title: Gene expression profile in pelvic organ prolapse

status: Public on Sep 20 2008

city: Honolulu

first: Shawna

last: Brizzolara

state: HI

address: 1319 Punahou St #824

department: Obstetrics and Gynecology

institute: Shawna Brizzolara, MD

summary:
Introduction and Hypothesis: Identify processes contributing to pelvic organ prolapse (POP) by transcriptional profiling of pelvic connective tissue in conjunction with light microscopy. Methods: We performed a frequency matched case-control study of women undergoing hysterectomy. Total RNA, extracted from uterosacral and round ligament samples used to generate labeled cRNA, was hybridized to microarrays and analyzed for the expression of 32,878 genes. Significance Analysis of Microarrays, (Stanford University, CA), identified differentially expressed genes used for ontoanalysis, and quantitative PCR (qPCR) confirmed results. Light microscopy confirmed tissue type and assessed inflammatory infiltration. Results:  The analysis of thirty-four arrays revealed 249 differentially expressed genes with fold changes larger than 1.5 fold and false discovery rates ≤5.2%. Immunity and Defense was the most significant biological process differentially expressed in POP. Selected qPCR confirmed 4 genes. Light microscopy showed no inflammatory infiltrates. Conclusions: Genes enriched for Immunity and Defense contribute to POP independent of inflammatory infiltrates.

Keywords: whole tissue (endopelvic fascia) type comparison

overall_design: This was a group matched case control study of 8 women with pelvic organ prolapse versus 9 non-prolapse controls, both undergoing hysterectomy for benign conditions.  Two separate pelvic support tissues were collected from each patient.  The uterosacral ligament and round ligament tissue was removed at the time of hysterectomy, RNA was extracted and ABI whole genome chips used to identify differences in expression profiles of individual samples.  Various ethnic groups, age groups and menopausal status were included.

first: Johann

last: Urschitz

first: Jeffery

last: Killeen


title: Distinct differences in chromatin structure at telomeric X- and Y-elements in budding yeast.

status: Public on Jul 01 2009

city: Gothenburg

first: Claes

middle: M

last: Gustafsson

address: Medcinaregatan 9A

department: Department of Medical biochemistry and cell biology

country: Sweden

institute: Gothenburg University

type: Genome binding/occupancy profiling by genome tiling array

summary:
The telomeric repeat sequences (TGTGG) are the same at all chromosome ends, but the number and position of subtelomeric X and Y repeat elements vary. Using chromatin immunoprecipitation and genome-wide analyses, we here demonstrate that the subtelomeric X and Y elements have distinct structural and functional properties. Y elements are transcriptionally active and highly enriched in histones, whereas X elements are repressed and completely devoid of nucleosomes. In contrast to X elements, the Y elements lack the classical hallmarks of heterochromatin, e.g. high Sir3 and Rap1 occupancy as well as low levels of histone 4 lysine 16 acetylation, Our analyses refute the idea that subtelomeric silencing is a general phenomenon and suggest that the presence of X and Y elements govern the chromatin structure and transcription activtiy at individual chromosome ends.

Keywords: nucleosome position and Histone 4 lys 16 acetylation Chip on CHIP

overall_design: Two experiment series were included in this study. First is nucleosome position of wild type budding yeast strain. Second one is H4K16Ac in wild type strains. Genome background control was used as the control channel.

first: Xuefeng

last: Zhu


title: Transcriptional changes in blood from metabolic syndrome patients after a period of high intensity interval training

status: Public on Jun 01 2008

city: TRONDHEIM

first: Anja

last: Bye

address: Olav Kyrres gt 9

country: Norway

institute: NTNU

summary:
Abstract

The metabolic syndrome is a cluster of conditions that predispose for diabetes and cardiovascular disease. Nine metabolic syndrome patients were recruited to 48 workouts of interval training. At the end of the study, all patients significantly reduced their risk of cardiovascular disease (in terms of VO2max, blood pressure and plasma lipid). Exercise-induced transcriptional changes may provide new mechanistically insights in the area of improved health by exercise.

Aim: Determine whether transcriptional changes occurred in the blood cells of patients after the exercise program. We hypothesized that significantly altered biological processes in blood would include a set of genes at least partly responsible for the improvement seen in this patient group after the exercise programme.

Methods: Blood from five patients were collected in PAXgene tubes pre and post the exercise period. RNA was extracted and run on microarrays.

Results: Gene set enrichment analysis revealed twelve significantly enriched biological processes and molecular functions that were up-regulated post exercise. Seven processes and functions were down-regulated (nom p-value <0.05). Exercise induced a down-regulation of plasma mRNA- and protein levels of arginase-I and vWf, which might explain improved risk profile and endothelial function of the metabolic syndrome patients.

Conclusion: After the exercise period was completed, the metabolic syndrome patients had decreased transcription of genes associated with blood clotting and steroid metabolism. Arginase-I was decreased post exercise, which may explain the previous reported improved NO-availability and endothelial function.
Keywords: aerobic capacity, cardiovascular disease, microarray, arginase, blood clotting

overall_design:
Patient group
The patients included in this study were randomly selected from a population of patients described in our recently published study of the metabolic syndrome. Briefly, nine patients diagnosed with the metabolic syndrome, according to the WHO-criteria were recruited for aerobic interval training. All subjects provided written, informed consent, and the regional committee for medical research ethics approved the protocol. The participants’ diet was registered during the study period.

The blood from 5 of these patients were analyzed for whole-genome gene expression pre and post the exercise period, to study transcriptional alterations in blood that might be able to explain.

first: Arnt

middle: E

last: Tjønna

first: Tomas

last: Stølen

first: Ulrik

last: Wisløff


title: Comparison of gene expression between follicular thyroid adenoma and follicular thyroid carcinoma

status: Public on Feb 28 2009

city: Frankfurt

first: Nora

last: Hinsch

address: Theodor-Stern-Kai 7

department: Pathology

country: Germany

institute: University hospital Frankfurt/Main

summary:
Follicular thyroid tumours were investigated using global gene expression analysis. Aim of this study was the identification of new markers for follicular thyroid carcinoma.

