiling of the Borrelia burgdorferi bb0647 mutant

city: Dallas

first: zhiming

last: ouyang

address: 6000 Harry Hines Blvd

department: Department of Microbiology

institute: UT Southwestern Medical Center

summary: RNA was isolated from late-log pahse wild-type Borrelia burgdorferi B31 and the bb0647 mutant grown in BSKH media at 37 degree, 5% CO2. cDNA was synthesized, labeled and hybridized to the 70mer oligonucleotide B. burgdorferi array. Slides were scanned using axon scanner and image were analyzed using GenePix 6.0. Dta were further analysed using the professional software Acuity 4.o, based on a ratio-based normalization.

overall_design: Samples were labeled by either Cy3 or Cy5. 5 hybs were performed, including dye-swaps.

first: Zhiming

last: Ouyang

last: Norgard


title: Transcriptomic and phenotypic responses of L. monocytogenes with different growth efficiencies under acidic conditions

city: Hobart

first: John

middle: Phillip

last: Bowman

state: Tasmania

address: College Road, Sandy Bay

department: School of Agricultural Science

country: Australia

institute: University of Tasmania

summary:
A screening process was used to determine the aciduric capacities of a diverse set of Listeria monocytogenes strains and was based on the capacity to grow at pH 5.0 in the presence 4 different organic acids. Food and clinical isolates tended to be more resistant but strain genetic groupings were found to be only weakly linked to sodium diacetate (SDA)-resistance. Representative and comparatively aciduric food isolates FW04/0023 and FW04/0025 were found to accumulate reduced levels of acetate anion and K+ ion during growth in the presence of SDA, compared to more acid sensitive reference strains EGD (ATCC BAA-739) and ATCC 19111. The aciduric nature of FW04/0023 and FW04/0025 was also reflected by their comparatively high tolerance to pH 2.4 acid challenge; a property boosted by the presence of SDA, and growth-phase dependent acid tolerance. Transcriptomic analyses revealed increased levels of SDA did not activate or promulgate the response of any of the known acid protective mechanisms, except for acetoin biosynthesis.  Elevated levels of unprotonated SDA (20 mM SDA at pH 5.0) was found to have broad effects on gene expression that could be differentiated from effects caused by mildly acidic conditions (pH 5.0) and substantial strain variation was also found. Collated SDA mainly effect genes associated with virulence, cell wall processes, DNA repair, and cofactor and lipid biosynthesis. Strain FW04/0025 was more responsive to elevated unprotonated SDA increasing the expression of 222 genes (>2-fold change, p<0.05), compared to 112 genes for strain EGD. Key differences between the strains in relation to SDA-enhanced transcript abundance in FW04/0023 were primarily associated with the cell wall, oxidative stress management, intermediary metabolism, and several hypothetical proteins. This complement of different responses could potentially alter phenotypes resulting in different cell envelope properties and metabolic properties that effect accumulation of acetate anion. The data demonstrates that amongst different L. monocytogenes strains acetate has differing effects that may ultimately influence growth efficiency under stressful conditions relevant to acidic foods and the gastrointestinal environment.
The microarray component of the experiments had the aim of determining:  i) To examine the affect of elevated levels of unprotonated sodium diacetate (21 mM sodium diacetate added to cultures at pH 5.0, resulting in ~7.7 mM unprotonated acetate) compared to mild acid stress at pH 5.0 in which lower levels of acetate accumulates through natural end-product formation.  ii) To examine the gene expression responses of two L. monocytogenes strains that had different intrinsic acid resistances, including an organic acid resistant strain FW04/0025 and a more sensitive reference strain EGD on which genome the microarray oligonucleotide set is based.

overall_design: Two to  four  biologically replicated control cultures were labelled with Cy3 for each treatment sample with two sets of experimental conditions investigated for two different L. monocytogenes strains .  Condition 1) – Mildly acidic stress:   Control culture (Cy3-labelled RNA) - strains were grown to early- mid exponential growth phase at 25°C (in a shaking water bath) in brain-heart infusion broth at pH 7.3.   Test cultures (Cy5-labelled RNA) – strains grown to early- mid exponential growth at 25°C phase in BHI broth adjusted to pH 5.0 (in a shaking water bath).  Condition 2) – Organic acid (sodium diacetate) stress:   Control culture (Cy3-labelled RNA) - strains were grown to exponential growth phase at 25°C (in a shaking water bath) in BHI broth at pH 5.0.   Test cultures (Cy5-labelled RNA) – strains grown to exponential growth at 25°C phase in BHI broth adjusted to pH 5.0 and amended with 21 mM (0.3% w/v)sodium diacetate (in a shaking water bath).

