title: a1

status: Public on Aug 14 2008

type: RNA

city: Lausanne

first: Ruth

last: Luthi-Carter

address: AI 2138, Station 15

country: Switzerland

institute: EPFL

description: Uninfected

molecule: total RNA

organism: Rattus norvegicus

source_name: Primary rat striatal neuron cultures, 6 weeks in vitro


title: d5

description: Infected 6 weeks prior with Htt-N171-82Q-expressing recombinant lentivirus


title: b14

description: Infected 6 weeks prior with GFP-expressing recombinant lentivirus


title: b2


title: b6


title: c10

description: Infected 6 weeks prior with Htt-N171-19Q-expressing recombinant lentivirus


title: Salmonella enterica subsp. enterica serovar Pullorum, strain 10704, rep2

status: Public on Feb 27 2008

type: protein

city: London

first: Vesela

last: Encheva

state: Greater London

address: 61 Colindale Avenue

department: Centre for Infections

country: United Kingdom

institute: Health Protection Agency

description:
Rehydration and Isoelectric Focusing (IEF):
Protein samples containing a total of 150 ?g of protein were mixed with rehydration buffer (7 M urea, 2 M thiourea, 2% w/v CHAPS, 50 mM DTT, 0.5% v/v IPG buffer pH 3-10 NL, (Amersham BioSciences)). The rehydration of the IPG strips was performed by the in-gel rehydration method (Rabilloud et al, 1994) using 18 cm IPG strips (pH 3-10 NL, Amersham BioSciences). The samples were left to rehydrate for 18 h at 20?C. IEF was performed using an Investigator 5000 (Genomic Solutions) power supply for 24 h, for a total of 85 000 Vh at a maximum voltage of 5000 V and a maximum current of 110 ?A.

Equilibration of the IPG strips and second dimension separation:
The focused IPG strips were equilibrated in Tris/acetate equlibration buffer (Genomic Solutions) containing of 0.5% w/v SDS, 30% v/v glycerol and 6 M urea. The equilibration was performed in two consecutive steps of 30 min each. The first step was conducted in the presence of 50 mM DTT, and the second in the presence of 135 mM iodoacetamide.
The second dimension separation was performed on 10% Duracryl ® gels (Proteomic Solutions). The gels were run for 5h, at a maximum voltage of 500 V and maximum power of 20 000 mW per gel using the Investigator 5000 power supply (Genomic Solutions).
The gels were post-stained with fluorescent dye SYPRO Ruby (Molecular Probes) for 18 h on constant shaking and destained for 2 h in 10 % v/v methanol. The resulting 2D GE profiles were visualised using the Typhoon Scanner (Amersham Biosciences). To excise the spots the gels were double stained with Colloidal G and destained in 25 % v/v methanol.

Analysis of the 2D GE profiles:
For the comparative analysis duplicate experimental samples from two different batches of each isolate were used (two technical replicates and two biological replicates for each isolate). The density/volume of the SYPRO Ruby-stained protein spots were determined using the software package Proteome Weaver (Definiens). For a protein to be reported as differentially expressed, the density values measured in Proteome Weaver were further subjected to the Student?s T-test, and only changes with p< 0.01 were considered significant.
In gel trypsin digestion and peptide analysis using LC/MS/MS:
In-gel digestion with trypsin was performed with a robotic digestion system (Investigator ProGest, Genomic Solutions).Colloidal G was removed by sequential washing with 50 mM ammonium hydrogen carbonate buffer and acetonitrile. Cysteine residues were reduced with DTT and derivatised by treatment with iodoacetamide. After further washing with ammonium hydrogen carbonate buffer, the gel pieces were again dehydrated with acetonitrile and dried at 60°C, prior to the addition of modified trypsin (Promega) (10 µl of a 6.5 ng µl-1 solution in 25 mM ammonium hydrogen carbonate). Digestion proceeded for 8 h at 37°C, and the products were recovered by sequential extractions with 25 mM ammonium hydrogen carbonate, 5% v/v formic acid, and acetonitrile. The pooled extracts were lyophilised, and redissolved in 0.1% v/v formic acid for mass spectrometry.
Tandem electrospray mass spectra were recorded using a Q-TOF hybrid quadrupole / orthogonal acceleration time of flight spectrometer (Micromass) interfaced to a Micromass CapLC capillary chromatograph. Samples were dissolved in 0.1% v/v formic acid, and 6 µl was injected onto a Pepmap C18 trap column (300 µm ? 0.5 cm; LC Packings), and washed for 3 min with 0.1 % v/v formic acid. The flow rate was then reduced to 1 µl per min, and the peptides were eluted into the mass spectrometer with an acetonitrile / 0.1% v/v formic acid gradient (5% to 70 % v/v acetonitrile over 20 minutes).
The capillary voltage was set to 3,500 V, and data-dependent MS/MS acquisitions were performed on precursors with charge states of 2, 3 or 4 over a survey mass range of 540-1200 Da. Known trypsin autolysis products and keratin-derived precursor ions were automatically excluded. The collision voltage was varied between 18 and 45 V depending on the charge and mass of the precursor. Product ion spectra were charge state de-encrypted and de-isotoped with a maximum entropy algorithm (MaxEnt 3, Micromass). Proteins were identified by correlation of uninterpreted tandem mass spectra to entries in SwissProt/TREMBL using ProteinLynx Global Server (Version 1.1, Micromass) and NCBInr using MASCOT. One missed cleavage per peptide was allowed, and an initial mass tolerance of 50 ppm was used in all searches. Cysteines were assumed to be carbamidomethylated, but other potential modifications were not considered in the first pass search.
All matching spectra were reviewed manually, but in cases where the score reported by ProteinLynx global server was less than 100, additional searches were performed against the NCBI nr database using MASCOT, which utilises a robust probalistic scoring algorithm. Identifications based on a single matching peptide were verified manually using the MassLynx program Pepseq (Micromass).

molecule: protein

growth_protocol: The bacterial cells were grown on Colombia Blood Agar (CBA; Oxoid) for 18 h at 37 °C. Extracts were prepared from the second inoculum of each strain and each biological replicate was prepared independently on different day under identical growth conditions.

extract_protocol: Cell lysates were obtained by harvesting the growth of three plates of S. enterica and resuspending the cells in 20 mM Tris (pH=7.5). The cells were washed and collected by centrifugation at 5 100 ? g, 10 min. The resulting cell pellet was resuspended in 100 ?l of lysis solution containing 0.3% w/v SDS, 200 mM DTT and 50 mM Tris (pH=8.0), followed by the addition of 50 ?l of 0.5 M Tris (pH=8.0), 50 mM MgCl2, 1 mg ml-1 DNase 1 and 0.25 mg ml-1 RNase A. The samples were vortexed with 0.3 g of glass beads (? 105 ?m diameter) and homogenised five times for 1 min using the Mickle Cell Desintegrator (Mickle Laboratory Engineering) with 1 min cooling time on ice in between runs. The unbroken cells and cell debris were removed by centrifugation at 21 000 ? g for 30 min, 4?C, and the resulting supernatant was used for further analysis.


title: Salmonella enterica subsp. enterica serovar Pullorum, strain 10704, rep1

data_processing: The density/volume of the SYPRO Ruby-stained protein spots were determined using the software package Proteome Weaver (Definiens). For a protein to be reported as differentially expressed, the density values measured in Proteome Weaver were further subjected to the Student?s T-test, and only changes with p< 0.01 were considered significant.


title: Salmonella enterica subsp. enterica serovar Choleraesuis, strain B7, rep1

tag: clinical isolate


title: Salmonella enterica subsp. enterica serovar Choleraesuis, strain B5, rep2


title: Salmonella enterica subsp. enterica serovar Choleraesuis, strain B5, rep1


title: a3

description: Uninfected


title: d11.2


title: a7

description: Uninfected


title: d12


title: c8.2


title: Salmonella enterica subsp. enterica serovar Enteritidis, strain 12694, rep2


title: d10.2


title: c4


title: Salmonella enterica subsp. enterica serovar Typhimurium, strain 74, rep2


title: Salmonella enterica subsp. enterica serovar Typhimurium, strain 204, rep2


title: Salmonella enterica subsp. enterica serovar Typhimurium, strain 204, rep1


title: Mouse calvaria, sErbB3-treated

status: Public on May 01 2008

city: Houston

first: Nanyue

last: Chen

state: TX

address: 1515 Holcombe Blvd

country: USA

institute: UT-MD Anderson Cancer Center

description: Performed by SuperArray Bioscience

hyb_protocol: Performed by SuperArray Bioscience

scan_protocol: Performed by SuperArray Bioscience

data_processing:
The value column shows the analyzed annotated data after background subtraction and normalization.
We chose Interquartile value for normalization. The minimal intensity was used for background subtraction.
The detection calls for each gene are described as a spot is considered "Absent" if the average density of the spot is less then the mean value of the local backgrounds of the lower 75 percentile of all non-bleeding spots. All other spots are considered Present.
http://www.superarray.com

label: Performed by SuperArray Bioscience

organism: Mus musculus

source_name: mouse calvaria, sErbB3-treated, 4 days

label_protocol: Performed by SuperArray Bioscience

extract_protocol: Total RNA was extracted with Trizol (Invitrogen) and was further purified by the use of a Mini-RNA extraction kit (Stragagene). RNAs were used to probe Osteogenesis cDNA GEarrays containg skeletal-related genes (SuperArray Bioscience).

treatment_protocol: Calvaria were excised from 4-days-old CD1 mice. 5 calvaria were placed in one well of 6-well plate, and incubated in 2 ml of BGJb medium (Invitragen) containing 0.1% bovine serum albumin (BSA). At 4th day after treated with or without BF-1 (p45-sErbB3), calvaria were collected for RNA extraction.

tag: Strain

value: CD1 white mouse;

tag: Age

value: 4 days

tag: Tissue

value: calvaria


title: Salmonella enterica subsp. enterica serovar Typhimurium, strain 12023, rep2


title: Salmonella enterica subsp. enterica serovar Pullorum, strain B52, rep2


title: Salmonella enterica subsp. enterica serovar Pullorum, strain B52, rep1


title: Mouse calvaria, untreated

description: Performed by SuperArray Bioscience

source_name: mouse calvaria, control


title: c7.2


title: FACS-sorted cells with photoconverted Kaede vs. FACS-sorted cells with non-photoconverted Kaede_ZF1351

status: Public on Feb 26 2008

city: Bethesda

first: Christopher

last: Pan

state: MD

address: Rm 5228, Bldg 50, South Dr.

department: NHGRI

institute: NIH

description: Zebrafish embryos

hyb_protocol: 200 picomoles of Cy3 and 100 picomoles of Cy5 labeled cRNAs are put in a total volume of 24 ul (qsp with DEPC H20) and 6 microliter of 5x fragmentation buffer from Affymetrix. the mixture is incubated at 94 ¡C for 15 minutes. Then, 25 ul of 2X  hybridization buffer (250 _l Formamide, 250 _l 20xSSC and 20 _l 10%SDS). The mixture is then denatured for 2 minutes and applied on the Maui (BioMicro Systems, Inc) hybridization chamber/ array slide sandwich. The hybridization is performed at 45 degre for 20 hours.

scan_protocol: Agilent Scanner

data_processing: IPLab/DeArray

organism: Danio rerio

label_protocol: 0.5-1.0 microgram of total RNA was reverse transcribed with T7 Oligo-dT, followed by a second strand synthesis and overhight i9n vitro transcription in the presence of  Aminoallyl UTPs using the Amino Allyl MessageAmpª II aRNA Amplification kit from Ambion (Kit Cat#1753). The Aminoallyl UTP labeled cRNAs are then coupled with either Cy3 or Cy5 mono NHS Ester Dyes. The CyDyes labeled cRNAs are then purified with columns and reagents provided by the Kit (Cat#1753).

