title: IR29 root, control, biological replicate 1

description: IR29 control

source_name: root, IR29, control


title: FL478 root, salt-treated, biological replicate 3

status: Public on Jul 16 2009

type: RNA

city: Glen Osmond

first: Ute

last: Baumann

state: SA

address: Hartley Grove

department: Australian Centre for Plant Functional Genomics

country: Australia

institute: University of Adelaide

description: FL478 salt-treated

hyb_protocol: Standard Affymetrix procedures.

scan_protocol: Standard Affymetrix procedures.

data_processing: The data were normalized using the Robust Multichip Average method of background correction, quantile normalisation, and summarisation of cell signal intensities (Irrizary et al., 2003).

label: biotin

molecule: total RNA

organism: Oryza sativa Indica Group

source_name: root, FL478, salt-treated

label_protocol: Approx. 8 µg of total RNA was processed to produce biotinylated cRNA targets.

growth_protocol: Plants were grown in sand-filled tanks and irrigated with nutrient solution (for details see Walia et al (2005), Plant Physiol. 139:822-35) for 30 days.

extract_protocol: RNA was extracted using TRIzol and subsequently purified using an RNeasy spin column (Qiagen) and an on-column DNase treatment.

treatment_protocol: On day 22 after germination, salt treatment commenced by adding NaCl and CaCl2 (5:1 molar concentration) in two steps over a period of 3 days to the nutrient solution reservoir. Salinity in the treatment reservoir reached 7.4 dS/m. Plants were harvested on day 30.

tag: salt-tolerant


title: FL478 root, salt-treated, biological replicate 2

description: FL478 salt-treated


title: FL478 root, control, biological replicate 2

description: FL478 control

source_name: root, FL478, control


title: IR29 root, control, biological replicate 2

description: IR29 control

tag: salt-intolerant


title: Dog Biceps femoris Unexercised Control Pool (n=4) vs. Biceps femoris; 1450 min (24h Post Exercise); Dog #05

status: Public on Jul 01 2009

city: Rahway

first: Olivia

last: Fong

state: NJ

department: Molecular Profiling

country: USA

institute: Merck & Co.

description: Dog muscle

hyb_protocol: Microarrays are incubated at 40°C for 48 hours in a rotating carousel.  Hybridizations to custom Agilent microarrays are completed as previously described (Hughes et al.Nat Biotech (2001), 19(4):342-7).  Microarrays are washed to remove non-specific hybridized sample. Afterwards, microarrays are dried in an ozone-free nitrogen chamber.

scan_protocol: Microarrays are scanned using the Agilent LP2 laser scanner. The scanner output is a Tiff file, which contains the quantitative hybridization data from each individual microarray. The Tiff files are then processed using Rosetta custom feature extraction software.

data_processing: Data were processed using the Rosetta Resolver® system. Rosetta's custom feature extraction software performs error modeling before data are loaded into the Resolver system. The Resolver system performs a squeeze operation that combines replicates of the same sequence in an array while applying error weighting. The error weighting consists of adjusting for additive and multiplicative noise. A P-value is generated and propagated throughout the system. The P-value represents the probability that a gene is expressed. The Resolver system allows users to set thresholds, below which genes of a P-value are considered to be significantly expressed. The Resolver system also combines multiple arrays using a squeezing process. If multiple spots reference one sequence, summarization is performed using an error-weighted average as described in Roland Stoughton and Hongyue Dai, Statistical Combining of Cell Expression Profiles. US Patent #6,351,712, February 26, 2002.

organism: Canis lupus familiaris

label_protocol: custom automated version of the aminoallyl MessageAmp II kit from Ambion.

extract_protocol: Qiagen RNeasy spin columns with DNAse treatment

tag: tissue

value: Biceps femoris

tag: activity

value: unexercised

value: exercised, 24 hrs


title: FL478 root, control, biological replicate 1

description: FL478 control


title: Dog Biceps femoris Unexercised Control Pool (n=4) vs. Biceps femoris; 0735 min (12h Post Exercise); Dog #07

description: Dog muscle

value: exercised, 12 hrs


title: FL478 root, salt-treated, biological replicate 1

description: FL478 salt-treated


title: Dog Biceps femoris Unexercised Control Pool (n=4) vs. Biceps femoris; 1495 min (24h Post Exercise); Dog #13

description: Dog muscle

value: Biceps femoris,


title: Dog Biceps femoris Unexercised Control Pool (n=4) vs. Biceps femoris; 0095 min (02h Post Exercise); Dog #11

description: Dog muscle

value: exercised, 2 hrs


title: Dog Biceps femoris Unexercised Control Pool (n=4) vs. Biceps femoris; 1476 min (24h Post Exercise); Dog #14

description: Dog muscle


title: FL478 root, control, biological replicate 3

description: FL478 control


title: Dog Biceps femoris Unexercised Control Pool (n=4) vs. Biceps femoris; 0762 min (12h Post Exercise); Dog #06

description: Dog muscle


title: Dog Biceps femoris Unexercised Control Pool (n=4) vs. Biceps femoris; 0415 min (06h Post Exercise); Dog #10

description: Dog muscle

value: exercised, 6 hrs


title: Gene expression in the pancreatic lymph node of Deaf1 knock out mice vs. BALB/c controls-sample 1

status: Public on Jul 15 2009

city: Stanford

first: Linda

last: Yip

state: CA

address: 269 Campus Drive West, CCSR Building Rm. 2240

department: Medicine

institute: Stanford University

description: no additional description.

hyb_protocol: two color microarrays were performed using the whole mouse genome (4x44K) Oligo microarray kit.  Hybridization was performed using the Agilent Gene Expression Hybridization Kit and the Agilent Microarray Hybridization Chamber as instructed by the manufacturer. Samples were hybridized for 17 h at 65C, and slides were washed with Gene Expression Wash Buffer (Agilent) according to maufacturer's instructions.

scan_protocol: Microarray chips were scanned using the DNA microarray scanner (Agilent Technologies) witht he following parameters:  2 color scan; scan resolution 5 um, single pass, red and green dye channels.  Data was extracted using Feature Extraction software.

data_processing: Data was process using GeneSpring GX 7.3 Software.  Data was imported using the enhanced Agilent FE Import setting. Spot information in Feature Extraction was used to flag data, and normalization was performed using "per spot and per chip: intensity dependent (Lowess) normalization".

organism: Mus musculus

source_name: pancreatic lymph node of BALB/c control mice

label_protocol: RNA was labeled using the Agilent low RNA input fluorescence linear amplification kit (Agilent Technologies) according to manufacturer's instructions.

growth_protocol: mice were maintained under pathogen-free conditions according to institutional guidelines under approved protocols in the Stanford Medical Center’s Department of Comparative Medicine (Stanford, CA)

extract_protocol: pancreatic lymph nodes were removed and immediately homogenized in Trizol Reagent.  RNA was extracted in Trizol and then purified using the RNeasy kit (Qiagen). RNA quality was assessed using the Agilent RNA 6000 Nano Reagents, RNA Nano chips, and the Agilent 2100 bioanalyzer (Agilent Technologies), according to manufacturer’s instructions.  RNA was amplified, labeled with Cy3 and combined with spike A  using the Agilent low RNA input fluorescence linear amplification kit (Agilent Technologies).  The amplified cRNA was purified using the RNeasy kit (Qiagen), and specific activity was determined with the NanoDrop 1000 spectrophotometer (Thermo Scientific).

treatment_protocol: no treatment

tag: strain

value: BALB/c

tag: gender

value: Female

tag: age

value: 4 wk

value: Pancreatic lymph node

tag: Pool of tissue from 4 individuals

source_name: Pancreatic lymph node of Deaf1 Knock out mice

extract_protocol: pancreatic lymph nodes were removed and immediately homogenized in Trizol Reagent.  RNA was extracted in Trizol and then purified using the RNeasy kit (Qiagen). RNA quality was assessed using the Agilent RNA 6000 Nano Reagents, RNA Nano chips, and the Agilent 2100 bioanalyzer (Agilent Technologies), according to manufacturer’s instructions.  RNA was amplified, labeled with Cy5 and combined with spike B  using the Agilent low RNA input fluorescence linear amplification kit (Agilent Technologies).  The amplified cRNA was purified using the RNeasy kit (Qiagen), and specific activity was determined with the NanoDrop 1000 spectrophotometer (Thermo Scientific).

value: Deaf1 Knockout mice on BALB/c background

tag: Tissue from Deaf1-Knockout mouse A


title: Dog Biceps femoris Unexercised Control Pool (n=4) vs. Biceps femoris; 0070 min (02h Post Exercise); Dog #08

description: Dog muscle


title: Gene expression in the pancreatic lymph node of Deaf1 knock out mice vs. BALB/c controls-sample 2

description: no additional description.

tag: Tissue from Deaf1-Knockout mouse B


title: Dog Biceps femoris Unexercised Control Pool (n=4) vs. Biceps femoris; 0395 min (06h Post Exercise); Dog #09

description: Dog muscle


title: Dog Biceps femoris Unexercised Control Pool (n=4) vs. Biceps femoris; 0000 min (Not Exercised); Dog #03

description: Dog muscle


title: Dog Biceps femoris Unexercised Control Pool (n=4) vs. Biceps femoris; 0000 min (Not Exercised); Dog #04

description: Dog muscle


title: Dog Biceps femoris Unexercised Control Pool (n=4) vs. Biceps femoris; 0000 min (Not Exercised); Dog #02

description: Dog muscle


title: Dog Biceps femoris Unexercised Control Pool (n=4) vs. Biceps femoris; 0000 min (Not Exercised); Dog #01

description: Dog muscle


title: PLC_24hrs_replica3 (PLC3_24hrsCy5)

city: Denver

first: Ivana

middle: V

last: Yang

state: CO

address: 1400 Jackson Street

institute: National Jewish Health

hyb_protocol: Agilent hyb reagents and protocol.

scan_protocol: Agilent scanner - default settings.

data_processing: Feature Extraction Version A.7.5.1

source_name: Universal mouse reference RNA

label_protocol: Aminoallyl labeling (TIGR protocol).

growth_protocol: RAW 264.7 cells were maintained in RPMI 1640 supplemented with 10% FBS and 200 μg/ml gentamicin (Invitrogen Life Technologies).  For stimulation of the RAW 264.7 cell, cells were plated in 6-well culture plates (3 x 10^6/well) overnight. The overnight growth was performed to allow resting before stimulation.

extract_protocol: Cells were harvested 6 or 24 hours post-treatment and RNA was extracted using Trizol and following manufacturers protocols (Invitrogen Life Technologies)

treatment_protocol: Cells were washed twice with Opti-MEM (Invitrogen Life Technologies) and stimulated with 0.5 μg/ml Escherichia coli 0111:B4 Ultrapure LPS (List Biological Laboratories), 0.25 μg/ml poly(I:C) (InvivoGen) or 1.5 μM CpG ODN 1826 (Midland Certified Reagents).

tag: sample

value: Stratagene cat number 740100

tag: cell line

value: RAW264.7 macrophages

tag: treatment

value: poly(I:C) (Invivogen)

tag: amount

tag: time

value: 24 hrs


title: PLC_6hrs_replica2 (PLC2_6hrsCy5)

value: 6 hrs


title: PLC_6hrs_replica2 (PLC2_6hrsCy3)


title: PLC_6hrs_replica3 (PLC3_6hrsCy3)


title: media_24hrs_replica5 (M5_24hrsCy3)

value: media control


title: media_6hrs_replica3 (M3_6hrsCy5)


title: media_24hrs_replica6 (M6_24hrsCy5)


title: PLC_6hrs_replica1 (PLC1_6hrsCy5)


title: PLC_24hrs_replica3 (PLC3_24hrsCy3)


title: media_24hrs_replica4 (M4_24hrsCy5)


title: media_24hrs_replica3 (M3_24hrsCy5)


title: PLC_24hrs_replica1 (PLC1_24hrsCy5)


title: media_24hrs_replica6 (M6_24hrsCy3)


title: PLC_6hrs_replica1 (PLC1_6hrsCy3)


title: PLC_6hrs_replica3 (PLC3_6hrsCy5)


title: PLC_24hrs_replica2 (PLC2_24hrsCy5)


title: media_6hrs_replica3 (M3_6hrsCy3)


title: PLC_24hrs_replica1 (PLC1_24hrsCy3)


title: media_24hrs_replica3 (M3_24hrsCy3)


title: media_24hrs_replica4 (M4_24hrsCy3)


title: media_6hrs_replica2 (M2_6hrsCy3)


title: PLC_24hrs_replica2 (PLC2_24hrsCy3)


title: media_6hrs_replica2 (M2_6hrsCy5)


title: media_24hrs_replica5 (M5_24hrsCy5)


title: media_24hrs_replica2 (M2_24hrsCy3)


title: media_6hrs_replica1 (M1_6hrsCy5)

value: media contro

value: time:6 hrs


title: LPS_6hrs_replica3 (LPS3_6hrsCy3)

value: Ultrapure LPS (LIST)

value: 0.5 ug/mL


title: media_6hrs_replica1 (M1_6hrsCy3)


title: media_24hrs_replica1 (M1_24hrsCy3)