middle: P


title: mRNA profiling in rectus abdominis muscle from patients with upper GI cancer

city: Stirling

first: Iain

last: Gallagher

address: Room 4B133 Cottrell Building, University of Stirling, Airthrey Road

department: Faculty of Health Sciences and Sport

institute: University of Stirling

summary: Rectus abdominis muscle biopsies were obtained from 65 upper gastrointestinal (UGI) cancer patients during open surgery and RNA profiling was performed on a subset of this cohort (n=21) using the Affymetrix U133+2 platform with the aim of identifying biomarkers of cancer related muscle wasting.

overall_design: N = 21 (18 UGI cancer; 3 non cancer surgery)

first: James

last: Timmons


title: growth dependent expression of s. aureus mssa476

status: Public on Jan 25 2010

city: Bilthoven

first: Tessa

middle: E

last: Pronk

address: A.v Leeuwenhoeklaan 9

department: GBO

institute: RIVM

summary:
Survival and pathogenesis of Staphylococcus aureus in the host requires the ability to respond to changes. Therefore, tight regulation of gene expression via various regulators is essential. Also, the organization of genes in operons is of influence on the regulation of gene expression.  Knowledge of gene expression under different conditions and the ability to accurately predict operons are important steps towards understanding the transcriptional regulation, function and pathogenesis.
A whole genome Agilent microarray was developed for the highly virulent, community-acquired MSSA476.During standard growth in a defined medium, we were able to determine four basic gene expression patterns of S. aureus for both virulence and non-virulence genes. In addition, we predicted operon structures by calculating Pearson correlation coefficients of the transcriptional profiles for all adjacent probes over all time points and replicas.
In this study, we have set a basis for the knowledge on gene expression of MSSA476 during growth. Moreover, the correlation of time-dependent transcriptional profiles of adjacent probes seems to be a promising approach to predict operon structures.

sample_organism: Staphylococcus aureus subsp. aureus MSSA476

overall_design: Five separate cultures of S. aureus mssa476 were grown. Of each replicate culture, samples were taken at 1, 2, 3, 4, 5, 6 and 9 h post inoculation (p.i.). In total, this amounts to 35 samples (7 time points in 5 replicates).

first: Nicole

last: Smits

first: Wim

middle: C

last: de Leeuw

first: Ad

last: Fluit

first: Edwin

last: Boel

first: Timo

last: Breit


title: Gene Expression Profile of Hyperoxic/hypoxic Retinas in Mouse Model of Oxygen-induced Retinopathy

status: Public on Jan 15 2010

city: Fukuoka

first: Keijiro

last: Ishikawa

address: 3-1-1  Maidashi, Higashi-ku

department: Ophthalmology

institute: Kyushu University

summary: Retinal ischemia is believed to initiate a series of events that leads to retinal neovascularization observed in several retinal diseases such as diabetetic retinopathy and retinal vein occlusion. Although some molecules such as VEGF and MCP-1 have been implicated in the process, its underlying mechanisms remain elusive. In order to identify genes associated with retinal ischemia, we performed gene expression analyses in retinas of mouse model of oxygen-induced retinopathy using DNA microarray technology.

overall_design: C57BL/6N pups were placed in a 75% oxygen environment on postnatal day 7 (P7) for 5 days and then returned to room air.  Retinas were removed from mice at 0 (P12), and 12 hours (P12.5) after ischemia. To isolate sufficient total RNA for labeling protocols, 2 retinas from one mouse were pooled. The results of three experiments for each condition (biological replicates) were used for the statistical analyses.

first: Shigeo

last: Yoshida


title: Alternative splicing and gene expression in a mouse model of myotonic dystrophy

city: Santa Cruz

first: Manny

last: Ares

state: CA

address: 1125 High St

department: Molecular and Cellular Biology

institute: UCSC

summary:
Distinct RNA-mediated impacts on alternative splicing and extracellular matrix gene expression in a mouse model of myotonic dystrophy.