extract_protocol: RNA was extracted with Trizol/Chloroform method according to manifecturer protocol and the RNA is further purified by the Qiagen RNAeazy columns

tag: Zebrafish embryos

source_name: FACS-sorted cells with non-photoconverted Kaede,


title: FACS-sorted cells with photoconverted Kaede vs. FACS-sorted cells with non-photoconverted Kaede_ZF1346

description: Zebrafish embryos


title: FACS-sorted cells with photoconverted Kaede vs. FACS-sorted cells with non-photoconverted Kaede_ZF1349

description: Zebrafish embryos


title: FACS-sorted cells with photoconverted Kaede vs. FACS-sorted cells with non-photoconverted Kaede_ZF1348

description: Zebrafish embryos


title: FACS-sorted cells with non-photoconverted Kaede vs. whole embryos_ZF1345

description: Zebrafish embryos

source_name: whole embryos


title: Salmonella enterica subsp. enterica serovar Choleraesuis, strain B7, rep2


title: FACS-sorted cells with non-photoconverted Kaede vs. whole embryos_ZF1343

description: Zebrafish embryos


title: Salmonella enterica subsp. enterica serovar Dublin, strain 193, rep1


title: Salmonella enterica subsp. enterica serovar Dublin, strain 193, rep2


title: Salmonella enterica subsp. enterica serovar Dublin, strain 12709, rep2


title: Salmonella enterica subsp. enterica serovar Dublin, strain 12709, rep1


title: FACS-sorted cells with non-photoconverted Kaede vs. whole embryos_ZF1342

description: Zebrafish embryos


title: Salmonella enterica subsp. enterica serovar Enteritidis, strain 97, rep1


title: Salmonella enterica subsp. enterica serovar Enteritidis, strain 97, rep2


title: Salmonella enterica subsp. enterica serovar Enteritidis, strain 12694, rep1


title: MEF vs. NEF_ZF1336

description: Zebrafish embryos

source_name: Mesendoderm precursors

source_name: neurectoderm precursors


title: MEF vs. NEF_ZF1330

description: Zebrafish embryos


title: MEF vs. NEF_ZF1334

description: Zebrafish embryos


title: FoxH1 morpholino-injected 40% epiboly-stage embryos vs. control morpholino-injected 40% epiboly-stage embryos_ZF1221

description: Zebrafish embryos

hyb_protocol: The combined Cy3 and Cy5 labeled cDNAs are put in a total volume of 250 ul DEPC H20 and 250 ul of 2X Agilent hybridization buffer. The mixture is then denatured for 2 minutes and applied on the Agillent hybridization chamber/gasket slide in sandwich with the array slide. The hybridization is performed at 65 degre for 16 hours.

source_name: control morpholino-injected 40% epiboly-stage zebrafish embryos

label_protocol: 10-15 microgram of total RNA was reverse transcribed with Anchor Oligo-dT and Aminoallyl dUTP, after NaOH RNA hydrolysis, the labeled Aminoallyl dUTP cDNA is purified with Qiagen, QIAQuick PCR Purification Kit (Cat. # 28104). The Aminoallyl dUTP cDNAs are then coupled with either Cy3 or Cy5 mono NHS Ester Dyes. The CyDyes labeled cDNAs are then purified with Qiagen, QIAQuick PCR Purification Kit (Cat. # 28104).

source_name: FoxH1 morpholino-injected 40% epiboly-stage zebrafish embryos


title: MEF vs. NEF_ZF1337

description: Zebrafish embryos


title: FoxH1 morpholino-injected 40% epiboly-stage embryos vs. control morpholino-injected 40% epiboly-stage embryos_ZF1222

description: Zebrafish embryos


title: Salmonella enterica subsp. enterica serovar Typhimurium, strain 227, rep2


title: NBTII-FGFR1 transfected cells, FGF1 stimulation 48h rep2

status: Public on Feb 20 2008

city: Villejuif cedex

first: Marianne

last: Tuefferd

address: 16 avenue Paul Vaillant Couturier

country: France

institute: INSERM

description: Gene expression data from NBTII-FGFR1 cells activated by FGF-1 ligand

hyb_protocol: Affymetrix protocol

scan_protocol: GeneChips were scanned using the Hewlett-Packard GeneArray Scanner G2500A.

data_processing: The data were analyzed with RMA global normalization procedure

label: biotin

source_name: NBTII-FGFR1 cells

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 6 microg total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).

extract_protocol: RNA PLUS™ extraction Kit (Bioprobes Systems)

treatment_protocol: FGF1 stimulation (20ng/ml) + 50µl/ml heparine

tag: strain

value: NBTII-FGFR1 ; stimulation :FGF1 ; time :48 hours


title: NBTII cells FGF1 stimulation 48 hours rep2

description: Gene expression data from NBTII cells activated by FGF-1 ligand

source_name: NBTII cells

value: NBTII ; stimulation :FGF1 ; time :48 hours


title: NBTII cells FGF1 stimulation 24 hours rep2

value: NBTII ; stimulation :FGF1 ; time :24 hours


title: Salmonella enterica subsp. enterica serovar Typhimurium, strain 227, rep1


title: NBTII-FGFR1 transfected cells, FGF1 stimulation 6h rep2

value: NBTII-FGFR1 ; stimulation :FGF1 ; time :6 hours


title: Salmonella enterica subsp. enterica serovar Typhimurium, strain 12023, rep1


title: NBTII-FGFR1 transfected cells, FGF1 stimulation 6h rep1


title: FACS-sorted cells with photoconverted Kaede vs. FACS-sorted cells with non-photoconverted Kaede_ZF1347

description: Zebrafish embryos


title: Salmonella enterica subsp. enterica serovar Typhimurium, strain 74, rep1

description:
Rehydration and Isoelectric Focusing (IEF):
Protein samples containing a total of 150 ?g of protein were mixed with rehydration buffer (7 M urea, 2 M thiourea, 2% w/v CHAPS, 50 mM DTT, 0.5% v/v IPG buffer pH 3-10 NL, (Amersham BioSciences)). The rehydration of the IPG strips was performed by the in-gel rehydration method (Rabilloud et al, 1994) using 18 cm IPG strips (pH 3-10 NL, Amersham BioSciences). The samples were left to rehydrate for 18 h at 20?C. IEF was performed using an Investigator 5000 (Genomic Solutions) power supply for 24 h, for a total of 85 000 Vh at a maximum voltage of 5000 V and a maximum current of 110 ?A.

Equilibration of the IPG strips and second dimension separation:
The focused IPG strips were equilibrated in Tris/acetate equlibration buffer (Genomic Solutions) containing of 0.5% w/v SDS, 30% v/v glycerol and 6 M urea. The equilibration was performed in two consecutive steps of 30 min each. The first step was conducted in the presence of 50 mM DTT, and the second in the presence of 135 mM iodoacetamide.
The second dimension separation was performed on 10% Duracryl ® gels (Proteomic Solutions). The gels were run for 5h, at a maximum voltage of 500 V and maximum power of 20 000 mW per gel using the Investigator 5000 power supply (Genomic Solutions).
The gels were post-stained with fluorescent dye SYPRO Ruby (Molecular Probes) for 18 h on constant shaking and destained for 2 h in 10 % v/v methanol. The resulting 2D GE profiles were visualised using the Typhoon Scanner (Amersham Biosciences). To excise the spots the gels were double stained with Colloidal G and destained in 25 % v/v methanol.

Analysis of the 2D GE profiles:
For the comparative analysis duplicate experimental samples from two different batches of each isolate were used (two technical replicates and two biological replicates for each isolate). The density/volume of the SYPRO Ruby-stained protein spots were determined using the software package Proteome Weaver (Definiens). For a protein to be reported as differentially expressed, the density values measured in Proteome Weaver were further subjected to the Student?s T-test, and only changes with p< 0.01 were considered significant.

In gel trypsin digestion and peptide analysis using LC/MS/MS:
In-gel digestion with trypsin was performed with a robotic digestion system (Investigator ProGest, Genomic Solutions).Colloidal G was removed by sequential washing with 50 mM ammonium hydrogen carbonate buffer and acetonitrile. Cysteine residues were reduced with DTT and derivatised by treatment with iodoacetamide. After further washing with ammonium hydrogen carbonate buffer, the gel pieces were again dehydrated with acetonitrile and dried at 60°C, prior to the addition of modified trypsin (Promega) (10 µl of a 6.5 ng µl-1 solution in 25 mM ammonium hydrogen carbonate). Digestion proceeded for 8 h at 37°C, and the products were recovered by sequential extractions with 25 mM ammonium hydrogen carbonate, 5% v/v formic acid, and acetonitrile. The pooled extracts were lyophilised, and redissolved in 0.1% v/v formic acid for mass spectrometry.
Tandem electrospray mass spectra were recorded using a Q-TOF hybrid quadrupole / orthogonal acceleration time of flight spectrometer (Micromass) interfaced to a Micromass CapLC capillary chromatograph. Samples were dissolved in 0.1% v/v formic acid, and 6 µl was injected onto a Pepmap C18 trap column (300 µm ? 0.5 cm; LC Packings), and washed for 3 min with 0.1 % v/v formic acid. The flow rate was then reduced to 1 µl per min, and the peptides were eluted into the mass spectrometer with an acetonitrile / 0.1% v/v formic acid gradient (5% to 70 % v/v acetonitrile over 20 minutes).
The capillary voltage was set to 3,500 V, and data-dependent MS/MS acquisitions were performed on precursors with charge states of 2, 3 or 4 over a survey mass range of 540-1200 Da. Known trypsin autolysis products and keratin-derived precursor ions were automatically excluded. The collision voltage was varied between 18 and 45 V depending on the charge and mass of the precursor. Product ion spectra were charge state de-encrypted and de-isotoped with a maximum entropy algorithm (MaxEnt 3, Micromass). Proteins were identified by correlation of uninterpreted tandem mass spectra to entries in SwissProt/TREMBL using ProteinLynx Global Server (Version 1.1, Micromass) and NCBInr using MASCOT. One missed cleavage per peptide was allowed, and an initial mass tolerance of 50 ppm was used in all searches. Cysteines were assumed to be carbamidomethylated, but other potential modifications were not considered in the first pass search.
All matching spectra were reviewed manually, but in cases where the score reported by ProteinLynx global server was less than 100, additional searches were performed against the NCBI nr database using MASCOT, which utilises a robust probalistic scoring algorithm. Identifications based on a single matching peptide were verified manually using the MassLynx program Pepseq (Micromass).


title: FACS-sorted cells with photoconverted Kaede vs. FACS-sorted cells with non-photoconverted Kaede_ZF1350

description: Zebrafish embryos


title: NBTII cells FGF1 stimulation 2 hours rep1

value: NBTII ; stimulation :FGF1 ; time :2 hours


title: NBTII cells FGF1 stimulation 2 hours rep2


title: NBTII-FGFR1 transfected cells, FGF1 stimulation 2h rep2

value: NBTII-FGFR1 ; stimulation :FGF1 ; time :2 hours


title: NBTII-FGFR1 transfected cells, FGF1 stimulation 30min rep1

value: NBTII-FGFR1 ; stimulation :FGF1 ; time :30 min


title: FACS-sorted cells with non-photoconverted Kaede vs. whole embryos_ZF1344

description: Zebrafish embryos


title: NBTII-FGFR1 transfected cells, FGF1 stimulation Omin rep2

value: NBTII-FGFR1 ; stimulation :FGF1 ; time :0 min ; control cells


title: FACS-sorted cells with non-photoconverted Kaede vs. whole embryos_ZF1340

description: Zebrafish embryos


title: MEF vs. NEF_ZF1335

description: Zebrafish embryos


title: MEF vs. NEF_ZF1333

description: Zebrafish embryos


title: FACS-sorted cells with non-photoconverted Kaede vs. whole embryos_ZF1341

description: Zebrafish embryos


title: NBTII cells FGF1 stimulation Omin rep1

value: NBTII ; stimulation :FGF1 ; time :0 min ; control cells


title: 045_10_01

status: Public on Jan 31 2008

city: Pennington

first: Shirin

middle: Khambata

last: Ford

state: NJ

address: 311 Pennington-Rocky Hill Road, 3B-2.06

institute: Bristol-Myers Squibb Pharmaceutical Research Institute

description: Gene expression data from pre-cetuximab monotherapy treatment biopsies of advanced metastatic colorectal cancer patients

scan_protocol: GeneChips were scanned using the Affymetrix GeneChip Scanner 3000

data_processing: Data scaled to 1500 mean signal intensity value using GCOS 1.1.1.052

organism: Homo sapiens

source_name: Pre-treatment metastatic biopsy of CRC

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 1 microg total RNA RNA!Sample_hyb_protocol = Standard Affymetrix procedures

extract_protocol: RNA isolated from core-needle biopsy stored in RNAlater at RT using RNeasy Mini Kit (Qiagen)according to the manufacturer's instructions.

