title: LOXIMVI hg133b21

description:
Background Avg = 38.66
Background Max = 41.7
Background Min = 35.3
Background Stdev = 1.43
Central- Avg = 1224
Central- Count = 9
Corner+ Avg = 30
Corner+ Count = 32
Corner- Avg = 1160
Corner- Count = 32
Noise Avg = 1.98
Noise Max = 2.4
Noise Min = 1.6
Noise Stdev = 0.13
RawQ = 2.00
Columns = 712
Number of Cells = 506944
Rows = 712
OrganismPart: skin

source_name: amelanotic melanoma

value: amelanotic melanoma

value: malignant amelanotic melanoma


title: COLO205 hg133b31

status: Public on Mar 26 2007

type: RNA

city: Bethesda

first: Uma

middle: T

last: Shankavaram

state: MD

address: 9000 Rockville Pike

department: NCI

country: USA

institute: NIH

description:
Background Avg = 49.62
Background Max = 51.6
Background Min = 47.4
Background Stdev = 0.86
Central- Avg = 1395
Central- Count = 9
Corner+ Avg = 32
Corner+ Count = 32
Corner- Avg = 1217
Corner- Count = 32
Noise Avg = 1.97
Noise Max = 2.3
Noise Min = 1.7
Noise Stdev = 0.09
RawQ = 2.00
Columns = 712
Number of Cells = 506944
Rows = 712
OrganismPart: colon

scan_protocol: Affymetrix CEL analysis (Percentile); Protocol Type = feature_extraction; Parameter CellMargin = 2; Parameter OutlierHigh = 1.500; Parameter OutlierLow = 1.004; Parameter Percentile = 75; Software: Affymetrix MicroArraySuite, type: feature_extraction_software; Protocol Type = image_acquisition; Software: Affymetrix MicroArraySuite, type: image_acquisition_software; Hardware: ;

data_processing: ExpressionStat; Protocol Type = bioassay_data_transformation; Parameter Alpha1 = 0.05; Parameter Alpha2 = 0.065; Parameter BG = 4; Parameter Epsilon = 0.5; Parameter Gamma1H = 0.0045; Parameter Gamma1L = 0.0045; Parameter Gamma2H = 0.006; Parameter Gamma2L = 0.006; Parameter HZ = 4; Parameter NF = 1.000000000000; Parameter Perturbation = 1.1; Parameter SF = 1.000000000000; Parameter SmoothFactorBG = 100; Parameter Tau = 0.015; Parameter VZ = 4; Software: Affymetrix GeneChip Operating Software, type: bioassay_data_transformation_software;

label: biotin

molecule: total RNA

organism: Homo sapiens

source_name: adenocarcinoma

label_protocol: Affymetrix Labeling Protocol; The exact protocol is not available. But Standard Operating Proceadure can be found at          http://www.affymetrix.com/support/technical/manual/expression_manual.affx         or         http://www.affymetrix.com/Auth/support/downloads/manuals/expression_ever_manual.zip         Manual number: 701025 Revision 6, page 2.1.3; Protocol Type = labeling;

growth_protocol: NCI60Adherent; RNA harvesting protocol for adherent cells           Media used:         RPMI 1640 500 ml          FBS 25 ml -use the DTP serum if possible, from Bio Whittaker, not heat         inactivated          200 mM Glutamine 5 ml           1 flask = ~15 x 106 cells yields ~ 100 ugr RNA. Grow 10 flasks.                    Cells: start from growing cells from Frederick (not frozen). Started before at         passage #8-12. Do not use past passage 20.                    Growth schedule prior to harvest: Grow cells to         ~80 confluencey. Trypsinize cells with 5 ml try-EDTA per T162, 15 min., 370C. Pipet up and         down several times w 10 ml pipet to get good dispersment of cells. Count cells. Pass cells         to as many flasks as there are cells for. When passing cells, combine flasks into a single         pool. Pass 1x106 cells into each T162 w 30 ml media.  Repeat growth cycle until 10 flasks         are available.   Refeed Refeed cells the day prior to harvest without harvesting).  Draw off         media (wo cells). Add back media, 30 ml per T162. Add back to T-162âs. 37 deg C, ON  Harvest         Target confluency 80%  # of flasks = 10   Draw off media from 1st flask. Lyse cells in 15         ml lysis buffer (w 10 ul fresh BME per ml). Scrape cells. Draw off media from 2nd flask.         Pipet lysis buffer from flask 1 into flask 2. Repeat lysis with up to 4 T162âs. Pipet into         50 ml tube.  Repeat w next set of 4 flasks.  When done, vortex 10 sec.. Draw lysate thru a         20 guage needle 12xâs. Freeze at ö800C.  Purify using Quiagen Midi Kit.  Use a maximun of         100 x 106 cells per column. More will not bind to the column.                           Company Info         ------------           Quiagen Midi Kit Cat # 75144  RLT Buffer (lysis buffer) Cat. # 79216 $60-list, $51- discounted 1-800-362-7737                    Fetal Bovine Serum 500 ml, list $261- Cat #14-502F Lot # 9S083F Cambrex (old Bio Whittaker) 1-800-638-8174                   1x PBS pH 7.3-7.5 Bio Whittaker cat #17-516F $5.30 per 500 ml bottle (1-11)                    250 ml conical tubes Corning cat #25350-250                    RPMI-1640 wo L-glutamine cat # 12-167F  $13.25 per 500 ml bottle (for 1-11 bottles) Cambrex (old Bio Whittaker) Walkersville, Md. 21793 301-898-7025 1-800-638-8174                  Trypsin (0.05%)-EDTA (0.1%) In phosphate buffered saline without calcium and magnesium. cat # 118-087-061 $4.20 per 100 ml bottle Quality Biological, Inc. 301-840-9331                    L-glutamine 200 mM, 100x Gibco-BRL cat # 25030-149 20 ml 1-800-828-6686                    162 cm2 flask cat # 3150                    Costar Tissue Culture Cell Scrapper, 25 cm Sarstedt Cat. # 83.1830                    Cells are received from Nick Scudiero,  E-mail: SCUDIERO@dtpax2.ncifcrf.gov          Also trypsin, FBS and glutamine have been coming from there as well.; Protocol Type = grow;

extract_protocol: NCI60 Extraction Protocol; Harvest         Target confluency  80%            # of flasks = 10                  Draw off media from 1st flask.         Lyse cells in 15 ml lysis buffer (w 10 ul fresh BME per ml).         Scrape cells.         Draw off media from 2nd flask.         Pipet lysis buffer from flask 1 into flask 2.         Repeat lysis with up to 4 T162’s.         Pipet into 50 ml tube.                  Repeat w next set of 4 flasks.                  When done, vortex 10 sec..         Draw lysate thru a 20 guage needle 12x’s.         Freeze at –800C.                  Purify using Quiagen Midi Kit.          Use a maximun of 100 x 106 cells per column. More will not bind to the column.         ; Protocol Type = nucleic_acid_extraction;

tag: age

value: adenocarcinoma

tag: sex

value: male


title: Diseased 1

city: Catania

first: Sebastiano

last: Cavallaro

address: Viale Regina Margherita, 6

department: Institute of Neurological Sciences

country: Italy

institute: Italian National Research Council

description: Complementary RNAs (cRNAs) labelled with Cy5-CTP (Perkin-Elmer) were synthesized from 1 µg of total RNA of each sample using the Low RNA Input Fluorescent Linear Amplification Kit (Agilent Technologies) following the manufacturer’s protocol. A reference cRNA, labelled with Cy3-CTP (Perkin-Elmer), was synthesized from 1 µg of sample Diseased 2 RNA. Aliquots (750 ng) of Cy3 and Cy5 labelled cRNA targets were co-hybridized on Whole Human Genome Oligo Microarrays (Agilent Technologies). Microarray hybridization and washing were performed using reagents and instruments (hybridization chambers and rotating oven) as indicated by the manufacturer (Agilent Technologies). Microarrays were scanned at 10-μm resolution using a GenePix Personal 4100A microarray scanner and the GenePix Pro 6.0 acquisition and data-extraction software (Molecular Devices, Corp.).

data_processing: No Normalization

source_name: motor cortex, reference

tag: Sample ID

value: Diseased 2; Sample Type:tissue; Order Code:BTB3292 [Brain and Tissue Bank for Developmental Disorders (Miami, FL, USA)]; Chief Disease:ALS; Cause of Death:ALS; Age [years / days]:76 / 197; Gender:Male; Race:Caucasian; PMI [h]:24.

source_name: motor cortex, diseased 1

value: Diseased 1; Sample Type:tissue; Order Code:BTB3235 [Brain and Tissue Bank for Developmental Disorders (Miami, FL, USA)]; Chief Disease:ALS; Cause of Death:respiratory failure; Age [years / days]:73 / 106; Gender:Male; Race:Caucasian; PMI [h]:5.5.


title: Diseased 5

source_name: motor cortex, diseased 5

value: Diseased 5; Sample Type:tissue; Order Code:BTB3583 [Brain and Tissue Bank for Developmental Disorders (Miami, FL, USA)]; Chief Disease:ALS; Cause of Death:respiratory distress; Age [years / days]:52 / 54; Gender:Female; Race:Hispanic White; PMI [h]:20.5.


title: CCRF-CEM hg133a21

description:
Background Avg = 53.75
Background Max = 63.7
Background Min = 48.1
Background Stdev = 3.31
Central- Avg = 778
Central- Count = 9
Corner+ Avg = 29
Corner+ Count = 32
Corner- Avg = 1035
Corner- Count = 32
Noise Avg = 1.82
Noise Max = 2.7
Noise Min = 1.5
Noise Stdev = 0.18
RawQ = 2.01
Columns = 712
Number of Cells = 506944
Rows = 712
OrganismPart: blood

source_name: lymphoblastic leukemia

growth_protocol: NCI60Suspended; RNA harvesting protocol for suspended cells:           Media used:          RPMI 1640 500 ml          FBS 25 ml          200 mM Glutamine 5 ml                    Cells:          start from growing cells from Frederick (not frozen). Started before at passage #8-12. Do not use past passage 20. Grow 10         flasks. 1 flask = ~15 x 106 cells yields ~ 100 ugr RNA.  Growth schedule prior to harvest         Count cells as necessary to follow growth. Grow cells to ~0.52 x 106 cells /ml  Pass 1x106         cells into each T162 w 30 ml media. Pass cells to as many flasks as there are cells for.         When passing cells, combine into a single pool.  Repeat growth cycle until 10 flasks are         available.  Refeed Refeed cells the day prior to harvest (without passsing). To do so: Spin         down cells 1.7k rpm, 10 min., in a 50 ml tube. Draw off media. Resuspend in 30 ml media.         Return to flask.  Harvest Harvest at 0.52 x 106cells per ml when 10 flasks are available.         Spin  4 flasks of cells at  1.7k rpm (~ 500 x g), 10 min (speed setting at 3.5), in a 250 ml         centrifuge tube.   Draw off media, leaving ~0.5 ml behind. Flick centirifuge tube to break         up cells. Lyse cells in 15 ml lysis buffer (w 10 ul fresh BME per ml) per 4 T162âs.  Vortex         10 sec. Draw thru a 20 guage needle 12xâs. Freeze at ö800C.   Purify using Quigen Midi         Kit.Use a maximum of 100 x 106 cells per column. More will not bind to the column.                  Company Info:           Fetal bovine serum Do not heat inactivate. Bio Whittaker Cat# 14-501F Lot #9S083F 301-898-7025                    Quiagen Midi Kit Cat # 75144                    DTT Fluka or Gallard  1x PBS pH 7.3-7.5                  Bio Whittaker cat #17-516F $5.30 per 500 ml bottle (1-11)                    250 ml conical centrifuge tubes Corning cat # 25350-250                    RPMI-1640 wo L-glutamine cat # 12-167F  $13.25 per 500 ml bottle         (for 1-11 bottles) Cambrex (old Bio Whittaker) Walkersville, Md. 21793 301-898-7025         1-800-638-8174                    Trypsin (0.05%)-EDTA (0.1%) In phosphate buffered saline without calcium and         magnesium. cat # 118-087-061 Quality Biological, Inc. 301-840-9331                    L-glutamine 200 mM, 100x Gibco-BRL cat # 25030-081  162 cm2 flask cat # 3150                   Costar Tissue Culture Cell Scrapper, 25cm Sarstedt Cat. # 83.1830                    Cells are recieved from Nick Scudiero,  E-mail:SCUDIERO@dtpax2.ncifcrf.gov          Also trypsin, FBS and glutamine have been coming from there as well.; Protocol Type = grow;

value: lymphoblastic leukemia

value: acute lymphoblastic leukemia

value: female


title: Control 9

source_name: motor cortex, control 9

value: Control 9; Sample Type:RNA; Order Code:6782 [Ambion, Inc. (Austin, TX, USA)]; Chief Disease:thrombosis and coronary artherosclerosis; Cause of Death:heart attack; Age [years / days]:68 / -; Gender:Male; Race:Caucasian; PMI [h]:n/a.