Keywords: cell type comparison

overall_design: Gene expression analysis of 4 follicular thyroid adenomas, 4 follicular thyroid carcinomas, and 4 microinvasive follicular thyroid carcinomas.

first: Frank

last: Matthias

first: Claudia

last: Döring

first: Christian

last: Vorländer

first: Martin-Leo

last: Hansmann


title: New specific molecular targets for radiochemotherapy in colorectal cancer

status: Public on Apr 23 2009

city: Tromsø

first: Ruth

middle: Hracky

last: Paulssen

address: MH-Bygget Breivika

department: Institute of Clinical Medicine

institute: Universitetet i Tromsø

summary: A promising treatment for patients with advanced colorectal cancer is preoperative radiochemotherapy. The early side effects of this treatment have been considered to be acceptable. The aim of this study was to identify the effects of preoperative radiochemotherapy (PRT) on gene expression in tumour and normal colon rectal tissue form the same patients, before and after PRT.  For that purpose, tissue samples from ten patients with operable rectal adenocarcinomas were collected for use in whole genome–microarray based gene expression analysis. A factorial experimental design allowed us to look solely at the radiation effect on tumours. This resulted in 4496 differentially expressed genes in tumour tissue with p<0.05. In addition to known markers for radiochemotherapy, a Gene Set Enrichment Analysis (GSEA) showed a significant enrichment in gene sets associated with cell adhesion and leukocyte transendothelial migration (TEM).  We conclude that radiochemotherapy has a greater effect in tumour tissue gene expression than normal tissue.  Not only is the effect on normal tissue limited compared to tumour, but significantly different gene sets are enriched. The profound change of cell adhesion molecule expression in tumour tissue could either increase the risk of metastasis, or decrease the tumours invasive potential. Further characterization of genes involved in cell adhesion and leukocyte TEM may give new insights into the molecular responses to radiochemotherapy in colorectal cancer.

overall_design: Factorial experimental design

middle: H

first: Christopher

middle: G

last: Fenton

first: Kristin

last: Snipstad

first: Jørn

last: Kjæve

first: Endre

last: Anderssen


title: Annotating Low Abundance and Transient RNAs in Yeast using Tiling Microarrays

status: Public on Dec 18 2008

city: Stanford

first: Gavin

last: Sherlock

state: CA

address: 300 Pasteur Drive

department: Genetics

country: USA

institute: Stanford University

type: Expression profiling by genome tiling array

summary:
A complete description of the transcriptome of an organism is crucial for a comprehensive understanding of how it functions, how its transcriptional networks are controlled, and my provide insights into the organism's evolution.  Despite the status of Saccharomyces cerevisiae as arguably the most well understood model eukaryote, we still do not have a full catalog and understanding of all its genes.  In order to interrogate the transcriptome of S. cerevisiae for low abundance or rapidly turned over transcripts, we deleted elements of the RNA degradation machinery with the aim of increasing the relative abundance of such transcripts.  We then used high-resolution tiling microarrays and ultra high-throughput sequencing (UHTS) to identify and map the locations of unannotated transcripts that are more abundant in the RNA degradation mutants relative to wild-type cells, revealing 540 currently unannotated, presumably low abundance or short-lived RNAs, of which 231 are previously unknown and unique to this study.  It is likely that many of these represent cryptic unstable transcripts (CUTs) which are rapidly degraded and whose function(s) within the cell are still unclear, while others may represent novel functional transcripts.  Of the 271 transcripts we identified in current intergenic regions, greater than 90 percent have lower conservation scores amongst closely related yeast species than 95 percent of the verified ORFs in S. cerevisiae; such regions of the genome have typically been less well studied, and by definition encode transcripts that distinguish S. cerevisiae from these closely related species.

Keywords: Saccharomyces cerevisiae, transcriptome, salt shock, xrn1, rrp6, lsm1, pat1, RNA degradation

overall_design: Two samples, a wild-type reference and a mutant containing 4 deletions, were subjected to high salt shock, and total RNA was harvested.  PolyA RNA was purified, and both polyA and total RNAs were hybridized to Forward and Reverse Affymetrix Tiling Microarrays.  No replicates were done for any of the hybridizations.

first: Albert

last: Lee

first: Kasper

middle: D

last: Hansen

first: James

last: Bullard

first: Sandrine

last: Dudoit


title: Gene Expression Signature in Peripheral Blood Detects Thoracic Aortic Aneurysm

status: Public on Jan 29 2008

city: Foster City

first: Catalin

last: Barbacioru

address: 850 Lincoln Centre Dr

institute: Applied Biosystems

type:
Expression profiling by array
Expression profiling by RT-PCR

summary:
This study identified informative gene expression signatures in peripheral blood cells that can characterize TAA status and subtypes of TAA. Moreover, a 41-gene classifier based on expression signature can identify TAA patients with high accuracy. The transcriptional programs in peripheral blood leading to the identification of these markers also provide insights into the mechanism of development of aortic aneurysms, and highlight potential targets for therapeutic intervention. The classifier genes identified in this study and validated by TaqMan® real-time PCR assays define a set of promising potential diagnostic markers, setting the stage for a blood-based gene expression test to facilitate early detection of TAA.
Keywords: Gene Expression Signature

overall_design:
Whole Genome gene expression profiles from a total of 94 peripheral blood samples (collected from 58 individuals diagnosed with TAA and 36 normal controls) were analyzed in this study. Significance Analysis of Microarray (SAM) analysis identified potential signature genes characterizing TAA vs. normal, ascending vs. descending TAA, as well as sporadic vs. familial TAA. Using a training set containing 36 TAA patients
and 25 controls, a 41-gene classification model was constructed for detecting TAA status and an overall accuracy of 78 ± 6% was achieved. Testing this classifier on an independent validation set containing 22 TAA samples and 11 controls yielded an overall classification accuracy of 78%. These 41 classifier genes were further validated by TaqMan® real-time PCR assays. Classification based on the TaqMan® data  eplicated the microarray results and achieved 80% classification accuracy on the testing set.