value: Liver


title: 045_10_02


title: 045_10_09


title: 045_10_04


title: 045_09_08


title: MEF vs. NEF_ZF1331

description: Zebrafish embryos


title: MEF vs. NEF_ZF1332

description: Zebrafish embryos


title: FoxH1 morpholino-injected 40% epiboly-stage embryos vs. control morpholino-injected 40% epiboly-stage embryos_ZF1223

description: Zebrafish embryos


title: NBTII-FGFR1 transfected cells, FGF1 stimulation 48h rep1


title: 045_09_04


title: 045_09_06


title: 045_09_05


title: NBTII cells FGF1 stimulation 48 hours rep1


title: NBTII-FGFR1 transfected cells, FGF1 stimulation 24h rep1

value: NBTII-FGFR1 ; stimulation :FGF1 ; time :24 hours


title: 045_08_10


title: NBTII-FGFR1 transfected cells, FGF1 stimulation 24h rep2


title: NBTII cells FGF1 stimulation 6 hours rep1

value: NBTII ; stimulation :FGF1 ; time :6 hours


title: NBTII cells FGF1 stimulation 6 hours rep2


title: NBTII-FGFR1 transfected cells, FGF1 stimulation 2h rep1


title: NBTII cells FGF1 stimulation 30min rep1

value: NBTII ; stimulation :FGF1 ; time :30 min


title: NBTII-FGFR1 transfected cells, FGF1 stimulation Omin rep1


title: 045_10_03

value: Abdomen (possibly rectal)


title: 045_10_06


title: 045_10_10

value: Rib


title: 045_07_06

value: Cervical-Neck node


title: 045_06_10

value: Rectal


title: NBTII cells FGF1 stimulation 24 hours rep1


title: 045_10_05


title: 045_08_09


title: 045_06_07

value: Adrenal gland; Kidney


title: 045_06_05


title: 045_09_01


title: 045_08_08


title: 045_06_02


title: 045_08_07


title: 045_08_04

value: Scalp


title: 045_09_03

value: Abdomen


title: 045_08_01


title: 045_05_09


title: 045_07_04


title: 045_07_08


title: 045_05_04


title: 045_07_03


title: 045_07_01


title: 045_07_10


title: 045_07_02

value: Abdomen (soft tissue mass)


title: 045_05_03


title: 045_08_02


title: 045_06_08


title: 045_06_03


title: 045_06_06


title: 045_06_01


title: 045_08_03


title: 045_03_03


title: 045_05_10


title: 045_03_04

value: Bone


title: 045_03_09


title: 045_05_07


title: 045_02_10


title: 045_05_01


title: 045_04_10


title: 045_05_05


title: 045_05_02


title: 045_04_09


title: 045_04_07


title: 045_04_06


title: 045_02_04


title: 045_04_04


title: 045_02_01


title: 045_04_05

value: Pancreas


title: 045_02_02


title: 045_04_01


title: 045_03_10


title: 045_03_08


title: 045_04_03


title: 045_01_09


title: 045_03_05


title: 045_03_06

value: Colon


title: 045_03_02


title: 045_03_01


title: 045_02_08

value: Lung


title: NICD Injected 1

status: Public on Jan 18 2008

city: New York

first: Curtis

middle: R

last: Altmann

state: NY

address: 599 Lexington Avenue

institute: Goodwin Procter LLP

description: St 20, NICD injected animal pole tissue explant

hyb_protocol: standard Affymetrix procedures

scan_protocol: standard Affymetrix procedures

data_processing: Affymetrix Microarray Suite version 5.0

organism: Xenopus laevis

source_name: Animal pole explant

label_protocol: 1 µg of total RNA was processed using MessageAmp™ aRNA Amplification Kit (AMBION) to produce biotinylated cRNA targets.

extract_protocol: Animal pole tissue explants were isolated embryonic stage 8 and cultured until sibling embryos reached stage 20.

tag: Wild type, NICD RNA Injected Stage 20 explant


title: 045_01_01


title: 045_02_09


title: Untreated animal pole explant 3

description: St 20, untreated animal pole tissue explant

tag: Wild type, Uninjected Stage 20 explant


title: 045_02_06


title: Untreated animal pole explant 2


title: 045_01_08


title: Female BALB/c Meibomian Estrogen+Progesterone, biological rep 1

status: Public on Jan 01 2008

city: Boston

first: David

middle: A

last: Sullivan

state: MA

address: 20 Staniford Street

institute: Schepens Eye Research Institute

description: Meibomian gland gene expression from ovarectomized mice treated with estrogen+progesterone for 14 days.

hyb_protocol: Following fragmentation, 10 microg of cRNA were hybridized for 18 hr at 37C on GE Healthcare/Amersham Biosciences CodeLink UniSet Mouse I Bioarray. Bioarrays were washed and stained with streptavidin-Alexa 647 and washed again.

scan_protocol: Bioarrays were scanned with a ScanArray Express HT scanner using ScanArray Express software.

data_processing: Data were analyzed with the CodeLink Image Analysis software using the default settings and normalized using global scaling to a median target intensity  on each array of 1.0.

source_name: Meibomian glands from estrogen+progesterone treated female BALB/c mice (glands pooled from 5-8 mice)

label_protocol: Biotinylated cRNA were prepared according to the standard Codelink protocol from 2 microg total RNA using a CodeLink Expression Assay Reagent Kit

extract_protocol: RNA was purified with Trizol and RNAqueous spin columns following the manufacturer's instructions.


title: 045_02_03


title: 045_01_07


title: Female BALB/c Meibomian Estrogen+Progesterone, biological rep 2

source_name: Meibomian glands from estrogen treated female BALB/c mice (glands pooled from 5-8 mice)


title: 045_02_05


title: Female BALB/c Meibomian Progesterone, biological rep 2

description: Meibomian gland gene expression from ovarectomized mice treated with progesterone for 14 days.

source_name: Lacrimal glands from progesterone treated female BALB/c mice (glands pooled from 5-8 mice)


title: Untreated animal pole explant 1


title: NICD Injected 2


title: 045_01_03

value: Rectum


title: Female BALB/c Meibomian Estrogen, biological rep 1

description: Meibomian gland gene expression from ovarectomized mice treated with estrogen for 14 days.


title: Female BALB/c Meibomian Placebo, biological rep 3

description: Meibomian gland gene expression from ovarectomized mice treated with placebo for 14 days.

source_name: Meibomian glands from placebo treated female BALB/c mice (glands pooled from 5-8 mice)


title: 045_01_04

value: Liver


title: 045_01_10


title: 045_01_11


title: Female BALB/c Meibomian Estrogen, biological rep 3


title: 045_01_05


title: Female BALB/c Meibomian Placebo, biological rep 1


title: Female BALB/c Meibomian Estrogen, biological rep 2


title: Female BALB/c Meibomian Progesterone, biological rep 3

description: Meibomian gland gene expression from ovarectomized mice treated with progesterone  for 14 days.

source_name: Meibomian glands from progesterone treated female BALB/c mice (glands pooled from 5-8 mice)


title: Female BALB/c Meibomian Placebo, biological rep 2


title: Female BALB/c Lacrimal Progesterone, biological rep 2

description: Lacrimal gland gene expression from ovarectomized mice treated with progesterone for 14 days.


title: Female BALB/c Meibomian Progesterone, biological rep 1


title: Female BALB/c Lacrimal Progesterone, biological rep 3

description: Lacrimal gland gene expression from ovarectomized mice treated with progesterone  for 14 days.


title: Female BALB/c Meibomian Estrogen+Progesterone, biological rep 3


title: P21 Cochlear Nucleus, Deafferented P21_48h_2X

status: Public on Dec 26 2007

city: Seattle

first: Julie

last: Harris

state: WA

department: Otolaryngology-HNS

institute: University of Washington

description: Ipsilateral CN 48 hours after deafferentation.  Replicate 2.

hyb_protocol: Affymetrix standard protocol

scan_protocol: Affymetrix standard protocol

data_processing: Normalization using GC-RMA

label: Biotin

source_name: P21 Cochlear Nucleus, Deafferented

label_protocol: Affymetrix standard protocol

extract_protocol: Qiagen lipid RNeasy kit with on column DNAse digestion

treatment_protocol: Deafferented, ipsilateral to cochlear removal + 48h, CN dissected fresh & snap frozen

tag: Age P21, strain C57Bl6/j


title: Replicate_1_dyeswap_B_slide_21

city: Chapel Hill

first: John

middle: F

last: Rawls

state: NC

address: 111 Mason Farm Rd.

department: Department of Cell and Molecular Physiology

institute: University of North Carolina at Chapel Hill

hyb_protocol: Competing cDNA samples for each hybridization were paired, purified and concentrated using Microcon YM-30 microconcentrators (Millipore), and then resuspended in hybridization buffer (MWG Biotech).  Four hybridizations were carried out in a sequential manner.  The primary hybridization was performed by adding 40 μl of sample to the DNA microarray under a supported glass coverslip (Erie Scientific) with subsequent incubation at 42oC for 16-20 h at high humidity in the dark.  Prior to the secondary hybridization, slides were gently submerged in 2X SSC/0.2% SDS, transferred to 2X SSC, and then to 0.2X SSC (12 min/wash; room temperature) with subsequent drying by centrifugation.  Secondary hybridization was performed using the complementary capture reagents provided in the 3DNA Array 350 kit (Genisphere). 40 μL reaction mixtures were assembled containing 2.5 μL 3DNA capture reagent with Cy3, 2.5 μL 3DNA capture reagent with Cy5, 20 μL MWG hybridization buffer (containing a 1:100 dilution Genisphere anti-fade reagent) plus 15 μL DEPC-treated water, and then incubated in the dark at 80oC for 10 min, followed by 15 min at 50oC. 40μL of the treated solution was then added to the slide under a supported glass coverslip and incubated at 50oC for 3 h at high humidity in the dark. Following hybridization, the DNA microarray was washed in 2X SSC/0.2% SDS, followed by 2X SSC and 0.2X SSC (12 min/cycle; room temperature), and then dried by centrifugation.

scan_protocol: Slides were scanned in a ScanArray Express HT (Perkin Elmer) machine to detect Cy3 and Cy5 fluorescence at 543nm and 633nm, respectively.  Laser power was kept constant for Cy3/Cy5 scans and PMT values were set for each experiment based on background fluorescence. Gridding and analysis of images was performed with ScanArray software (Perkin Elmer).

data_processing: Each spot was defined on a pixel-by-pixel basis using a modified Mann-Whitney statistical test.  Normalization of Cy3/Cy5 channel intensities was performed using a LOWESS procedure.  After normalization, expression ratios were calculated for each spot and transformed to log values.

source_name: Whole 6dpf zebrafish larvae mock-infected at 1dpf; biological replicate 1

label_protocol: First strand cDNA was generated by reverse transcription utilizing the 3DNA Array 350 kit (Genisphere) and modified oligo-dT primers containing a 5’ fluorophore/dendrimer-specific sequence. The fluorophore-specific oligo-dT primer (1 μL/reaction) was added to 5 μg total RNA in 10 μL DEPC-treated water. The 11 μL solution was incubated at 80oC for 10 min, cooled on ice for 2 min and the following materials added to a total volume of 20μL: 4 μL 5X first strand buffer (Invitrogen), 1 μL RNase inhibitor (Genisphere), 1 μL dNTP mix (10mM dATP, dCTP, dGTP, and dTTP), 2 μL 0.1M DTT, and 1 μL Superscript II RNase H- Reverse Transcriptase (10 U/μL; Invitrogen). Reverse transcription was carried out at 42oC for 2 h and terminated by adding 0.5M NaOH / 50mM EDTA (65oC for 10 min), followed by neutralization with 1M Tris-HCl pH 7.5.

extract_protocol: Total RNA was purified from pooled intact zebrafish larvae (n=180 larvae) and digested with DNase I using an Absolutely RNA Microprep Kit (Stratagene) according to manufacturers' protocols. RNA quality was confirmed using an Agilent 2100 Bioanalyzer.

value: AB, Age:6 days post-fertilization (6dpf), Tissue:whole larvae, Rearing conditions:conventional, Microbial status:mock-infected, Days post-infection:5

source_name: Whole 6dpf zebrafish larvae infected with Mycobacterium marinum wild-type strain at 1dpf; biological replicate 1

value: AB, Age:6 days post-fertilization (6dpf), Tissue:whole larvae, Rearing conditions:conventional, Microbial status:infected with Mycobacterium marinum wild-type strain, Days post-infection:5


title: Untreated animal pole explant 4


title: P21 Cochlear Nucleus, Deafferented P21_24h_3X

description: Ipsilateral CN 24 hours after deafferentation.  Replicate 3.

treatment_protocol: Deafferented, ipsilateral to cochlear removal + 24h, CN dissected fresh & snap frozen


title: P21 Cochlear Nucleus, Deafferented P21_48h_3X

description: Ipsilateral CN 48 hours after deafferentation.  Replicate 3.