title: Control 7

source_name: motor cortex, control 7

value: Control 7; Sample Type:tissue; Order Code:HCtEU [Brain and Tissue Bank for Developmental Disorders (Miami, FL, USA)]; Chief Disease:coronary artery disease; Cause of Death:n/a; Age [years / days]:79 / -; Gender:Female; Race:Caucasian; PMI [h]:7.5.


title: Control 8

source_name: motor cortex, control 8

value: Control 8; Sample Type:tissue; Order Code:HCtFM [Brain and Tissue Bank for Developmental Disorders (Miami, FL, USA)]; Chief Disease:n/a; Cause of Death:n/a; Age [years / days]:70 / -; Gender:Female; Race:Caucasian; PMI [h]:8.5.


title: ACHN hg133b21

description:
Background Avg = 47.14
Background Max = 54.8
Background Min = 43.9
Background Stdev = 1.87
Central- Avg = 1734
Central- Count = 9
Corner+ Avg = 50
Corner+ Count = 32
Corner- Avg = 1744
Corner- Count = 32
Noise Avg = 2.28
Noise Max = 2.6
Noise Min = 2.0
Noise Stdev = 0.11
RawQ = 2.23
Columns = 712
Number of Cells = 506944
Rows = 712
OrganismPart: kidney

source_name: renal cell carcinoma

value: renal cell carcinoma

value: renal cell carcinoma-poor or moderately differentiated


title: Diseased 11

source_name: motor cortex, diseased 11

value: Diseased 11; Sample Type:RNA; Order Code:6166 [Ambion, Inc. (Austin, TX, USA)]; Chief Disease:ALS; Cause of Death:respiratory failure; Age:[years / days]:70 / -; Gender:Male; Race:Caucasian; PMI [h]:n/a.


title: Control 6

source_name: motor cortex, control 6

value: Control 6; Sample Type:tissue; Order Code:HCtAV [Brain and Tissue Bank for Developmental Disorders (Miami, FL, USA)]; Chief Disease:n/a; Cause of Death:n/a; Age [years / days]:84 / -; Gender:Female; Race:Caucasian; PMI [h]:2.75.


title: OVCAR-4 hg133b21

description:
Background Avg = 43.19
Background Max = 45.2
Background Min = 40.9
Background Stdev = 1.02
Central- Avg = 1651
Central- Count = 9
Corner+ Avg = 35
Corner+ Count = 32
Corner- Avg = 1685
Corner- Count = 32
Noise Avg = 2.25
Noise Max = 2.7
Noise Min = 2.0
Noise Stdev = 0.11
RawQ = 2.06
Columns = 712
Number of Cells = 506944
Rows = 712
OrganismPart: ovary


title: HCT-116 hg133b21

description:
Background Avg = 42.18
Background Max = 43.2
Background Min = 41.2
Background Stdev = 0.35
Central- Avg = 1441
Central- Count = 9
Corner+ Avg = 33
Corner+ Count = 32
Corner- Avg = 1441
Corner- Count = 32
Noise Avg = 1.67
Noise Max = 1.9
Noise Min = 1.5
Noise Stdev = 0.06
RawQ = 1.78
Columns = 712
Number of Cells = 506944
Rows = 712
OrganismPart: colon

source_name: carcinoma very poorly differentiated

value: unknown

value: carcinoma very poorly differentiated

value: carcinoma


title: HL-60 hg133b21

description:
Background Avg = 47.03
Background Max = 49.5
Background Min = 45.2
Background Stdev = 0.83
Central- Avg = 2158
Central- Count = 9
Corner+ Avg = 44
Corner+ Count = 32
Corner- Avg = 2576
Corner- Count = 32
Noise Avg = 1.51
Noise Max = 1.7
Noise Min = 1.4
Noise Stdev = 0.06
RawQ = 1.84
Columns = 712
Number of Cells = 506944
Rows = 712
OrganismPart: blood

source_name: promyelocytic leukemia

value: promyelocytic leukemia

value: acute promyelocytic leukemia

value: male


title: Control 4

value: Diseased 2; Sample Type:tissue; Order Code:BTB3292 [Brain and Tissue Bank for Developmental Disorders (Miami, FL, USA)]; Chief Disease:ALS; Cause of Death:ALS; Age [years / days]:76 / 197; Gender:Male; Race:Caucasian; PMI [h]:24.

source_name: motor cortex, control 4

value: Control 4; Sample Type:tissue; Order Code:BTB3688 [Brain and Tissue Bank for Developmental Disorders (Miami, FL, USA)]; Chief Disease:ascending colon adenocarcinoma; Cause of Death:ascending colon adenocarcinoma; Age [years / days]:60 / -; Gender:Male; Race:Hispanic White; PMI [h]:17.


title: Control 5

source_name: motor cortex, control 5

value: Control 5; Sample Type:tissue; Order Code:HCtAN [Brain and Tissue Bank for Developmental Disorders (Miami, FL, USA)]; Chief Disease:n/a; Cause of Death:n/a; Age:[years / days]:72 / -; Gender:Male; Race:Caucasian; PMI [h]:11.


title: MDA-MB-231/ATCC hg133b21

description:
Background Avg = 49.82
Background Max = 58.6
Background Min = 44.5
Background Stdev = 3.05
Central- Avg = 6256
Central- Count = 9
Corner+ Avg = 78
Corner+ Count = 32
Corner- Avg = 5963
Corner- Count = 32
Noise Avg = 2.33
Noise Max = 2.8
Noise Min = 1.9
Noise Stdev = 0.14
RawQ = 2.15
Columns = 712
Number of Cells = 506944
Rows = 712
OrganismPart: breast

source_name: breast epithelial adenocarcinoma

value: breast epithelial adenocarcinoma

value: adenocarcinoma

value: female


title: Control 2

source_name: motor cortex, control 2

value: Control 2; Sample Type:tissue; Order Code:BTB3576 [Brain and Tissue Bank for Developmental Disorders (Miami, FL, USA)]; Chief Disease:hypertension; Cause of Death:ruptured abdominal aortic aneurism; Age [years / days]:59 / 260; Gender:Male; Race:Caucasian; PMI [h]:15.


title: UACC-62 hg133b21

description:
Background Avg = 44.93
Background Max = 46.3
Background Min = 43.0
Background Stdev = 0.64
Central- Avg = 883
Central- Count = 9
Corner+ Avg = 24
Corner+ Count = 32
Corner- Avg = 799
Corner- Count = 32
Noise Avg = 1.85
Noise Max = 2.0
Noise Min = 1.7
Noise Stdev = 0.07
RawQ = 2.08
Columns = 712
Number of Cells = 506944
Rows = 712
OrganismPart: skin

source_name: melanoma

value: melanoma


title: Control 3

source_name: motor cortex, reference

source_name: motor cortex, control 3

value: Control 3; Sample Type:tissue; Order Code:BTB3577 [Brain and Tissue Bank for Developmental Disorders (Miami, FL, USA)]; Chief Disease:n/a; Cause of Death:n/a; Age [years / days]:59 / 118; Gender:Male; Race:Caucasian; PMI [h]:21.


title: Wt_larvae_rep2

status: Public on Mar 22 2007

city: Umea

first: Jan

last: Larsson

address: Umea University

department: Molecular Biology

country: Sweden

institute: Umea University

description: Gene expression data from whole 1st instar larvae (Wt and pof mutant).

data_processing: Raw data was normalized using RMA (R and Bioconductor)

label: Biotin

molecule: polyA RNA

organism: Drosophila melanogaster

source_name: Wt 1st instar larvae

tag: Gender

value: mixed

tag: Tissue


title: Wt_larvae_rep3


title: Mutant_larvae_rep4

source_name: pof mutant 1st instar larvae


title: Control 1

data_processing: No normalization

source_name: motor cortex, control 1

value: Control 1; Sample Type:tissue; Order Code:BTB3428 [Brain and Tissue Bank for Developmental Disorders (Miami, FL, USA)]; Chief Disease:drowning; Cause of Death:drowning; Age [years / days]:56 / 275; Gender:Female; Race:Caucasian; PMI [h]:17.


title: Mutant_larvae_rep2


title: Genotype RL4452 developing seed at 5 days after anthesis, biological rep1 location SB

status: Public on Mar 25 2007

city: Winnipeg

first: Mark

last: Jordan

state: Manitoba

address: 195 Dafoe Rd.

department: Cereal research Centre

country: Canada

institute: Agriculture and Agri-Food Canada

description: Gene expression data from field grown developing seeds 5 days after anthesis

hyb_protocol: Following fragmentation, the labelled samples were mixed with 0.1 mg/ml sonicated herring sperm DNA in hybridization buffer (100mM 2-N-morpholino-ethane-sulphonic acid (MES), 1 M NaCL, 20mM EDTA, 0.01% Tween 20, 0.5 mg/ml BSA, 10% DMSO), denatured at 99°C for 5 min then added to GeneChip Wheat Genome Array (Affymetrix) according to manufacturer instructions and hybridized for 16 h at 45°C in a rotisserie oven at 60 rpm. Following hybridization the arrays were washed and stained in the Affymetrix fluidics station 450 according to standard protocol (Section 2.3.3 of the Expression Analysis Technical Manual

scan_protocol: GeneChips were scanned using the Affymetrix scanner 3000.

data_processing: The raw data in the Affymetrix CEL files of all 4 parental arrays representing two biological replicates of each of two parental genotypes were subjected to quantile normalization using Robust Multi-Array Average (RMA) (Bolstad, B.M., Irizarry, R.A., Astrand, M., Speed, T.P. 2003. A comparison of normalization methods for high density oligonucleotide array data based on variance and bias. Bioinformatics 19:185-193) followed by normalization to the median.

organism: Triticum aestivum

source_name: developing seed collected from field grown material 5 days after anthesis

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol (Expression Analysis Technical Manual, Affymetrix). mRNA was used for first and second strand cDNA synthesis using the One Cycle cDNA Synthesis Kit (Affymetrix) following the manufacturers instructions. The resulting cDNA was purified using the GeneChip Sample Cleanup Module (Affymetrix) according to instructions and then used to make purified biotin labelled cRNA using the GeneChip IVT labelling kit and the GeneChip Sample Cleanup Module (Affymetrix). The resulting labelled cRNA was quantified and the yield and size distribution of labelled fragments checked by electrophoresis using the Agilent Bioanalyzer. The labelled cRNA was fragmented using the GeneChip Sample Cleanup Module (Affymetrix) and stored at -20°C.