title: LPS_24hrs_replica3 (LPS3_24hrsCy5)


title: media_24hrs_replica1 (M1_24hrsCy5)


title: media_24hrs_replica2 (M2_24hrsCy5)


title: LPS_6hrs_replica3 (LPS3_6hrsCy5)


title: LPS_6hrs_replica2 (LPS2_6hrsCy5)


title: LPS_6hrs_replica2 (LPS2_6hrsCy3)


title: LPS_24hrs_replica1 (LPS1_24hrsCy3)


title: CpG_6hrs_replica3 (CpG3_6hrsCy3)

value: CpG ODN 1826 (Midland)


title: LPS_24hrs_replica3 (LPS3_24hrsCy3)


title: CpG_24hrs_replica3 (CpG3_24hrsCy5)


title: LPS_6hrs_replica1 (LPS1_6hrsCy5)


title: CpG_24hrs_replica3 (CpG3_24hrsCy3)


title: CpG_6hrs_replica3 (CpG3_6hrsCy5)


title: LPS_24hrs_replica2 (LPS2_24hrsCy5)


title: LPS_24hrs_replica1 (LPS1_24hrsCy5)


title: LPS_6hrs_replica1 (LPS1_6hrsCy3)


title: LPS_24hrs_replica2 (LPS2_24hrsCy3)


title: CpG_6hrs_replica2 (CpG2_6hrsCy3)


title: CpG_6hrs_replica1 (CpG1_6hrsCy3)


title: CpG_24hrs_replica2 (CpG2_24hrsCy5)


title: CpG_6hrs_replica2 (CpG2_6hrsCy5)


title: CpG_24hrs_replica2 (CpG2_24hrsCy3)


title: CpG_24hrs_replica1 (CpG1_24hrsCy3)


title: CpG_6hrs_replica1 (CpG1_6hrsCy5)


title: 016699

status: Public on Jun 01 2009

city: Valbonne - Sophia-Antipolis

first: Kevin

last: Lebrigand

address: 660 route des lucioles

country: France

institute: IPMC/CNRS

description:
K562 CT exp. 1 vs PDR CT exp. 2
Date:2006/08/08

hyb_protocol: Agilent : 750.0 ng at 62 degree_C during 17 hours

scan_protocol: Scanned on an GenePix 4000B fluorescent scanner. Image intensity data were extracted with GenePix Pro 6.0 analysis software.

data_processing: Interslide norm. : quantile, Intraslide norm. : printtiploess, Background norm. : none, Software used : limmagui --> VALUE data obtained from log of processed Red signal/processed Green signal.

organism: Homo sapiens

source_name: K562 CT exp. 1

label_protocol: Ambion

extract_protocol: Qiagen

tag: Labeling_reference:K562 CT exp. 1

value: K562 CT exp. 1

tag: Tissue

value: Cell line

source_name: PDR CT exp. 2

tag: Labeling_reference:PDR CT exp. 2

value: PDR CT exp. 2


title: 016612

description:
PDR CT exp. 1 vs ImaR CT exp. 2
Date:2006/08/08

source_name: PDR CT exp. 1

tag: Labeling_reference:PDR CT exp. 1

value: PDR CT exp. 1

source_name: ImaR CT exp. 2

tag: Labeling_reference:ImaR CT exp. 2

value: ImaR CT exp. 2


title: treatment 8, rep4

city: Richland

first: Katrina

middle: M

last: Waters

state: WA

address: 902 Battelle Blvd; MSIN J4-18

department: Biological Sciences Division

institute: Pacific Northwest National Laboratory

description: Farmed soil

hyb_protocol: Bioprimed cDNA was hybridized to gene specific probes printed on epoxysilane coated 6-well microarray slides.  The signal was amplified using an optimized TSA kit (Perkin Elmer, Waltham, MA) protocol.   First, slides were pre-hybridized with TNB blocking buffer for 30 min at 22°C.  The target cDNA was mixed with hybridization buffer (2X SSC, 0.02% Tween 20, and 1 ul per well of 1 ng µl-1 salmon sperm DNA) and denatured at 95°C for 2 min.  After removal of TNB blocking buffer, the denatured cDNA was added to the microarray slide.  Lip01 and mnp02 control DNA was added at a concentration of 100 ng/well, P. chrysosporium was added at 250 ng/well, and soil DNA was added at a concentration of 1000 ng/well in a 75 µl per well volume. Hybridizations were incubated for 3 hours in a humidified chamber at 21°C (plus or minus 1 degree), and then washed with TNT buffer for 2 min with agitation.  The biotin in the biotin labeled target cDNA was amplified by incubating microarray slides with 1:100 SA-HRP (Strepavidin-horse radish peroxidase) in TNB for 30 min at 22°C. The SA-HRP was removed by washing slides in TNT buffer.  Background biotin binding was reduced by incubating slides with 10% equine serum in 2X SSC for 30 min at 22°C.  Slides were washed once more with TNT buffer and incubated with 1:50 Biotinyl Tyramide (amplification reagent in amplification diluent) for 10 min at 22°C and washed in TNT buffer.  A SA-Alexa 546 (Molecular Probes, Invitrogen Corporation, Carlsbad, CA) fluorescent indicator (1:500 SA-Alexa in 1x SSC, 5X Denhardt’s solution) was added to each array to bind to the amplified biotin labeled target cDNA and incubated in the dark for 1 hour, at 22°C.  Subsequent to fluorescent binding, slides were washed in TNT buffer, quickly dipped in 0.6X SSC, and dried with compressed nitrogen.

scan_protocol: ScanArray, Perkin Elmer, Waltham, MA

data_processing: The microarray spot intensities were extracted with an automated microarray image analysis toolbox for MatLAB (White, A. M., D. S. Daly, A. R. Willse, M. Protic, and D. P. Chandler. 2005. Automated Microarray Image Analysis Toolbox for MATLAB. Bioinformatics 21:3578-9.) Spot intensities were background corrected and log transformed prior to normalization and statistical analysis.

organism: unidentified

source_name: Farmed soil

label_protocol: Superscript II (Invitrogen Corporation, Carlsbad, CA) was used to generate first strand cDNA products from the soil RNA extract.  Following cDNA generation, products were labeled with biotin (Bioprime, Invitrogen, Carlsbad, CA).  In the first bioprime reaction, the entire cDNA reaction and 20µL of 2.5X Random Prime solution was mixed, denatured for 5 min at 95°C and snap cooled on ice.  While the denatured cDNA mixture was on ice 1 µL of non-biotinylated 10 mM dNTP’s, 7 µL of water and 2 µL Klenow were mixed and incubated at 37°C for 4 hours.   A second bioprime reaction was created by using a 10 µL aliquot of the first bioprime reaction as template.  To the template 10 µL water, and 20 µL of 2.5X Random Prime solution were added, mixed, and denatured at 95°C for 5 min.  Products were snap cooled and 5 µL 10x dNTP’s, 3 µL water, and 2 µL of Klenow were added, mixed, and incubated at 37°C for 4 hours.  After incubation 5 µL stop buffer was added to stop polymerase function and the reaction was purified using a min-elute PCR purification kit (Qiagen, Inc., Valencia, CA).

growth_protocol: Prior to RNA extraction, soils (50 g) were conditioned at room temperature and 0.03 MPa water content.

extract_protocol: RNA was extracted from soil using a mechanically-assisted (bead beating) method.  Briefly, 0.5 g moist soil, 0.25 g 0.1-mm zirconia/silica beads, 0.3 mL 50 mM sodium acetate + 10 mM EDTA (pH 5.1), 0.3 mL 10% sodium dodecylsulphate, and 0.6 mL phenol:chloroform:isoamyl alcohol (25: 24:1) were added to a screw top tube.  Cell lysis was accomplished by bead beating (Biospec products) for 2 min at homogenize speed, heating to 60ºC for 10 min, followed by an additional two min bead beating treatment.  The RNA extract was centrifuged for 10 minutes at 15000 x g to collect the aqueous layer.  RNA was purified from the aqueous phase by isopropanol precipitation, then resuspended in 50 µL nuclease free H2O and quantified on a Nanodrop spectrophotometer (Nanodrop Technologies, Wilmington, DE.)

treatment_protocol: After approximately 4 days of conditioning, ground wheat straw was added to the soils and the soils incubated another 7 days.

tag: Farmed soil


title: 016552

description:
ImaR CT exp. 1 vs K562 CT exp. 2
Date:2006/08/08

source_name: ImaR CT exp. 1

tag: Labeling_reference:ImaR CT exp. 1

value: ImaR CT exp. 1

source_name: K562 CT exp. 2

tag: Labeling_reference:K562 CT exp. 2

value: K562 CT exp. 2


title: treatment 8, rep3

description: Farmed soil


title: 016641

description:
PDR CT exp. 1 vs K562 CT exp. 2
Date:2006/08/08


title: Thyroid_IFNg_transgenic_CD45depeleted

type: SAGE

anchor: NlaIII

city: Baltimore

first: Patrizio

last: Caturegli

state: MD

address: 720 Rutland Avenue

department: Pathology

institute: Johns Hopkins University

molecule: polyA RNA

source_name: thyroid-specific IFNg transgenic mouse thyrocytes isolated by enzymatic digestion followed by CD45 magnetic beads separation to deplete hematopoietic cells

extract_protocol:
Velculescu, V.E. et al. (1995) Science 270: 484-487
Saha, S. et al. (2002) Nature Biotechnology 19: 508-512

tag: strain

tag: gender

value: female

tag: transgene

value: thyroid-specific IFNg transgenic mice

tag: age

value: 60 days

tag: tissue

value: thyroid

tag: preparation

value: thyroid lobes were digested to make single suspension. CD45 positive cells (hematopoietic cells) were depleted by CD45 magnetic beads separation.


title: 016611

description:
ImaR CT exp. 1 vs PDR CT exp. 2
Date:2006/08/08


title: 016551

description:
K562 CT exp. 1 vs ImaR CT exp. 2
Date:2006/08/08


title: Thyroid_wildtype_CD45depleted

source_name: wildtype BL6 mouse thyrocytes isolated by enzymatic digestion followed by CD45 magnetic beads separation to deplete hematopoietic cells

value: wild type controls for thyroid-specific IFNg transgenic mice


title: CpG_24hrs_replica1 (CpG1_24hrsCy5)

tag: sample

value: Stratagene cat number 740100

tag: cell line

value: RAW264.7 macrophages

tag: treatment

value: 24 hrs


title: treatment 7, rep8

description: Prairie soil inoculated with P. chrysosporium prior to RNA extraction


title: treatment 7, rep7

source_name: Prairie soil inoculated with P. chrysosporium prior to RNA extraction


title: treatment 7, rep4

tag: Prairie soil inoculated with P. chrysosporium prior to RNA extraction


title: treatment 8, rep1

description: Farmed soil


title: treatment 7, rep5


title: treatment 7, rep3


title: treatment 7, rep6


title: treatment 7, rep2


title: treatment 6, rep5

description: P. chrysosporium cDNA mixed with prairie soil cDNA, then biotin-labeled


title: treatment 6, rep6

source_name: P. chrysosporium cDNA mixed with prairie soil cDNA, then biotin-labeled


title: treatment 6, rep7

tag: P. chrysosporium cDNA mixed with prairie soil cDNA, then biotin-labeled


title: treatment 6, rep2


title: treatment 7, rep1


title: treatment 6, rep4


title: treatment 6, rep3


title: treatment 5, rep12

description: Biotin-labeled P. chrysosporium cDNA combined with labeled prairie soil cDNA


title: treatment 5, rep11

source_name: Biotin-labeled P. chrysosporium cDNA combined with labeled prairie soil cDNA


title: treatment 6, rep1


title: treatment 5, rep10

tag: Biotin-labeled P. chrysosporium cDNA combined with labeled prairie soil cDNA


title: treatment 5, rep9


title: treatment 5, rep7


title: treatment 5, rep8


title: treatment 5, rep4


title: treatment 5, rep5


title: treatment 8, rep2

description: Farmed soil


title: treatment 5, rep3


title: treatment 5, rep1


title: treatment 5, rep2


title: treatment 4, rep7

type: other

description: Biotin-labeled lip01 and mnp02 amplicons combined with labeled prairie soil cDNA

molecule: other

organism:
synthetic construct
unidentified


title: treatment 4, rep8

source_name: Biotin-labeled lip01 and mnp02 amplicons combined with labeled prairie soil cDNA


title: treatment 4, rep6

tag: Biotin-labeled lip01 and mnp02 amplicons combined with labeled prairie soil cDNA


title: treatment 5, rep6


title: treatment 4, rep4


title: treatment 4, rep2


title: treatment 4, rep1


title: treatment 4, rep5


title: treatment 3, rep6

description: Tallgrass prairie soil

source_name: Tallgrass prairie soil

tag: Tallgrass prairie soil


title: treatment 4, rep3


title: treatment 3, rep5

description: Tallgrass prairie soil


title: Case 3

type: genomic

first: Jonathan

last: Pevsner

address: 707 N. Broadway

department: Neurology

institute: Kennedy Krieger Institute

description: SNP genotyping was performed at the Johns Hopkins University Genetic Resources Core Facility SNP Center.