title: Female BALB/c Lacrimal Estrogen+Progesterone, biological rep 1

description: Lacrimal gland gene expression from ovarectomized mice treated with estrogen+progesterone for 14 days.

source_name: Lacrimal glands from estrogen+progesterone treated female BALB/c mice (glands pooled from 5-8 mice)


title: P21 Cochlear Nucleus, Deafferented P21_48h_1X

description: Ipsilateral CN 48 hours after deafferentation.  Replicate 1.


title: Replicate_2_dyeswap_B_slide_23

source_name: Whole 6dpf zebrafish larvae mock-infected at 1dpf; biological replicate 2

extract_protocol: Total RNA was purified from pooled intact zebrafish larvae (n=154 larvae) and digested with DNase I using an Absolutely RNA Microprep Kit (Stratagene) according to manufacturers' protocols. RNA quality was confirmed using an Agilent 2100 Bioanalyzer.

source_name: Whole 6dpf zebrafish larvae infected with Mycobacterium marinum wild-type strain at 1dpf; biological replicate 2

extract_protocol: Total RNA was purified from pooled intact zebrafish larvae (n=176 larvae) and digested with DNase I using an Absolutely RNA Microprep Kit (Stratagene) according to manufacturers' protocols. RNA quality was confirmed using an Agilent 2100 Bioanalyzer.


title: P21 Cochlear Nucleus, Control  P21_48h_3

description: Contralateral CN 48 hours after ipsilateral deafferentation.  Replicate 3.

source_name: P21 Cochlear Nucleus, Control

treatment_protocol: Control, contralateral to cochlear removal + 48h, CN dissected fresh & snap frozen


title: Female BALB/c Lacrimal Estrogen+Progesterone, biological rep 2


title: Female BALB/c Lacrimal Estrogen+Progesterone, biological rep 3


title: Replicate_2_dyeswap_A_slide_22


title: Female BALB/c Lacrimal Estrogen, biological rep 3

description: Lacrimal gland gene expression from ovarectomized mice treated with estrogen for 14 days.

source_name: Lacrimal glands from estrogen treated female BALB/c mice (glands pooled from 5-8 mice)


title: Replicate_1_dyeswap_A_slide_20


title: Female BALB/c Lacrimal Estrogen, biological rep 2


title: Female BALB/c Lacrimal Placebo, biological rep 3

description: Lacrimal gland gene expression from ovarectomized mice treated with placebo for 14 days.

source_name: Lacrimal glands from placebo treated female BALB/c mice (glands pooled from 5-8 mice)


title: Female BALB/c Lacrimal Placebo, biological rep 2


title: Female BALB/c Lacrimal Estrogen, biological rep 1


title: Female BALB/c Lacrimal Placebo, biological rep 1


title: P21 Cochlear Nucleus, Control  P21_12h_2

description: Contralateral CN 12 hours after ipsilateral deafferentation.  Replicate 2.

treatment_protocol: Control, contralateral to cochlear removal + 12h, CN dissected fresh & snap frozen


title: P21 Cochlear Nucleus, Control  P21_24h_3

description: Contralateral CN 24 hours after ipsilateral deafferentation.  Replicate 3.

treatment_protocol: Control, contralateral to cochlear removal + 24h, CN dissected fresh & snap frozen


title: P21 Cochlear Nucleus, Deafferented P21_12h_1X

description: Ipsilateral CN 12 hours after deafferentation.  Replicate 1.

treatment_protocol: Deafferented, ipsilateral to cochlear removal + 12h, CN dissected fresh & snap frozen


title: P21 Cochlear Nucleus, Control  P21_48h_2

description: Contralateral CN 48 hours after ipsilateral deafferentation.  Replicate 2.


title: P21 Cochlear Nucleus, Control  P21_48h_1

description: Contralateral CN 48 hours after ipsilateral deafferentation.  Replicate 1.


title: P21 Cochlear Nucleus, Deafferented P21_12h_2X

description: Ipsilateral CN 12 hours after deafferentation.  Replicate 2.


title: Female BALB/c Lacrimal Progesterone, biological rep 1


title: P21 Cochlear Nucleus, Deafferented P21_24h_2X

description: Ipsilateral CN 24 hours after deafferentation.  Replicate 2.


title: P7 Cochlear Nucleus, Control  P7_48h_1

source_name: P7 Cochlear Nucleus, Control

tag: Age P7, strain C57Bl6/j


title: P21 Cochlear Nucleus, Deafferented P21_24h_1X

description: Ipsilateral CN 24 hours after deafferentation.  Replicate 1.


title: P21 Cochlear Nucleus, Control  P21_24h_2

description: Contralateral CN 24 hours after ipsilateral deafferentation.  Replicate 2.


title: P21 Cochlear Nucleus, Control  P21_24h_1

description: Contralateral CN 24 hours after ipsilateral deafferentation.  Replicate 1.


title: P7 Cochlear Nucleus, Deafferented P7_24h_2X

source_name: P7 Cochlear Nucleus, Deafferented


title: P21 Cochlear Nucleus, Control  P21_12h_3

description: Contralateral CN 12 hours after ipsilateral deafferentation.  Replicate 3.


title: P7 Cochlear Nucleus, Control  P7_24h_3


title: P21 Cochlear Nucleus, Deafferented P21_12h_3X

description: Ipsilateral CN 12 hours after deafferentation.  Replicate 3.


title: P7 Cochlear Nucleus, Deafferented P7_24h_1X


title: P7 Cochlear Nucleus, Deafferented P7_48h_1X


title: P7 Cochlear Nucleus, Control  P7_48h_2


title: P7 Cochlear Nucleus, Control  P7_48h_3


title: P7 Cochlear Nucleus, Control  P7_12h_3


title: P21 Cochlear Nucleus, Control  P21_12h_1

description: Contralateral CN 12 hours after ipsilateral deafferentation.  Replicate 1.


title: P7 Cochlear Nucleus, Deafferented P7_48h_3X

description: Ipsilateral CN 48 hours after deafferentation.  Replicate 3.


title: P21 Cochlear Nucleus, Deafferented P21_6h_2X

description: Ipsilateral CN 6 hours after deafferentation.  Replicate 2.

treatment_protocol: Deafferented, ipsilateral to cochlear removal + 6h, CN dissected fresh & snap frozen


title: P7 Cochlear Nucleus, Deafferented P7_6h_1X

description: lpsilateral CN 6 hours after deafferentation.  Replicate 1.


title: P7 Cochlear Nucleus, Control  P7_24h_1


title: P7 Cochlear Nucleus, Control  P7_6h_2

description: Contralateral CN 6 hours after ipsilateral deafferentation.  Replicate 2.

treatment_protocol: Control, contralateral to cochlear removal + 6h, CN dissected fresh & snap frozen


title: P7 Cochlear Nucleus, Deafferented P7_6h_2X

description: lpsilateral CN 6 hours after deafferentation.  Replicate 2.


title: P7 Cochlear Nucleus, Control  P7_12h_1


title: P7 Cochlear Nucleus, Deafferented P7_12h_1X


title: P7 Cochlear Nucleus, Deafferented P7_48h_2X

description: Ipsilateral CN 48 hours after deafferentation.  Replicate 2.


title: P21 Cochlear Nucleus, Control P21_6h_1

description: Contralateral CN 6 hours after ipsilateral deafferentation.  Replicate 1.


title: P7 Cochlear Nucleus, Control  P7_6h_1


title: P7 Cochlear Nucleus, Control  P7_12h_2

description: Contralateral CN 12 hours after ipsilateral deafferentation.  Replicate 2.


title: P21 Cochlear Nucleus, Control  P21_6h_2


title: P7 Cochlear Nucleus, Deafferented P7_12h_2X


title: P7 Cochlear Nucleus, Control  P7_24h_2


title: C3H sighted - eye rep 6

status: Public on Dec 01 2007

city: Neuherberg

first: Martin

last: Irmler

state: Bayern

address: Ingolstaedter Landstrasse 1

department: Institute of Experimental Genetics

country: Germany

institute: Helmholtz Zentrum München GmbH

description: RNA isolation, Labelling, Hybridisation and Scanning procedure are described in the fields above

hyb_protocol: This hybridization mixture was placed on a prehybridised microarray, under a cover slip, placed into a hybridization chamber (Genetix) and immersed in a thermostatic bath at 42°C for at least 16 hours. After hybridization slides were washed in 40ml of 3x , 1x, 0.25x and 0.1x SSC at room temperature. For drying slides were placed in an empty 50 ml Falcon tube (Becton Dickinson, USA) and centrifuged at 4000m/s2. Beckers, J., Herrmann, F., Rieger, S., Drobyshev, A., Horsch, M., Hrabé de Angelis, M. and Seliger, B. (2005): Identification and validation of novel ERBB2 (Her2, NEU) targets including genes involved in angiogenesis. Int. J. Cancer 114: 590-597. Greenwood AD, Horsch M, Stengel A, Vorberg I, Lutzny G, Maas E, Schädler S, Erfle V, Beckers J, Schätzl H and Leib-Mösch C (2005): Cell Line Dependent RNA Expression Profiles of Prion-infected Mouse Neuronal Cells. JMB 349: 487-500 Seltmann M, Horsch M, Drobyshev A, Chen Y, Hrabé de Angelis M, Beckers J (2005): Assessment of a Systematic Expression Profiling. Approach in ENU-Induced Mouse Mutant Lines. Mam Genome 16 (1): 1-10

scan_protocol: Dried slides were scanned with a GenePix 4000A microarray scanner and the images were analyzed using the GenePix Pro3.0 image processing software (Axon Instruments, USA) Greenwood AD, Horsch M, Stengel A, Vorberg I, Lutzny G, Maas E, Schädler S, Erfle V, Beckers J, Schätzl H and Leib-Mösch C (2005): Cell Line Dependent RNA Expression Profiles of Prion-infected Mouse Neuronal Cells. JMB 349: 487-500 Seltmann M, Horsch M, Drobyshev A, Chen Y, Hrabé de Angelis M, Beckers J (2005): Assessment of a Systematic Expression Profiling. Approach in ENU-Induced Mouse Mutant Lines. Mam Genome 16 (1): 1-10

data_processing: Normalisation with GenePix Pro 6.0: Ratio based normalized data in each image so that the mean of ratio of means of all of the features is equal to 1. The VALUE column contains the log2 ratio of the result file after normalisation

source_name: C3H sighted eye

label_protocol:
For labeling 20 µg of total RNA were used for reverse transcription and indirectly labeled with Cy3 or Cy5 fluorescent dye according the TIGR protocol (Hedge et al., 2000). Labeled cDNA was dissolved in 30 µl hybridization buffer (6x SSC, 0.5% SDS, 5x Denhardt’s solution and 50% formamide) and mixed with 30 µl of reference cDNA solution (pool from six control animals) labeled with the second dye.
HegdeP, Qi R, Abernathy R, Gay C, Dharap S, et al (2000): A concise guide to cDNA microarray analysis-II. Biotechniques 29: 548-562

extract_protocol: Total RNA was isolated just before processing for expression profiling. For preparation of total RNA individual organs were thawed in buffer containing chaotropic salt (RLT buffer, Qiagen) and homogenised using a Polytron homogeniser. Total RNA from individual samples was obtained according to manufacturer’s protocols using RNeasy Midi kits (Qiagen). The concentration was calculated from OD260/280 measurement and 2 µg RNA aliquots were run on a formaldehyde agarose gel to check for RNA integrity. The RNA was stored at -80°C in RNase free water (Qiagen).

tag: C3H.pde6b+, male, eye, Mouse ID 	30059625

source_name: C3H eye

tag: C3H.pde6b-, male, eye, RNA pool including Mouse ID


title: C3H sighted - eye rep 8

tag: C3H.pde6b+, male, eye, Mouse ID 	30059655


title: C3H sighted - eye rep 7


title: C3H sighted - eye rep 10

tag: C3H.pde6b+, male, eye, Mouse ID 	30059656


title: C3H sighted - eye rep 12

tag: C3H.pde6b+, male, eye, Mouse ID 	30059659


title: Neuro2a_transfected_60H_Rep3

status: Public on Dec 08 2007

city: New Delhi

first: TANAY

last: GHOSH

email: tghosh@igib.res.in, gtanay@yahoo.com

state: Delhi

address: Mall Road

department: Functional Genomics Unit

country: India

institute: Institute of Genomics and Integrative Biology

description: Not applicable

hyb_protocol: Hybridization buffer: DIG Easy Hyb buffer (Roche); Hybridization temprature: 37degree centigrade; Hybridization time: 16 H; Blocking, washing and post hybridization steps: according to the manufacturer instruction given in Micromax NEN TSA labeling system (Perkin-Elmer Life Sciences, USA).