growth_protocol: The 39 lines and parental lines were grown as 1.47 m rows in three replications in a 7x7 (49 entry) lattice design at Winnipeg, Manitoba in 2004. Each replicate had a unique randomization

extract_protocol: Total RNA was extracted from frozen seed using a modification of the protocol described at Montana State University Plant Sciences website. The content of a 2 mL tube full of frozen seeds was ground to a fine powder in liquid nitrogen using a mortar and pestle. Ground seed was transferred to 50 mL Oak Ridge tubes and 7.5 mL of extraction buffer was added. [50 mL Extraction Buffer contained 35 mL nuclease-free water, 2.5 mL 1M Tris, pH 9.0. 5 mL 2M NaCl, 5 mL 10% Sarcosyl, 2 mL 0.5M EDTA,  pH 8.0, 250 μL 1M DTT]. After vortexing well, 3.75 mL phenol and 3.75 mL chloroform/isoamyl alcohol (49:2) were added and samples vortexed again. Samples were centrifuged 10 min at 13,000 rpm in a Sorvall model RC5C at 4°C using an SS-34 rotor. A total of 7.5 mL of the upper aqueous phase was transferred to a clean Oak Ridge tube, 15 mL Trizol was added and the sample was vortexed. A volume of 3 mL of choloroform was added and the sample was vortexed again. Samples were centrifuged 10 min at 13,000 rpm in the Sorvall at 4°C. A volume of 15 mL of the upper aqueous phase was transferred to a fresh Oak Ridge tube and 10 mL of chloroform was added and the sample was vortexed. Samples were centrifuged again in the Sorvall at 13,000 rpm for 10 min at 4°C. A volume of 12 mL of the upper phase was transferred to a fresh Oak Ridge tube and the RNA precipitated by adding 1,200 uL of 3M sodium acetate and 24 mL of absolute ethanol. Samples were left at -80°C overnight then centrifuged at 14,000 rpm at 4°C in the Sorvall for 20-30 minutes to precipitate the RNA. The pellet was rinsed with 70% ethanol, lightly dried and re-suspended in 1 mL of nuclease-free water. RNA concentration was quantified using a spectrophotometer. All solutions and equipment used in this procedure were RNAse-free. Messenger RNA was enriched from total RNA using the Poly(A) Purist MAG kit  (Ambion) according to the protocol provided by the manufacturer. Messenger RNA was checked for quality and quantified by micro-chromatography using an Agilent 2000 Bioanalyzer and the RNA 6000 Nano Assay Kit both purchased from Agilent Technologies . Quantification was carried out according to the instructions provided with the kit.

treatment_protocol: spikes were tagged when anthers were visible, enough developing seeds to fill a 2mL tube were isolated from each spike 5 days after tagging and flash frozen in liquid nitrogen

tag: Genotype

value: RL4452, Age:developing seed 5 days after anthesis


title: Mutant_larvae_rep3


title: Mutant_larvae_rep1


title: mouse brain ipsilateral LPS.RU486, biological rep5

city: Buenos Aires

first: Ariel

last: Chernomoretz

address: Ciudad Universitaria Pab I

department: Physics Department

country: Argentina

institute: University of Buenos Aires

description: mouse brain ipsilateral LPS.RU486

hyb_protocol: cRNA probes were hybridized to an MOE430A Genechip (Affymetrix, Santa Clara, CA). Fifteen micrograms of fragmented cRNA was hybridized for 16 h at 45°C with constant rotation (60 rpm). Microarrays were processed in an Affymetrix GeneChip Fluidic Station 400 protocol EukGE-WS2v4. Staining was made with streptavidin-conjugated phycoerythrin (SAPE) followed by amplification with a biotinylated anti-streptavidin antibody and a second round of SAPE.

scan_protocol: GeneChips were scanned using the Agilent GeneArray Scanner (Agilent Technologies) ; Pixel Size : 2.5 ; Filter : 570 ;Scanner Type M10.

data_processing: The data were analyzed with the RMA summarization algorithm as implemented in Bioconductors package: affy, with deault analysis settings.

organism: Mus musculus

source_name: mouse brain ipsilateral LPS.RU486

label_protocol: 10 micrograms of total RNA was used for cDNA synthesis. Converted to cDNA with SuperScript reverse transcriptase (Gibco-BRL by Invitrogen), using T7-oligo-d(T)24 as a primer. Second-strand synthesis was performed using T4 DNA polymerase and E. Coli DNA ligase followed by blunt ending by T4 polynucleotide kinase. cDNA was isolated by phenol-chloroform extraction using phase lock gels (Brinkmann). cDNA was in vitro transcribed using the T7 BioArray High Yield RNA Transcript Labeling Kit (Enzo Biochem, New York, N.Y.) to produce biotinylated cRNA. Labelled cRNA was isolated using an RNeasy Mini Kit column (Qiagen). Purified cRNA was fragmented to 200–300 mer cRNA using a fragmentation buffer (100 mM potassium acetate-30 mM magnesium acetate-40 mM Tris-acetate, pH 8.1), for 35 min at 94°C. The quality of total RNA, cDNA synthesis, cRNA amplification and cRNA fragmentation was monitored by micro-capillary electrophoresis (Bioanalizer 2100 by Bioanalyser 2100, Agilent Technologies).

extract_protocol: The RNA was isolated using Trizol reagent (Invitrogen) following manufacture’s protocol.

treatment_protocol: C57Bl/6 mouse received an i.p. injection of vehicle (RU486 – 50 mg/kg in 50 microliters) and were submitted to surgery 4 h later. The mice receiving intraparenchymal injections were anesthetized with a mixture of ketamine and xylazine and placed in a stereotaxic apparatus (David Kopf Instruments). The right caudate putamen was reached, using a small cannula (28 gauge; Plastic One) at the coordinates 0.0 mm anteroposterior, -2.0 mm lateral, and -3.0 mm dorsoventral according to a mouse brain atlas [Paxino, G., and Franklin, K. B. J. (2001). The mouse brain in stereotaxic coordinates., 2nd ed (San Diego: Academic Press)]. The animal received an infusion of LPS (from Eschericia coli; serotype O55:B5; Sigma L2880 - 2.5 micrograms - 1 microliter) over 2 min by means of a microinjection 18 pump. Animal was killed 12 h after the intracerebral infusion. The mouse was anesthetized under isofluorane and blood was drawn via cardiac puncture before head decapitation. The brain was removed rapidly from the skull and placed in cold phosphate buffered saline (PBS) solution. A brain region limited at plane anteroposterior +1.5 to -1.5 and dorsoventral -4.0 was dissected, separated in ipsilateral side and quickly immersed in liquid nitrogen. The tissue was stored at -80 oC until RNA extraction was performed.

tag: Age

value: 60 days

value: brain

tag: Brain dissection

value: limited at plane anteroposterior +1.5 to -1.5 and dorsoventral -4.0 (Paxino and Franklin, 2001)


title: M14 hg133b21

description:
Background Avg = 36.85
Background Max = 38.5
Background Min = 35.5
Background Stdev = 0.64
Central- Avg = 2196
Central- Count = 9
Corner+ Avg = 28
Corner+ Count = 32
Corner- Avg = 2029
Corner- Count = 32
Noise Avg = 1.36
Noise Max = 1.6
Noise Min = 1.2
Noise Stdev = 0.05
RawQ = 1.81
Columns = 712
Number of Cells = 506944
Rows = 712
OrganismPart: skin

source_name: melanoma-melanotic

value: unknown

value: melanoma-melanotic


title: Genotype Domain developing seed at 5 days after anthesis, biological rep3 location SB

data_processing: The raw data in the Affymetrix CEL files of all 4 parental arrays representing two biological replicates of each of 2 parental genotypes were subjected to quantile normalization using Robust Multi-Array Average (RMA) (Bolstad, B.M., Irizarry, R.A., Astrand, M., Speed, T.P. 2003. A comparison of normalization methods for high density oligonucleotide array data based on variance and bias. Bioinformatics 19:185-193) followed by normalization to the median.

value: Domain, Age:developing seed 5 days after anthesis,


title: Genotype Domain developing seed at 5 days after anthesis, biological rep1 location SB

value: Domain,  Age:developing seed 5 days after anthesis,


title: mouse brain ipsilateral saline.RU486, biological rep1

description: mouse brain ipsilateral RU486

source_name: mouse brain ipsilateral RU486

treatment_protocol: C57Bl/6 mouse received an i.p. injection of vehicle (RU486 – 50mg/kg in 50 microliters) and were submitted to surgery 4 h later. The mice receiving intraparenchymal injections were anesthetized with a mixture of ketamine and xylazine and placed in a stereotaxic apparatus (David Kopf Instruments). The right caudate putamen was reached, using a small cannula (28 gauge; Plastic One) at the coordinates 0.0 mm anteroposterior, -2.0 mm lateral, and -3.0 mm dorsoventral according to a mouse brain atlas [Paxino, G., and Franklin, K. B. J. (2001). The mouse brain in stereotaxic coordinates., 2nd ed (San Diego: Academic Press)]. The animal received an infusion of sterile pyrogen-free saline (1 microliter) over 2 min by means of a microinjection 18 pump. Animal was killed 12 h after the intracerebral infusion. The mouse was anesthetized under isofluorane and blood was drawn via cardiac puncture before head decapitation. The brain was removed rapidly from the skull and placed in cold phosphate buffered saline (PBS) solution. A brain region limited at plane anteroposterior +1.5 to -1.5 and dorsoventral -4.0 was dissected, separated in ipsilateral side and quickly immersed in liquid nitrogen. The tissue was stored at -80 oC until RNA extraction was performed.


title: mouse brain ipsilateral LPS.RU486, biological rep4

description: mouse brain ipsilateral LPS.RU486


title: mouse brain ipsilateral saline.RU486, biological rep3

description: mouse brain ipsilateral RU486


title: mouse brain ipsilateral saline.RU486, biological rep2

description: mouse brain ipsilateral RU486


title: mouse brain ipsilateral LPS.RU486, biological rep2

description: mouse brain ipsilateral LPS.RU486


title: mouse brain ipsilateral saline.RU486, biological rep4

description: mouse brain ipsilateral RU486


title: mouse brain ipsilateral LPS.RU486, biological rep1

description: mouse brain ipsilateral LPS.RU486


title: mouse brain ipsilateral LPS.RU486, biological rep3

description: mouse brain ipsilateral LPS.RU486


title: mouse brain ipsilateral LPS.Dmso, biological rep5

description: mouse brain ipsilateral LPS

source_name: mouse brain ipsilateral LPS

growth_protocol: Before and after intracerebral infusion the mouse was kept under standard contidions: 12 h :12 h light : dark cycle; free access to mouse chow; one mouse per cage.

treatment_protocol: C57Bl/6 mouse received an i.p. injection of vehicle (DMSO – 50 microliters) and were submitted to surgery 4 h later. The mice receiving intraparenchymal injections were anesthetized with a mixture of ketamine and xylazine and placed in a stereotaxic apparatus (David Kopf Instruments). The right caudate putamen was reached, using a small cannula (28 gauge; Plastic One) at the coordinates 0.0 mm anteroposterior, -2.0 mm lateral, and -3.0 mm dorsoventral according to a mouse brain atlas [Paxino, G., and Franklin, K. B. J. (2001). The mouse brain in stereotaxic coordinates., 2nd ed (San Diego: Academic Press)]. The animal received an infusion of LPS (from Eschericia coli; serotype O55:B5; Sigma L2880 - 2.5 micrograms - 1 microliter) over 2 min by means of a microinjection 18 pump. Animal was killed 12 h after the intracerebral infusion. The mouse was anesthetized under isofluorane and blood was drawn via cardiac puncture before head decapitation. The brain was removed rapidly from the skull and placed in cold phosphate buffered saline (PBS) solution. A brain region limited at plane anteroposterior +1.5 to -1.5 and dorsoventral -4.0 was dissected, separated in ipsilateral side and quickly immersed in liquid nitrogen. The tissue was stored at -80 oC until RNA extraction was performed.