Myotonic dystrophy (DM1) is associated with expression of expanded CTG DNA repeats as RNA (CUGexp RNA). To test whether CUGexp RNA creates a global splicing defect, we compared skeletal muscle of two mouse DM1 models, one expressing a CTGexp transgene, and another homozygous for a defective Mbnl1 gene.  Strong correlation in splicing changes for ~100 new Mbnl1-regulated exons indicates loss of Mbnl1 explains >80% of the splicing pathology due to CUGexp RNA.  In contrast, only about half of mRNA level changes can be attributed to loss of Mbnl1, indicating CUGexp RNA has Mbnl1-independent effects, particularly on mRNAs for extracellular matrix (ECM) proteins.  We propose that CUGexp RNA causes two separate effects: loss of Mbnl1 function, disrupting splicing, and loss of another function that disrupts ECM mRNA regulation, possibly mediated by MBNL2.  These findings reveal unanticipated similarities between DM1 and other muscular dystrophies.

overall_design:
MBNL1 knockout and HSALR mice on FVB background.
To test whether CUGexp RNA creates a global splicing defect, we compared skeletal muscle of two mouse DM1 models, one expressing a CTGexp transgene, and another homozygous for a defective Mbnl1 gene. These samples were compared to the skeletal muscle of a wildtype mouse.

first: Charles

last: Thornton

first: Maurice

last: Swanson

first: Hongqing

last: Du

first: Lily

last: Shiue

first: Robert

last: Osborne

first: Daniel

last: Tuttle

first: Tyson

last: Clark

last: Donohue


title: Osmotic stress in Listeria monocytogenes serotype 4b strain F2365 grown in BHI medium and BHI medium with 1.2% NaCl

city: Mississippi State

first: Chinling

last: Wang

state: MS

address: College of Veterinary Medicine P.O. box 6100

department: Basic Sciences

institute: Missippi State University

summary: Gene expression profiles of L. monocytogenes strain F2365 to osmotic stress was examined. Based on a log2 scale, four and 24 genes were upregulated and downregulated by the stress, respectively.The expression level of ATP-binding cassette superfamily and ribosomal protein L2 genes were increased, whereas genes associated with PTS system, its metabolic enzymes, pathogenesis, and hypothetical protein were downregulated.

overall_design: To examine the gene expression profiles of L. monocytogenes grown in the medium added with or without 1.2% NaCl , L. monocytogenes strains 4b F2365 aminosilane-coated cDNA microarray slide was used. Cy3 and Cy5 dyes were used for cDNA labelling. Data were obtained from three biological and four technical replicates (n = 12). Each biological replicate included a flip dye assay.

first: Dongryeoul

last: Bae


title: Salmonella Typhiumurium LT2, wild type versus rfaC mutant.

status: Public on Jan 14 2010

city: Brno

first: Magdalena

last: Crhanova

address: Hudcova 70

department: Bacteriology

country: Czech Republic

institute: Veterinary research institute

summary:
Transcriptional analysis of rfaC mutant strain compared to an isogenic wild type strain Salmonella Typhimurium.

Keywords: Genetic modification.

sample_organism: Salmonella enterica subsp. enterica serovar Typhimurium str. LT2

overall_design: Three independent biological replicates.

first: Ivan

last: Rychlik


title: Genome-wide direct targets of PIL5 (ChIP-chip analysis of PIL5)

status: Public on Jan 12 2010

city: Daejeon

first: Giltsu

last: Choi

address: 335 Gwahangno

department: Biological Sciences

country: South Korea

institute: KAIST

summary:
PIL5 is a key negative regulator of phytochrome mediated seed germination and PIL5 protein is degraded by red light irradiation through phytochrome.
The ChIP-chip analysis aimed to find various PIL5 direct targets in the Arabidopsis genome.

overall_design: PIL5-myc over-expressed seeds were used for ChIP-chip analysis. Conventional ChIP DNA was amplified by using LM-PCR method. Input DNA (before immuno-precipitation) was used as control DNA.Total three biological replicates were used for the ChIP-chip analysis.