scan_protocol: Fluorescent array images were scanned for both Cy3 and Cy5 with GenePix 4200A scanner and image intensities were analyzed with GenePixpro 6.0 software.

data_processing: Background subtracted median intensity of each channel was taken to represent the actual spot intensity. log2 transformed spot intensities were used for Z score normalization described by Cheadle C et al., (J Mol Diagn 2003;5:73-81). Briefly, Z score was computed for each CH1 and CH2 spot according to the mathematical expression: Z score=(x-a)/b where, x=log2 transformed spot intensity, a=mean and b=standard deviation of all log2 transformed spot intensity in a particular channel. Z score result for each control channel (here CH1) spot was then subtracted from the treated channel (here CH2) to get Z difference. Z differences were then Z score normalized for computing the Z-ratio for each spot.

source_name: Neuro 2a cells, transfected with vector construct contains GFP (pEGFPN3;Clontech).

label_protocol: Total RNA labeled by Micromax NEN TSA labeling system (Perkin-Elmer Life Sciences, USA) according to manufacturer instruction.

growth_protocol: Neuro-2a were maintained in Minimum essential medium (MEM) (GIBCO-BRL) supplemented with 10% fetal calf serum [Biological Industries, Israel], 2mM L-glutamine [Sigma], 1mM sodium pyruvate [Sigma] and antibiotic-antimycotic solution (100X stock) [Sigma] at 370C humidified incubator with 5% CO2. Approximately 0.3 million cells were seeded in each well of 6 well plate 24 H prior to transient transfection when 50-80% confluency was reached. Cells were washed once with Opti-MEM (GIBCO-BRL) and maintained in 1 ml Opti-MEM. After transfection cells were kept in Co2 incubator (mentioned above) for 60H.

extract_protocol: After 60 hs. of transfection total RNA was isolated from the cells using TRIzol reagent (GIBCO-BRL) according to the manufacturer’s instruction. RNA was cleaned using RNeasy Mini Kit (Qiagen) following the protocol by the manufacturer.

treatment_protocol: Neuro 2a cells were transfected with vector construct contains GFP. 1µg of transfection quality DNA (Endo free plasmid maxi kit; Qiagen) and 3µl of fugene 6 transfection reagent (Roche) were used for preparation of transfection complex. Transfection was performed following the protocol from the manufacturer.

source_name: Neuro 2a cells, transfected with GFP tagged human TBP containing CAG repeat length 59.

growth_protocol: Neuro-2a were maintained in Minimum essential medium (MEM) (GIBCO-BRL) supplemented with 10% fetal calf serum [Biological Industries, Israel], 2mM L-glutamine [Sigma], 1mM sodium pyruvate [Sigma] and antibiotic-antimycotic solution (100X stock) [Sigma] at 37degree Centigrade humidified incubator with 5% CO2. Approximately 3×105 cells were seeded in each well of 6 well plate 24 H prior to transient transfection when 50-80% confluency was reached. Cells were washed once with Opti-MEM (GIBCO-BRL) and maintained in 1 ml Opti-MEM. After transfection cells were kept in Co2 incubator (mentioned above) for 60H.

treatment_protocol: Neuro 2a cells were transfected with a construct containing GFP tagged human TBP bearing CAG repeat length 59. 1µg of transfection quality DNA (Endo free plasmid maxi kit; Qiagen) and 3µl of fugene 6 transfection reagent (Roche) were used for preparation of transfection complex. Transfection was performed following the protocol from the manufacturer.

tag: Murine neuroblastoma cells (ATCC number CCL-131; Neuro-2a or N2a).


title: Neuro2a_transfected_60H_Rep5

description: Not applicable

growth_protocol: Neuro-2a were maintained in Minimum essential medium (MEM) (GIBCO-BRL) supplemented with 10% fetal calf serum [Biological Industries, Israel], 2mM L-glutamine [Sigma], 1mM sodium pyruvate [Sigma] and antibiotic-antimycotic solution (100X stock) [Sigma] at 37 degree Centigrade humidified incubator with 5% CO2. Approximately 0.3 million cells were seeded in each well of 6 well plate 24 H prior to transient transfection when 50-80% confluency was reached. Cells were washed once with Opti-MEM (GIBCO-BRL) and maintained in 1 ml Opti-MEM. After transfection cells were kept in Co2 incubator (mentioned above) for 60H.

tag: Murine neuroblastoma cells (ATCC number CCL-131; Neuro-2a or N2a)


title: C3H sighted - eye rep 11


title: P21 Cochlear Nucleus, Deafferented P21_6h_1X

description: Ipsilateral CN 6 hours after deafferentation.  Replicate 1.


title: Neuro2a_transfected_60H_Rep6

description: Not applicable.

data_processing: Microsoft Excel was used for data processing. Background subtracted median intensity of each channel was taken to represent the actual spot intensity. log2 transformed spot intensities were used for Z score normalization described by Cheadle C et al., (J Mol Diagn 2003;5:73-81). Briefly, Z score was computed for each CH1 and CH2 spot according to the mathematical expression: Z score=(x-a)/b where, x=log2 transformed spot intensity, a=mean and b=standard deviation of all log2 transformed spot intensity in a particular channel. Z score result for each control channel (here CH1) spot was then subtracted from the treated channel (here CH2) to get Z difference. Z differences were then Z score normalized for computing the Z-ratio for each spot.


title: Neuro2a_transfected_60H_Rep4

description: Not applicable

growth_protocol: Neuro-2a were maintained in Minimum essential medium (MEM) (GIBCO-BRL) supplemented with 10% fetal calf serum [Biological Industries, Israel], 2mM L-glutamine [Sigma], 1mM sodium pyruvate [Sigma] and antibiotic-antimycotic solution (100X stock) [Sigma] at 37degree Centigrade humidified incubator with 5% CO2. Approximately 0.3 million cells were seeded in each well of 6 well plate 24 H prior to transient transfection when 50-80% confluency was reached. Cells were washed once with Opti-MEM (GIBCO-BRL) and maintained in 1 ml Opti-MEM. After transfection cells were kept in Co2 incubator (mentioned above) for 60H.


title: C3H sighted - eye rep 5


title: P7 Cochlear Nucleus, Deafferented P7_24h_3X

description: Ipsilateral CN 24 hours after deafferentation.  Replicate 3.


title: Neuro2a_transfected_60H_Rep1

description: Not applicable

scan_protocol: Fluorescent array images were scanned for both Cy3 and Cy5 with a ScanArray scanner (Packard Biochip Technologies) and image intensities were analyzed with QuantArray version 3.0.0.0 (Packard Biochip Technologies) analysis software.

data_processing: Background subtracted mean intensity of each channel was taken to represent the actual spot intensity. log2 transformed spot intensities were used for Z score normalization described by Cheadle C et al., (J Mol Diagn 2003;5:73-81). Briefly, Z score was computed for each CH1 and CH2 spot according to the mathematical expression: Z score=(x-a)/b where, x=log2 transformed spot intensity, a=mean and b=standard deviation of all log2 transformed spot intensity in a particular channel. Z score result for each control channel (here CH2) spot was then subtracted from the treated channel (here CH1) to get Z difference. Z differences were then Z score normalized for computing the Z-ratio for each spot.

growth_protocol: Neuro-2a were maintained in Minimum essential medium (MEM) (GIBCO-BRL) supplemented with 10% fetal calf serum [Biological Industries, Israel], 2mM L-glutamine [Sigma], 1mM sodium pyruvate [Sigma] and antibiotic-antimycotic solution (100X stock) [Sigma] at 370C humidified incubator with 5% CO2. Approximately 3ױ05 cells were seeded in each well of 6 well plate 24 H prior to transient transfection when 50-80% confluency was reached. Cells were washed once with Opti-MEM (GIBCO-BRL) and maintained in 1 ml Opti-MEM. After transfection cells were kept in Co2 incubator (mentioned above) for 60H.

extract_protocol: After 60 hs. of transfection total RNA was isolated from the cells using TRIzol reagent (GIBCO-BRL) according to the manufacturer?s instruction. RNA was cleaned using RNeasy Mini Kit (Qiagen) following the protocol by the manufacturer.

treatment_protocol: Neuro 2a cells were transfected with a construct containing GFP tagged human TBP bearing CAG repeat length 59. 1ug of transfection quality DNA (Endo free plasmid maxi kit; Qiagen) and 3ul of fugene 6 transfection reagent (Roche) were used for preparation of transfection complex. Transfection was performed following the protocol from the manufacturer.

treatment_protocol: Neuro 2a cells were transfected with vector construct contains GFP. 1ug of transfection quality DNA (Endo free plasmid maxi kit; Qiagen) and 3ul of fugene 6 transfection reagent (Roche) were used for preparation of transfection complex. Transfection was performed following the protocol from the manufacturer.


title: Neuro2a_transfected_60H_Rep2

description: Not applicable

data_processing: Background subtracted mean intensity of each channel was taken to represent the actual spot intensity. log2 transformed spot intensities were used for Z score normalization described by Cheadle C et al., (J Mol Diagn 2003;5:73-81). Briefly, Z score was computed for each CH1 and CH2 spot according to the mathematical expression: Z score=(x-a)/b where, x=log2 transformed spot intensity, a=mean and b=standard deviation of all log2 transformed spot intensity in a particular channel. Z score result for each control channel (here CH1) spot was then subtracted from the treated channel (here CH2) to get Z difference. Z differences were then Z score normalized for computing the Z-ratio for each spot.

growth_protocol: Neuro-2a were maintained in Minimum essential medium (MEM) (GIBCO-BRL) supplemented with 10% fetal calf serum [Biological Industries, Israel], 2mM L-glutamine [Sigma], 1mM sodium pyruvate [Sigma] and antibiotic-antimycotic solution (100X stock) [Sigma] at 37 degree  centigrade humidified incubator with 5% CO2. Approximately 0.3 million cells were seeded in each well of 6 well plate 24 H prior to transient transfection when 50-80% confluency was reached. Cells were washed once with Opti-MEM (GIBCO-BRL) and maintained in 1 ml Opti-MEM. After transfection cells were kept in Co2 incubator (mentioned above) for 60H.

extract_protocol: After 60 hs. of transfection total RNA was isolated from the cells using TRIzol reagent (GIBCO-BRL) according to the manufacturer instruction. RNA was cleaned using RNeasy Mini Kit (Qiagen) following the protocol by the manufacturer.

growth_protocol: Neuro-2a were maintained in Minimum essential medium (MEM) (GIBCO-BRL) supplemented with 10% fetal calf serum [Biological Industries, Israel], 2mM L-glutamine [Sigma], 1mM sodium pyruvate [Sigma] and antibiotic-antimycotic solution (100X stock) [Sigma] at 37 degree centigrade humidified incubator with 5% CO2. Approximately 3ױ05 cells were seeded in each well of 6 well plate 24 H prior to transient transfection when 50-80% confluency was reached. Cells were washed once with Opti-MEM (GIBCO-BRL) and maintained in 1 ml Opti-MEM. After transfection cells were kept in Co2 incubator (mentioned above) for 60H.

treatment_protocol: Neuro 2a cells were transfected with a construct containing GFP tagged human TBP bearing CAG repeat length 59. 1ug of transfection quality DNA (Endo free plasmid maxi kit; Qiagen) and 3u of fugene 6 transfection reagent (Roche) were used for preparation of transfection complex. Transfection was performed following the protocol from the manufacturer.