hyb_protocol: Hybridization was performed at the Johns Hopkins University Genetic Resources Core Facility SNP Center.

scan_protocol: Scans were performed at the Johns Hopkins University Genetic Resources Core Facility SNP Center.

data_processing: Data processing was performed using BeadStudio software (Illumina)(v3.0).

label: SNPs were detected using a two-color single base extension reaction.

molecule: genomic DNA

source_name: transformed lymphoblasts

label_protocol: SNPs were analyzed at the Johns Hopkins University Genetic Resources Core Facility SNP Center.

extract_protocol: Genomic DNA was isolated using an extraction kit.

value: male


title: treatment 3, rep3

description: Tallgrass prairie soil


title: treatment 3, rep4

description: Tallgrass prairie soil


title: treatment 3, rep2

description: Tallgrass prairie soil


title: treatment 2, rep8

description: P. chrysosporium

organism: Phanerodontia chrysosporium

source_name: P. chrysosporium

tag: P. chrysosporium


title: Case 1

source_name: transformed lymphoblasts from Case 1

tag: Gender


title: treatment 3, rep8

description: Tallgrass prairie soil


title: treatment 3, rep1

description: Tallgrass prairie soil


title: treatment 2, rep7

description: P. chrysosporium


title: treatment 2, rep5

description: P. chrysosporium


title: treatment 3, rep7

description: Tallgrass prairie soil


title: treatment 2, rep4

description: P. chrysosporium


title: treatment 2, rep2

description: P. chrysosporium


title: Father of Case 4


title: treatment 2, rep1

description: P. chrysosporium


title: treatment 2, rep3

description: P. chrysosporium


title: treatment 1, rep7

description: lip01 and mnp02 amplicons

organism: synthetic construct

source_name: lip01 and mnp02 amplicons

tag: lip01 and mnp02 amplicons


title: treatment 2, rep6

description: P. chrysosporium


title: Mother of Case 1


title: treatment 1, rep6

description: lip01 and mnp02 amplicons


title: Father of Case 1


title: Case 4


title: treatment 1, rep5

description: lip01 and mnp02 amplicons


title: treatment 1, rep4

description: lip01 and mnp02 amplicons


title: treatment 1, rep2

description: lip01 and mnp02 amplicons


title: treatment 1, rep3

description: lip01 and mnp02 amplicons


title: buffer only, rep6

description: buffer only

source_name: buffer only

tag: buffer only


title: treatment 1, rep1

description: lip01 and mnp02 amplicons


title: Mother of Case 4

tag: gender

value: mother


title: buffer only, rep5

description: buffer only


title: buffer only, rep4

description: buffer only


title: buffer only, rep3

description: buffer only


title: buffer only, rep1

description: buffer only


title: buffer only, rep2

description: buffer only


title: Case 2


title: HSF1.NSC

city: Piscataway

first: Ronald

middle: P.

last: Hart

address: 604 Allison Rd Rm B430

department: Cell Biology & Neuroscience

institute: Rutgers University

description: NSC stage from HSF1 cultures.

hyb_protocol: Standard Illumina hybridization protocol.

scan_protocol: Standard Illumina scanning protocol.

data_processing: The data were normalised using quantile normalisation with BeadStudio.

source_name: HSF1, NSC

label_protocol: Biotinylated cRNA were prepared with the Ambion MessageAmp kit for Illumina arrays.

extract_protocol: RNA was extracted with Trizol reagent and alcohol precipitated. Quality control was performed with Agilent Bioanalyser.

tag: stage

value: NSC


title: H1.NSC

description: NSC stage from H1 cultures.

source_name: H1, NSC


title: HSF6.ESC

description: ESC stage from HSF6 cultures.

source_name: HSF6, ESC

value: ESC


title: HSF1.ESC

description: ESC stage from HSF1 cultures.

source_name: HSF1, ESC


title: H1.ESC

description: ESC stage from H1 cultures.

source_name: H1, ESC


title: HSF6.NSC

description: NSC stage from HSF6 cultures.

source_name: HSF6, NSC


title: LLC-Moil-24h-rep1, quantile

city: Athens

first: Fragiskos

middle: N

last: Kolisis

address: 48 Vasileos Constantinou ave.

department: Institute of Biological Research and Biotechnology

country: Greece

institute: National Hellenic Research Foundation

description:
Replicate 1 of 2.
Mastic oil 24h - 1

data_processing: Data were normalized with quantile normalization, and statistical analysis was perfomed using ANOVA and clustering analysis with k-means clustering. Prior to quantile normalization, probesets absent to all conditions and replicates were removed.

source_name: Lewis lung carcinoma cells, Mastic Oil, 24h treatment

label_protocol: Biotinylated cRNA was prepared with Illumina TotalPrep RNA Amplification Kit.

growth_protocol: Cells were cultured in DMEM supplemented with 10% FCS L-glutamine and antibiotics (Gibco, Grand Island, NY).

extract_protocol: Total RNA was isolated with the Trizol Plus kit (Invitrogen) according to manufacturer's instructions.

treatment_protocol: Cells were treated with 0.01% v/v mastic oil or 0.1% DMSO vehicle for the indicated periods of time (3 to 48h).

value: Lung adenocarcinoma (LLC)

tag: treatment time

tag: strain


title: LLC-DMSO-48h-rep2, quantile

description:
Replicate 2 of 2.
Vehicle 48h - 2

source_name: Lewis lung carcinoma cells, DMSO, 48h treatment


title: LLC-Moil-12h-rep2, quantile

description:
Replicate 2 of 2.
Mastic oil 12h - 2

source_name: Lewis lung carcinoma cells, Mastic Oil, 12h treatment


title: LLC-Moil-24h-rep2, quantile

description:
Replicate 2 of 2.
Mastic oil 24h - 2


title: LLC-DMSO-48h-rep1, quantile

description:
Replicate 1 of 2.
Vehicle 48h - 1


title: LLC-Moil-48h-rep2, quantile

description:
Replicate 2 of 2.
Mastic oil 48h - 2

source_name: Lewis lung carcinoma cells, Mastic Oil, 48h treatment


title: LLC-Moil-3h-rep1, quantile

description:
Replicate 1 of 2.
Mastic oil 3h - 1

source_name: Lewis lung carcinoma cells, Mastic Oil, 3h treatment


title: LLC-DMSO-12h-rep1, quantile

description:
Replicate 1 of 2.
Vehicle 12h - 1

source_name: Lewis lung carcinoma cells, DMSO, 12h treatment


title: LLC-Moil-3h-rep2, quantile

description:
Replicate 2 of 2.
Mastic oil 3h - 2


title: LLC-Moil-6h-rep2, quantile

description:
Replicate 2 of 2.
Mastic oil 6h - 2

source_name: Lewis lung carcinoma cells, Mastic Oil, 6h treatment


title: LLC-DMSO-24h-rep1, quantile

description:
Replicate 1 of 2.
Vehicle 24h - 1

source_name: Lewis lung carcinoma cells, DMSO, 24h treatment


title: LLC-Moil-48h-rep1, quantile

description:
Replicate 1 of 2.
Mastic oil 48h - 1


title: LLC-DMSO-6h-rep1, quantile

description:
Replicate 1 of 2.
Vehicle 6h - 1

source_name: Lewis lung carcinoma cells, DMSO, 6h treatment


title: LLC-DMSO-6h-rep2, quantile

description:
Replicate 2 of 2.
Vehicle 6h - 2


title: LLC-DMSO-24h-rep2, quantile

description:
Replicate 2 of 2.
Vehicle 24h - 2


title: LLC-DMSO-3h-rep1, quantile

description:
Replicate 1 of 2.
Vehicle 3h - 1

source_name: Lewis lung carcinoma cells, DMSO, 3h treatment


title: LLC-Moil-6h-rep1, quantile

description:
Replicate 1 of 2.
Mastic oil 6h - 1


title: LLC-DMSO-12h-rep2, quantile

description:
Replicate 2 of 2.
Vehicle 12h - 2


title: LLC-Moil-12h-rep1, quantile

description:
Replicate 1 of 2.
Mastic oil 12h - 1


title: LLC-Moil-48h-rep2, rank invariant

description:
Replicate 2 of 2.
Mastic oil 48h - 2

data_processing: Data were normalized with rank invariant normalization, and statistical analysis was perfomed using ANOVA and clustering analysis with k-means clustering.


title: LLC-Moil-48h-rep1, rank invariant

description:
Replicate 1 of 2.
Mastic oil 48h - 1


title: LLC-DMSO-3h-rep2, quantile

description:
Replicate 2 of 2.
Vehicle 3h - 2


title: LLC-Moil-24h-rep1, rank invariant

description:
Replicate 1 of 2.
Mastic oil 24h - 1


title: LLC-Moil-6h-rep2, rank invariant

description:
Replicate 2 of 2.
Mastic oil 6h - 2


title: LLC-Moil-12h-rep1, rank invariant

description:
Replicate 1 of 2.
Mastic oil 12h - 1


title: LLC-DMSO-48h-rep2, rank invariant

description:
Replicate 2 of 2.
Vehicle 48h - 2


title: LLC-Moil-24h-rep2, rank invariant

description:
Replicate 2 of 2.
Mastic oil 24h - 2


title: LLC-Moil-3h-rep2, rank invariant

description:
Replicate 2 of 2.
Mastic oil 3h - 2


title: LLC-DMSO-48h-rep1, rank invariant

description:
Replicate 1 of 2.
Vehicle 48h - 1


title: LLC-Moil-12h-rep2, rank invariant

description:
Replicate 2 of 2.
Mastic oil 12h - 2


title: LLC-Moil-6h-rep1, rank invariant

description:
Replicate 1 of 2.
Mastic oil 6h - 1


title: LLC-DMSO-12h-rep1, rank invariant

description:
Replicate 1 of 2.
Vehicle 12h - 1


title: LLC-DMSO-24h-rep1, rank invariant

description:
Replicate 1 of 2.
Vehicle 24h - 1


title: LLC-DMSO-24h-rep2, rank invariant

description:
Replicate 2 of 2.
Vehicle 24h - 2


title: LLC-Moil-3h-rep1, rank invariant

description:
Replicate 1 of 2.
Mastic oil 3h - 1


title: LLC-DMSO-12h-rep2, rank invariant

description:
Replicate 2 of 2.
Vehicle 12h - 2


title: LLC-DMSO-6h-rep1, rank invariant

description:
Replicate 1 of 2.
Vehicle 6h - 1


title: LLC-DMSO-3h-rep2, rank invariant

description:
Replicate 2 of 2.
Vehicle 3h - 2


title: TAF4b heterozygote 1

city: Providence

first: Lindsay

last: Lovasco

state: RI

address: 70 Ship St.

department: MCB

institute: Brown University

hyb_protocol: per instructions of GeneChip Mouse gene 1.0 ST array

scan_protocol: Affymetrix Expression Console v1.1

data_processing: RMA-sketch

source_name: TAF4b heterozygote

label_protocol: Affymetrix Whole-transcript Sense Target Labeling Assay (Rev3)

extract_protocol: RNA was isolated using Trizol reagent

tag: genotype

value: TAF4b heterozygote


title: Wild type 3

source_name: Wild type

value: Wild type


title: Wild type 2


title: LLC-DMSO-3h-rep1, rank invariant

description:
Replicate 1 of 2.
Vehicle 3h - 1


title: Wild type 1


title: TAF4b knockout 1

source_name: TAF4b knockout

value: TAF4b knockout


title: TAF4b heterozygote 2


title: LLC-DMSO-6h-rep2, rank invariant

description:
Replicate 2 of 2.
Vehicle 6h - 2


title: RT101-monolayer, rep2

hyb_protocol: Biotin-labeled cRNA (15ug) was fragmented to a size range between 50-200 bases and used for array hybridization with Affymetrix reagents.  After hybridization, the arrays were washed and stained with streptavidin-phycoerythrin.

scan_protocol: Arrays were scanned at a resolution of 2.5 microns using an Affymetrix GeneChip Scanner 3000.

data_processing: Data were processed with RMA background correction and quantile normalization using Bioconductor packages

source_name: RT101 monolayer cells

label_protocol: cDNA was synthesized from 5ug of total RNA in the presence of an oligo-dT primer containing a T7 RNA polymerase promoter, and an in vitro transcription reaction was performed in the presence of a mixture of biotin-labeled ribonucleotides to produce biotinylated cRNA using Affymetrix reagents.

growth_protocol: JB6 cells were cultured in Eagle’s minimal essential medium (EMEM; BioWhittaker, Walkersville, MD) supplemented with 4% fetal bovine serum (FBS), 2 mM L-glutamine and 25 mg/mL gentamicin (Life Technologies/Gibco, Gaithersburg, MD)

extract_protocol: Total RNA was extracted and purified using the Qiagen RNeasy kit (Qiagen, Valencia, CA)

treatment_protocol: Cells were seeded over soft agar in 10 cm dishes (5 x 104 cells/dish) to induce colony formation.  RT101 cells (tumor variant) received no treatment and parental JB6 clone 41-5A cells were treated with 10 ng/ml bFGF. Colonies were harvested on day 7 to minimize selection of large colonies with necrotic centers.