title: mouse brain ipsilateral LPS.Dmso, biological rep3

description: mouse brain ipsilateral LPS


title: Genotype RL4452 developing seed at 5 days after anthesis, biological rep3 location SB


title: MDA-N hg133b21

description:
Background Avg = 38.75
Background Max = 39.9
Background Min = 37.0
Background Stdev = 0.47
Central- Avg = 4061
Central- Count = 9
Corner+ Avg = 37
Corner+ Count = 32
Corner- Avg = 3920
Corner- Count = 32
Noise Avg = 1.34
Noise Max = 1.6
Noise Min = 1.2
Noise Stdev = 0.05
RawQ = 1.70
Columns = 712
Number of Cells = 506944
Rows = 712
OrganismPart: skin

source_name: MDA-N was derived from MDA-MB435 by transfection with a plasmid designed to express erbB2. MDA-N and MDA-MB435 are very similar under non selective conditions.


title: mouse brain ipsilateral LPS.Dmso, biological rep1

description: mouse brain ipsilateral LPS


title: mouse brain ipsilateral LPS.Dmso, biological rep4

description: mouse brain ipsilateral LPS


title: mouse brain ipsilateral control, biological rep1

description: mouse brain ipsilateral control

source_name: mouse brain ipsilateral control

treatment_protocol: C57Bl/6 mouse received an i.p. injection of vehicle (DMSO - 50 microliters) and were submitted to surgery 4 h later. The mice receiving intraparenchymal injections were anesthetized with a mixture of ketamine and xylazine and placed in a stereotaxic apparatus (David Kopf Instruments). The right caudate putamen was reached, using a small cannula (28 gauge; Plastic One) at the coordinates 0.0 mm anteroposterior, -2.0 mm lateral, and -3.0 mm dorsoventral according to a mouse brain atlas [Paxino, G., and Franklin, K. B. J. (2001). The mouse brain in stereotaxic coordinates., 2nd ed (San Diego: Academic Press)]. The animal received an infusion of sterile pyrogen-free saline (1 microliter) over 2 min by means of a microinjection 18 pump. Animal was killed 12 h after the intracerebral infusion. The mouse was anesthetized under isofluorane and blood was drawn via cardiac puncture before head decapitation. The brain was removed rapidly from the skull and placed in cold phosphate buffered saline (PBS) solution. A brain region limited at plane anteroposterior +1.5 to -1.5 and dorsoventral -4.0 was dissected, separated in ipsilateral side and quickly immersed in liquid nitrogen. The tissue was stored at -80 oC until RNA extraction was performed.


title: mouse brain ipsilateral control, biological rep5

description: mouse brain ipsilateral control


title: Wt_larvae_rep1


title: mouse brain contralateral saline.RU486, biological rep1

description: mouse brain contralateral RU486

source_name: mouse brain contralateral RU486

treatment_protocol: C57Bl/6 mouse received an i.p. injection of vehicle (RU486 – 50mg/kg in 50 microliters) and were submitted to surgery 4 h later. The mice receiving intraparenchymal injections were anesthetized with a mixture of ketamine and xylazine and placed in a stereotaxic apparatus (David Kopf Instruments). The right caudate putamen was reached, using a small cannula (28 gauge; Plastic One) at the coordinates 0.0 mm anteroposterior, -2.0 mm lateral, and -3.0 mm dorsoventral according to a mouse brain atlas [Paxino, G., and Franklin, K. B. J. (2001). The mouse brain in stereotaxic coordinates., 2nd ed (San Diego: Academic Press)]. The animal received an infusion of sterile pyrogen-free saline (1 microliter) over 2 min by means of a microinjection 18 pump. Animal was killed 12 h after the intracerebral infusion. The mouse was anesthetized under isofluorane and blood was drawn via cardiac puncture before head decapitation. The brain was removed rapidly from the skull and placed in cold phosphate buffered saline (PBS) solution. A brain region limited at plane anteroposterior +1.5 to -1.5 and dorsoventral -4.0 was dissected, separated in contralateral side and quickly immersed in liquid nitrogen. The tissue was stored at -80 oC until RNA extraction was performed.


title: mouse brain ipsilateral control, biological rep3

description: mouse brain ipsilateral control


title: mouse brain contralateral LPS.RU486, biological rep3

description: mouse brain contralateral LPS.RU486

source_name: mouse brain contralateral LPS.RU486

treatment_protocol: C57Bl/6 mouse received an i.p. injection of vehicle (RU486 – 50 mg/kg in 50 microliters) and were submitted to surgery 4 h later. The mice receiving intraparenchymal injections were anesthetized with a mixture of ketamine and xylazine and placed in a stereotaxic apparatus (David Kopf Instruments). The right caudate putamen was reached, using a small cannula (28 gauge; Plastic One) at the coordinates 0.0 mm anteroposterior, -2.0 mm lateral, and -3.0 mm dorsoventral according to a mouse brain atlas [Paxino, G., and Franklin, K. B. J. (2001). The mouse brain in stereotaxic coordinates., 2nd ed (San Diego: Academic Press)]. The animal received an infusion of LPS (from Eschericia coli; serotype O55:B5; Sigma L2880 - 2.5 micrograms - 1 microliter) over 2 min by means of a microinjection 18 pump. Animal was killed 12 h after the intracerebral infusion. The mouse was anesthetized under isofluorane and blood was drawn via cardiac puncture before head decapitation. The brain was removed rapidly from the skull and placed in cold phosphate buffered saline (PBS) solution. A brain region limited at plane anteroposterior +1.5 to -1.5 and dorsoventral -4.0 was dissected, separated in contralateral side and quickly immersed in liquid nitrogen. The tissue was stored at -80 oC until RNA extraction was performed.


title: mouse brain ipsilateral LPS.Dmso, biological rep2

description: mouse brain ipsilateral LPS


title: mouse brain contralateral saline.RU486, biological rep2

description: mouse brain contralateral RU486


title: mouse brain ipsilateral control, biological rep2

description: mouse brain ipsilateral control


title: mouse brain contralateral LPS.RU486, biological rep4

description: mouse brain contralateral LPS.RU486


title: mouse brain contralateral LPS.Dmso, biological rep6

description: mouse brain contralateral LPS

source_name: mouse brain contralateral LPS

treatment_protocol: C57Bl/6 mouse received an i.p. injection of vehicle (DMSO – 50 microliters) and were submitted to surgery 4 h later. The mice receiving intraparenchymal injections were anesthetized with a mixture of ketamine and xylazine and placed in a stereotaxic apparatus (David Kopf Instruments). The right caudate putamen was reached, using a small cannula (28 gauge; Plastic One) at the coordinates 0.0 mm anteroposterior, -2.0 mm lateral, and -3.0 mm dorsoventral according to a mouse brain atlas [Paxino, G., and Franklin, K. B. J. (2001). The mouse brain in stereotaxic coordinates., 2nd ed (San Diego: Academic Press)]. The animal received an infusion of LPS (from Eschericia coli; serotype O55:B5; Sigma L2880 - 2.5 micrograms - 1 microliter) over 2 min by means of a microinjection 18 pump. Animal was killed 12 h after the intracerebral infusion. The mouse was anesthetized under isofluorane and blood was drawn via cardiac puncture before head decapitation. The brain was removed rapidly from the skull and placed in cold phosphate buffered saline (PBS) solution. A brain region limited at plane anteroposterior +1.5 to -1.5 and dorsoventral -4.0 was dissected, separated in contralateral side and quickly immersed in liquid nitrogen. The tissue was stored at -80 oC until RNA extraction was performed.


title: mouse brain contralateral saline.RU486, biological rep4

description: mouse brain contralateral RU486


title: mouse brain contralateral saline.RU486, biological rep3

description: mouse brain contralateral RU486


title: mouse brain ipsilateral control, biological rep4

description: mouse brain ipsilateral control


title: Gonad Fish7

city: Exeter

first: Eduarda

middle: M

last: Santos

state: Devon

address: Prince of Wales Road

department: School of Biosciences

country: United Kingdom

institute: University of Exeter

description: Gonad Fish7

hyb_protocol: The reference and sample, labeled with Cy5 and Cy3, respectively, were combined and denatured for 3 minutes at 95oC. An equal volume of hybridization buffer (50% formamide, 10X SSC, 0.2% SDS) was added to each sample, and the mixture was added to the arrayed slides. The slides were hybridized in a humidified airtight box for 36 hours at 42oC.

scan_protocol: Fluorescent array images were collected for both Cy3 and Cy5 with a Array Express fluorescent scanner and image intensity data were extracted and analyzed with Imagene V5 analysis software.

data_processing: Pre-processed in R using the LimmaGUI package with background subtraction, followed by a pin based-Lowess within array normalization and a scaled between chip normalization.  For each experimental group, data were filtered on ‘flags’ and on intensity of the raw data, and each spot was required to have flag=1 and above 0.01 in at least 40% of the conditions to be considered for further analysis

organism: Danio rerio

source_name: Gonad Fish7

label_protocol: 5ug of aaRNA were labelled with Cy3 using FluoroLink  Cy3 monofunctional dye 5-pack (Amersham), according to the instructions provided within the Amino Allyl MessageAmp aRNA amplification kit (Ambion, USA).

growth_protocol: Gonads extracted from adult breeding zebrafish kept in 15L breeding tanks ( 10 L of water were displaced daily), inder a 12 light/dark cycle, and fed twice daily with commercial flake food and live Artemia.

extract_protocol: RNA was extracted using the TRI reagent method (Sigma, UK) followed by a LiCl precipitation. A RNA amplification step was carried out using the Amino Allyl MessageAmp aRNA amplification kit (Ambion, USA).

tag: Gonad Tissue

source_name: Pool of Danio rerio gonadal RNA

label_protocol: 5ug of aaRNA Pool was labeled with Cy5 using FluoroLink Cy5 monofunctional dye 5-pack (Amersham), according to the instructions provided within the Amino Allyl MessageAmp aRNA amplification kit (Ambion, USA).

tag: Pool of 5ug of aRNA from each gonadal extract included in this experiment


title: mouse brain contralateral LPS.Dmso, biological rep4

description: mouse brain contralateral LPS


title: Gonad Fish5

description: Gonad Fish5

source_name: Gonad Fish5


title: mouse brain contralateral LPS.Dmso, biological rep3

description: mouse brain contralateral LPS


title: Gonad Fish8

description: Gonad Fish8

source_name: Gonad Fish8


title: Gonad Fish4

description: Gonad Fish4

source_name: Gonad Fish4


title: mouse brain contralateral LPS.Dmso, biological rep5

description: mouse brain contralateral LPS


title: Gonad Fish3

description: Gonad Fish3

source_name: Gonad Fish3


title: mouse brain contralateral LPS.Dmso, biological rep2

description: mouse brain contralateral LPS


title: mouse brain contralateral LPS.Dmso, biological rep1

description: mouse brain contralateral LPS


title: mouse brain contralateral LPS.RU486, biological rep1

description: mouse brain contralateral LPS.RU486


title: Gonad Fish2

description: Gonad Fish2

source_name: Gonad Fish2


title: Gonad Fish12

description: Gonad Fish12

source_name: Gonad Fish12


title: mouse brain contralateral LPS.RU486, biological rep2

description: mouse brain contralateral LPS.RU486


title: Pain spinal cord right carrageenan 3

status: Public on Mar 21 2007

city: Baltimore

first: Svetlana

middle: I

last: Novikova

address: 666 W. Baltimore st

department: Biomedical Sciences

institute: University of Maryland

description:
Neonatal noxious insult produces a long-term effect on pain processing in adults. Rats subjected to
carrageenan (CAR) injection in one hindpaw within the sensitive period develop bilateral
hypoalgesia as adults. In the same rats, inflammation of the hindpaw, which was the site of the
neonatal injury, induces a localized enhanced hyperalgesia limited to this paw. To gain an insight
into the long-term molecular changes involved in the above-described long-term nociceptive effects
of neonatal noxious insult at the spinal level, we performed DNA microarray analysis (using
microarrays containing oligo-probes for 205 genes encoding receptors and transporters for
glutamate, GABA, and amine neurotransmitters, precursors and receptors for neuropeptides, and
neurotrophins, cytokines and their receptors) to compare gene expression profiles in male rats that received neonatal CAR treatment within (at postnatal day 3; P3) the sensitive period.