first: Wang

last: Yulei

middle: C

first: Dov

last: Shiffman

first: Sriram

last: Balasubramanian

first: Olga

last: Iakoubova

first: Maryann

last: Tranquilli

first: Gonzalo

last: Albornoz

first: Julie

last: Blake

first: Necip

middle: N

last: Mehmet

first: Dewi

last: Ngadimo

first: Karen

last: Poulter

first: Frances

last: Chan

first: Raymond

middle: R

last: Samaha

first: John

middle: A

last: Elefteriades


title: EBER2 RNA-induced transcriptome changes identify cellular processes likely targeted during Epstein Barr Virus infection.

status: Public on Oct 02 2008

city: Bures-sur-Yvette

first: Arndt

last: Benecke

address: 35 route des Chartres

department: Integrative Biology

country: France

institute: Institut des Hautes Etudes Scientifiques & I.R.I.-CNRS

summary:
Little is known about the physiological role of the EBER1 and 2 nuclear RNAs during Epstein Barr viral infection. The cellular transcription response to EBER2 expression using the wild-type and an internal deletion mutant was determined. Significant changes in gene expression patterns were observed. A functional meta-analysis of the regulated genes points to inhibition of stress and immune responses, as well as activation of cellular growth and cytoskeletal reorganization as potential targets for EBER2 RNA. Different functions can be assigned to different parts of the RNA. These results provide new avenues to the understanding of EBER2 and EBV biology.
Keywords: Overexpression of non-translated RNA

overall_design: Biological relicates, 3 conditions: empty expression vector (pUC-18),  pUC-18-EBER2, pUC-18-EBER2-L2

first: Sebastian

last: Eilebrecht

first: François-Xavier

last: Pellay

first: Peter

last: Odenwälder

first: Guillaume

last: Brysbaert

first: Bernd-Joachim


title: Identification of JA/COI1-regulated genes in tomato reproductive tissue and JA-treated roots.

status: Public on Sep 01 2007

city: Rockville

first: Jia

last: Liu

state: MD

address: 9712 Medical Center Drive

institute: Plant Genomics

summary:
Jasmonic acid (JA) is a fatty-acid derived signaling molecule that regulates a wide variety of plant developmental and stress-related responses. We identified a jasmonate-insensitive1 (jai1) mutant of tomato that is defective in the LeCOI1 gene, which plays an essential role in induced resistance of tomato to a broad spectrum of pests, and is also required for maternal control of seed and fruit development.  The present study is aimed at identifying genes involved in two JA-signaled processes that have not been studied in other plants. The first specific aim is to identify the set of JA/COI1-regulated genes in tomato reproductive tissues, with emphasis on genes expressed in maternal tissues. We hypothesize that JA/COI1 signaling regulates maternal processes that nurture the developing embryo and seed. The second aim of the study is to identify JA/COI1-regulated genes that are specifically expressed in tomato roots. Although it has been shown that JA promotes root defenses against various pests, virtually nothing is known about the JA-regulated transcriptome in roots.  Tomato (Solanum lycopersicum) cv Micro-Tom wild-type and jai1 seed were grown in peat pots and maintained in growth chambers. To identify COI1-regulated genes in reproductive tissues, flower buds, open flowers and immature fruit at 7 and 17 days post-pollination were collected from 8 to 10 week old plants. To identify COI1-regulated genes in roots, 3 week old plants were exposed to exogenous JA by a single soil drench with 0.1mM JA, and roots were collected after 1, 6, 12 and 24 hours. Plants treated with water were used for the 0hr time point. All tissue was snap-frozen in liquid nitrogen, and RNA was extracted and analyzed as described in the TIGR Potato Microarray protocol (phenol method). Three biological replicates were collected for this study.
Keywords: Direct comparison

overall_design: 26 hybs total

first: Howe

last: Gregg


title: Collateral Flow Evokes Distinct Endothelial Gene Expression

status: Public on Jan 01 2009

city: Boston

middle: J

last: Mack

state: MA

address: 77 Avenue Louis Pasteur, NRB-752

institute: Harvard Medical School

summary: Biomechanical forces influence vascular function, but the molecular mechanisms regulating many of these mechano-activated cellular events remain largely uncharacterized.  In particular, the vascular endothelium can sense alterations in hemodynamic shear stress which, in the context of adaptive remodeling or arteriogenesis, has been shown to lead to the generation of endothelial-derived signals critical for controlling blood vessel structure and function.  Here, we sought to define the arteriogenic responses evoked in endothelial cells exposed to flow using transciptional profiling.  Analysis of these transcriptional programs identified several genes previously shown to be important for endothelial-smooth muscle interactions and vascular remodeling, including the transcription factor kruppel-like factor 2 (KLF2).

overall_design: A lumped parameter model of the human coronary collateral circulation was used to simulate previously uncharacterized normal (NCC) and adaptive remodeling (ACC) coronary collateral shear stress waveforms.  These waveforms were then applied to cultured human endothelial cells (EC) and the resulting differences in EC gene expression were assessed by genome-wide transcriptional profiling to identify genes distinctly regulated by collateral flow.