tag: RT101 cells


title: TAF4b knockout 2


title: RT101-colonies, rep1

source_name: RT101 colonies


title: TAF4b heterozygote 3


title: bFGF-colonies, rep3

source_name: bFGF-treated colonies, JB6 cells

tag: naïve JB6 cells


title: TAF4b knockout 3


title: RT101-monolayer, rep1


title: stationary phase_3

city: Warsaw

first: Izabela

last: Sitkiewicz

address: Chełmska 30/34

department: Epidemiology and Clinical Microbiology

country: Poland

institute: National Medicines Institute

description: Part of series of experiments performed to determine response of S. agalactiae to human amniotic fluid

hyb_protocol: Overnight 45°C, standard Affymetrix protocol

scan_protocol: Affymetrix Genechip scanner 3000

data_processing: Raw data normalized to total signal intensity of S. agalactiae probes present on the array

organism: Streptococcus agalactiae

source_name: stationary phase

label_protocol: Standard Affymetrix

extract_protocol: Each amniotic fluid sample with bacteria used for RNA isolation was treated with 2 volumes of RNAprotect Bacteria Reagent (Qiagen, Valencia, CA), collected by centrifugation, and stored at –80°C until processing. RNA was extracted using the Lysing Matrix B microtubes containing 0.1 mm silica spheres (Qbiogene, Carlsbad, CA) and 1 mL Trizol (Invitrogen) with the FastPrep FP120 cell disrupter (Qbiogene). RNA extraction was completed with the RNeasy Mini kit (Qiagen) according to the manufacturer’s instructions.

tag: Strain NEM316 grown in human amniotic fluid to stationary phase


title: bFGF-monolayer, rep1

source_name: bFGF-treated monolayer JB6 cells


title: RT101-colonies, rep2


title: RT101-colonies, rep3


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

city: Beijing

first: Ruifu

address: No.20, DOngdajie, Fengtai

country: China

institute: Institute of Microbiology and Epidemiology

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

hyb_protocol: For hybridization, the slides were incubated in 50 µl of prehybridization buffer (5X SSC, 0.1% SDS and 0.1% BSA) at 42°C for at least 1 hr. The slides were washed twice in water for 1 min and once in 95% ethanol for 1 min, and then blown to dry with hot air. Dual-fluorescence labeled dried probes were resuspended in 35 µl of hybridization solution (50% deionized formamide, 5×SSC, 0.1% SDS, 5×Denhardt’s solution, and 0.5 µg/µl of sheared salmon sperm DNA). The mixture was denatured at 98°C for 5 min, and then was applied to the microarray slide followed by covering a 24×50 mm glass coverslip. The slides were hybridized in a moisture chamber at 42°C for 18 to 20 hr in a hybridization incubator (Robbins Scintific). After hybridization, the slides were washed in 1×SSC with 0.06% SDS for 2 min, then in 0.06×SSC for 2 min and finally in ethanol for 1 min. The slides were blown dry for scanning.

scan_protocol: The slides were scanned at a resolution of 10 µm by using a GenePix Personal 4100A Microarray Scanner (Axon Instruments).

data_processing: The scanning images were processed and the data were further analyzed by using GenePix Pro 4.1 software (Axon Instruments) combined with Microsoft Excel software. Spots were analyzed by adaptive quantitation, and the local background was subsequently subtracted. Spots with background-corrected signal intensity (median) in both channels less than two folds of background intensity (median) were rejected from further analysis. Data normalization was performed on the remaining spots by total intensity normalization methods. The normalized log2 ratio of WT/Δzur signal for each spot was recorded. Genes with less than four data points were considered unreliable, and their data points were discarded as well. The averaged log2 ratio for each remaining gene on the eight replicate slides was ultimately calculated. Significant changes of gene transcription level were identified with SAM software using one class mode. After the SAM analysis, only genes with at least 2-fold changes in expression were collected for further analysis.

organism: Yersinia pestis

source_name: Wild-type Y. pestis strain 201, 5mM ZnCl2, 26°C, 30 min

label_protocol: Each RNA preparation was used to prepare fluorescence-labeled probes for three separate slides. For the total of eight replicate slides, the incorporated dye was reversed to make up three flip dye pairs. 20 µg of total RNA was mixed with 5 µg of GDPs and 5 µg random hexamer primers, heated to 70°C for 10 min, and cooled quickly on ice. 1 µl of RNasin RNase inhibitor (Promega), 8 µl of 5×reverse transcription buffer, 4 µl of 0.1 M DDT, 1.6 µl of unmodified dNTP mixture (12.5 mM of each dATP, dGTP, and dCTP, plus 7.5 mM dTTP), 2 µl of 4 mM aminoallyl-dUTP (Sigma) and 200 U of SuperScript II reverse transcriptase (Invitrogen) were added to a final volume of 40 µl. The reaction occurred at 42°C for 1 hr, after which an additional 200 U of Superscript II reverse transcriptase was added. The mixture was then incubated at 42°C for an additional hour. The reaction was stopped by the addition of 20 mM EDTA, and the RNA was degraded by the addition of NaOH to a final concentration of 30 mM, followed by incubation at 65°C for 20 min. The mixture was neutralized by adding HCl to a final concentration of 30 mM. Free amines were removed by using Qiaquick PCR purification columns (Qiagen). The purified cDNA was air dried at 50°C, and then was labeled by the addition of 1/10 of one reaction vial of Cy3 monofunctional dye (Amersham) in 0.05 M sodium bicarbonate (pH9.3) and incubated for 2 hr at room temperature in the dark.

extract_protocol: To minimize bacterial RNA degradation and preserve gene transcription profile, double-volume of RNAprotect Bacteria Reagent (Qiagen) was added immediately to each aliquot, mixed thoroughly and placed at room temperature for 5 min. Total RNA extraction was performed by using MasterPureTM RNA Purification kits (Epicenter), exactly following the kit’s manual. Briefly, cell pellets were collected from the bacteria cultures by centrifugation. The lysis of cell samples was performed by Protease K digestion at 65°C for 15min, and protein was then removed from the lysed sample with the protein precipitation reagent provided by the kit. Isopropanol was added to precipitate nucleic acid and the residue DNA was removed with RNase-free DNase I. The RNA was then recovered with isopropanol. The RNAs from each four aliquots were pooled together. RNA quality was monitored by agarose gel electrophoresis and RNA quantity was measured by spectrophotometer.

value: wild-type strain 201

tag: growth

value: middle exponential growth phase

source_name: Zur mutant, 5mM ZnCl2, 26°C, 30 min

label_protocol: Each RNA preparation was used to prepare fluorescence-labeled probes for three separate slides. For the total of eight replicate slides, the incorporated dye was reversed to make up three flip dye pairs. 20 µg of total RNA was mixed with 5 µg of GDPs and 5 µg random hexamer primers, heated to 70°C for 10 min, and cooled quickly on ice. 1 µl of RNasin RNase inhibitor (Promega), 8 µl of 5×reverse transcription buffer, 4 µl of 0.1 M DDT, 1.6 µl of unmodified dNTP mixture (12.5 mM of each dATP, dGTP, and dCTP, plus 7.5 mM dTTP), 2 µl of 4 mM aminoallyl-dUTP (Sigma) and 200 U of SuperScript II reverse transcriptase (Invitrogen) were added to a final volume of 40 µl. The reaction occurred at 42°C for 1 hr, after which an additional 200 U of Superscript II reverse transcriptase was added. The mixture was then incubated at 42°C for an additional hour. The reaction was stopped by the addition of 20 mM EDTA, and the RNA was degraded by the addition of NaOH to a final concentration of 30 mM, followed by incubation at 65°C for 20 min. The mixture was neutralized by adding HCl to a final concentration of 30 mM. Free amines were removed by using Qiaquick PCR purification columns (Qiagen). The purified cDNA was air dried at 50°C, and then was labeled by the addition of 1/10 of one reaction vial of Cy5 monofunctional dye (Amersham) in 0.05 M sodium bicarbonate (pH9.3) and incubated for 2 hr at room temperature in the dark. The Cy3 and Cy5 reaction mixtures were combined, and the unincorporated dye was removed by using MinElute reaction cleanup columns (Qiagen). The purified probe was air dried at 50°C, and was stored at -20°C until use.

value: Zur mutant


title: bFGF-monolayer, rep2


title: bFGF-colonies, rep1


title: stationary phase_1


title: stationary phase_2


title: late log phase_3

source_name: late log phase

tag: Strain NEM316 grown in human amniotic fluid to late log phase


title: bFGF-monolayer, rep3


title: bFGF-colonies, rep2


title: late log phase_2


title: late log phase_1


title: mid log phase_1

source_name: mid log phase

tag: Strain NEM316 grown in human amniotic fluid to mid log phase


title: mid log phase_3


title: mid log phase_2


title: Expression of S. agalactiae genes in blood after 30 minutes at 37°C (donor A)

description: Part of series of experiments performed to determine response of S. agalactiae to human blood

label: Biotin

source_name: Growth in blood

extract_protocol: Each blood with bacteria sample was treated with 2 volumes of RNAprotect Bacteria Reagent (Qiagen, Valencia, CA), collected by centrifugation, and stored at –80°C until processing. Red cells were lyzed by Erythocyte Lysis (EL) buffer (Qiagen). RNA was extracted using the Lysing Matrix B microtubes containing 0.1 mm silica spheres (Qbiogene, Carlsbad, CA) and 1 mL Trizol (Invitrogen) with the FastPrep FP120 cell disrupter (Qbiogene). RNA extraction was completed with the RNeasy Mini kit (Qiagen) according to the manufacturer’s instructions.

tag: Strain NEM316 grown in blood


title: Expression of S. agalactiae genes in blood after 30 minutes at 40°C (donor A)


title: Expression of S. agalactiae genes in blood after 90 minutes at 40°C (donor A)


title: Expression of S. agalactiae genes in blood after 90 minutes at 37°C (donor A)


title: Expression of S. agalactiae genes in blood at time 0 (donor A)


title: Recovered5

status: Public on May 31 2009

city: San Francisco

first: Michael

last: DeSalvo

department: Department of Anesthesia

institute: UC San Francisco

hyb_protocol: Prior to hybridization, microarrays were post-processed by: 1) UV crosslinking at 60 mJ; 2) a “shampoo” treatment (3x SSC, 0.2% SDS at 65oC); 3) blocking with 5.5g succinic anhydride dissolved in 335mL 1-methyl-2-pyrrilidinone and 15mL sodium borate; and 4) drying via centrifugation. Dye-coupled cDNAs were cleaned (Qiagen MinElute), and appropriate Cy3 and Cy5 labeled cDNAs were mixed together in a hybridization buffer containing 0.25% SDS, 25mM HEPES, and 3x SSC. The hybridization mixtures were boiled for 2min at 99oC then allowed to cool at room temperature for 5min. The cooled hybridization mixtures were pipetted under an mSeries Lifterslip (Erie Scientific), and hybridization took place overnight at 63oC. Microarrays were washed twice in 0.6x SSC and 0.01% SDS followed by a rinse in 0.06x SSC and dried via centrifugation.

scan_protocol: All microarrays were scanned using an Axon 4000B scanner (Molecular Devices) where care was taken to balance photomultiplier tube (PMT) settings.

data_processing: TIFF images were generated with GenePix Pro 6.0, and gridding was performed using TIGR Spotfinder 3.1.1 with the Otsu segmentation method and background correction (the top 25% of background pixels were discarded prior to local background estimation). Using TIGR MIDAS 2.19, background-corrected data were LOWESS normalized, and in-slide duplicates were averaged. Genes were included in statistical analyses only if there were data for three out of five hybridizations for a given category (i.e., control, bleaching, or recovery).

organism: Orbicella faveolata

label_protocol: 1ug of total RNA was amplified using the MessageAmp II aRNA kit (Ambion), and 3ug of aRNA per sample were primed with 5ug/uL random nonamer for 10min at 70oC. Reverse transcription (RT) lasted for 2hr at 50oC using a master mix containing a 4:1 ratio of aminoallyl-dUTP to TTP. Following RT, single-stranded RNA was hydrolyzed by incubating the RT reactions in 10uL 0.5M EDTA and 10uL 1M NaOH for 15min at 65oC. After hydrolysis, RT reactions were cleaned using Qiagen MinElute reaction purification columns. Cy3 and Cy5 dyes (GE Healthcare) were dissolved in 17uL DMSO, and the coupling reactions lasted for 2hr at room temperature in the dark.

extract_protocol: Total RNA from all frozen coral fragments was isolated using Qiazol lysis reagent (QIAGEN). Live tissue was chiseled off each coral fragment and homogenized using a pre-chilled mortar and pestle embedded in dry ice. Frozen coral powder was transferred directly to Qiazol. Two chloroform extractions were performed, followed by isopropanol precipitation and 2 washes in 70% ethanol. RNA pellets were re-dissolved in nuclease-free water and cleaned further over RNeasy Mini columns (QIAGEN). RNA quality and quantity were assessed with a NanoDrop ND-1000 spectrophotometer and an Agilent 2100 Bioanalyzer.