title: C3H sighted - heart rep 7

source_name: C3H sighted heart

tag: C3H.pde6b+, male, heart, Mouse ID 	   30053126

source_name: C3H heart

tag: C3H.pde6b-, male, heart, RNA pool including Mouse ID


title: C3H sighted - eye rep 9


title: C3H sighted - heart rep 8


title: C3H sighted - heart rep 4

tag: C3H.pde6b+, male, heart, Mouse ID 	   30053078


title: C3H sighted - eye rep 3

tag: C3H.pde6b+, male, eye, Mouse ID 	30059624


title: C3H sighted - eye rep 1

tag: C3H.pde6b+, male, eye, Mouse ID 	30059404


title: C3H sighted - brain rep 7

source_name: C3H sighted brain

tag: C3H.pde6b+, male, brain, Mouse ID 	   30053078

source_name: C3H brain

tag: C3H.pde6b-, male, brain, RNA pool including Mouse ID


title: C3H sighted - heart rep 6

tag: C3H.pde6b+, male, brain, Mouse ID 	   30053080


title: C3H sighted - heart rep 5


title: C3H sighted - brain rep 8


title: C3H sighted - brain rep 9

tag: C3H.pde6b+, male, brain, Mouse ID 	  30053080


title: C3H sighted - eye rep 4


title: C3H sighted - heart rep 3


title: C3H sighted - heart rep 2

tag: C3H.pde6b+, male, heart, Mouse ID 	   30053079


title: C3H sighted - eye rep 2


title: C3H sighted - brain rep 12

tag: C3H.pde6b+, male, brain, Mouse ID 	   30053126


title: P7 Cochlear Nucleus, Deafferented P7_12h_3X

tag: Age P7, strain C57Bl6/j


title: C3H sighted - brain rep 11


title: C3H sighted - brain rep 1

tag: C3H.pde6b+, male, brain, Mouse ID 	   30053134


title: C3H sighted - brain rep 10


title: C3H sighted - brain rep 2


title: Yeast-control-3

status: Public on Oct 14 2007

city: Indianapolis

first: Mark

last: Wagner

state: IN

address: 950 West Walnut Street

department: Medicine/Nephrology

institute: Indiana University Medical Ctr

description: control

hyb_protocol: Fifteen ug of biotinylated cRNA is added to a total hybridization cocktail of 300ul, and 200ul is hybridized to a GeneChip after adding control oligonucleotides.  Hybridization is at 45C for 17 h with constant rotation.  The hybridization mixture is then removed and the GeneChips are washed, stained with phycoerythrin-labeled Streptavidin, washed, incubated with biotinylated anti-strepavidin, and then restained with phycoerythrin-labeled Strepavidin to amplify the signals; these steps are carried out using the Affymetrix Fluidics Station.  To reduce non-random error, balanced groups of samples are handled in parallel.

scan_protocol: Arrays are then scanned using the dedicated scanner, controlled by Affymetrix GCOS software.  Images are examined for defects.

data_processing: The Affymetrix Microarray Suite version 5 (MAS5) algorithm analyzes the hybridization intensity data from GeneChip expression probe arrays and calculates a set of metrics that describe probe set performance. The average intensity on each array is normalized by global scaling to a target intensity of 1000.

organism: Saccharomyces cerevisiae

source_name: Yeast, untreated-3

label_protocol:
Protocols recommended by Affymetrix were used,  cDNA was synthesized using a T7 promoter-dT24 oligonucleotide as primer with the Invitrogen Life technologies SuperScript Choice system.  Following second strand cDNA synthesis and incubation with T4 DNA polymerase, the products are purfied using an Affymetrix Cleanup Module.
Biotinylated cRNA is made using the Affymetrix IVT kit.  The cRNA is purified using Qiagen RNeasy columns, quantitated and then fragmented by incubation at high temperature with magnesium.

growth_protocol: Cell were grown overnight in YPD +/- gentamicin, diluted to 5x106 cells/ml and grown to mid-log with and without gentamicin 500ug/ml

extract_protocol: Total RNA was isolated using the Epicentre Masterpure Yeast RNA Purification kit (Madison,WI)

treatment_protocol: Control cells received no gentamicin, Gent treated cells were acclimated to growth in gentamicin, 500ug/ml and grown with gentamicin throughout the experiment

tag: Strain 4073


title: C3H sighted - brain rep 5

source_name: sighted

tag: C3H.pde6b+, male, brain, Mouse ID 	   30053125


title: C3H sighted - brain rep 6


title: Epithelial cells pre-exposed with 70 nM 17beta-estradiol, immunopptd with 5-methylcytosine antibody, rep1

type: genomic

city: Columbus

first: Alfred

last: Cheng

state: OH

address: OSU Biomedical Research Tower 460 W. 12th Ave.

institute: The Ohio State University

description: Immunoprecipitated DNA (Cy5) was co-hybridized with total genomic DNA (Cy3). The ratio between Cy5 and Cy3 signal was calculated for all high quality features.

hyb_protocol: After dye coupling, about 200pmol of each Cy dye were denatured and hybridized onto the microarray for 20 hr at 65C. Microarray were washed 1 time with wash buffer I (6x SSPE, 0.005% sacrosine) and 1 time with wash buffer II (0.06x SSPE) and finally dip into acetonitrile.

scan_protocol: microarrays were scanned using the Axon GenePix 4000B Scanner

data_processing: The acquired microarray image was analyzed with GenePix Pro 6.0 software. After excluding the spots flagged for bad quality, signal ratios were log2 transformed and normalized using intensity-dependent normalization. The VALUE column reports Lowess-normalized log2 (IP/input) ratios

molecule: genomic DNA

source_name: Pre-exposed cells immunoprecipitated

label_protocol: DNA were amino-allyl labelled using randomly-primed, Klenow-based extension protocol. Cy5 dye was then incorporated into the DNA using in-direct labeling by incubating the dye with DNA for 3hrs.

extract_protocol: genomic DNA were extracted using standard pheno/chloroform extraction

tag: Pre-exposed cells, anti-5-methylcytosine immunoprecipitated DNA

source_name: Pre-exposed cells_input

label_protocol: DNA were amino-allyl labelled using randomly-primed, Klenow-based extension protocol. Cy3 dye was then incorporated into the DNA using in-direct labeling by incubating the dye with DNA for 3hrs.

tag: Pre-exposed cells, total input DNA


title: C3H sighted - brain rep 3

tag: C3H.pde6b+, male, brain, Mouse ID 30053079


title: Yeast-control-1

description: control

source_name: Yeast, untreated-1


title: Yeast-control-4

description: control

source_name: Yeast, untreated-4


title: Yeast-Gent-8

description: gentamicin-treated

source_name: Yeast, Gent-8


title: Melanoma xenograft, MED1_Sub_B (LeiFidler)

status: Public on Oct 02 2007

city: cambridge

first: steve

last: shen

address: 77 mass ave

institute: MIT

description: human melanoma tumors grown  in mice

hyb_protocol: Following fragmentation, 15 ug of cRNA were hybridized on one U133A human chip and one test chip. GeneChips were washed and stained in the Affymetrix Fluidics Station 400.

data_processing: raw data (CEL file) were normalized with RMA and further analyzied with GenePatter comparative marker selection module.

source_name: Melanoma metastatic derivatives, S.C

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol (701025, Revision 3).

extract_protocol: Rneasy mini kit

tag: Genotype

value: highly metastatic

tag: Age


title: C3H sighted - brain rep 4

tag: C3H.pde6b+, male, brain, Mouse ID 	   30053079


title: Yeast-Gent-5

description: gentamicin-treated

source_name: Yeast, Gent-5


title: propane vs pyruvate, rep1

status: Public on Nov 01 2007

city: Vancouver

first: Justin

middle: Christian

last: LeBlanc

state: British Columbia

address: 2350 Health Sciences Mall

department: Microbiology & Immunology

country: Canada

institute: University of British Columbia

description: Rhodococcus sp. RHA1 grown on propane versus pyruvate, replicate 1

hyb_protocol: The microarray slides were pre-hybridized using 5X SSC containing 0.1% SDS and 0.2% BSA for 45 minutes at 48°C and used immediately for hybridization in a GeneTac HybStation (Genomic Solution). Equal amounts of differentially labelled cDNA – 50 million pixels as measured by ImageQuant 5.2 (Molecular Dynamics) – from propane- and pyruvate-grown cells were hybridized at 42°C for 17 h with agitation using 120 ?L per slide of SlideHyb#1 hybridization solution (Ambion). The post-hybridization washing consisted of 3 cycles of 20 second incubations with each of the following solutions:  2X SSC plus 0.1% SDS (medium stringency) at 42°C; 0.1X SSC plus 0.05% SDS (high stringency) at 25°C; and 0.1X SSC (low stringency) at 25°C.  Lastly, the slides were dipped in 0.2X SSC and dried by centrifugation at 225 x g for 5 min at room temperature.

scan_protocol: Microarray slides were scanned with a GenePix 4000B scanner (Axon Instruments) and spot intensities were quantified using Imagene 6.0 (BioDiscovery, Inc.).

data_processing: Expression ratios were normalized using GeneSpring version 7.2 (Silicon Genetics) by the intensity-dependent Lowess method, with 20% of the data used for smoothing.  Replicate spots of the same type were combined to yield a single normalized ratio, which is output for the first ID_REF number representing that spot type.

organism: Rhodococcus jostii RHA1

source_name: late exponential phase, propane-grown Rhodococcus sp. RHA1

label_protocol: Cy-labelled cDNA was prepared by indirect labelling.  For one of the three hybridizations, the Cy3/Cy5 dyes were reversed (i.e., cDNA from the propane treatment was labelled with Cy5 rather than Cy3) to control for dye bias.

growth_protocol: Grown at 30°C and 150 rpm in basal salts medium amended with a 20% headspace volume addition of 99.5% purity propane (Matheson - Newark, CA) in sealed flasks.

extract_protocol: Cell pellets were vortexed with glass beads, hot phenol, and SDS at final concentrations of 14.3% and 0.9% (vol/vol) respectively.  Debris was precipitated with acetate followed by the addition of 4.0 mL phenol-chloroform (1:1, vol/vol).  Nucleic acids were precipitated with acetate plus isopropanol, treated with DNase, and purified with an RNeasy mini column (Qiagen).

source_name: late exponential phase, 23 mM pyruvate-grown Rhodococcus sp. RHA1

growth_protocol: Grown at 30°C and 150 rpm in basal salts medium amended with 23 mM pyruvate (Fisher Scientific -Fair Lawn, NJ).


title: Yeast-Gent-6

description: gentamicin-treated

source_name: Yeast, Gent-6


title: Yeast-Gent-7

description: gentamicin-treated

source_name: Yeast, Gent-7


title: C3H sighted - heart rep 1

source_name: C3H sighted heart

tag: C3H.pde6b-, male, heart, RNA pool including Mouse ID


title: Melanoma xenograft, MED1_Sub_C (LeiFidler)

description: human melanoma tumors grown  in mice


title: Yeast-control-2

description: control

source_name: Yeast, untreated-2


title: Melanoma xenograft, MED1_Lung_C (LeiFidler)

description: human melanoma tumors grown  in mice

source_name: Melanoma metastatic derivatives, lung


title: Control epithelial cells, immunopptd with 5-methylcytosine antibody, rep2 (dye-swap)

description: Total genomic DNA (Cy5) was co-hybridized with immunoprecipitated DNA (Cy3). The ratio between Cy5 and Cy3 signal was calculated for all high quality features.

source_name: Control cells_input

tag: Control cells, total input DNA

source_name: Control cells immunoprecipitated

tag: Control cells, anti-5-methylcytosine immunoprecipitated DNA


title: Melanoma xenograft, MEA3_Sub_C (LeiFidler)

description: human melanoma tumors grown  in mice


title: Control epithelial cells, immunopptd with 5-methylcytosine antibody, rep1


title: propane vs pyruvate, rep2

description: Rhodococcus sp. RHA1 grown on propane versus pyruvate, replicate 2

tag: late exponential phase, propane-grown Rhodococcus sp. RHA1

tag: late exponential phase, 23 mM pyruvate-grown Rhodococcus sp. RHA1


title: propane vs pyruvate, rep3

description: Rhodococcus sp. RHA1 grown on propane versus pyruvate, replicate 3


title: Melanoma xenograft, MED1_Sub_A (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MEA3_Sub_A (LeiFidler)

description: human melanoma tumors grown  in mice


title: Epithelial cells pre-exposed with 70 nM 17beta-estradiol, immunopptd with 5-methylcytosine antibody, rep2 (dye-swap)


title: Melanoma xenograft, MEC1_Lung_C (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MEC2_Sub_C (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MEC2_Sub_B (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MEC2_Lung_A (LeiFidler)

description: human melanoma tumors grown  in mice

source_name: Melanoma metastatic derivatives, Lung


title: Melanoma xenograft, MEC2_Lung_B (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MEC1_Lung_A (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MEA2_Sub_A (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MEC1_Sub_B (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MEA2_Lung_B (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MEC1_Sub_A (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, SM_Lung_A (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MEA2_Sub_B (LeiFidler)

description: human melanoma tumors grown  in mice

source_name: Melanoma metastatic derivatives, S.C.