first: Yuzhi

last: Zhang

first: Roger

last: Kamm

first: Guillermo

last: Garcia-Cardena


title: Induction of tobacco genes by Agrobacterium oncogene 6b

summary:
Tumor induction by Agrobacterium is due to transfer of T-DNA genes. We study the 6b T-DNA oncogene that induces tumors on species like Nicotiana glauca, but for which no precise growth induction mechanism has been found sofar. The 6b gene belongs to a larger group of T-DNA genes including rolB, rolC and orf13, no homologies are known with other genes. In N. tabacum, 6b modifies responses to auxins and cytokinins and induces strong morphological changes including cell enlargement, abnormal division, double flowers, enations and root shortening (Helfer et al., 2003, Grémillon et al., 2004). Others have localized 6b and rolB proteins in the nucleus, where they could modify transcription (Kitakura et al., 2002, Moriuchi et al., 2004).  Recently, we developed a system to study early  6b-induced growth changes, using a dexamethasone-inducible 6b gene (dex-T-6b). Leaves of homozygous dex-T-6b plants are infiltrated with a 10 mM MgSO4 solution, with or without 20 micromolar dex. In the presence of dex, a reproducible increase in glucose and fructose concentrations is observed. Such changes are also found in cultured leaf discs and in that case accompanied by an increase in expansion. 6b-induced changes are detected as early as 7 hours after dex infiltration. Western analysis shows that dex-T-6b plants have undetectable 6b protein levels without induction, and rapidly accumulate 6b protein upon induction. We propose to study gene expression in induced and non-induced leaves at different times of induction: 0-3-6-9 and 12 hours. The data will be used to establish whether the 6b gene induces specific genes, and whether these genes are involved in hexose changes and cell expansion. We will also test the effect of infiltrating tobacco leaves with 10 mM MgSO4 by comparison with non-infiltrated leaf tissues in order to correct for changes due to the infiltration procedure.
Keywords: Reference design

overall_design: 30 hybs total

first: Otten

last: Léon


title: Nucleosome occupancy in wild type and histone H3(A111G) yeast

status: Public on Jul 19 2008

city: Albany

first: Randall

middle: Heywood

last: Morse

state: NY

institute: Wadsworth Center

summary:
We screened for histone H3 mutations that cause derepression of the uninduced CHA1 promoter and identified six mutants, including one, H3(A111G), that causes a constitutively active chromatin configuratin at the uninduced CHA1 promoter.  We examined genomewide nucleosome occupancy in the A111G mutant and compared it to wild type yeast; that is the data deposited here.

Keywords: Nucleosomal DNA from wild type and histone H3(A111G) mutant yeast

overall_design: Three A111G mutant and two wild type samples were analyzed.  These were compared to three genomic DNA samples (Lee et al., Nature Genetics doi:10.1038/ng2117) to generate the .bar files deposited here.

first: Qiye

last: He


title: Histone H3 N-terminus Regulates Higher Order Structure of Yeast Heterochromatin

status: Public on Aug 01 2009

city: Los Angles

first: Adam

middle: Samuel

last: Sperling

address: 332 Boyer Hall

department: Biological Chemistry

institute: UCLA

summary: In budding yeast, telomeres and the mating type (HM) loci are found in a heterochromatin-like silent structure initiated by Rap1 and extended by the interaction of Sir (Silencing Information Regulator) proteins with histones. Binding data demonstrate that both the H3 and H4 N terminal domains required for silencing in vivo interact directly with Sir3 and Sir4 in vitro. The role of H4 lysine 16 deacetylation is well established in Sir3 protein recruitment, however that of the H3 N terminal tail has remained unclear. In order to characterize the role of H3 in silent chromatin formation and compare it to H4 we have generated comprehensive high resolution genome-wide binding maps of heterochromatin proteins. We find that H4 lysine 16 deacetylation is required for the recruitment and spreading of heterochromatin proteins at all telomeres and HM loci. In contrast the H3 N terminus is required for neither recruitment nor spreading of Sir proteins. Instead, deletion of the H3 tail leads to increased accessibility within heterochromatin of an ectopic bacterial dam methylase and the decreased mobility of an HML heterochromatic fragment in sucrose gradients. These findings indicate an altered chromatin structure. We propose that Sir proteins recruited by the H4 tail then interact with the H3 tail to form a higher order silent chromatin structure.

overall_design: Sir3, Sir4, and Rap1 were ChIP'd from wild type budding yeast strains and normalized to input. Sir3 and Rap1 were ChIP'd from mutant budding yeast strains and normalized to input.

middle: S

first: Michael

last: Grunstein


title: Wound response of LapA-silenced tomato leaves

summary:
LapA encodes an acidic leucine aminopeptidase that is wound-inducible and found only in tomato (Solanum lycopersicum) and a small subset of the Solanaceae.  LapA is expressed in developing flowers, fruit and in foliage after wounding, insect feeding (Manduca sexta), Pseudomonas syringae pv. tomato infection, salinity, and water-deficit stress. Using Agrobacterium tumefaciens, UC82b plants were transformed  with a chimeric gene containing the CaMV 35S promoter and LapA cDNA in the sense orientation. Kanamycin-resistant plants were screened by RNA and protein blots to  identify plants that did not accumulate and LAP-A protein upon wounding; these plants were designated LapA-silenced (LapA-SI) lines. The LapA-SI plants have an impaired wound response and do not accumulate wound response transcripts (proteinase inhibitors and polyphenol oxidase) to wild-type levels. LAP-A is the first aminopeptidase to be shown to have a regulatory role in plants.   LapA-SI and UC82b plants were germinated on filter paper for 7 days before being planted in soil. Plants were grown in a growth chamber, (16 hour days at 28°C, 8 hour nights at 24°C, 50% humidity) for three weeks. Plants were wounded by crushing each leaflet on each of two lower leaves with pliers. Wounded leaves were collected at 0, 1 and 8 hr (local wounded tissue). Two upper, unwounded leaves were collected from the same plants at the same times (systemically wounded tissue). Five to seven plants were pooled for each time point. The experiment was repeated three times. RNA was extracted using a hot phenol/LiCl protocol and purified and DNase treated using the SV Total RNA Isolation Kit from Promega.
Keywords: Reference design