treatment_protocol: On the night of 1 September, control time point samples from five different colonies were collected from each tank (Figure 1), before one 200-Watt aquarium heater was turned on in the treatment aquarium. A second heater was turned on 3 days later. During the thermal stress experiment, the control aquarium received mean water temperature of 28.8 ± 1.2oC; the heated aquarium, 31.5 ± 1.1oC. During the thermal stress experiment, tanks received mean PAR of 420 ± 152 µmol m-2 s-1. On 11 September, and 19 October, one sample each from five different colonies was collected from each tank for the bleaching and recovery time point, respectively.

tag: sample

value: Pool of all control samples included in the experiment

value: Fragment that had regained symbionts following experimental bleaching


title: 32D cells time 4 hr after actinomycin D, Low IL3,  Experiment 1

city: Pittsburgh

first: Richard

middle: A

last: Steinman

state: PA

address: 5117 Centre Avenue

institute: University of Pittsburgh

description: This sample is part of a series in which kinetic analysis of transcript expression after actinomycin D exposure was performed. Time 0 samples were collected immediately before exposure.

hyb_protocol: Affymetrix FS 450_000Q performed per manufacturer

scan_protocol: Affymetrix 6G scanner, automated scanning

data_processing: Microarray normalization was done with dChip software with the default settings.

source_name: 32D myeloblasts

label_protocol: Genechip expression 3' amplification 1 cycle target labeling kit (Affymetrix)

treatment_protocol: 32Dcl3 cells were washed free of IL-3 and resuspended in IL-3-free medium. Actinomycin-D (Sigma-Aldrich) was titrated to 10 µg/mL and the culture was divided into two equal parts. IL-3 was titrated to either 1 ng/mL (high IL3) or 10 pg/mL (low IL3) and RNA was prepared from 1x107 cells at 0, 2, 4, and 6 hour time points using RNeasy total RNA purification kits (Qiagen, Valencia, CA). For half-life assessments made in G-CSF, cells were prepared as noted above and incubated for 18 hours. Actinomycin-D was titrated to 10 µg/mL and cell cultures were divided into two equal parts. Cells received either 1 ng/mL IL-3 or an equal volume of water as a control. RNA was prepared from cells harvested at 0, 2, 4 and 6 hour time points. Actinomycin-D was titrated to 10 µg/mL and cells were harvested at 0, 1, 2, 4 and 6 hour time points.

tag: 32D cells proliferate in IL3 and differentiate into neutrophils in GCSF


title: 32D cells time 2 hr after actinomycin D, Low IL3,  Experiment 1


title: 32D cells time 4 hr after actinomycin D, High IL3,  Experiment 1


title: Recovered4


title: Recovered3


title: 32D cells time 2 hr after actinomycin D, High IL3,  Experiment 2

description: This sample is part of a series in which kinetic analysis of transcript expression after actinomycin D exposure was performed.  Time 0 samples were collected immediately before exposure.

source_name: 32Dcl3 myeloblasts

tag: 32D myeloblasts proliferate in IL3, differentiate in GCSF.


title: 32D cells time 0 after actinomycin D Experiment 2

treatment_protocol: 32Dcl3 cells were washed free of IL-3 and resuspended in IL-3-free medium. Actinomycin-D (Sigma-Aldrich) was titrated to 10 µg/mL and the culture was divided into two equal parts. IL-3 was titrated to either 1 ng/mL or 10 pg/mL and RNA was prepared from 1x107 cells at 0, 2, 4, and 6 hour time points using RNeasy total RNA purification kits (Qiagen, Valencia, CA). For half-life assessments made in G-CSF, cells were prepared as noted above and incubated for 18 hours. Actinomycin-D was titrated to 10 µg/mL and cell cultures were divided into two equal parts. Cells received either 1 ng/mL IL-3 or an equal volume of water as a control. RNA was prepared from cells harvested at 0, 2, 4 and 6 hour time points. Actinomycin-D was titrated to 10 µg/mL and cells were harvested at 0, 1, 2, 4 and 6 hour time points.

tag: Murine myelopblast cell line that proliferates in IL-3 and differentiates into neutrophils when IL3 is replaced with G-CSF


title: 32D cells time 4 hr after actinomycin D, Low IL3,  Experiment 2

tag: 32Dcl3 myeloblast cells proliferate in IL3, differentiate into neutrophils in GCSF


title: 32D cells time 2 hr after actinomycin D, Low IL3,  Experiment 2

tag: 32Dcl3 cells proliferate in IL3, differentiate into neutrophils in GCSF


title: 32D cells time 2 hr after actinomycin D, High IL3,  Experiment 1

tag: 32Dcl3 cells proliferate in IL3 and differentiate into neutrophils in GCSF


title: 32D cells time 4 hr after actinomycin D, High IL3,  Experiment 2

tag: 33D cells proliferate in IL3, differentiate in GCSF


title: Recovered2


title: 32D cells time 0 after actinomycin D Experiment 1

tag: 32D cells proliferate in IL3, differentiate into neutrophils in GCSF


title: Brassica_Globular-Cotyledon_BR2-TR4

city: Saskatoon

first: Jitao

last: Zou

state: Saskatchewan

address: 110 Gymnasium Place

country: Canada

institute: Plant Biotechnology Institute, National Research Council Canada

description: Endosperm at the globular-shape embryo stage (6 days after flowering (DAF)) and  cotyledon stage (14 DAF).

hyb_protocol: After a second purification as above, both Cy5- and Cy3-labeled cDNAs were pooled. The mixture was concentrated using a vacuum dryer and resuspended in 30 µl hybridization buffer. The pre-hybridization and hybridization procedures were performed as previously described (Huang et al, Plant Mol Biol, 2006, 62, 579-591).

scan_protocol: Hybridized arrays were scanned using a Genepix 4000B microarray scanner (Axon Instruments, CA, USA) at 5 µm resolution, 100% laser power, and different PMT values to obtain a similar green and red overall intensity. Raw spot fluorescence intensities were collected using GenePix Pro version 6.0 (Axon Instruments, CA, USA). A quality control filter was used to flag questionable spots on the array so they could be removed from analysis.

data_processing: Before normalization, a basic pre-processing was performed. The outlier spots with median intensity larger than that of (median ± 10×SD) were removed. Channel intensity was considered low if its background-corrected intensity value was less than a preset low-intensity threshold (LIT) of 0.5; low channel intensities were replaced by the LIT. The spot was marked as bad if either of the two channels was low. Spots with background-subtracted median intensity greater than median background intensity in at least one channel were selected and used for normalization and further analysis. Array features annotated as “DMSO,” “Blank” or “Empty” were flagged and excluded from analysis. A spot was removed if the fold change of the Cy3/Cy5 group was in the opposite direction of that of the Cy5/Cy3 group. For example, if spot A was up-regulated for the Cy3/CY5 group and down-regulated for the Cy5/Cy3 group, then spot A was removed because of its conflicting data.  Foreground intensities of selected genes were background corrected using the normexp background measure, and the signal intensities were normalized using ‘global loess’ for within-array normalization and quantile for between-array normalization using R-package Limma. The signal intensity (background-corrected) of each spot on the microarray was combined by averaging two biological duplicates using Excel 2003 (Microsoft).   After the normalization process, the signal intensities and log2 ratio of the two channels of each replicate were obtained. These two sets of data were processed through rank product implemented in the BioMiner software suite with a 5% false discovery rate. Genes with very inconsistent signal intensities and log2 ratio were excluded from this process. Then we combined the results from all three stages. The genes that ranked high in both datasets were considered differentially expressed genes and classified into patterns based on the clustering approach.  The DNA sequence for each of the unisequences spotted on the array was searched for in the TAIR protein database (version 7) using BLASTX programs at an E value less than 10-5. The best match was extracted using an in-house Perl script and used as a basis for obtaining annotations for each probe based on sequence identity. The best matches were compared to terms of the Gene Ontology (GO) Consortium (GO). Categories were assigned based on biological, functional, and molecular annotations available from GO.

organism: Brassica napus

source_name: Endosperm at globular-shape-embryo stage

label_protocol: Approximately 1 µg total RNA was treated with DNase I and reverse transcribed into aRNA with MessageTM II aRNA Amplification Kit (Ambion, Austin, TX, USA) according to the manufacturer’s protocol. The aRNA was selected as the template for fluorescent target preparation for microarray experiments. The quality of amplification was identified using agarose gel electrophoresis, and the aRNA yield was quantified using a spectrophotometer DU®7400 (Beckman, USA).  An aliquot of 3 μg of aRNA was reverse transcribed using an oligo(dT)20 primer and 400 units of Superscript III reverse transcriptase (Invitrogen, Cat. No.18080-044) in the presence of 25 mM dATP, 25 mM dCTP, 25 mM dGTP, 15 mM aminoallyl-dUTP (Ambion, Cat. No. AM8439) and 10 mM dTTP. The single stranded cDNA was purified using YM30 filter column (Millipore, Billerica, MA) and labeled with CyDye post-labeling dye (Amersham, UK) for 1 hr.

growth_protocol: B. napus L. cv DH12075 was grown in 4-liter pots containing Terra-lite Redi-earth mix (W. R. Grace and Co., Ajax, ON, Canada). A total of 10 g of time-release (100-day) 14-14-14-fertilizer pellets were applied to the surface of each pot at the time of seeding. Each pot was watered daily with 200 ml of nutrient solution (150 μg ml−1 N, 62.5 μg ml−1 P, and 125 μg ml−1 K) at the early growth stage, with 400 ml of the solution from 3 weeks after sowing, and with 200 ml of the solution during seed maturation. The plants were grown in a greenhouse with 20/16°C (d/n) temperature, 16/8 h photoperiod, and 250 μmol m−2 s−1 light intensity.

extract_protocol: Total RNA was extracted from frozen tissues at each embryo developmental stage using RNeasy Plant Mini Kit (Qiagen, Mississauga, ON) and on-column DNase digestion with RNase-free DNase Set (Qiagen) was performed for each sample during RNA isolation according to the protocol provided by the manufacturer. The yield and RNA purity were determined using a Nanodrop ND-1000 Spectrophotometer, and the intactness of RNA was verified by gel electrophoresis.

treatment_protocol: We collected endosperm tissues by first dissecting out the embryos, which were examined to determine the developmental stage. Then, using a glass pipette with a fine tip, the endosperm was extracted and collected into pools based on whether embryo development was at the globular-shape, heart-shape or cotyledon stage. During this process, care was taken to avoid tissue from the integument, but limited contamination cannot be ruled out. Amongst different sections of the endosperm, our tissue collections were enriched with micropylar endosperm surrounding the zygote.

tag: cultivar

source_name: Endosperm at cotyledon stage


title: Brassica_Globular-Cotyledon_BR2-TR3


title: Brassica_Globular-Cotyledon_BR2-TR2


title: Brassica_Globular-Cotyledon_BR2-TR1


title: Bleached5

value: Thermal stressed fragment taken after a significant bleaching response


title: Recovered1


title: Brassica_Globular-Cotyledon_BR1-TR2


title: Brassica_Globular-Cotyledon_BR1-TR1


title: Bleached4


title: Bleached3


title: Brassica_Globular-Cotyledon_BR1-TR4


title: Brassica_Globular-Cotyledon_BR1-TR3


title: Control4

value: Control fragment taken prior to thermal stress


title: Bleached2


title: Bleached1


title: Control5


title: Control3


title: Control2


title: Brassica_Heart-Cotyledon_BR2-TR1

description: Endosperm at the heart-shape embryo stage (8 DAF) and cotyledon stage (14 DAF).

source_name: Endosperm at heart-shape-embryo stage


title: Brassica_Heart-Cotyledon_BR2-TR2


title: Brassica_Heart-Cotyledon_BR2-TR3


title: Mfav_tmb_top5

description: Six fragments (9.5 ± 3.5cm2) from the top (2.7m), middle (3.7m), and bottom (5.2m) of one massive colony of Montastraea faveolata were collected with a hammer and chisel at “La Bocana” reef near Puerto Morelos, Quintana Roo, Mexico on 31 July 2007.  The fragments were divided evenly between two aquaria (50L) that received a constant flow of seawater (~0.64L/min).  Each aquarium was fit with a water pump connected to a spray-bar to provide constant water movement and aeration.  Both aquaria were placed in a common pond with flowing water to buffer diurnal temperature fluctuations, and both aquaria were exposed to shaded ambient light.  All coral fragments were mounted on plasticene and kept at a depth of ~7cm. On the night of 1 September, one 200-Watt aquarium heater was turned on in the treatment aquarium, and a second heater was turned on 3 days later.  During the thermal stress experiment, the control aquarium received an average water temperature of 28.8 ± 1.2oC; the heated aquarium, 31.5 ± 1.1oC. PAR present during the thermal stress experiment averaged 420 ± 152 uE.  On the night of 7 September, all fragments were frozen in liquid nitrogen.