title: Melanoma xenograft, MEA3_Lung_A (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MEA3_Lung_C (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, SM_Sub_B (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MEA3_Lung_B (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MA1_Lung_A (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MEA2_Lung_C (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MA2_Lung_A (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MC1_Lung_C (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MA2_Sub_B (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, SM_Lung_C (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, SM_Sub_A (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MA2_Sub_A (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MEA2_Lung_A (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MC2_Sub_C (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MC2_Sub_B (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MA1_Sub_A (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MA2_Sub_C (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MC1_Sub_C (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MED1_Lung_B (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MEC2_Sub_A (LeiFidler)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MC1_Sub_A (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MA2_Lung_C (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MC2_Sub_A (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MA1_Sub_B (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MEC1_Sub_C (LeiFidler)

description: human melanoma tumors grown  in mice


title: Tonsil tissue AGM_tonsil

status: Public on Oct 01 2007

city: Paris cedex 15

first: Beatrice

last: JACQUELIN

address: 25 rue du Docteur Roux

department: Virologie

institute: Institut Pasteur

description: Tonsil from 1 AGM.

hyb_protocol: 15µg of the labeled cRNA were hybridized to the ABI Human Whole Genome Arrays v2.0 Array using the Applied Biosystems Chemiluminescence Detection Kit accordingly to the protocol.

scan_protocol: Array processing, chemiluminescence detection, image acquisition, and analysis were performed using Applied Biosystems Chemiluminescence Detection Kit and Applied Biosystems 1700 Chemiluminescent Microarray Analyzer following manufacturer’s protocol.

data_processing: Images were auto-gridded and the chemiluminescent signals were quantified, corrected for background, and finally, spot- and spatially-normalized using the Applied Biosystems 1700 Chemiluminescent Microarray Analyzer software.

label: Chemiluminescence / DIG-UTP

organism: Chlorocebus sabaeus

source_name: Tonsil tissue

label_protocol: Digoxigenin-UTP labeled cRNA was generated and amplified from 0.8µg of total RNA using Applied Biosystems NanoAmp RT-IVT Labeling Kit according to the manufacturer’s protocol.

extract_protocol: The tissue was mechanically homogenized (X620 CAT M Zipperer GmbH, Germany) in RLT buffer prior to the extraction with the RNeasy® Mini Kit (Qiagen, France) following the manufacturer's instructions.

treatment_protocol: Tonsil tissues were collected on sacrified AGM from a former experiment (Gueye, A., et al. (2004) J Med Primatol 33, 83) and kept in OCT-buffer at -80°C until use.

tag: African Green Monkey cells (chlorocebus sabaeus from Senegal) isolated from  tonsil.


title: Melanoma xenograft, MC2_Lung_B (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MC2_Lung_A (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MC1_Lung_B (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MA1_Lung_C (LeiATCC)

description: human melanoma tumors grown  in mice


title: Unstimulated Circulating CD4+ T cells HUM_unstim_1

description: A pool of total RNA was prepared from enriched blood unstimulated CD4+ T lymphocytes isolated from 4 donors.

source_name: unstimulated circulating CD4+ T cells

label_protocol: Digoxigenin-UTP labeled cRNA was generated and amplified from 0.8µg of total RNA using Applied Biosystems Chemiluminescent RT-IVT Labeling Kit v2.0 according to the manufacturer’s protocol.

extract_protocol: Total RNA was extracted from 3 to 10 x 10^6 cells by using the RNeasy® Mini extraction Kit (Qiagen, Courtaboeuf, France) following the manufacturer's instructions.

treatment_protocol: PBMC were isolated by Ficoll-Hypaque (Pharmacia Biotech) density gradient centrifugation. The CD4+ cell subset was further isolated using the MACS magnetic labeling system (Miltenyi Biotec, Cologne, Germany) according to instructions of the manufacturer.

tag: Human CD4+ cells isolated from Peripheral Blood Mononuclear Cells (PBMC) consists of >95% of CD4+ T lymphocytes (CD3+ CD4+ cells).


title: Brain tissue AGM_brain

description: Brain from 1 AGM.

source_name: Brain tissue

treatment_protocol: Brain tissues were collected on sacrified AGM from a former experiment (Gueye, A., et al. (2004) J Med Primatol 33, 83) and kept in OCT-buffer at -80°C until use.

tag: African Green Monkey cells (chlorocebus sabaeus from Senegal) isolated from  brain.


title: Melanoma xenograft, MC1_Sub_B (LeiATCC)

description: human melanoma tumors grown  in mice


title: Melanoma xenograft, MA2_Lung_B (LeiATCC)

description: human melanoma tumors grown  in mice


title: Circulating CD4+ T cells stimulated with ConA AGM_stim_1_AB

description: A pool of total RNA was prepared from enriched blood CD4+ T lymphocytes isolated from 5 AGM. The cells were previously activated with ConA for 48H. A unique sample was obtained and then divided to generate 4 aliquots used to hybridize 2 technical replicates for each of the ABI and Affymetrix platforms.

hyb_protocol: 15µg of the labeled cRNA were hybridized to the ABI Human Whole Genome Arrays v1.0 Array using the Applied Biosystems Chemiluminescence Detection Kit accordingly to the protocol.

source_name: circulating CD4+ T cells stimulated with ConA

label_protocol: Digoxigenin-UTP labeled cRNA was generated and amplified from 0.8µg of total RNA using Applied Biosystems Chemiluminescent RT-IVT Labeling Kit v1.0 according to the manufacturer’s protocol.

growth_protocol: For activation, 20 × 10^6 cells were plated in RPMI-1640 with 5% fetal calf serum (FCS) with ConA. The cells were harvested after 48H. The CD4+ cell subset was further isolated using the MACS magnetic labeling system (Miltenyi Biotec, Cologne, Germany) according to instructions of the manufacturer.

treatment_protocol: PBMC were isolated by Ficoll-Hypaque (Pharmacia Biotech) density gradient centrifugation and activated with 10µg/ml of ConA.

tag: African Green Monkey cells (chlorocebus sabaeus from Senegal).

tag: Cells

value: CD4+ cells isolated from Peripheral Blood Mononuclear Cells (PBMC) consists of >95% of CD4+ T lymphocytes (CD3+ CD4+ cells).

tag: Stimulus

value: 48H with Concanavalin A (ConA)


title: Unstimulated Circulating CD4+ T cells AGM_unstim_2

description: A pool of total RNA was prepared from enriched blood unstimulated CD4+ T lymphocytes isolated from 4 AGM. A unique sample was obtained and then divided to generate 4 aliquots used to hybridized 4 ABI microarrays with 4 distinct RNA input quantities.


title: Circulating CD4+ T cells stimulated with ConA MAC_stim_2

description: A pool of total RNA was prepared from enriched blood CD4+ T lymphocytes isolated from 5 rhesus macaques. The cells were previously activated with ConA for 48H. A unique sample was obtained and then divided to generate 2 aliquots used to hybridize 2 technical replicates.

organism: Macaca mulatta

label_protocol: Digoxigenin-UTP labeled cRNA was generated and amplified from 1.2µg of total RNA using Applied Biosystems Chemiluminescent RT-IVT Labeling Kit v1.0 according to the manufacturer’s protocol.

tag: Rhesus macaque cells (M. mulatta from China).


title: Unstimulated Circulating CD4+ T cells AGM_unstim_4

label_protocol: Digoxigenin-UTP labeled cRNA was generated and amplified from 2µg of total RNA using Applied Biosystems Chemiluminescent RT-IVT Labeling Kit v2.0 according to the manufacturer’s protocol.


title: Human bronchial epithelial cells_passage 0 vs passage 3_2

status: Public on Aug 31 2007

city: San Francisco

first: Yuanyuan

last: Xiao

state: CA

address: 185 Berry Street, Lobby4, Suite 5700

department: Epidemiology and Biostatistics

institute: UCSF

hyb_protocol: Hybridization Protocol: Prior to hybridization, excess oligonucleotide was removed from the arrays by gently shaking them for 10 minutes in 0.2% SDS.  Arrays were then washed 5 times with distilled, filtered water and dried for 5 minutes at 600 rpm in a centrifuge.  A solution of 0.1% SDS, 5X SSC and 1% BSA was applied to the arrays under a cover slip (LifterSlip, Erie Scientific, Portsmouth, NH) for 1-2 hours at 42oC to block nonspecific binding.  Arrays were then washed 5 times with distilled, filtered water and dried for 5 minutes at 600 rpm in a centrifuge. The labeled aRNA targets were fragmented for 15 min at 70C with Ambion fragmentation buffer. (cat#8740). The fragmented, labeled aRNA targets were added to Ambion’s Hybridization buffer (cat #8861), denatured for 5 minutes at 95oC, and applied to the arrays under a cover slip (LifterSlip, Erie Scientific, Portsmouth, NH).  Arrays were placed in a SciGene Hybex hybridization chamber (SciGene, Sunnyvale, CA) at 55oC for 48 hours.  After hybridization, arrays were washed successively with 1X SSC at 55C for 3 minutes, and then 0.03% SDS, 0.2X SSC, and 0.05X SSC for 3 minutes at room temperature and dried for 5 minutes at 600 rpm in a centrifuge.

scan_protocol: Arrays were scanned using an Axon 4000B laser scanner (Molecular Devices Corp., Union City, CA). Image analyses were performed using the software package GenePix 6.1 (Molecular Devices Corp).

data_processing: Quality analysis of the hybridizations was performed using the Bioconductor software package arrayQuality (http://www.bioconductor.org and http://arrays.ucsf.edu/software ). Data were normalized using the print-tip loess method.

source_name: Human bronchial epithelial cells, passage 0

label_protocol: labeling, and array hybridizations were performed according to standard protocols from the UCSF Shared Microarray Core Facilities (Barczak et al, Gen Res. 2003; 13(7): 1775-1785 and http://arrays.ucsf.edu/spottedarrayarchives/protocols ). 10uf of total RNA was amplified with the Amino-Allyl MessageAmp II aRNA kit (Ambion, Austin, TX), according to the manufacturer’s protocol. The amplified RNAs were coupled to N-hydroxysuccinimidyl esters of Cy5 dye (CyScribe, Amersham Biosciences) for 1 h in the dark. The labeled aRNAs were purified and concentrated using Zymo RNA cleanup columns (Zymo, Orange, CA).

growth_protocol: Vitrogen (10 mg/ml coating concentration) coated 10cm tissue culture plates; BEGM (Clonetics); 7.5% CO2

extract_protocol: Total RNA was extracted using the Qiiagen RNeasy Kit, according to the manufacturer’s protocol (Qiagen Inc, Valencia, Ca).

tag: Normal adult non-smoker

source_name: Human bronchial epithelial cells, passage 3

label_protocol: Channel 1 labeling, and array hybridizations were performed according to standard protocols from the UCSF Shared Microarray Core Facilities (Barczak et al, Gen Res. 2003; 13(7): 1775-1785 and http://arrays.ucsf.edu/spottedarrayarchives/protocols ). 10uf of total RNA was amplified with the Amino-Allyl MessageAmp II aRNA kit (Ambion, Austin, TX), according to the manufacturer’s protocol. The amplified RNAs were coupled to N-hydroxysuccinimidyl esters of Cy3 dye (CyScribe, Amersham Biosciences) for 1 h in the dark. The labeled aRNAs were purified and concentrated using Zymo RNA cleanup columns (Zymo, Orange, CA).

growth_protocol: Vitrogen (10 mg/ml coating concentration) coated 10cm tissue culture plates; BEGM (Clonetics); 7.5% C02


title: Human bronchial epithelial cells_passage 3 vs. passage 0_1


title: upf1_control.3

city: Santa Cruz

first: Manny

last: Ares

address: 1125 High St

department: Molecular and Cellular Biology

institute: UCSC

description: Mouse N2A cells were transfected with control siRNA

hyb_protocol: Targets were hybridized to chips in 7% DMSO solution for 16 hrs overnight at 50. Microarrays were washed, processed with anti-biotin antibodies and streptavidin-phycoerythrin according to the standard Affymetrix protocol.

scan_protocol: Standard Affymetrix procedures

data_processing: Intensity values from the DNA arrays were normalized using a quantile normalization (Bolstad et al., 2003), and probe set summaries were derived using the Robust Multi-chip Analysis (RMA) procedure (Irizarry et al., 2003a; Irizarry et al., 2003b) with two modifications. The first modification was to remove all probes with 17 or more continuous bases that match to any other mouse transcript in order to minimize cross-hybridization issues. The second modification was to use the mode of the probe intensity values of similar GC content probes for the background estimate of a particular probe.