hyb_protocol: The resultant cDNA was then hybridized to oligonucleotide microarrays custom designed and printed for us by TeleChem International (Sunnyvale, CA). The cDNA hybridization was performed at 55°C for 16 h in 2 × SDS-Based Hybridization Buffer (3DNA Array Detection Kit, Genesphere, Hartfield, PA). The hybridization was followed by washing with 2 × SSC and 0.2% SDS at 42°C, 2 × SSC at room temperature, and 0.2 × SSC at room temperature, 10 min each. Washed slides were air-dried for 30 sec. For fluorescent labeling of microarray-bound cDNA, the slides were incubated for 3 h at 65°C with a Capture Reagent Hybridization Mixture from a 3DNA Array Detection Kit (Genesphere Inc, Hatfield, PA).The labeling reaction was terminated by washing the microarray slides at room temperature for 10 min with 0.005% Triton in 2 × SSC and for another 10 min in 0.2 × SSC. After that, the slides were air-dried for 30 sec.

scan_protocol: The processed microarray slides were scanned on a GenePix 4100A scanner (Axon Instrument, Union City, CA) with the laser excitation at 532 nm [emission filter 575DF35 (green); photomultiplier voltage 550] for Alexa 546 (control samples) and the laser excitation at 635 nm [emission filter 670DF40 (red); photomultiplier voltage 695] for Alexa 647 (experimental samples). The densitometry was performed with GenePix Pro 4.1 (Axon Instruments, Union City, CA). Background was subtracted using the 'Local Background Correction' procedure and quality control was achieved by 'Quality Control' feature of the GenePix software.

data_processing:
For each
array, the data were normalized with Acuity 2.0 (Axon
Instruments, Union City, CA) by LOWESS normalization method.

label: Alexa 647

organism: Rattus norvegicus

source_name: lumbar dorsal horn right region, carrageenan

label_protocol:
The resultant cDNA was
then hybridized to oligonucleotide microarrays custom
designed and printed for us by TeleChem International
(Sunnyvale, CA). For fluorescent
labeling of microarray-bound cDNA, the slides
were incubated for 3 h at 65°C with a Capture Reagent
Hybridization Mixture from a 3DNA 300 Array Detection Kit
(Genesphere Inc, Hatfield, PA).

extract_protocol:
For the analysis, the total sample RNA was isolated with
TRIzol protocol (Invitrogen, Carsbad, CA). The RNA yield
was determined using a DU 640 Spectrophotometer
(Beckman, Coulter, Fullerton, CA). The 260/280 nm
ratios of the samples were 1.8. The RNA isolated from a
specific pairs of samples (5 μg RNA per sample) was
reverse transcribed utilizing a 3DNA Array Detection Kit
(Genesphere, Hartfield, PA); with primers containing different
capture sequences for CONT and experimental
tissue

treatment_protocol:
Litters with ≥10
pups were selected for this study. Pups received a single injection of 0.25% CAR in sterile 0.9% saline (1 μl/g) in to the plantar surface of their left hindpaw at 9:00 pm on P3 . The injection was conducted with a microliter syringe with a 31-gauge needle. After weaning at P21, male pups were housed in groups of three with animals in any given cage receiving the same neonatal treatment. Animals were grown till P59.

tag: p59 male, lumbar dorsal horn right, at P3 rats were injected in their left hind paws with 0.25 % carrageenan

label: Alexa 546

source_name: lumbar dorsal horn right region, control

label_protocol: Same with Alexa 546

extract_protocol: same as channel 1

treatment_protocol: Same as channel 1, but no injections.

tag: p59 male, lumbar dorsal horn right region, intact animals


title: Gonad Fish11

description: Gonad Fish11

source_name: Gonad Fish11


title: Pain spinal cord right carrageenan 1


title: Pain spinal cord right carrageenan 4


title: mouse brain contralateral control, biological rep2

description: mouse brain contralateral control

data_processing: The data were analyzed with the RMA summarization algorithm as implemented in Bioconductor package: affy, with deault analysis settings.

source_name: mouse brain contralateral control

treatment_protocol: C57Bl/6 mouse received an i.p. injection of vehicle (DMSO - 50 microliters) and were submitted to surgery 4 h later. The mice receiving intraparenchymal injections were anesthetized with a mixture of ketamine and xylazine and placed in a stereotaxic apparatus (David Kopf Instruments). The right caudate putamen was reached, using a small cannula (28 gauge; Plastic One) at the coordinates 0.0 mm anteroposterior, -2.0 mm lateral, and -3.0 mm dorsoventral according to a mouse brain atlas [Paxino, G., and Franklin, K. B. J. (2001). The mouse brain in stereotaxic coordinates., 2nd ed (San Diego: Academic Press)]. The animal received an infusion of sterile pyrogen-free saline (1 microliter) over 2 min by means of a microinjection 18 pump. Animal was killed 12 h after the intracerebral infusion. The mouse was anesthetized under isofluorane and blood was drawn via cardiac puncture before head decapitation. The brain was removed rapidly from the skull and placed in cold phosphate buffered saline (PBS) solution. A brain region limited at plane anteroposterior +1.5 to -1.5 and dorsoventral -4.0 was dissected, separated in contralateral side and quickly immersed in liquid nitrogen. The tissue was stored at -80 oC until RNA extraction was performed.


title: Gonad Fish1

description: Gonad Fish1

source_name: Gonad Fish1


title: Pain spinal cord left carrageenan 5

source_name: lumbar dorsal horn left region, carrageenan

tag: p59 male, RVM, at P3 rats were injected in their left hind paws with 0.25 % carrageenan

source_name: lumbar dorsal horn left region, control

tag: p59 male, lumbar dorsal horn left region, intact animals


title: Pain spinal cord left carrageenan 3


title: Pain spinal cord right carrageenan 2

tag: p59 male, dorsal horan lumbar right, at P3 rats were injected in their left hind paws with 0.25 % carrageenan


title: Pain spinal cord left carrageenan 2


title: mouse brain contralateral control, biological rep3

description: mouse brain contralateral control


title: Pain spinal cord left carrageenan 1


title: mouse brain contralateral control, biological rep1

description: mouse brain contralateral control


title: Pain RVM saline 3

source_name: RVM brain region, saline

treatment_protocol:
Litters with ≥10
pups were selected for this study. Pups received a single injection of saline in to the plantar surface of their left hindpaw at 9:00 pm on P3 . The injection was conducted with a microliter syringe with a 31-gauge needle. After weaning at P21, male pups were housed in groups of three with animals in any given cage receiving the same neonatal treatment. Animals were grown till P59.

tag: p59 male, RVM, at P3 rats were injected in their left hind paws with saline

source_name: RVM brain region, control

tag: p59 male, RVM, intact animals


title: Gonad Fish9

description: Gonad Fish9

source_name: Gonad Fish9


title: Pain spinal cord left carrageenan 4


title: Pain RVM saline 4


title: Gonad Fish10

description: Gonad Fish10

source_name: Gonad Fish10

source_name: Pool of Danio rerio gonadal RNA


title: mouse brain contralateral control, biological rep4

description: mouse brain contralateral control


title: Pain RVM saline 2


title: Pain RVM carrageenan 3

source_name: RVM brain region, carrageenan


title: Pain RVM carrageenan 5


title: Pain RVM saline 1


title: Pain RVM carrageenan 2


title: Pain PAG saline 3

description:
Neonatal noxious insult produces a long-term effect on pain processing in adults. Rats subjected to
carrageenan (CAR) injection in one hindpaw within the sensitive period develop bilateral
hypoalgesia as adults. In the same rats, inflammation of the hindpaw, which was the site of the
neonatal injury, induces a localized enhanced hyperalgesia limited to this paw. To gain an insight
into the long-term molecular changes involved in the above-described long-term nociceptive effects
of neonatal noxious insult at the spinal level, we performed DNA microarray analysis (using
microarrays containing oligo-probes for 205 genes encoding receptors and transporters for
glutamate, GABA, and amine neurotransmitters, precursors and receptors for neuropeptides, and
neurotrophins, cytokines and their receptors) to compare gene expression profiles in male rats that received neonatal CAR treatment within (at postnatal day 3; P3)  the sensitive period.

source_name: PAG brain region, saline

extract_protocol:
For the analysis, the total sample RNA was isolated with
TRIzol protocol (Invitrogen, Carsbad, CA). The RNA yield
was determined using a DU 640 Spectrophotometer
(Beckman, Coulter, Fullerton, CA). The 260/280 nm
ratios of the samples were 1.8. The RNA isolated from a
specific pairs of samples (5 μg RNA per sample) was
reverse transcribed utilizing a 3DNA Array Detection Kit
(Genesphere, Hartfield, PA); with primers containing different
capture sequences for CONT and experimental
 tissue

treatment_protocol:
Litters with ≥10
pups were selected for this study. Pups received a single injection of  saline in to the plantar surface of their left hindpaw at 9:00 pm on P3 . The injection was conducted with a microliter syringe with a 31-gauge needle. After weaning at P21, male pups were housed in groups of three with animals in any given cage receiving the same neonatal treatment. Animals were grown till P59.

tag: p59 male, PAG, at P3 rats were injected in their left hind paws with saline

source_name: PAG brain region, control

tag: p59 male, PAG, intact animals


title: Pain PAG carr 5

source_name: PAG brain region, carrageenan

tag: p59 male, PAG, at P3 rats were injected in their left hind paws with 0.25 % carrageenan


title: Pain PAG saline 1


title: Pain PAG saline 2


title: Pain RVM carrageenan 1


title: Pain PAG carr 3


title: Pain PAG carr 2


title: Pain PAG carr 4


title: Pain PAG saline 5


title: Pain PAG saline 4


title: Pain RVM carrageenan 4


title: Pain PAG carr 1


title: ChIP-chip examination of H3me2K9 in S. pombe histone demethylase mutant lsd1

type: genomic

city: Austin

first: Matthew

middle: Wayne

last: Vaughn

state: TX

address: 10100 Burnet Rd

department: Texas Advanced Computing Center

institute: University of Texas at Austin

description: A reference design including a dye reversal was used to examine H3me2K9 localization in a genomic context in the histone demethylase mutant lsd1. Genomic DNA was extracted after crosslinking from whole cell-extract and chromatin immunoprecipitated using an anti-H3me2K9 antibody

hyb_protocol: Hybridization performed by NimbleGen Systems, Inc.

scan_protocol: Scanning performed by NimbleGen Systems, Inc.

data_processing: Nimblegen-supplied data files were imported into the Bioconductor computing environment (http://bioconductor.org/). After performing quantile normalization, per-probe enrichments were estimated for each probe by fitting a fixed linear model accounting for array, dye, and treatment effects to the data using the limma package (Smyth 2004). Moderated t-statistics and the log-odds score for ChIP-chip enrichment were computed by empirical Bayes shrinkage of the standard errors with the False Discovery Rate controlled to 0.05 using the method of Benjamini and Hochberg.

label: Cy5 and Cy3

molecule: genomic DNA

organism: Schizosaccharomyces pombe

source_name: H3me2K9 immunoprecipitated DNA

label_protocol: Sample labeling performed by NimbleGen Systems, Inc.

extract_protocol: 7 ul of the immunoprecipitated DNA or a 1/100 dilution of input DNA were subjected to two rounds of sequenase primer extension with primer A, and later PCR amplified for 40 cycles with primer B. The PCR product was purified using Qiaquick PCR purification columns, eluted in water and concentrated by vacuum centrifugation to 0.5 ug/ul.