hyb_protocol: Prior to hybridization, microarrays were post-processed by: 1) UV crosslinking at 60 mJ; 2) a “shampoo” treatment (3x SSC, 0.2% SDS at 65oC); 3) blocking with 5.5g succinic anhydride dissolved in 335mL 1-methyl-2-pyrrilidinone and 15mL sodium borate; and 4) drying via centrifugation. Dye-coupled cDNAs were cleaned (QIAGEN MinElute), and appropriate Cy3 and Cy5 labeled cDNAs were mixed together in a hybridization buffer containing 0.25% SDS, 25mM HEPES, and 3x SSC. The hybridization mixtures were boiled for 2min at 99oC then allowed to cool at room temperature for 5min. The cooled hybridization mixtures were pipetted under an mSeries Lifterslip (Erie Scientific), and hybridization took place in Corning hybridization chambers overnight at 63oC. Microarrays were washed twice in 0.6x SSC and 0.01% SDS followed by a rinse in 0.06x SSC and dried via centrifugation. Slides were immediately scanned using an Axon 4000B scanner.

scan_protocol: Scanned using Axon 4000B scanner with GenePix 6.0 software.

data_processing: All microarrays were scanned using an Axon 4000B scanner (Molecular Devices), and TIFF images were generated with GenePix Pro 6.0 software.  Gridding was performed using Spotfinder 3.1.1 (TIGR) with the Otsu segmentation method and background correction (the top 25% of background pixels were discarded prior to local background estimation).  Using MIDAS 2.19 (TIGR), background-corrected data were LOWESS normalized, and in-slide duplicates were averaged.  Log2 ratios were manually calculated, and data from all 18 hybridizations were compiled into a single file.  Genes were included in statistical analyses only if there were data for two out of three hybridizations for a given category (i.e. top-control, top-treatment, middle-control, middle-treatment, etc…).  All statistical analyses and clustering were performed in TMEV 4.0 (TIGR).  A two-factor ANOVA was performed with “location” as factor one (3 groups, n=6 per group) and “treatment” as factor two (2 groups, n=9 per group).  Differentially expressed genes were chosen at α = 0.05. To look for relatedness in gene expression, array trees were generated with complete linkage clustering according to Manhattan distance.  Support values for tree nodes were generating by bootstrapping the gene expression data over 1000 replicates.  Additionally, t-tests between control (n=3) and heat-stressed (n=3) samples were performed to identify differentially expressed genes within each region of the colony (top, middle, and bottom).  One-way ANOVAs were also performed to look for differences between control top, middle, and bottom fragments, and again for heat-treated top, middle, and bottom fragments.

label_protocol:
Labeled using Amersham CyDye Post-labeling kit.
http://cat.ucsf.edu/pdfs/hybCourseProtocols_v2.7.pdf
Specifically, 15ug of total RNA were primed with 10ug/uL of Oligo-dT primer for 10min at 70oC. Reverse transcription (RT) lasted for 2hr at 42oC using a master mix containing a 4:1 ratio of aminoallyl-dUTP to TTP. Following RT, single-stranded RNA was hydrolyzed by incubating the RT reactions in 10uL 0.5M EDTA and 10uL 1M NaOH for 15min at 65oC. After hydrolysis, RT reactions were cleaned using QIAGEN MinElute Reaction Purification columns. Cy3 and Cy5 dyes (GE Healthcare) were dissolved in 20uL DMSO, and the coupling reactions lasted for 2hr at room temperature in the dark.

extract_protocol:
Qiazol used to extract total RNA.
RNeasy Mini kit used to clean RNA.
Details: Total RNA from all frozen coral fragments was isolated using Qiazol lysis reagent (QIAGEN). Live tissue was chiseled off each coral fragment and homogenized using a pre-chilled mortar and pestle embedded in dry ice. Frozen coral powder was transferred directly to Qiazol. Two chloroform extractions were performed, followed by isopropanol precipitation and 2 washes in 70% ethanol. RNA pellets were re-dissolved in nuclease-free water and cleaned further over RNeasy Mini columns (QIAGEN). RNA quality and quantity were assessed with a NanoDrop ND-1000 spectrophotometer and an Agilent 2100 Bioanalyzer. Microarray protocols followed those established by the Center for Advanced Technology at the University of California, San Francisco (http://cat.ucsf.edu/).

value: Pool of all coral samples included in the experiment.

treatment_protocol: Heat-treated coral fragment.

value: Fragment taken from top of colony(~2m).


title: Mfav_tmb_top6


title: Control1


title: Brassica_Heart-Cotyledon_BR1-TR3


title: Brassica_Heart-Cotyledon_BR2-TR4


title: Brassica_Heart-Cotyledon_BR1-TR4


title: Brassica_Heart-Cotyledon_BR1-TR2


title: Brassica_Globular-Heart_BR2-TR4

description: Endosperm at the globular-shape embryo stage (6 days after flowering (DAF)) and  heart-shape embryo stage (8 DAF).


title: Brassica_Globular-Heart_BR2-TR2


title: Brassica_Heart-Cotyledon_BR1-TR1


title: Mfav_tmb_top4


title: Mfav_tmb_top3

treatment_protocol: Control fragment -- not thermal stressed.


title: Brassica_Globular-Heart_BR2-TR3


title: Mfav_tmb_top1


title: Mfav_tmb_middle6

value: Fragment taken from middle of colony (~3m).


title: Mfav_tmb_middle4


title: Mfav_tmb_top2


title: Mfav_tmb_middle1


title: Brassica_Globular-Heart_BR1-TR4


title: Mfav_tmb_bottom3

value: Fragment taken from bottom of colony (~5.5m).


title: Brassica_Globular-Heart_BR1-TR3


title: Brassica_Globular-Heart_BR2-TR1


title: Brassica_Globular-Heart_BR1-TR1


title: Mfav_tmb_middle2


title: Mfav_tmb_bottom5


title: Mfav_tmb_bottom6


title: Mfav_tmb_bottom2


title: Mfav_tmb_bottom1


title: Brassica_Globular-Heart_BR1-TR2


title: Mfav_tmb_middle5


title: Mfav_tmb_bottom4


title: Mfav_tmb_middle3


title: MfAg-72Hrs

status: Public on May 15 2009

city: Bethesda

first: Sasisekhar

last: Bennuru

address: 10 Center Dr, Bldg 10, Rm 11N208

department: LPD

institute: NIH/NIAID

hyb_protocol: Oligo arrays were hybridized to the cDNA probes in formamide based-hybridization buffer (MWG’s “Coverslips Hybridization buffer”) at 42C overnight and washed in a series of room temperature baths (2X SSC, 0.2% SDS; then 2X SSC, then 0.2X SSC) for 12 minutes each. Fluorescent Cy3- and Cy5- capture reagents were combined in MWG’s “Coverslips hybridization buffer” and added to each array. The arrays were incubated for 3 hours at 50C and washed as above. Each experiment consisted of a pair wise competitive hybridization of cDNA samples from female and male worms

scan_protocol: Using Axon's GenePix 4000B fluorescent scanner.

data_processing: Axon's GenePix Pro software was used for extracting the data from the scanned images of the microarrays. Normalization method: 50th Percentile (Median) using all spot filtered Genes. Signal Values were floored at 50 (Chan A and Chan B). Data was extracted and aligned by the Inventory Well ID. Within array multiple occurrences of Well ID were reduced to a single instance by selecting the one with the strongest signal (Chan A + Chan B).

source_name: LEC-MfAg, 72hrs

label_protocol: Amino-allyl amplified cDNA was labeled using the Amersham Pharmacia Dye labeling kit.

growth_protocol: LEC were cultured in EGM-2MV medium that is supplemented with growth factors

extract_protocol: Total RNA was extracted using Qiagen RNAeasy kit

treatment_protocol: LECs seeded into separate well were stimulated with Brugia malayi adult crude extract (BmA) or Brugia malayi microfilarial antigen (MfAg) for 24-72 hrs

tag: cell type

value: adult skin derived primary lymphatic endothelial cells

source_name: LEC-unexposed, 72hrs


title: Control-72Hrs

source_name: LEC-unexposed-pooled, 72hrs


title: BmA-48Hrs

source_name: LEC-BmA, 48hrs

source_name: LEC-unexposed, 48hrs


title: MfAg-48Hrs

source_name: LEC-MfAg, 48hrs


title: MfAg-24Hrs

source_name: LEC-MfAg, 24hrs

source_name: LEC-unexposed, 24hrs


title: BmA-72Hrs

source_name: LEC-BmA, 72hrs


title: Control-48Hrs

source_name: LEC-unexposed-pooled, 48hrs


title: Control-24Hrs

source_name: LEC-unexposed-pooled, 24hrs


title: BmA-24Hrs

source_name: LEC-BmA, 24hrs


title: Cardiac Progenitor MUT3

status: Public on May 13 2009

first: Chulan

last: Kwon

address: 1650 Owens Street

institute: Gladstone Institutes

description: YFP+ cells marking precardiac mesoderm were FACS-sorted (w/wo stabilized Beta-catenin), and their total RNA was isolated, amplified and labeled with the NUGENE pico kit for microarray analyses.  Mutant samples were obtained from YFP+ cells isolated from embryos with Isl1-cre, Rosa-YFP, Beta-catenin (loxP ex3). The mutants are conditional knockout mouse embryos that activates stable Beta-catenin in cardiac progenitors.

hyb_protocol: Performed as manufacturer's protocol (Affymetrics)

data_processing: Raw data generated from at least three independent experiments were further analyzed by the group of Dr. Ru-Fang Yeh at the Center for Informatics and Molecular Biostatistics, UCSF.

source_name: Embryonic Cardiac Progenitors - mutant

label_protocol: Total RNA was amplified with the WT-Ovation Pico RNA Amplification System, fragmented and labeled with the FL-Ovation cDNA Biotin Module V2 (Nugen).

extract_protocol: Trizol extraction

value: E9.0 embryonic cardiac progenitors


title: Cardiac Progenitor MUT1

scan_protocol: Performed as manufacturer's protocol (Affymetrics)


title: Subcutaneous adipose tissue from morbid obese patients female 11

status: Public on May 10 2009

city: Baronissi (SA)

first: Rosanna

last: Martinelli

address: Via Allende

department: Medicine and Surgery

country: Italy

institute: Salerno University

description: No further additional informations

hyb_protocol: For each sample 1 μg of total RNA was labeled and samples from the Hy3 and Hy5 labeling reactions were combined in a total volume of 220 μL,.added to 220 μL of 2X Hybridization buffer incubated at 95°C for 5 min. Mix were applied to the arrays and incubate at 60° C for 16h using the Agilent microarray hybridization chambers SureHyb.  in a hybridization oven with rotation. Microarray slide were washed twice at 60° C for 1 min. in Wash buffer A,  twice at 23° C for 2 min. in Wash Buffer B, twice at 23° C for 2 min in Wash Buffer C.

scan_protocol: Microarrays were scanned on a Agilent DNA Microarray Scanner (Agilent Technologies) and raw data extracted using Feature Extraction software (Agilent Technologies).

data_processing: Data were analyzed using GeneSpring® version 7.2 (Agilent Technologies) and statistical analysis was performed using background-corrected mean signal intensities from each dye channel. Microarray data were normalized using intensity-dependent global normalization (LOWESS). Differentially expressed miRNAs were identified using a filtering by standard deviation (0-2) and the Benjamini and Hochberg False Discovery Rate (p-Value 0.05) to minimize selection of false positives. Of the significantly differentially expressed miRNA, only those with greater than log2 1.5-fold increase or decrease in expression of obese subject compared with non obese controls were used for further analysis.

source_name: subcutaneous adipose tissue from non obese females

label_protocol: Total RNAs of control were labeled with Cy3 using the miRCURY™  LNA microRNA Array labeling kit (Exiqon) following instruction manual.

extract_protocol:
Total RNA was extracted from frozen subcutaneous adipose tissue samples by homogenization in guanidine thiocyanate-phenol solution (QIAzolTM Lysis Reagent; Qiagen) and then purified using mirVana miRNA Isolation Kit (Ambion), according to the manufacturer's protocol.
RNA concentrations and purity was evaluated by NanoDrop ND-1000 UV-Vis Spectrophotometer (NanoDrop Technologies, Wilmington, DE, USA). RNA integrity was verified using the Agilent 2100 Bioanalyzer (Agilent Technologies).

tag: Subcutaneous adipose tissue (SAT) from a pool of 5 non obese females. SAT aliquots (about 1 cm2 of fat, average 100-600 mg), not in contact with abdominal district, were excised during surgery. All biopsies were snap frozen and stored in liquid nitrogen until RNA isolation

source_name: subcutaneous adipose tissue from obese female

label_protocol: Total RNAs of obese female were labeled with Cy5 using the miRCURY™  LNA microRNA Array labeling kit (Exiqon) following instruction manual.

tag: Subcutaneous adipose tissue (SAT) from a obese female. SAT aliquot not in contact with abdominal district, were excised during surgery. The biopsy was snap frozen and stored in liquid nitrogen until RNA isolation


title: Subcutaneous adipose tissue from morbid obese patients female 12

description: No further additional informations

tag: Subcutaneous adipose tissue (SAT) from a pool of 5 non obese females. SAT aliquots (about 1 cm2 of fat, average 100-600 mg), not in contact with abdominal district, were excised during surgery. All biopsies were snap frozen and stored in liquid nitrogen until RNA isolation.

tag: Subcutaneous adipose tissue (SAT) from a obese female. SAT aliquot not in contact with abdominal district, were excised during surgery. The biopsy was snap frozen and stored in liquid nitrogen until RNA isolation.