title: miRNA expression signatures for human stomach biopsy samples, H. pylori positive versus negative

status: Public on Jan 08 2010

city: Nagasaki

first: Kayoko

last: Matsushima

address: Sakamoto 1-7-1

department: Gastroenterology and Hepatology

institute: Nagasaki university

summary: We have employed whole microRNA microarray with the potential to distinguish H.pylori infection.Among the 470 human miRNAs represented on the array chip, 228 were undetectable or expressed below the background and so were eliminated, leaving 242 miRNAs for the supervised analysis. When comparing 10 H. pylori-negative and nine H. pylori-positive subjects, 55 miRNAs were deemed significantly different on the basis of microRNA arrays.

overall_design: During endoscopy, a biopsy specimen was obtained from the gastric antrum along the lesser curvature. Biopsies of 10 H. pylori-negative and nine H. pylori-positive patients' subjects were performed.Exclusion criteria were: age <18 or >80 y, pregnancy, body mass index (BMI) >30 kg/m2, diabetes mellitus, cachectic state (including cancer), systemic infection, liver disease, renal impairment, use of medications effective against H. pylori during the preceding 3 months, alcohol abuse, drug addiction, and chronic corticosteroid or nonsteroidal anti-inflammatory drug use. None of the subjects had undergone gastrointestinal surgery.

first: Hajime

last: Isomoto


title: Growth phase-dependent activation of the DccRS regulon of Campylobacter jejuni

status: Public on Jan 09 2010

city: Albany

first: Craig

middle: T

last: Parker

address: 800 Buchanan St

department: PSM

institute: USDA ARS

summary: Two-component systems are widespread prokaryotic signal transduction devices which allow the regulation of cellular functions in response to changing environmental conditions. The two-component system DccRS (Cj1223-Cj1222) of Campylobacter jejuni is important for the colonization of chickens. Here we dissected the DccRS system in more detail and provide evidence that the sensor DccS selectively phosphorylates the cognate effector DccR. Microarray expression profiling, real-time RT-PCR, EMSA, and primer extension analyses revealed that the DccRS regulon of strain 81116 consists of five promoter elements all containing the consensus direct repeat sequence WTTCAC-N6-TTCACW covering the putative -35 promoter regions. One of these promoters is located in front of an operon encoding a putative macrolide efflux pump while the others are in front of genes coding for putative periplasmic or membrane proteins. The DccRS regulated genes in C. jejuni strain 81116 are needed to enhance early in vivo growth of C. jejuni in 7-day old chicken. The DccRS system is activated in the late stationary bacterial growth phase, probably via the release of metabolic products. Whole genome mRNA profiling and real-time RT-PCR analysis under these conditions demonstrated that the system has no influence on the transcription of genes outside the DccRS regulon.

sample_organism: Campylobacter jejuni subsp. jejuni 81116

overall_design: Two-condition experiment, C. jejuni strain WT81116 vs. C. jejuni strain 81116dccR::cm. Technical replicates: 3 WT81116, 3 81116dccR::cm, independently grown to stationary phase and harvested. One replicate per array.   A type 2 gene expression experimental design was used, with fluorescently labeled genomic DNA as a reference channel in each experiment as described by Lucchini, S., et al. 2005.  Infect Immun 73:88-102.

first: M

last: Wösten

first: L

last: Heijmen-van Dijk

first: C

last: Guilhabert

first: Y

last: Janssen

first: J

last: Wagenaar

last: van Putten


title: Molecular profiling identifies prognostic subgroups of pediatric glioblastoma and shows increased tumor YB-1 expression

city: Montreal

first: Andre

last: Nantel

state: QC

address: 6100 Royalmount

department: Biotechnology Research Institute

institute: National Research Council of Canada

summary: Our aim was to gain insight into the molecular pathways that distinguish pediatric- from adult-onset glioblastomas.

overall_design: Amplified RNA from Type IV Glioblastoma brain tumor cells compared to amplified RNA from a pool of 3 normal brain samples. Pediatric and adult patients. 2-6 replicate hybridizations per patient.

first: Damien

last: Faury

first: Takrima

last: Haque

first: Nada

last: Jabado