first: Walling

last: Linda


title: Basis of CP12 phenotype via gene expression patterns in life cycle of CP12 antisense Nicotiana tabacum

summary:
CP12 is a small nuclear encoded chloroplast protein that has been shown to form a complex with the Calvin cycle enzymes, PRK and GADPH. We have taken an antisense approach in order to address the importance of this regulatory protein on carbon assimilation. Transgenic Nicotiana tabacum cv. Samsun with reduced levels of the CP12 protein, produced by leaf disc transformation with a full length tobacco CP12 cDNA, regulated by the cauliflower mosaic virus 35S promoter. Agrobacterium tumefaciens-mediated transformation and selection on kanamycin produced primary transformants whose progeny segregate for severity of the antisense CP12 phenotype. Because of the impact of the antisense reductions in CP12 on carbon assimilation, these plants have abnormal morphology of leaves, floral organs, greatly reduced fertility and markedly slower growth rates. Our hypothesis is that the phenotype is exposing the regulatory nature of the CP12 protein. Intermediates leaving the Calvin cycle are essential for the synthesis of hormones and perturbation of hormone balances is a potential source of the morphological and development effects observed in the CP12 antisense plants. Our objective is to understand the CP12 phenotype by identifying changes in gene expression occurring at key stages in the life cycle of the CP12 antisense plants.  CP12 antisense and wild type control plants for microarray analysis were germinated in a growth chamber in agar medium with 1% sucrose and 1/2 strength MS; with a 16 h light/8 h dark cycle at 25°C. At 20 days plants were transferred to a controlled environment greenhouse and grown in Fisons Levington F2 compost at 25°C, with light levels above 500 umol m-2 s-1 ;using supplementary lighting in a 16 h light/8 h dark cycle. 3 independent sets of plants (both wild type and CP12 antisense) were grown for preparation of three independent biological replicate RNA samples. RNA was extracted according to the TIGR protocol from plants in the early in all samples.
Keywords: Direct comparison

overall_design: 13 hybs total

first: Raines

last: Christine


title: Potato cyst nematode repsonses in potato

summary:
Sets of seven 2-week old potato plants carrying the nematode resistance gene H1, grown from tuber ‘chits’ in sandy loam at a constant temperature of 20 ºC and a light cycle of 16 hour light/8 hour dark, were each inoculated in the roots evenly with 2000 juveniles of the virulent potato cyst nematode Globodera pallida, or with the avirulent G. rostochiensis pathotype Ro1, or with water. Plants were manually watered throughout the duration of the experiment.   5, 17 and 33 days after inoculation, the roots were carefully washed and root tissue samples were individually flash-frozen in liquid nitrogen and stored at –80 ºC. Total RNA isolations were performed using the Qiagen RNeasy kit. All samples were treated with DNase. The experiment was replicated twice.
Keywords: Direct comparison

first: Poch

last: Hector


title: Role of oxylipins in pathogen defense of potato

summary:
Oxylipins play a role in the response of plants to pathogens both as antimicrobial compounds and as signaling molecules. In potato, pathogen infection leads to the stimulation of the 9-lipoxygenase pathway (Göbel et al. 2001; 2002; Stumpe et al. 2001). In order to analyze whether 9-LOX-derived oxylipins such as colnele(n)ic acid (products of the 9-divinyl ether synthase reaction) act as signaling molecules for the activation of defense responses, transgenic potato plants (Solanum tuberosum cv. Désirée) were generated which express an RNAi construct directed against the pathogen-induced 9-lipoxygenase (Göbel et al. 2003) or against the pathogen-induced 9-divinyl ether synthase (Stumpe et al. 2001). Highly reduced transcript levels correlate with low levels of 9-lipoxygenase-derived oxylipins (Göbel et al. 2003).  Transgenic and wild type plants were grown as sterile plants on MS medium supplemented with agar in a phytochamber with 16 h of light [200 µE] at 22 °C. After transfer to soil, plants were kept in a phytochamber with 16 h of light [200 µE], 18 °C and 60 % humidity for four weeks. Lower leaves were infiltrated with a suspension of Pseudomonas syringae pv. maculicola at a concentration of 108 cfu/ml MgCl2 or, as a control, 10 mM MgCl2-solution. RNA was isolated both from the infiltrated leaves and the upper, non-treated leaves using the trizol method. Subsequently, RNA was digested with DNase and purified via Qiagen RNeasy Kit (Qiagen, Hilden, Germany) according to the manufacturer´s instructions.
Keywords: Direct comparison

overall_design: 17 hybs total

first: Rosahl

last: Sabine


title: Identification of Gene Expression Patterns During Somatic Embryogenesis in Potato