tag: lsd1 deletion; h- strain

source_name: lsd1- mutant control input DNA


title: Primary Lung SCC Carcinoma Sample_R18

city: Brisbane

first: Jill

middle: Everland

last: Larsen

state: Queensland

address: Rode Road, Chermside

department: Thoracic Medicine

country: Australia

institute: The Prince Charles Hospital

description: Synthesis of the labelled first strand cDNA was conducted using Amersham CyScribe Post-Labelling kit with starting material of 20 ug of total RNA. The amino-allyl labeled dNTP mix was added to the reaction to generate amino-allyl labelled second strand cDNA. Following the hydrolysis reaction, single-stranded cDNA probes were purified using Amersham GFX columns. Dye coupling reactions were performed by mixing the cDNA samples with Amersham Cy3 (reference) or Cy5 (tumour) and incubating in the dark. The reactions were purified with Amersham GFX columns to remove the unincorporated/quenched dyes. After the purification samples were combined for hybridization, the labeled cDNAs were co-hybridized to 22K oligo microarrays. Slides were scanned on GMS418 confocal scanner (Agilent).

data_processing: Raw images were imported into Imagene V5.1 (BioDiscovery, CA, USA) to extract pixel intensities and to flag spots with poor/absent signal. The raw background subtracted signal median for each probe was imported into GeneSpring GX V7.3 (Agilent Technologies, Inc., CA, USA) for analysis. Data was normalized (Lowess: per spot and per chip; per chip to the 50th percentile) and probe signals filtered on pixel intensity (for both red and green channels any spots less than 20 units or greater than 65,000 units were excluded) and consistent spot morphology. Only spots flagged present in at least 80% of samples were included. For each probe, the logarithm to the base 2 of the ratio between the intensity in the tumor sample (red) channel and the reference (green) channel was used as the expression value for the probe.

source_name: Universal Reference RNA

tag: The reference sample used for this experiment was commercially available Universal Reference RNA (Stratagene)

source_name: Tumour sample R18

tag: Primary SCC carcinoma of the lung. Total RNA extracted from fresh-frozen tissue using Trizol and RNeasy clean-up.


title: Primary Lung SCC Carcinoma Sample_R17

source_name: Tumour sample R17


title: Primary Lung SCC Carcinoma Sample_R16

source_name: Tumour sample R16


title: Primary Lung SCC Carcinoma Sample_R19

source_name: Tumour sample R19


title: Primary Lung SCC Carcinoma Sample_R14

source_name: Tumour sample R14


title: ChIP-chip examination of H3me2K4 in S. pombe histone demethylase mutant lsd1

description: A reference design including a dye reversal was used to examine H3me2K9 localization in a genomic context in the histone demethylase mutant lsd1. Genomic DNA was extracted after crosslinking from whole cell-extract and chromatin immunoprecipitated using an anti-H3me2K4 antibody

source_name: H3me2K4 immunoprecipitated DNA


title: ChIP-chip examination of H3me2K9 in S. pombe wild-type

description: A reference design including a dye reversal was used to examine H3me2K9 localization in a genomic context in wild-type cells. Genomic DNA was extracted after crosslinking from whole cell-extract and chromatin immunoprecipitated using an anti-H3me2K9 antibody.

tag: Wild-type; h- strain

source_name: Wild-type control input DNA


title: Primary Lung SCC Carcinoma Sample_R13

source_name: Tumour sample R13


title: Primary Lung SCC Carcinoma Sample_R15

source_name: Tumour sample R15


title: ChIP-chip examination of H3me2K4 in S. pombe wild-type


title: ChIP-chip localization of S. pombe LSD2 histone demethylase

description: A reference design including a dye reversal was used to examine LSD2 localization in a genomic context. Genomic DNA was extracted after crosslinking from whole cell-extract and chromatin immunoprecipitated using an anti-LSD2 antibody

data_processing: Nimblegen-supplied data files were imported into the Bioconductor computing environment (http://bioconductor.org/). After performing quantile normalization, per-probe enrichments were estimated for each probe by fitting a fixed linear model accounting for array, dye, and treatment effects to the data using the limma package (Smyth 2004). Moderated t-statistics and the log-odds score for differential expression or ChIP enrichment was computed by empirical Bayes shrinkage of the standard errors with the False Discovery Rate controlled to 0.05

source_name: LSD2 immunoprecipitated DNA


title: ChIP-chip localization of S. pombe LSD1 histone demethylase

description: A reference design including a dye reversal was used to examine LSD1 localization in a genomic context. Genomic DNA was extracted after crosslinking from whole cell-extract and chromatin immunoprecipitated using an anti-LSD1 antibody.

source_name: LSD1 immunoprecipitated DNA


title: Primary Lung SCC Carcinoma Sample_R8

source_name: Tumour sample R8


title: Primary Lung SCC Carcinoma Sample_R10

source_name: Tumour sample R10


title: Primary Lung SCC Carcinoma Sample_R6

source_name: Tumour sample R6


title: Primary Lung SCC Carcinoma Sample_R7

source_name: Tumour sample R7


title: Primary Lung SCC Carcinoma Sample_R3

source_name: Tumour sample R3


title: Primary Lung SCC Carcinoma Sample_R5

source_name: Universal Reference RNA

source_name: Tumour sample R5


title: Primary Lung SCC Carcinoma Sample_R9

source_name: Tumour sample R9


title: Primary Lung SCC Carcinoma Sample_N30

source_name: Tumour sample N30


title: Primary Lung SCC Carcinoma Sample_R11

source_name: Tumour sample R11


title: Primary Lung SCC Carcinoma Sample_R12

source_name: Tumour sample R12


title: Primary Lung SCC Carcinoma Sample_N31

source_name: Tumour sample N31


title: Primary Lung SCC Carcinoma Sample_R1

source_name: Tumour sample R1


title: Primary Lung SCC Carcinoma Sample_N32

source_name: Tumour sample N32


title: Primary Lung SCC Carcinoma Sample_R4

source_name: Tumour sample R4


title: Primary Lung SCC Carcinoma Sample_R2

source_name: Tumour sample R2


title: Primary Lung SCC Carcinoma Sample_N28

source_name: Tumour sample N28


title: Primary Lung SCC Carcinoma Sample_N29

source_name: Tumour sample N29


title: Primary Lung SCC Carcinoma Sample_N20

source_name: Tumour sample N20


title: Primary Lung SCC Carcinoma Sample_N25

source_name: Tumour sample N25


title: Primary Lung SCC Carcinoma Sample_N24

source_name: Tumour sample N24


title: Primary Lung SCC Carcinoma Sample_N27

source_name: Tumour sample N27


title: Primary Lung SCC Carcinoma Sample_N26

source_name: Universal Reference RNA

source_name: Tumour sample N26


title: Primary Lung SCC Carcinoma Sample_N22

source_name: Tumour sample N22


title: Primary Lung SCC Carcinoma Sample_N23

source_name: Tumour sample N23


title: Primary Lung SCC Carcinoma Sample_N19

source_name: Tumour sample N19


title: Primary Lung SCC Carcinoma Sample_N21

source_name: Tumour sample N21


title: Primary Lung SCC Carcinoma Sample_N16

source_name: Tumour sample N16


title: Primary Lung SCC Carcinoma Sample_N17

source_name: Tumour sample N17


title: Primary Lung SCC Carcinoma Sample_N18

source_name: Tumour sample N18


title: Transcriptional profilng of the S. pombe histone demethylase mutant lsd1

description: A reference design including a dye reversal was used to examine genome-wide expression levels in the histone demethylase mutant lsd1.

data_processing: Nimblegen-supplied data files were imported into the Bioconductor computing environment (http://bioconductor.org/). After performing quantile normalization, per-probe differences relative to wild-type were estimated for each probe by fitting a fixed linear model accounting for array, dye, and treatment effects to the data using the limma package (Smyth 2004). Moderated t-statistics and the log-odds score for differential expression was computed by empirical Bayes shrinkage of the standard errors with the False Discovery Rate controlled to 0.05 using the Benjamini-Hochberg method.

source_name: lsd1- mutant

extract_protocol: Total RNA from wild type and mutant cells cultured to mid-logaritmic phase was isolated using standard procedures. Copy DNA for labeling was produced from 10 ug of total RNA using the Superscript II Double Stranded cDNA synthesis kit (Invitrogen) with the provided OligodT primer, purified by Phenol:chloroform:isoamyl alchohol extraction and ethanol precipitation, then resuspended in water

tag: Wild-type; h- strain versus lsd1 deletion; h- strain

source_name: Wild-type total RNA


title: Primary Lung SCC Carcinoma Sample_N15

source_name: Tumour sample N15


title: Primary Lung SCC Carcinoma Sample_N14

source_name: Tumour sample N14


title: Primary Lung SCC Carcinoma Sample_N13

source_name: Tumour sample N13


title: Primary Lung SCC Carcinoma Sample_N11

source_name: Tumour sample N11


title: Primary Lung SCC Carcinoma Sample_N8

source_name: Tumour sample N8


title: UVA irradiated chemostat for 1 h, run 3

status: Public on Mar 20 2007

city: Bronx

first: Michael

last: Berney

state: NY

address: 1301 Morris Park Avenue

department: Department of Microbiology and Immunology

institute: Albert Einstein College of Medicine

description: see publication

data_processing: Jaguar

organism: Escherichia coli

source_name: Control chemostat 1 h not irradiated

tag: E. coli K12 MG1655

source_name: Chemostat 1 h irradiated


title: Primary Lung SCC Carcinoma Sample_N6

source_name: Universal Reference RNA

source_name: Tumour sample N6


title: Primary Lung SCC Carcinoma Sample_N9

source_name: Tumour sample N9


title: Primary Lung SCC Carcinoma Sample_N7

source_name: Tumour sample N7


title: UVA irradiated chemostat for 50 h, run 3

description: see publication

source_name: Control chemostat 50 h not irradiated

source_name: Chemostat 50 h irradiated


title: UVA irradiated chemostat for 1 h, run 1

description: see publication


title: UVA irradiated chemostat for 1 h, run 2

description: see publication

source_name: Control chemostat 1h not irradiated

source_name: Chemostat 1h irradiated


title: Primary Lung SCC Carcinoma Sample_N3

source_name: Tumour sample N3


title: Primary Lung SCC Carcinoma Sample_N2

source_name: Tumour sample N2


title: UVA irradiated chemostat for 1 h, run 2 dye swap

description: see publication


title: UVA irradiated chemostat for 50 h, run 1

description: see publication


title: Primary Lung SCC Carcinoma Sample_N10

source_name: Tumour sample N10


title: Primary Lung SCC Carcinoma Sample_N1

source_name: Tumour sample N1


title: Caffeine treated 3rd instar larvae - Array 1

city: Parkville

first: Lee

last: Willoughby

state: Victoria

address: 30 Flemington Road

department: Genetics

institute: University of Melbourne

description: The Drosophila melanogaster "detox" microarray contained the cytochrome P450, glutathione-S-transferase and esterase gene families and also incorporated both individual control genes and control dilutions, to account for the large number of "detox" genes that could have potentially been differentially regulated in response to different treatments.

hyb_protocol: Microarray pre-hybridisation preparation Microarray slides were prepared for use by incubation for an hour at 42 degrees C in a 50ml centrifuge tube containing the following solution: 0.5g 0.5ml 12.5ml 12.5ml 24.5ml The microarray slide was then immersed in a 50ml centrifuge tube containing H2O and then placed into an empty 50ml centrifuge tube and dried by centrifugation for 5 minutes at 1500rpm. Hybridisation Labeled samples were pooled and made up to a volume of 27Âul either by drying down (as described above) or by the addition of NFW. The hybridization mixture was made up by addition of the following: 27Âul 7Âul 1.2Âul 2Âul 2Âul 1Âul This mixture was incubated for 2 minutes at 98Â°C and then allowed to reach room temperature for 10 minutes. 22mm x 25mm Lifter slip was placed over the microarray and the hybridisation mix was applied to the microarray.  The microarray was then placed into a hybridization chamber, which was wrapped in aluminium foil and then incubated in a 65Â°C water bath for 20 hours. Washing Wash 1: Wash 2: Wash 3 and 4: All washes were performed at room temperature. The microarray was added to 50ml Wash 1 in a tray and agitated until the coverslips floated off, which were then removed.  This tray was then placed into a light-tight box and agitated for 5 minutes.  The microarray was then transferred to a tray containing 50ml Wash 2, placed into a light-tight box and agitated for 5 minutes.  The microarray was then transferred to a tray containing 50ml Wash 3, placed into a light-tight box and agitated for 5 minutes.  The microarray was then transferred into a 50ml centrifuge tube wrapped in aluminium foil, containing Wash 4 and agitated for 3 minutes, before being briefly immersed in water and then dried by centrifugation for 5 minutes at 1500rpm in a fresh 50ml centrifuge tube.