title: Cardiac Progenitor WT3

source_name: Embryonic Cardiac Progenitors - wild type


title: Subcutaneous adipose tissue from morbid obese patients female 9

description: No further additional informations


title: Cardiac Progenitor WT1

description: YFP+ cells marking precardiac mesoderm were FACS-sorted (w/wo stabilized Beta-catenin), and their total RNA was isolated, amplified and labeled with the NUGENE pico kit for microarray analyses. Mutant samples were obtained from YFP+ cells isolated from embryos with Isl1-cre, Rosa-YFP, Beta-catenin (loxP ex3). The mutants are conditional knockout mouse embryos that activates stable Beta-catenin in cardiac progenitors.


title: Subcutaneous adipose tissue from morbid obese patients female 7

description: No further additional informations


title: Subcutaneous adipose tissue from morbid obese patients female 8

description: No further additional informations


title: Cardiac Progenitor MUT2


title: Cardiac Progenitor WT2


title: Subcutaneous adipose tissue from morbid obese patients female 6

description: No further additional informations


title: Subcutaneous adipose tissue from morbid obese patients male 8

description: No further additional informations

source_name: subcutaneous adipose tissue from non obese males

tag: Subcutaneous adipose tissue (SAT) from a pool of 3 non obese males. SAT aliquots (about 1 cm2 of fat, average 100-600 mg), not in contact with abdominal district, were excised during surgery. All biopsies were snap frozen and stored in liquid nitrogen until RNA isolation

source_name: subcutaneous adipose tissue from obese male

label_protocol: Total RNAs of obese male were labeled with Cy5 using the miRCURY™  LNA microRNA Array labeling kit (Exiqon) following instruction manual.

tag: Subcutaneous adipose tissue (SAT) from a obese male. SAT aliquot not in contact with abdominal district, were excised during surgery. The biopsy was snap frozen and stored in liquid nitrogen until RNA isolation


title: Subcutaneous adipose tissue from morbid obese patients female 2

description: No further additional informations


title: Subcutaneous adipose tissue from morbid obese patients male 7

description: No further additional informations


title: Subcutaneous adipose tissue from morbid obese patients female 3

description: No further additional informations


title: Subcutaneous adipose tissue from morbid obese patients male 9

description: No further additional informations


title: Subcutaneous adipose tissue from morbid obese patients female 5

description: No further additional informations


title: Subcutaneous adipose tissue from morbid obese patients female 1

description: No further additional informations


title: Subcutaneous adipose tissue from morbid obese patients male 10

description: No further additional informations


title: Subcutaneous adipose tissue from morbid obese patients male 6

description: No further additional informations


title: Subcutaneous adipose tissue from morbid obese patients male 2

description: No further additional informations

data_processing: Data were analyzed using GeneSpring® version 7.2 (Agilent Technologies) and statistical analysis was performed using background-corrected mean signal intensities from each dye channel. Microarray data were normalized using intensity-dependent global normalization (LOWESS). Differentially expressed miRNAs were identified using a filtering by standard deviation (0-2) and the Benjamini and Hochberg False Discovery Rate (p-Value 0.05) to minimize selection of false positives. Of the significantly differentially expressed miRNA, only those with greater than log2 1.5-fold increase or decrease in expression of obese subject compared with non obese controls were used for further analysis

label_protocol: Total RNAs of obese male were labeled with Cy5 using the miRCURY™  LNA microRNA Array labeling kit (Exiqon) following instruction manual


title: CI#243.12_replicate2

status: Public on May 01 2009

city: Tuebingen

middle: H.

last: Walter

address: Calwer Str. 7

department: Department of Medical Genetics

country: Germany

institute: University of Tuebingen

description: not applicable

hyb_protocol: Affymetrix standard protocol

scan_protocol: Affymetrix standard protocol

data_processing: Normalization, background correction and signal summarization was done using RMA

source_name: Dppa4 knockout ES cells

label_protocol: Affymetrix standard protocol (One-Cycle Target Labeling Kit)

extract_protocol: Qiagen RNeasy Mini Kit

tag: cell line

value: Dppa4 knockout ES cells


title: CI#243.12_replicate3

description: not applicable


title: CI#243.12_replicate1

description: not applicable


title: Hepatocellular carcinoma_Mdr2-/-,IkB-SR_HCC10 Cy5

first: Daniel

last: Haag

state: Santa Clara

address: 265 Campus Drive

department: ISCBRM

institute: Stanford School of Medicine

description: Tumor model: mice carrying the Mdr2-/- allele were bred to transgenic mice (IkB-SR) carrying two transgenes: the IkB-superrepressor controlled by a tetracycline-regulated promoter and the tetracycline-transactivator under control of the hepatocyte-specific C/EBP? promoter (TALAP1) (Pikarsky et al., Nature, 2004).

hyb_protocol: Purified, dye-labelled cDNA was mixed with 120 µl UltraHyb hybridization buffer (Ambion), agitated for 30–60 min at 60°C, then for 10 min at 70°C on a thermo mixer and subsequently applied to pre-heated (60°C) microarrays mounted in a GeneTAC Hybridization Station (Genomic Solutions, Ann Arbor). Hybridizations were performed for 23 h at 42°C with gentle agitation. Thereafter, the arrays were automatically washed at 36°C with (i) 0.5× SSC, 0.1% (w/v) SDS for 5 min; (ii) 0.05× SSC, 0.1% (w/v) SDS for 3 min; and (iii) 0.05× SSC for 2 min. Flow time was set to 40 s. Immediately after the completion of the final washing step, the arrays were unmounted, immersed in 0.05× SSC, 0.1% (w/v) Tween 20 and dried by centrifugation in 50 ml Falcon tubes (30 s at 500, 1000 and 1500 r.p.m., respectively, followed by a final step of 240 s at 2000 r.p.m.).

scan_protocol: Hybridized microarrays were scanned in a two-color Agilent Scanner G25505B (Agilent, Santa Clara, CA) at 5 µm resolution and automatically adjusting PMT voltages according to manufactures specification. Recorded images were subsequently analyzed using GenePix Pro 6.0 software.

data_processing: Data tables of raw intensity values were filtered excluding those features that were either user-flagged, exhibited a foreground intensity mean-to-median ration of more than Q3+3 * Interquartile range or having an overall foreground intensity of below 250. For normalization of filtered features, the variance stabilization normalization algorithm (Huber et al., 2002) was applied. The “VALUE”-column contains the Log-ratio in Cy5 (channel 2) over Cy3 (channel 1) orientation for each microarray.

source_name: Universal Mouse Reference RNA (Stratagene)

label_protocol:
For reverse transcription:
RNA/primer-mix: 2 µg total RNA, 500 ng (dT)-T7 primer, total volume 6 µl
* denature RNA/primer 4' at 70°C, chill on ice;
RT-mix: 2 µl M-MuLV RT buffer (Fermentas), 0.5 µl 10 mM dNTP-mix, 0.5 µl T4gp32, 0.5 µl RNase Inhibitor (Promega) , 0.5 µl reverse transcriptase (RevertAid, Fermentas), total volume 4 µl
* add ice cold RT-mix, mix well
* incubate 1h at 50°C with heated lid, inactivate 15' at 65°C, chill on ice;
For second strand synthesis:
SSS-mix: 15 µl 5x 2nd strand buffer (Invitrogen), 1.5 µl 10 mM dNTP-mix, 2.22 µl DNA Pol. I (9 U/µl) (Promega), 0.1 µl RNase H (10 U/µl) (Epicentre), 0.5 µl DNA Ligase (10 U/µl) (USB), 45.68 µl RNase-free water, total volume 65 µl
* add ice cold SSS-mix to RT reaction, mix well
* incubate 2h at 14-16°C in thermal cycler
* add 3.33 µl T4 DNA-Pol. (3 U/µl) (NEB) , mix by flicking
* incubate 15' at 14-16°C in thermal cycler, heat inactivate 10' at 70°C
* add 75 µl phenol/chlorof./isoamylalc. (pH 8), mix vigorously by pipetting
* transfer to prespun (30'' at 16000g) PLG Heavy 0.5 ml (Eppendorf)
* spin 5' at 16000g
* transfer aq. phase to prepared (resuspend, drain by gravity, 2' at 1000g) P-6 MicroSpin column (BioRad)
* spin 4' at 1000g, recover eluate
* transfer eluate to 0.5 ml PCR tube
* add LPA, 3.5 µl 5 M NaCl, 220 µl 100% Ethanol, mix well
* precipitate 60' at -80°C over night
* spin tube 30' at 16000g, 4°C, remove supernatant
* wash pellet with 500 µl 70% ethanol, spin tube 5' at 16000g, remove supernatant
* resuspend dried cDNA-pellet in 10 µl water
For In Vitro transcription:
IVT-mix (Promega): 6 µl RNase free water, 8 µl 5x T7 Transcr. buffer, 3 µl each 100 mM ATP, GTP, CTP, UTP, 4 µl T7 Enzyme-mix, total volume 30 µl
* add IVT-mix to 10 µl cDNA, mix by pipetting
* incubate 12h at 37°C
For aRNA cleanup:
RLT / ß-ME – mix: 3.5 µl ß-mercapto-ethanol, 76.5 µl water, 350 µl RLT (Qiagen RNeasy), total volume 430 µl
* add RLT / ß-ME-mix to IVT product (aRNA), mix well
* add 250 µl 100% ethanol, apply to RNeasy Mini column (Qiagen)
* spin 15" at 8000g, discard flow-through, transfer column to new 2 ml tube
* wash with 500 µl RPE, spin 15" at 8000g, discard flow-through
* add 30 µl RNase-free water onto membrane, spin 1' at 8000g,repeat eluting step,
* precipitate aRNA with 1 µl LPA, 30 µl 7.5M ammonium-acetate and 227.5 µl 100% ethanol
* wash pellet with 500 µl 70% ethanol, spin tube 5' at 16000g, remove supernatant
* resuspend in RNase-free water (? 200 µg/µl)
For 2nd Round RT:
RNA/primer-mix: 5 µl aRNA (1 µg), 1 µg N6-random-primer, total volume 6 µl
* RT-mix: 2 µl M-MuLV RT buffer (Fermentas), 0.5 µl 10 mM dNTP-mix, 0.5 µl T4gp32, 0.5 µl RNase Inhibitor (Promega) , 0.5 µl reverse transcriptase (RevertAid, Fermentas), total volume 4 µl
* denature RNA/primer-mix 5' at 70°C, snap cool on ice 2', hold 5' at room temperature, mix well
* incubate:
20' at 37°C
20' at 42°C
10' at 50°C
10' at 55°C
15' at 65°C
hold at 37°C
* add 1 µl RNase H-dilution (1:10 in 5x 2nd strand buffer, (Invitrogen))
* incubate 30' at 37°C, 2' at 95°C
For Klenow-labeling (per channel):
Klenow-mix (Invitrogen): 40 µl 2.5X Random Primer Solution, 10 µl 10X dNTP-(low dTTP)-mix*, 3 µl Cy-labeled dUTP (Cy3 or Cy5), 34 µl water, 2 µl Klenow fragment Exo- (40 U/µl), total volume 89 µl
* add to cDNA (11 µl), mix gently but thoroughly
* incubate 16h at 37°C
* add 450 µl TE to labeled cDNA, apply to Microcon YM-30 column (Millipore)
* spin 13’ at 16000g, discard flow-through
* wash with 430 µl TE (measure dye incorporation on NanoDrop), add 18.75 µl blocking-mix**,
* spin 13’ at rpm, discard flow-through
* elute labeled cDNA by inverting column into new tube, spin 8’ at 16000g
* 2 mM dATP, 2mM dCTP, 2mM dGTP, 0.5 mM dTTP
**12.5 µl Cot-1 DNA (1 µg/µl), 2.5 µl poly-A RNA (5 µg/µl), 3.75 µl tRNA (10 µg/µl)

extract_protocol: Extraction of total RNA was accomplished by (Stratagene). According to manufacturer’s specification an Absolutely RNA Kit, based on a glassfiber method in combination with guanidine thiocyanate buffer was used.

tag: Gender

value: pooled

tag: Tissue

value: Liver, hepatocyte; Testis, leydig cell; Kidney; Mammary gland; Embryo; Lung, alveolar macrophage; Skin, epidermis; Thymus, T lymphocyte; Meyloma, B lymphocyte; Soleuse muscle, myoblast; Embryo, fibroblast

source_name: mouse hepatocellular carcinoma, Mdr2-KO, IkB-SR

growth_protocol: All animal experiments were performed in accordance with the guidelines of the institutional committe for the use of animals for research in Israel

value: unknown

value: liver

tag: Tumor stage

value: carcinoma

tag: Genetic background

value: mixed


title: Subcutaneous adipose tissue from morbid obese patients male 5

description: No further additional informations

tag: Subcutaneous adipose tissue (SAT) from a pool of 3 non obese males. SAT aliquots (about 1 cm2 of fat, average 100-600 mg), not in contact with abdominal district, were excised during surgery. All biopsies were snap frozen and stored in liquid nitrogen until RNA isolation.