summary:
This investigation was aimed at studying global gene expression during the process of somatic embryogenesis in potato using internodal segments (INS) as explants. The INS explants from 4 to 6 week old potato cv. Desiree in vitro cultures were cultured on somatic embryogenesis induction for 2 weeks (stage-I) and expression/regeneration modified MS media for 3 weeks, respectively with (stage-II) and without (stage-III) 2, 4-dichlorophenoxy acetic acid (5 µM). The culturing frequency was 10 INS/petridish with 25 ml medium.  The in vitro cultures were incubated at 19±1°C, 16/8 hrs light/dark cycles and 90 µmol/m2/s light intensity. To study the molecular events of SE as affected by the absence or presence of auxin, the INS explants were divided into 2 groups after the induction phase. One-half of the induced INS explants were transferred to auxin free medium to evoke SE, the other half were subjected to a prolonged induction phase. The total RNA from INS explants at these critical stages of SE progression was isolated using TRIzol (Invitrogen) as per the manufacturer’s instructions. The on-column DNase treatment and clean-up steps were performed using Qiagen RNase free DNase and RNeasy midi kits. The selected time points used for the RNA isolation were day 0 (freshly cut INS), day 14 (INS on induction medium) and day 21, 28 & 35 (INS on expression medium as well as from prolonged induced phase). Except day 0, where the whole INS were used for RNA isolation, all other time point RNA was isolated from the callused part of the INS distal end. Each time point was replicated 3 times. The number of explants for each biological replicate varied from 30 to 300 owing to variability in tissue availability and corresponding RNA yield from explants of each time-point. The entire process from tissue excision to RNA isolation was completed on the same day to avoid any unintended change in tissue condition by storage.
Keywords: Direct comparison

overall_design: 28 hybs total

first: Millam

last: Steven


title: Heat stress in 7 Solanaceae species, replicate 1 and 2

summary:
Seven different Solanaceae species, Potato (Solanum tuberosum), Tomato (Solanum lycopersicum), Eggplant (Solanum melongena), Pepper (Capsicum annuum), Tobacco (Nicotiana tabacum), Petunia and Nicotiana benthamiana were subjected to heat stress. Plants were grown for 4-6 weeks at 25 C after which heat stress was initiated by exposing the plants to 35 C for 6, 12, 24, 48 and 96 hours. Control samples were isolated from plants just before initiating the heat stress. RNA was isolated using Qiagen RNeasy.
Keywords: Direct comparison

sample_organism:
Capsicum annuum
Solanum lycopersicum
Nicotiana tabacum
Nicotiana benthamiana
Petunia x hybrida
Solanum tuberosum
Solanum virginianum

overall_design: 70 hybs total

first: Buell

last: Robin


title: Interactions between root knot nematodes and potato

last: Liu

summary:
Plant-parasitic nematodes and especially the root-knot nematodes are ubiquitous pathogens. Eggs of root-knot  nematodes (Meloidogyne javanica) were extracted from greenhouse cultures and second-stage juveniles (J2) were hatched in tap, sterile water on 30 µm sieves. Solanum tubersum cv. Desiree explants, i.e., stem including an axillary bud, were sectioned, under sterile conditions from in vitro growing seedlings and transferred to Magenta boxes containing Gamborg's media. Seedlings, 4 weeks post transferring to Gamborg's media were planted in pots containing autoclaved quartz sand. Three weeks later the plants were infected with 3000 M. javanica infective juveniles, applied to the soil in tap, sterile water. Control uninfected plants were mock-inoculated with tap, sterile water. Roots from 6 infected and 6 mock-inoculated plants were collected at a series of time points, including 5, 10 and 15 days post nematode infection/mock inoculation. Roots were observed under the microscope, and nematode feeding sites were selectively dissected, from young lateral roots. To control for the effect of tissue sectioning on gene expression, young lateral roots of the mock inoculated roots were dissected similarly to the infected roots, and collected. Dissected roots were snapped-freeze in liquid nitrogen and immediately stored in -80 C freezer. Total RNA was extracted using Qiagen RNAEasy kit. The experiment for each time point was duplicated, each duplicate derived from independent biological repeat. All RNA samples were amplified using the Ambion kit Message Amp catalog no. 1750, using as starting material 2.5 to 5 µg of total RNA.
Keywords: Reference design

first: Koltai

last: Hinanit


title: Cold stress in 7 Solanaceae species, replicate 1 and 2

first: Jia

summary:
Seven different Solanaceae species, Potato (Solanum tubersosum), Tomato (Lycopersicum esculentum), Eggplant (Solanum melangena), Pepper (Capsicum annuum), Tobacco (Nicotiana tabacum), Petunia and Nicotiana benthamiana were subjected to cold stress. Plants were grown at 25 C for 4-6 weeks after wich cold stress was initiated by exposing the plants to 4 C for 4, 8, 12, 24 and 48 hours. Control samples were isolated from  plants just before the cold stress was initated.  RNA was isolated using Qiagen RNeasy.
Keywords: Direct comparison

overall_design: 58 hybs total


title: Effect of two potato viruses, PLRV and PVY, on potato gene expression in Russet Burbank

summary:
Potato plants (cv. Russet Burbank) were grown as in vitro plantlets, and transplanted into 12-cm pots in Sunshine Mix #2 (peat/perlite mix) supplemented with slow release fertilizer (Osmocoat) in the greenhouse.  The experiment was conducted in the summer of 2004 at Moscow, ID.  Plantings were staggered to facilitate collection of volatiles and leaf samples during a short harvest window for all times of all treatments.  Infectious Green Peach Aphids were raised on PLRV-infected potato plants; test plants were infected by placing 10 infectious aphids on a leaf in a clip cage for 48 h.  The control plants for PLRV were similarly treated with aphids that had been raised on uninfected potato plants.  A local isolate of PVYO was inoculated mechanically using purified virus (5 ìg ml-1) in buffer (50 mM Na2HPO4, 20 mM Na2SO3 pH 7.0) lightly rubbed onto a leaf previously dusted with carborundum.  Control plants for PVY were treated in the same manner using buffer without PVY.  The RNA samples were made from systemically affected leaves, excluding the actual leaves inoculated by aphids or that received the mechanical infection.  Leaf volatiles were concurrently collected from equivalent plants; aliquots of frozen leaf samples were sent for metabolite analysis prior to RNA extraction.  Leaf harvests were made 1, 3, 7, 14, and 28 d post-inoculation.
Keywords: Direct comparison