scan_protocol: Microarray slides were scanned using a Genepix 4000B microarray scanner (Axon instruments) and signal quantitation was performed using Genepix pro software (Axon instruments).

data_processing: Data analysis was conducted with LimmaGUI (http://bioinf.wehi.edu.au/limmaGUI/), a graphical interface for Limma (Smyth, G. K. & Speed, T. P. (2003) Methods 31, 265-273).  The linear model was computed and the data was normalized with the print-tip loess method.  Genes were considered differentially expressed if they had P values < 0.05 after multiple comparison correction (Holm correction) and were greater than 2 fold up or down regulated.

label: Cy3-dCTP (Amersham)

source_name: Untreated 3rd instar larvae

label_protocol:
Labeling master mix
6ul
3ul
0.6ul
0.5ul
1.5ul
50x low-C dNTP
25ul
25ul
25ul
10ul
15ul
2ul of test/control Lucidea Universal Scorecard (Amersham) spike in RNA was added to 14.9ul 4ug/ul RNA sample.  1.5ul oligo dT (Invitrogen) was added to the sample and then incubated at 70°C for 10 minutes.
Samples were immeadiately transferred to ice and cooled for at least one minute.
11.6ul master mix was added to each sample, followed by 3ul Cy3-dCTP (Amersham), which was then incubated at 42°C for 2 hours.
1.5ul 1M NaOH was added to the sample, which was then incubated at 70°C for 10 minutes.
1.5ul 1M HCl was added to the sample and then the reaction mixture was concentrated to under 20ul (required for DyeEx columns).  The sample was concentrated by incubation at 80°C for 15-20 minutes, covered by aluminium foil.
Unincorporated Cy-dye was removed using the DyeEx 2.0 kit (QIAGEN), according to the manufacturer’s protocol.

growth_protocol: D. melanogaster y; cn bw sp strain adults were allowed to lay eggs for a 6 hour period on petri dishes containing standard fly food with 2x agar.  Petri dishes were then collected and incubated at 25°C.  Larvae were allowed to develop on the original petri dishes until they were to be exposed, so that all larvae would be 4 1/2 days old when frozen after exposure.

extract_protocol:
RNA extraction
RNA was extracted using trizol reagent (Invitrogen).  Eight samples (50 individuals per sample) were used.  Frozen samples were crushed on liquid nitrogen and then resuspended in 1ml of trizol reagent.  RNA extraction was then performed according to the manufacturer’s instructions and the pellet was resuspended in 50ul of nuclease free water (NFW).
RNA clean-up
Samples were pooled to generate four 100ul samples.  RNA was cleaned up using the column based RNeasy kit (QIAGEN).  RNA clean up was performed according to the manufacturer’s instructions with the RNA being eluted in 80ul NFW.  All four samples were pooled, with 5ul set aside for concentration determination using spectrophotometer analysis and an additional 5ul was set aside and run on a 1% agarose gel to assess RNA quality.
RNA concentration
23ul 3M NaOAc (pH 5.2) and 600ul ethanol was added to the RNA sample.  This was vortexed and then precipitated for an hour at -20°C.  After precipitation, the samples were centrifuged at maximum speed for 15 minutes at 4°C.  The pellet was aspirated and 500ul 75% ethanol was added.  The sample was vortexed and then centrifuged at maximum speed for 15 minutes at 4°C.  The pellet was aspirated and allowed to air dry for 10 minutes in a fume hood.  The pellet was then resuspended in NFW to give a final RNA concentration of 4ug/ul.

treatment_protocol: Fresh petri dishes containing standard fly food with 2x agar were prepared for exposure, containing either the compound or the appropriate control.  A final concentration of 1.5mg/ml Caffeine (Sigma) was used.  50 larvae were transferred to each petri dish.  After the appropriate 4 hours, the larvae were frozen in liquid nitrogen and stored at –80°C until RNA extraction.

tag: Strain

value: y cn bw sp

tag: Age

value: Four and a half days post laying, 3rd instar feeding larvae

tag: Treatment

value: unexposed control

source_name: Caffeine treated 3rd instar larvae

label_protocol:
Labeling master mix
6ul
3ul
0.6ul
0.5ul
1.5ul
50x low-C dNTP
25ul
25ul
25ul
10ul
15ul
2ul of test/control Lucidea Universal Scorecard (Amersham) spike in RNA was added to 14.9ul 4ug/ul RNA sample.  1.5ul oligo dT (Invitrogen) was added to the sample and then incubated at 70°C for 10 minutes.
Samples were immeadiately transferred to ice and cooled for at least one minute.
11.6ul master mix was added to each sample, followed by 3ul Cy5-dCTP (Amersham), which was then incubated at 42°C for 2 hours.
1.5ul 1M NaOH was added to the sample, which was then incubated at 70°C for 10 minutes.
1.5ul 1M HCl was added to the sample and then the reaction mixture was concentrated to under 20ul (required for DyeEx columns).  The sample was concentrated by incubation at 80°C for 15-20 minutes, covered by aluminium foil.
Unincorporated Cy-dye was removed using the DyeEx 2.0 kit (QIAGEN), according to the manufacturer’s protocol.

value: exposed to caffeine


title: PBO treated adults - detox microarray - slide 3

status: Public on Mar 18 2007

hyb_protocol: Microarray pre-hybridisation preparation Microarray slides were prepared for use by incubation for an hour at 42degC in a 50ml centrifuge tube containing the following solution: 0.5g BSA 0.5ml 10% SDS 12.5ml 20x SSC 12.5ml formamide 24.5ml H2O The microarray slide was then immersed in a 50ml centrifuge tube containing H2O and then placed into an empty 50ml centrifuge tube and dried by centrifugation for 5 minutes at 1500rpm. Hybridisation Labeled samples were pooled and made up to a volume of 27ul either by drying down (as described above) or by the addition of NFW. The hybridization mixture was made up by addition of the following: 27ul Pooled sample 7ul 20x SSC 1.2ul 10% SDS 2ul Salmon testes DNA (Sigma) 2ul Cot1 DNA (Invitrogen) 1ul Oligo-dA (Invitrogen) This mixture was incubated for 2 minutes at 98degC and then allowed to reach room temperature for 10 minutes. 22mm x 25mm Lifter slip was placed over the microarray and the hybridisation mix was applied to the microarray. The microarray was then placed into a hybridization chamber, which was wrapped in aluminium foil and then incubated in a 65degC water bath for 20 hours. Washing Wash 1: 1x SSC 0.2% SDS Wash 2: 0.1x SSC 0.2% SDS Wash 3 and 4: 0.1x SSC All washes were performed at room temperature. The microarray was added to 50ml Wash 1 in a tray and agitated until the coverslips floated off, which were then removed. This tray was then placed into a light-tight box and agitated for 5 minutes. The microarray was then transferred to a tray containing 50ml Wash 2, placed into a light-tight box and agitated for 5 minutes. The microarray was then transferred to a tray containing 50ml Wash 3, placed into a light-tight box and agitated for 5 minutes. The microarray was then transferred into a 50ml centrifuge tube wrapped in aluminium foil, containing Wash 4 and agitated for 3 minutes, before being briefly immersed in water and then dried by centrifugation for 5 minutes at 1500rpm in a fresh 50ml centrifuge tube.

data_processing: Data analysis was conducted with LimmaGUI at The Walter and Eliza Hall Institute (WEHI) bioinformatics web site, a graphical interface for Limma (Smyth, G. K. & Speed, T. P. (2003) Methods 31, 265-273). The linear model was computed and the data was normalized with the print-tip loess method. Genes were considered differentially expressed if they had P values < 0.05 after multiple comparison correction (Holm correction) and were greater than 2 fold up or down regulated.

source_name: PBO treated adult males

label_protocol:
Labeling master mix
6ul 5x Superscript II reaction buffer (Invitrogen)
3ul DTT (Invitrogen)
0.6ul 50x low-C dNTP (see below)
0.5ul RNase OUT (Invitrogen)
1.5ul Superscript II reverse transcriptase enzyme (Invitrogen)
50x low-C dNTP
25ul dATP (100mM)
25ul dGTP (100mM)
25ul dTTP (100mM)
10ul dCTP (100mM)
15ul NFW
2ul of test/control Lucidea Universal Scorecard (Amersham) spike in RNA was added to 14.9ul 4ug/ul RNA sample. 1.5ul oligo dT (Invitrogen) was added to the sample and then incubated at 70degC for 10 minutes.
Samples were immeadiately transferred to ice and cooled for at least one minute.
11.6ul master mix was added to each sample, followed by 3ul Cy3-dCTP (Amersham), which was then incubated at 42degC for 2 hours.
1.5ul 1M NaOH was added to the sample, which was then incubated at 70degC for 10 minutes.
1.5ul 1M HCl was added to the sample and then the reaction mixture was concentrated to under 20ul (required for DyeEx columns). The sample was concentrated by incubation at 80degC for 15-20 minutes, covered by aluminium foil.
Unincorporated Cy-dye was removed using the DyeEx 2.0 kit (QIAGEN), according to the manufacturer's protocol.

growth_protocol: Adult males of the y; cn bw sp strain were collected within a 24 hour window after emergence, sorted into groups of 50 males and then stored at 25oC for four days prior to exposure.

extract_protocol:
RNA extraction
RNA was extracted using trizol reagent (Invitrogen). Eight samples (50 individuals per sample) were used. Frozen samples were crushed on liquid nitrogen and then resuspended in 1ml of trizol reagent. RNA extraction was then performed according to the manufacturer's instructions and the pellet was resuspended in 50ul of nuclease free water (NFW).
RNA clean-up
Samples were pooled to generate four 100ul samples. RNA was cleaned up using the column based RNeasy kit (QIAGEN). RNA clean up was performed according to the manufacturer�s instructions with the RNA being eluted in 80ul NFW. All four samples were pooled, with 5ul set aside for concentration determination using spectrophotometer analysis and an additional 5ul was set aside and run on a 1% agarose gel to assess RNA quality.
RNA concentration
23ul 3M NaOAc (pH 5.2) and 600ul ethanol was added to the RNA sample. This was vortexed and then precipitated for an hour at -20degC. After precipitation, the samples were centrifuged at maximum speed for 15 minutes at 4degC. The pellet was aspirated and 500ul 75% ethanol was added. The sample was vortexed and then centrifuged at maximum speed for 15 minutes at 4degC. The pellet was aspirated and allowed to air dry for 10 minutes in a fume hood. The pellet was then resuspended in NFW to give a final RNA concentration of 4ug/ul.

treatment_protocol: 2µl PBO (Aldrich) was added to 150µl of acetone and immediately transferred to a 30ml scintillation vial, which was then rolled until the acetone had evaporated. 50 adult males were transferred to each scintillation vial. After 4 hours, the flies were frozen in liquid nitrogen and stored at -80°C until RNA extraction.

value: Four days post emergence from pupa

tag: Sex

value: Male

value: exposed to PBO

label: Cy5-dCTP (Amersham)

source_name: Untreated adult males

label_protocol:
Labeling master mix
6ul 5x Superscript II reaction buffer (Invitrogen)
3ul DTT (Invitrogen)
0.6ul 50x low-C dNTP (see below)
0.5ul RNase OUT (Invitrogen)
1.5ul Superscript II reverse transcriptase enzyme (Invitrogen)
50x low-C dNTP
25ul dATP (100mM)
25ul dGTP (100mM)
25ul dTTP (100mM)
10ul dCTP (100mM)
15ul NFW
2ul of test/control Lucidea Universal Scorecard (Amersham) spike in RNA was added to 14.9ul 4ug/ul RNA sample. 1.5ul oligo dT (Invitrogen) was added to the sample and then incubated at 70degC for 10 minutes.
Samples were immeadiately transferred to ice and cooled for at least one minute.
11.6ul master mix was added to each sample, followed by 3ul Cy5-dCTP (Amersham), which was then incubated at 42degC for 2 hours.
1.5ul 1M NaOH was added to the sample, which was then incubated at 70degC for 10 minutes.
1.5ul 1M HCl was added to the sample and then the reaction mixture was concentrated to under 20ul (required for DyeEx columns). The sample was concentrated by incubation at 80degC for 15-20 minutes, covered by aluminium foil.
Unincorporated Cy-dye was removed using the DyeEx 2.0 kit (QIAGEN), according to the manufacturer's protocol.