title: CI#84.2_replicate1

description: not applicable

source_name: Dppa4 hemizygous parental ES cell line

value: Dppa4 hemizygous parental ES cell line


title: CI#84.2_replicate3

description: not applicable


title: Subcutaneous adipose tissue from morbid obese patients male 1

description: No further additional informations

tag: Subcutaneous adipose tissue (SAT) from a obese male. SAT aliquot not in contact with abdominal district, were excised during surgery. The biopsy was snap frozen and stored in liquid nitrogen until RNA isolation.


title: CI#84.2_replicate2

description: not applicable


title: Subcutaneous adipose tissue from morbid obese patients male 3

description: No further additional informations


title: Subcutaneous adipose tissue from morbid obese patients male 4

description: No further additional informations


title: Liver_wildtype_Liver1 Cy3

description:
liver tissue from wildtype mice having the same genetic genetic background as the Mdr2-KO and combined Mdr2-KO, IkB-SR mice used in this experiment.
Wildtype liver tissue serves as a biological reference to build expression ratios with tumor tissue.

source_name: mouse liver, wildtype

extract_protocol: Total RNA was isolated from mouse skin using TRIzol (Invitrogen) following manufacturer’s protocol. (http://www.invitrogen.com/content.cfm?pageid=469)


title: Liver_wildtype_Liver1 Cy5


title: Hepatocellular carcinoma_Mdr2-/-,IkB-SR_HCC3 Cy5


title: Hepatocellular carcinoma_Mdr2-/-_HCC2 Cy5

description: Tumor model: mice carrying the Mdr2-/- allele spontaneously develop cholestatic hepatitis followed by dysplasia, dysplastic nodules and finally carcinoma (Mauad et al., Am J Pathol, 1994).

source_name: mouse hepatocellular carcinoma, Mdr2-KO


title: Hepatocellular carcinoma_Mdr2-/-,IkB-SR_HCC6 Cy5


title: Hepatocellular carcinoma_Mdr2-/-,IkB-SR_HCC8 Cy5


title: Hepatocellular carcinoma_Mdr2-/-,IkB-SR_HCC4 Cy5


title: Hepatocellular carcinoma_Mdr2-/-_HCC5 Cy5


title: Hepatocellular carcinoma_Mdr2-/-,IkB-SR_HCC9 Cy5


title: Hepatocellular carcinoma_Mdr2-/-_HCC7 Cy5


title: Hepatocellular carcinoma_Mdr2-/-,IkB-SR_HCC10 Cy3

hyb_protocol: Purified, dye-labelled cDNA was mixed with 120 μl UltraHyb hybridization buffer (Ambion), agitated for 30–60 min at 60°C, then for 10 min at 70°C on a thermo mixer and subsequently applied to pre-heated (60°C) microarrays mounted in a GeneTAC Hybridization Station (Genomic Solutions, Ann Arbor). Hybridizations were performed for 23 h at 42°C with gentle agitation. Thereafter, the arrays were automatically washed at 36°C with (i) 0.5× SSC, 0.1% (w/v) SDS for 5 min; (ii) 0.05× SSC, 0.1% (w/v) SDS for 3 min; and (iii) 0.05× SSC for 2 min. Flow time was set to 40 s. Immediately after the completion of the final washing step, the arrays were unmounted, immersed in 0.05× SSC, 0.1% (w/v) Tween 20 and dried by centrifugation in 50 ml Falcon tubes (30 s at 500, 1000 and 1500 r.p.m., respectively, followed by a final step of 240 s at 2000 r.p.m.).

scan_protocol: Hybridized microarrays were scanned in a two-color Agilent Scanner G25505B (Agilent, Santa Clara, CA) at 5 μm resolution and automatically adjusting PMT voltages according to manufactures specification. Recorded images were subsequently analyzed using GenePix Pro 6.0 software.

label_protocol:
For reverse transcription:
RNA/primer-mix: 2 µg total RNA, 500 ng (dT)-T7 primer, total volume 6 µl
* denature RNA/primer 4' at 70°C, chill on ice;
RT-mix: 2 µl M-MuLV RT buffer (Fermentas), 0.5 µl 10 mM dNTP-mix, 0.5 µl T4gp32, 0.5 µl RNase Inhibitor (Promega) , 0.5 µl reverse transcriptase (RevertAid, Fermentas), total volume 4 µl
* add ice cold RT-mix, mix well
* incubate 1h at 50°C with heated lid, inactivate 15' at 65°C, chill on ice;
For second strand synthesis:
SSS-mix: 15 µl 5x 2nd strand buffer (Invitrogen), 1.5 µl 10 mM dNTP-mix, 2.22 µl DNA Pol. I (9 U/µl) (Promega), 0.1 µl RNase H (10 U/µl) (Epicentre), 0.5 µl DNA Ligase (10 U/µl) (USB), 45.68 µl RNase-free water, total volume 65 µl
* add ice cold SSS-mix to RT reaction, mix well
* incubate 2h at 14-16°C in thermal cycler
* add 3.33 µl T4 DNA-Pol. (3 U/µl) (NEB) , mix by flicking
* incubate 15' at 14-16°C in thermal cycler, heat inactivate 10' at 70°C
* add 75 µl phenol/chlorof./isoamylalc. (pH 8), mix vigorously by pipetting
* transfer to prespun (30'' at 16000g) PLG Heavy 0.5 ml (Eppendorf)
* spin 5' at 16000g
* transfer aq. phase to prepared (resuspend, drain by gravity, 2' at 1000g) P-6 MicroSpin column (BioRad)
* spin 4' at 1000g, recover eluate
* transfer eluate to 0.5 ml PCR tube
* add LPA, 3.5 µl 5 M NaCl, 220 µl 100% Ethanol, mix well
* precipitate 60' at -80°C over night
* spin tube 30' at 16000g, 4°C, remove supernatant
* wash pellet with 500 µl 70% ethanol, spin tube 5' at 16000g, remove supernatant
* resuspend dried cDNA-pellet in 10 µl water
For In Vitro transcription:
IVT-mix (Promega): 6 µl RNase free water, 8 µl 5x T7 Transcr. buffer, 3 µl each 100 mM ATP, GTP, CTP, UTP, 4 µl T7 Enzyme-mix, total volume 30 µl
* add IVT-mix to 10 µl cDNA, mix by pipetting
* incubate 12h at 37°C
For aRNA cleanup:
RLT / β-ME – mix: 3.5 µl β-mercapto-ethanol, 76.5 µl water, 350 µl RLT (Qiagen RNeasy), total volume 430 µl
* add RLT / β-ME-mix to IVT product (aRNA), mix well
* add 250 µl 100% ethanol, apply to RNeasy Mini column (Qiagen)
* spin 15" at 8000g, discard flow-through, transfer column to new 2 ml tube
* wash with 500 µl RPE, spin 15" at 8000g, discard flow-through
* add 30 µl RNase-free water onto membrane, spin 1' at 8000g,repeat eluting step,
* precipitate aRNA with 1 µl LPA, 30 µl 7.5M ammonium-acetate and 227.5 µl 100% ethanol
* wash pellet with 500 µl 70% ethanol, spin tube 5' at 16000g, remove supernatant
* resuspend in RNase-free water ( 200 µg/µl)
For 2nd Round RT:
RNA/primer-mix: 5 µl aRNA (1 µg), 1 µg N6-random-primer, total volume 6 µl
* RT-mix: 2 µl M-MuLV RT buffer (Fermentas), 0.5 µl 10 mM dNTP-mix, 0.5 µl T4gp32, 0.5 µl RNase Inhibitor (Promega) , 0.5 µl reverse transcriptase (RevertAid, Fermentas), total volume 4 µl
* denature RNA/primer-mix 5' at 70°C, snap cool on ice 2', hold 5' at room temperature, mix well
* incubate:
20' at 37°C
20' at 42°C
10' at 50°C
10' at 55°C
15' at 65°C
hold at 37°C
* add 1 µl RNase H-dilution (1:10 in 5x 2nd strand buffer, (Invitrogen))
* incubate 30' at 37°C, 2' at 95°C
For Klenow-labeling (per channel):
Klenow-mix (Invitrogen): 40 µl 2.5X Random Primer Solution, 10 µl 10X dNTP-(low dTTP)-mix*, 3 µl Cy-labeled dUTP (Cy3 or Cy5), 34 µl water, 2 µl Klenow fragment Exo- (40 U/µl), total volume 89 µl
* add to cDNA (11 µl), mix gently but thoroughly
* incubate 16h at 37°C
* add 450 µl TE to labeled cDNA, apply to Microcon YM-30 column (Millipore)
* spin 13’ at 16000g, discard flow-through
* wash with 430 µl TE (measure dye incorporation on NanoDrop), add 18.75 µl blocking-mix**,
* spin 13’ at rpm, discard flow-through
* elute labeled cDNA by inverting column into new tube, spin 8’ at 16000g
* 2 mM dATP, 2mM dCTP, 2mM dGTP, 0.5 mM dTTP
**12.5 µl Cot-1 DNA (1 µg/µl), 2.5 µl poly-A RNA (5 µg/µl), 3.75 µl tRNA (10 µg/µl)


title: Hepatocellular carcinoma_Mdr2-/-_HCC1 Cy5


title: Hepatocellular carcinoma_Mdr2-/-,IkB-SR_HCC3 Cy3


title: Hepatocellular carcinoma_Mdr2-/-,IkB-SR_HCC6 Cy3


title: Hepatocellular carcinoma_Mdr2-/-_HCC7 Cy3


title: Hepatocellular carcinoma_Mdr2-/-,IkB-SR_HCC8 Cy3


title: Hepatocellular carcinoma_Mdr2-/-_HCC1 Cy3


title: Hepatocellular carcinoma_Mdr2-/-_HCC5 Cy3


title: CDR020

city: Montreal

first: Christian

middle: F

last: Deschepper

state: QC

address: 110 ave des Pins Ouest

institute: IRCM

description: replicate 4

hyb_protocol: Standard Illumina hybridization protocol

scan_protocol: Standard Illumina scanning protocol

data_processing: Variance was stabilized by log2 transformation, and data were normalised using quantile normalisation with lumi in R

source_name: cardiac left ventricle

label_protocol: Biotinylated probes were prepared from 50 ng of total RNA, using the Ambion Illumina TotalPrep RNA Amplification kit (Applied Biosystems, Streetsville, ON).

extract_protocol: Total RNA was extracted, using the RNeasy minikit (Qiagen Canada, Mississauga, ON), from cardiac left ventricles obtained from 12 week-old male mice.

tag: sex

value: male

tag: strain

value: C57BL/6J (or B)

tag: surgery

value: sham


title: Hepatocellular carcinoma_Mdr2-/-,IkB-SR_HCC9 Cy3

city: Stanford


title: CDR030

description: replicate 2

value: C57BL/6J-chrY<A> (or B.Y)

value: castration


title: CDR032

description: replicate 4


title: CDR023

description: replicate 3


title: CDR028

description: replicate 4


title: CDR031

description: replicate 3


title: CDR022

description: replicate 2


title: CDR024

description: replicate 4


title: CDR026

description: replicate 2


title: Hepatocellular carcinoma_Mdr2-/-_HCC2 Cy3


title: CDR029

description: replicate 1


title: CDR018

description: replicate 2


title: CDR025

description: replicate 1


title: CDR021

description: replicate 1


title: CDR017

description: replicate 1


title: Hepatocellular carcinoma_Mdr2-/-,IkB-SR_HCC4 Cy3


title: CDR019

description: replicate 3


title: CDR027

description: replicate 3


title: 143BTK- cell line, rho0, replicate C

city: Kansas City

first: Norman

last: Pavelka

state: MO

address: 1000 E. 50th Street

institute: Stowers Institute for Medical Research

description: Gene expression data from 143BTK- cells, depleted of mitochondrial DNA

hyb_protocol: Following fragmentation, cRNA was hybridized for 16 hr at 45C on GeneChip HG-U133A arrays. GeneChips were washed and stained in the Affymetrix Fluidics Station following the EukGE-WS2v4 protocol

scan_protocol: GeneChips were scanned using an Hewlett-Packard GeneArray Scanner

data_processing: Data were analyzed with Microarray Suite version 5.0 (MAS 5.0) using Affymetrix default analysis settings and global scaling as normalization method. The trimmed mean target intensity of each array was arbitrarily set to 100. Each subset relative to a particular cell line was then independently subjected to a quantile normalization in the R environment for statistical computing. Normalized data were finally rounded to the first decimal digit

source_name: 143BTK- cells, rho0

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 12 ug total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).

growth_protocol: Cells were grown for 3-4 weeks in Dulbecco’s modified Eagle medium (DMEM), high glucose (Euroclone), supplemented with 10% Fetal Calf Serum (FCS), 1mM NaPyruvate and 50ug/ml uridine

extract_protocol: Total RNA was isolated from 8 millions cells using the Qiagen RNeasy Mini kit (Qiagen)

treatment_protocol: To generate rho0 cells we added 50 ng/ml ethidium bromide (EthBr) to the growth medium, for a period of 3 weeks. No clone selection was performed but the cells were trypsinised and pooled together. One week after the removal of EthBr, absence of mtDNA was checked in rho0 cell lines by either PCR or by southern-blot

tag: mitochondria

value: depleted