overall_design: 31 hybs total

first: Lorenzen

last: Jim


title: Drought stress in 7 Solanaceae species, replicate 1 and 2

summary:
Seven different Solanaceae species, Potato (Solanum tubersosum), Tomato (Solanum lycopersicum), Eggplant (Solanum melongena), Pepper (Capsicum annuum), Tobacco (Nicotiana tabaccum), Petunia and Nicotiana benthiamana were subjected to drought stress. Drought stress was applied by stopping watering of the plants, control plants were normally watered with nutrient solution. Samples were collected at 0, 1, 3, 5, 7 and 10 days after the first application of the drought stress. RNA was isolated using Qiagen RNeasy.
Keywords: Direct comparison

overall_design: 79 hybs total


title: Defense signaling in potato

summary:
Potato cultivar Russet Norkotah was grown in a growth chamber at 16 hour light/8 hour dark at 22°C fertilized with Osmocote.  Plants were from tissue culture, transplanted to soil and used approximately 4 weeks after transplanting.  RNA was extracted from leaves using Trizol.  Plants were sprayed with one of four treatments, either 1mM salicylic acid, 100 µM methyl jasmonate, 1 mg/ml arachidonic acid or 100 µM methyl jasmonate plus 1 mM salicylic acid together.   Samples were harvested at 3, 24 and 72 hours after spraying.  Each salicylic acid or arachidonic acid treatment has 3 biological replicates done in parallel, using leaves pooled from 2 plants.  The control for SA or AA treated plants were water sprayed plants, from which leaves of 3 different plants were pooled at each timepoint, done side-by-side with the treated samples. Methyl jasmonate or MJ+SA treatments had 2 biological replicates each, using leaves pooled from 2 plants. A separate set of water sprayed plants (from the SA and AA control treatments) were used as controls, and pooled leaves from 3 plants were used at each time-point, and done in parallel with the treated samples.
Keywords: Direct comparison

first: Navarre

last: Roy


title: Whole genome survey of 32 Human Tissues

status: Public on Jun 04 2007

first: Julie

last: Blake

department: SDS/Arrays

summary:
The Applied Biosystems Tissue Gene Expression Database, known as the Human Body Map, is a collection of gene expression profiles from 31 normal tissues and a Universal Human Reference RNA (Stratagene). The Human BodyMap was created using the Applied Biosystems Expression Array System. There were three replicates per tissue for a total of 96 microarrays.
Keywords: Human tissue whole genome survey

overall_design: 31 human tissues from Clontech and 1 from Stratagene (UHR) were used to create the Applied Biosystems Human BodyMap (a.k.a.Tissue Gene Expression Database ). Digoxigenin-UTP-labeled cRNA was generated and linearly amplified from 1 µg of total RNA with three replicates per tissue, using the Applied Biosystems Chemiluminescent RT-IVT Labeling Kit v 2.0 and kit protocol. Microarray hybridization, chemiluminescence detection, image acquisition, and analysis were performed on the on the Applied Biosystems 1700 Chemiluminescent Microarray Analyzer using the Applied Biosystems Chemiluminescence Detection Kit and kit protocol.

first: Li

last: Kelly

first: Raymond

last: Samaha

first: Sun

last: Yongming

first: Chan

last: Frances

first: Blake

last: Julie

first: Roger

last: Canales


title: Tomato colonization by wt R. solanacearum and a hrp mutant

summary:
Tomato seeds (Solanum lycopersicum Mill., cultivar Marion) were germinated in vermiculite and 10-day old seedlings were transplanted into 4-inch plastic pots containing a mixture of composted pine bark and vermiculite (3:1) amended with lime and fertilizer. Plants were incubated in a growth chamber (constant 30oC, 14-h photoperiod; 180 ?mol s-1 m-2 light intensity) for 18-21 days before inoculation. Seedlings received a complete liquid fertilizer approximately 7 and 14 days after transplanting. Unwounded roots were inoculated with R. solanacearum by drenching the soil in each pot with 40 ml of water containing 1´107 CFU ml-1 of the wild type or 1´108 CFU ml-1 of the hrp mutant. Non inoculated plants received 40 ml of water. Four days after inoculation, a piece of each stem from about 1-cm above the soil line to just below the cotyledon node was excised, immediately plunged into liquid nitrogen, and stored at -80oC. A separate 0.5-cm stem section from the cotyledon node was removed aseptically from each plant and the number of bacteria in the section determined by dilution plating. Stem pieces from plants systemically colonized by R. solanacearum were pulverized in a mortar under liquid nitrogen, and total RNA was extracted with the Tri-Reagent (Molecular Research Center) and then treated with RNase-free recombinant DNase (Ambion) according to the recommended protocols from TIGR and stored at -70oC.
Keywords: Direct comparison

overall_design: 20 hybs total

first: Denny

last: Timothy


title: Herbivore-induced vaccination of N. attenuata and S. nigrum

summary:
Nicotiana attenuata and Solanum nigrum were challenged with different insects: Manduca sexta and Tupiocoris notatus in different combinations Goals of the study are first to haracterize the transcriptional response of two native Solanaceous plants (Nicotiana attenuata and Solanum nigrum) to attack from two herbivorous pests common on Solanaceous crops (Manduca sexta and Tupiocoris notatus). And second to identify genes involved in herbivore vaccination phenomenon. After germination, inbred and genetically characterized lines of N. attenuata DI92 and S. nigrum Sn30 originally collected from southwestern Utah in 1988, and Jena Germany in 2000, respectively, were grown in a greenhouse. Eggs of Manduca sexta (abbreviated M. s.) came from the North Carolina State University - Entomology Insectary and were hatched under 37°C. Nymphs and adults of Tupiocoris notatus (abbreviated T. n.) came from laboratory colonies started in 2000 with individuals collected from our Utah field sites. Plant were challenged with these different insect and combinations of them. All plants for a given treatment were enclosed in glass-and-mesh insect cages to avoid cross-infection. After 24 h or 5 days the herbivores and their frass were removed, and the above ground biomass of each plant was flash-frozen in liquid nitrogen and stored at -80 °C until RNA extraction. Three independent replicate cages (biological replicates) were used for each of the 7 treatments resulting in a total of 21 cages per species. RNA extraction was performed according to the protocol using Trizol. The RNA samples represent a pooled sample from 4 plants grown in a cage.
Keywords: Direct comparison