source_name: Neuroblastoma N2A cells

label_protocol: Total RNA was primed with random hexamers and reverse transcribed. After the reaction was completed, RNA was removed from the reaction by alkaline hydrolysis and the cDNA was purified using Qiagen PCR Quick Purification Kit. A typical reaction started with 5-6ug of total RNA usually yielded ~3ug of cDNA. The cDNA was then fragmented using DNAseI in an empirically controlled reaction that yields DNA fragments of 50-200 bases. This fragmented cDNA was then end labeled using terminal deoxynucleotidyl transferase and "DNA-Labeling-Reagent-1a (DLR-1a)", which is a biotinylated dideoxynucleoside triphosphate.

extract_protocol: Total RNA was extracted using Invitrogen Trizol reagent. RNA samples were quantified using NanoDrop ND-1000 Spectrophotometer.


title: Circulating CD4+ T cells stimulated with ConA MAC_stim_1


title: Unstimulated Circulating CD4+ T cells MAC_unstim_1

description: A pool of total RNA was prepared from enriched blood unstimulated CD4+ T lymphocytes isolated from 4 rhesus macaques.

label_protocol: Digoxigenin-UTP labeled cRNA was generated and amplified from 1.2µg of total RNA using Applied Biosystems Chemiluminescent RT-IVT Labeling Kit v2.0 according to the manufacturer’s protocol.


title: upf1.2

description: Mouse N2A cells were transfected with Upf1 siRNA in order to inhibit nonsense-mediated mRNA decay(NMD) pathway.


title: Unstimulated Circulating CD4+ T cells AGM_unstim_1

label_protocol: Digoxigenin-UTP labeled cRNA was generated and amplified from 0.5µg of total RNA using Applied Biosystems Chemiluminescent RT-IVT Labeling Kit v2.0 according to the manufacturer’s protocol.


title: Unstimulated Circulating CD4+ T cells AGM_unstim_3


title: Circulating CD4+ T cells stimulated with ConA AGM_stim_2_AB


title: emetine_control.1

description: Mouse N2A cells

data_processing: Intensity values from the DNA arrays were normalized using a quantile normalization (Bolstad et al., 2003), and probe set summaries were derived using the Robust Multi-chip Analysis (RMA) procedure (Irizarry et al., 2003a; Irizarry et al., 2003b) with two modifications. The first modification was to remove all probes with 17 or more continuous bases that match to any other mouse transcript in order to minimize cross-hybridization issues. The second modification was to use the mode of the probe intensity values of similar GC content probes for the background estimate of a particular probe. For example, if a probe has a GC count of 16, then the mode of the intensity of all the probes with a GC count of 16 was used as a background estimate as opposed to RMA in which the mode of all the probes is used as a background estimate for all the probes.

tag: Mouse N2A cells


title: Listeria_cold_LOG_37C vs 4C

city: Ithaca

first: Sarita

last: Raengpradub

address: 412 Stocking Hall

department: Food Science

institute: Cornell University

description: Expression profiles were measured from three independent RNA extractions, resulting in six replicate spots per gene (array contains duplicate spots).

hyb_protocol: Microarray slide blocking was performed prior to hybridization. Slides were incubated for 10 min in a solution containing methyl pyrrilidone, succinic anhydride and sodium borate, and then washed in fresh methyl pyrrilidone for 10 min. Successive 1-min washes in nanopure water (once), 0.1% SDS (3 times), nanopure water (5 times), and 95% EtOH (once). The slides were dried by centrifugation and immersed for 1 h in blocking solution (0.1% BSA, 5X SSC, 0.1% SDS) at 42°C. After blocking, the slides were washed for 1 min each in nanopure water (5 times) and 95% EtOH (2 times) and then dried by centrifugation. For the strains to be compared, labeled cDNAs were combined and dried, then resuspended in 50μl of hybridization buffer (5X SSC, 0.1% SDS, 0.1mM dithiothreitol (DTT), 0.5X formamide, 0.06 μg/μl salmon sperm DNA in DEPC-treated water), denatured at 95°C for 5 min then applied to the slides using mSeries LifterSlips™ (Erie Scientific). Following overnight hybridization at 42°C, the slides were washed in 2X SSC+0.1% SDS (5 min at 42˚C), with subsequent 5-min washes at room temperature in 2X SSC+0.1% SDS, 2X SSC, and 0.2X SSC, followed by final rinse in nanopure water. Slides were dried by centrifugation.

scan_protocol: Images were scanned using a GenePix 4000B scanner (Molecular Devices Corp.). Raw TIFF images were automatically gridded and analyzed using GenePix Pro 6.0 software.

data_processing: Analyses of microarray data were performed in R with Bioconductor using LIMMA software (Smyth, 2005). Background correction was performed using the “normexp” method. For normalization within arrays, the data were weighted for the genomic DNA controls and normalized using print-tip loess method (Smyth & Speed, 2003). The data were then scale normalized between arrays. The correlation between duplicate spots was calculated (Smyth et al., 2005) and a linear model was fitted to the normalized log ratios and empirical Bayes statistics were applied for differential expression analysis.

label: Alexa Fluor 635

organism: Listeria monocytogenes

source_name: Listeria monocytogenes 10403S

label_protocol: cDNA was synthesized from 10 μg purified total RNA (using random primers) and labeled with Alexa Fluor dyes using the SuperScript Plus Indirect cDNA Labeling System (Invitrogen).

growth_protocol: L. monocytogenes 10403S was streaked onto brain heart infusion (BHI) agar from -80°C glycerol stock cultures followed by incubation at 37°C for 24 h.  A single colony was subsequently inoculated into BHI broth and grown at 37°C overnight with shaking (250 rpm) and a 1 ml aliquot of this overnight culture was used to inoculate 99ml of BHI broth. After growth to mid-log phase (OD600 = 0.4), the L. monocytogenes suspension was used to inoculate 75 mL of pre-warmed (37°C) BHI broth (in a 300mL Erlenmeyer flask) to a starting OD600 of 0.15 ± 0.05.  Cells were then incubated at 37°C without shaking.  Bacterial cells were collected for RNA isolation at mid-log phase defined as OD600= 0.4 ± 0.05.

extract_protocol: RNA was isolated from frozen bacterial pellets with the Qiagen RNeasy Midi kit according to the manufacturer’s protocol for enzymatic and mechanical disruption, except that cells treated with lysozyme followed by sonication on ice for three 30 s bursts at 18–21 W (30 s pauses between bursts). RQ1 DNase treatment of total RNA in the presence of RNasin Plus RNA inhibitor (Promega) was performed according to the manufacturer’s protocol, followed by subsequent phenol:chloroform and chloroform extractions. RNA was ethanol-precipitated overnight, centrifuged and washed with 70% ethanol before resuspenson of the final RNA pellet in DEPC-treated water. Total nucleic acid concentration and purity were estimated using absorbance readings (260/280 nm, 260/230 nm) on a NanoDrop ND-1000 spectrophotometer. RNA integrity was checked with by agarose gel electrophoresis.

treatment_protocol: Prior to total RNA extraction, bacterial cultures were treated with RNAprotect™ Bacteria Reagent (Qiagen) according to the manufacturer’s protocol prior to RNA isolation to stabilize RNA.

tag: Listeria monocytogenes 10403S grown at 37°C to log phase

label: Alexa Fluor 532

growth_protocol: L. monocytogenes 10403S was streaked onto brain heart infusion (BHI) agar from -80°C glycerol stock cultures followed by incubation at 37°C for 24 h.  A single colony was subsequently inoculated into BHI broth and grown at 37°C overnight with shaking (250 rpm) and a 1 ml aliquot of this overnight culture was used to inoculate 99ml of BHI broth. After growth to mid-log phase (OD600 = 0.4), the L. monocytogenes suspension was used to inoculate 75 mL of pre-chilled (4°C) BHI broth (in a 300mL Erlenmeyer flask) to a starting OD600 of 0.15 ± 0.05.  Cells were then incubated at 4°C without shaking.  Bacterial cells were collected for RNA isolation at mid-log phase defined as OD600= 0.4 ± 0.05.

tag: Listeria monocytogenes 10403S grown at 4°C to log phase


title: emetine.1

description: Mouse N2A cells were treated with emetine in order to inhibit nonsense-mediated mRNA decay(NMD) pathway.

tag: Mouse N2A cells treated with 100(g/ml of emetine dihydrochloride hydrate (Fluka) 10 hours


title: upf1.3

tag: Mouse N2A cells were transfected with Upf1 siRNA


title: Listeria_cold_STATIONARY_37C vs 4C

growth_protocol: L. monocytogenes 10403S was streaked onto brain heart infusion (BHI) agar from -80°C glycerol stock cultures followed by incubation at 37°C for 24 h.  A single colony was subsequently inoculated into BHI broth and grown at 37°C overnight with shaking (250 rpm) and a 1 ml aliquot of this overnight culture was used to inoculate 99ml of BHI broth. After growth to mid-log phase (OD600 = 0.4), the L. monocytogenes suspension was used to inoculate 75 mL of pre-warmed (37°C) BHI broth (in a 300mL Erlenmeyer flask) to a starting OD600 of 0.15 ± 0.05.  Cells were then incubated at 37°C without shaking.  Bacterial cells were collected for RNA isolation at stationary phase defined as OD600= 1.0 ± 0.25 (4 h after inoculation for cells grown at 37°C).

tag: Listeria monocytogenes 10403S grown at 37°C to stationary phase

growth_protocol: L. monocytogenes 10403S was streaked onto brain heart infusion (BHI) agar from -80°C glycerol stock cultures followed by incubation at 37°C for 24 h.  A single colony was subsequently inoculated into BHI broth and grown at 37°C overnight with shaking (250 rpm) and a 1 ml aliquot of this overnight culture was used to inoculate 99ml of BHI broth. After growth to mid-log phase (OD600 = 0.4), the L. monocytogenes suspension was used to inoculate 75 mL of pre-chilled (4°C) BHI broth (in a 300mL Erlenmeyer flask) to a starting OD600 of 0.15 ± 0.05.  Cells were then incubated at 4°C without shaking.  Bacterial cells were collected for RNA isolation at stationary phase defined as OD600= 1.0 ± 0.25 (9 days after inoculation for cells grown at 4°C).

tag: Listeria monocytogenes 10403S grown at 4°C to stationary phase


title: upf1_control.1

tag: Mouse N2A cells were transfected with control siRNA


title: Human bronchial epithelial cells_passage 3 vs. passage 0_3

hyb_protocol: Prior to hybridization, excess oligonucleotide was removed from the arrays by gently shaking them for 10 minutes in 0.2% SDS.  Arrays were then washed 5 times with distilled, filtered water and dried for 5 minutes at 600 rpm in a centrifuge.  A solution of 0.1% SDS, 5X SSC and 1% BSA was applied to the arrays under a cover slip (LifterSlip, Erie Scientific, Portsmouth, NH) for 1-2 hours at 42oC to block nonspecific binding.  Arrays were then washed 5 times with distilled, filtered water and dried for 5 minutes at 600 rpm in a centrifuge. The labeled aRNA targets were fragmented for 15 min at 70C with Ambion fragmentation buffer. (cat#8740). The fragmented, labeled aRNA targets were added to Ambion’s Hybridization buffer (cat #8861), denatured for 5 minutes at 95oC, and applied to the arrays under a cover slip (LifterSlip, Erie Scientific, Portsmouth, NH).  Arrays were placed in a SciGene Hybex hybridization chamber (SciGene, Sunnyvale, CA) at 55oC for 48 hours.  After hybridization, arrays were washed successively with 1X SSC at 55C for 3 minutes, and then 0.03% SDS, 0.2X SSC, and 0.05X SSC for 3 minutes at room temperature and dried for 5 minutes at 600 rpm in a centrifuge.

extract_protocol: : Total RNA was extracted using the Qiiagen RNeasy Kit, according to the manufacturer’s protocol (Qiagen Inc, Valencia, Ca).

tag: Normal adult human non-smoker

label_protocol: labeling, and array hybridizations were performed according to standard protocols from the UCSF Shared Microarray Core Facilities (Barczak et al, Gen Res. 2003; 13(7): 1775-1785 and http://arrays.ucsf.edu/spottedarrayarchives/protocols ). 10uf of total RNA was amplified with the Amino-Allyl MessageAmp II aRNA kit (Ambion, Austin, TX), according to the manufacturer’s protocol. The amplified RNAs were coupled to N-hydroxysuccinimidyl esters of Cy3 dye (CyScribe, Amersham Biosciences) for 1 h in the dark. The labeled aRNAs were purified and concentrated using Zymo RNA cleanup columns (Zymo, Orange, CA).


title: upf1_control.2