sample_organism:
Solanum nigrum
Nicotiana attenuata

overall_design: 40 hybs total

first: Baldwin

last: Ian


title: Comparison of cross-species and species-specific hybridizations

status: Public on May 30 2006

city: Bet-Dagan

first: Carmiya

last: Bar-Or

address: Hamaccabim

department: Genomics

country: Israel

institute: ARO

summary:
Abstract
Background: One of the approaches for conducting genomics research in organisms that do not yet have a proper microarray template is to profile their expression patterns by using cross-species hybridization (CSH). Several different studies using spotted microarray for CSH resulted with contradicting conclusions as to the ability of CSH to reflect biological processes.
Results: We used a tomato spotted cDNA microarray to examine the ability of CSH to reflect species specific hybridization (SSH) data. Potato RNA was hybridized to spotted cDNA tomato and potato microarrays to generate heterologous and homologous hybridization data, respectively. The results revealed difficulties in obtaining transcriptomics data from CSH that reflected those obtained from SSH. Nevertheless, once the data was filtered for those corresponding to matching probe sets, by restricting proper cutoffs of probe homology, the CSH transcriptomics data better reflected those of the SSH, to an extent that was quantitated  by identification of differentially regulated genes.
Conclusions: This study enabled us to outline some considerations regarding evaluation of a microarray as candidate platform for CSH study, performance of CSH and proper data analysis that may allow CSH to reflect to some extent a biological process.
Keywords: cross-species hybridization; heterologous hybridization

platform_organism:
Solanum lycopersicum
Solanum tuberosum

overall_design: To examine whether cross species hybridization (CSH) can produce data that reflects species specific hybridization (SSH) data, we used the same RNA samples for both CSH and SSH microarray experiments. RNA samples were extracted from nematode-infected and non-infected (control) potato plants, sampled at two time points (at 5 and at 10 days of nematode infection). These RNA samples were co-hybridized with a reference sample by CSH to a tomato microarray and by SSH to a potato microarray. The reference sample was pooled from all 8 samples (2 time points of infected or non-infected plants; 2 biological replicates were performed). The CSHs were designated as 'PT' for potato samples on tomato microarrays; the SSHs were designated as 'PP' for potato samples on potato microarrays.

first: Meira

last: Bar-Eyal

first: Tali

middle: Z

last: Gal

first: Yoram

last: Kapulnik

first: Henryk

last: Czosnek

first: Hinanit

last: Koltai


title: Analysis of two plant-microbe interactions in N. benthamiana

first: Jia

last: Liu

summary:
Nicotiana benthamiana plants were grown to the 8th true leaf development stage with 16 h light at 150  mol s-1 m-2 with 25/17oC day/night temperatures. Colletotrichum orbiculare strain ATCC20767 and Pseudomonas syringae pv. tabaci strain BPIC4 was grown in pure cultures to obtain syringae pv. tabaci strain BPIC4 was grown in pure cultures to obtain conidia and cells, respectively. Leaves of N. benthamiana plants were inoculated by spraying a suspension of 2 X 106 spores/ml in sterile H2O for C. orbiculare as described by Shen et al. (2001) or by infiltrating with a needle-less syringe 2 X 105 CFU/ml in sterile 10 mM MgCl2 for P. syringae pv. tabaci according to Rommens et al. (1995). For a control, healthy leaves were sprayed with sterile H2O or infiltrated with sterile 10 mM MgCl2 and then incubated under the same conditions for the same period of time as the pathogen-inoculated plants. Total RNA was extracted at 12 hours and 1, 2, 3 and 4 days after inoculation. The first and second mature leaf was excised and immediately frozen at -70 C.
Keywords: Direct comparison

first: Goodwin

last: Paul


title: Profiling of zinc altered gene expression in human prostate normal (HPR-1) and malignant (PC-3) cells

status: Public on Sep 24 2006

city: Baltimore

first: Shufei

last: Lin

address: 666 W. Baltimore Street

department: Biomedical Sciences

institute: University of Maryland at Baltimore

summary:
To gain a better understanding of the zinc effect on prostate cells, gene expression profiling analyses were conducted using HPR-1 and PC-3 cells treated with or without zinc (1,500 ng/ml) after 24 hrs serum-depletion.  Zinc effect on gene expression was evaluated at 1, 3, and 6 hrs of zinc treatment compared to the control, using human genome survey microarray chips. A total of 6,110 identified genes exhibited three expression patterns in response to zinc a threshold of ±2.5 folds: suppressed (3,534 in PC-3; 1,953 in HPR-1), increased (571 in PC-3; 872 in HPR-1), and otherwise altered (16 in PC-3; 13 in HPR-1).
Keywords: Zinc effect, time course, cell type comparison

overall_design: Zinc effect on gene expression of HPR-1 and PC-3 cells was evaluated with a time-course study using human genomewise microarray chips (Applied Biosystems v 2.0). For each chip, 2µg total RNA from each sample was used.  A series of samples from each cell line was prepared as follows: control, 1 hr, 3 hrs, and 6 hrs zinc exposure. Synthesis of cRNA and subsequent hybridization was completed by the Core Facility at UMBI according to the ABI standard protocol. The ABI human genome survey microarray chip contained 32,878 probes for the interrogation of 29,098 known genes

first: Shu-fei

first: Hua

last: Wei

first: Yujuan

last: Hong

first: Pei

last: Feng