treatment_protocol: 150µl of acetone was transferred to a 30ml scintillation vial, which was then rolled until the acetone had evaporated. 50 adult males were transferred to each scintillation vial. After 4 hours, the flies were frozen in liquid nitrogen and stored at -80°C until RNA extraction.

value: unexposed


title: PBO treated adults - detox microarray - slide 4


title: UVA irradiated chemostat for 50 h, run 2

description: see publication


title: Primary Lung SCC Carcinoma Sample_N12

source_name: Tumour sample N12


title: PBO treated adults - detox microarray - slide 1


title: Rice coleoptiles, 4 days old, anoxic, REPLICATE 1 (Sample #11)

status: Public on Mar 19 2007

city: San Giuliano Terme

first: Pierdomenico

last: Perata

state: Pisa

address: Via Guidiccioni 8-10

department: Institute of Life Sciences

institute: Sant'Anna School of Advanced Studies

description: no additional description required

hyb_protocol: Affymetrix standard hybridization protocol

scan_protocol: Affymetrix standard scanning protocol

data_processing: GC-RMA algorithm

organism: Oryza sativa

source_name: Rice coleoptiles, 4 days old, anoxic

label_protocol: Affymetrix standard labeling protocol

growth_protocol: Rice (Oryza sativa L. cv Nipponbare) seeds were sterilized with diluted bleach (15-min incubation in 1.7% sodium hypochlorite, rinsing and washing in sterile water 10 times) and germinated for four days in ANOXIA on sterile filter paper at 28°C in the dark. All the air/anoxia treatments were performed in the dark at 28°C.

extract_protocol: Total RNA was extracted from the seedling samples using the Ambion RNAqueous kit (Ambion). RNA quality was assessed by agarose gel electrophoresis and spectrophotometry. RNA was processed for use on Affymetrix Rice Genome GeneChip arrays.

treatment_protocol: Rice seeds were germinated under anoxia for four days. An enclosed anaerobic work station (Anaerobic System model 1025; Forma Scientific, Marietta, OH) was used to provide an oxygen-free environment for seed germination or seedling incubation. This chamber uses palladium catalyst wafers and desiccant wafers to maintain strict anaerobiosis to less than 10 µg mL–1 O2 (according to the specifications provided by the manufacturer). High-purity N2 was used for purging the chamber initially and the working anaerobic gas mixture was N2:H2 proportioned at 90:10.

tag: Rice coleoptiles, free of leaf tissue. Rice cultivar

value: Nipponbare Age:4 days Germination:anoxic, 28°C, darkness


title: Rice coleoptiles, 4 days old, aerobic, replicate N.2 (sample#10)

description: no additional description required

scan_protocol: Affymetrix standard scan protocol

source_name: Rice coleoptiles, 4 days old, aerobic

growth_protocol: Rice (Oryza sativa L. cv Nipponbare) seeds were sterilized with diluted bleach (15-min incubation in 1.7% sodium hypochlorite, rinsing and washing in sterile water 10 times) and germinated for four days in air on sterile filter paper at 28°C in the dark. The primary leaf was removed from the coleoptiles of air-germinated seedlings (the leaf is absent in anoxia-germinated seedlings). All the air/anoxia treatments were performed in the dark at 28°C.

treatment_protocol: This is the control: aerobic samples to be compared to anoxic samples

value: Nipponbare Age:4 days Germination:aerobic, 28°C, darkness


title: Rice coleoptiles, 4 days old, anoxic, REPLICATE 2 (Sample #12)

description: no additional description required


title: Rice coleoptiles, 4 days old, aerobic, replicate N.1 (sample#9)

description: no additional description required


title: Nitenpyram treated 3rd instar larvae - Array 4

data_processing: Data analysis was conducted with LimmaGUI (http://bioinf.wehi.edu.au/limmaGUI/), a graphical interface for Limma (Smyth, G. K. & Speed, T. P. (2003) Methods 31, 265-273). The linear model was computed and the data was normalized with the print-tip loess method. Genes were considered differentially expressed if they had P values < 0.05 after multiple comparison correction (Holm correction) and were greater than 2 fold up or down regulated.

source_name: Nitenpyram treated 3rd instar larvae

growth_protocol: D. melanogaster y; cn bw sp strain adults were allowed to lay eggs for a 6 hour period on petri dishes containing standard fly food with 2x agar. Petri dishes were then collected and incubated at 25degC. Larvae were allowed to develop on the original petri dishes until they were to be exposed, so that all larvae would be 4 1/2 days old when frozen after exposure.

treatment_protocol: Fresh petri dishes containing standard fly food with 2x agar were prepared for exposure, containing either the compound or the appropriate control. A final concentration of 6ppm Nitenpyram  (Novartis) was used. 50 larvae were transferred to each petri dish. After the appropriate 4 hours, the larvae were frozen in liquid nitrogen and stored at -80degC until RNA extraction.

value: exposed to nitenpyram


title: Nitenpyram treated 3rd instar larvae - Array 1

growth_protocol: D. melanogaster y; cn bw sp strain adults were allowed to lay eggs for a 6 hour period on petri dishes containing standard fly food with 2x agar. Petri dishes were then collected and incubated at 25degC. Larvae were allowed to develop on the original petri dishes until they were to be exposed, so that all larvae would be 4 1/2 days old when frozen after exposure


title: Nitenpyram treated 3rd instar larvae - Array 2


title: Nitenpyram treated 3rd instar larvae - Array 3


title: Primary Lung SCC Carcinoma Sample_N5

source_name: Universal Reference RNA

source_name: Tumour sample N5


title: Primary Lung SCC Carcinoma Sample_N4

source_name: Tumour sample N4


title: Spinosad treated 3rd instar larvae - Array 1

treatment_protocol: Fresh petri dishes containing standard fly food with 2x agar were prepared for exposure, containing either the compound or the appropriate control. A final concentration of 6.0 x10-5% Spinosad  (Success - Dow Aggrosciences) was used. 50 larvae were transferred to each petri dish. After the appropriate 4 hours, the larvae were frozen in liquid nitrogen and stored at -80degC until RNA extraction.

source_name: Spinosad treated 3rd instar larvae

value: exposed to spinosad


title: Spinosad treated 3rd instar larvae - Array 3


title: Spinosad treated 3rd instar larvae - Array 4


title: Phenobarbital treated 3rd instar larvae - Array 3

treatment_protocol: Fresh petri dishes containing standard fly food with 2x agar were prepared for exposure, containing either the compound or the appropriate control. A final concentration of 10mM Phenobarbital (Sigma) was used. 50 larvae were transferred to each petri dish. After the appropriate 4 hours, the larvae were frozen in liquid nitrogen and stored at -80degC until RNA extraction.

source_name: Phenobarbital treated 3rd instar larvae

treatment_protocol: Fresh petri dishes containing standard fly food with 2x agar were prepared for exposure, containing either the compound or the appropriate control. A final concentration of 10mM Phehobarbital (Sigma) was used. 50 larvae were transferred to each petri dish. After the appropriate 4 hours, the larvae were frozen in liquid nitrogen and stored at -80degC until RNA extraction.

value: exposed to phenobarbital


title: Phenobarbital treated 3rd instar larvae - Array 1


title: Phenobarbital treated 3rd instar larvae - Array 2


title: Lufenuron treated 3rd instar larvae - Array 3

treatment_protocol: Fresh petri dishes containing standard fly food with 2x agar were prepared for exposure, containing either the compound or the appropriate control. A final concentration of 20ppm Lufenuron (Novartis) was used. 50 larvae were transferred to each petri dish. After the appropriate 4 hours, the larvae were frozen in liquid nitrogen and stored at -80degC until RNA extraction.

source_name: Lufenuron treated 3rd instar larvae

value: exposed to lufenuron


title: Phenobarbital treated 3rd instar larvae - Array 4


title: Lufenuron treated 3rd instar larvae - Array 4


title: Diazinon treated 3rd instar larvae - Array 4

source_name: Diazinon treated 3rd instar larvae

treatment_protocol: Fresh petri dishes containing standard fly food with 2x agar were prepared for exposure, containing either the compound or the appropriate control. A final concentration of 1.3 x10-2% Diazinon (Cooper's Dijet) was used. 50 larvae were transferred to each petri dish. After the appropriate 4 hours, the larvae were frozen in liquid nitrogen and stored at -80degC until RNA extraction.

value: exposed to diazinon


title: Dicyclanil treated 3rd instar larvae - Array 1

treatment_protocol: Fresh petri dishes containing standard fly food with 2x agar were prepared for exposure, containing either the compound or the appropriate control. A final concentration of 6.7 x10-2% Dicyclanil (Novartis) was used. 50 larvae were transferred to each petri dish. After the appropriate 4 hours, the larvae were frozen in liquid nitrogen and stored at -80degC until RNA extraction.

source_name: Dicyclanil treated 3rd instar larvae

value: exposed to dicyclanil


title: Lufenuron treated 3rd instar larvae - Array 2


title: Dicyclanil treated 3rd instar larvae - Array 4


title: Lufenuron treated 3rd instar larvae - Array 1


title: Diazinon treated 3rd instar larvae - Array 1


title: Phenobarbital treated adult males - Array 4

source_name: Phenobarbital treated adult males

treatment_protocol: 10µl 1M phenobarbital (Sigma) was added to 150µl of acetone and immediately transferred to a 30ml scintillation vial, which was then rolled until the acetone had evaporated. 50 adult males were transferred to each scintillation vial.  After 4 hours, the flies were frozen in liquid nitrogen and stored at -80Â°C until RNA extraction.

source_name: Untreated male adults


title: Spinosad treated 3rd instar larvae - Array 2

source_name: Untreated 3rd instar larvae


title: Dicyclanil treated 3rd instar larvae - Array 2


title: PBO treated adults - detox microarray - slide 2

source_name: PBO treated adult males


title: Dicyclanil treated 3rd instar larvae - Array 3


title: Phenobarbital treated adult males - Array 3

source_name: Untreated adult males

treatment_protocol: 150µl of acetone and immediately transferred to a 30ml scintillation vial, which was then rolled until the acetone had evaporated. 50 adult males were transferred to each scintillation vial.  After 4 hours, the flies were frozen in liquid nitrogen and stored at -80Â°C until RNA extraction.

source_name: Phenobarbital treated male adults


title: Diazinon treated 3rd instar larvae - Array 3


title: Diazinon treated 3rd instar larvae - Array 2


title: DDT treated adult males - Array 2

source_name: DDT treated adult males

treatment_protocol: 10µg DDT (Sigma) was added to 150µl of acetone and immediately transferred to a 30ml scintillation vial, which was then rolled until the acetone had evaporated. 50 adult males were transferred to each scintillation vial.  After 4 hours, the flies were frozen in liquid nitrogen and stored at -80Â°C until RNA extraction.

value: exposed to DDT


title: DDT treated adult males - Array 3

value: y cn bw sp


title: Phenobarbital treated adult males - Array 2


title: DDT treated adult males - Array 4


title: DDT treated adult males - Array 1

value: Five days post emergence from pupa


title: Phenobarbital treated adult males - Array 1


title: Caffeine treated 3rd instar larvae - Array 2

source_name: Caffeine treated 3rd instar larvae

label_protocol:
Labeling master mix
6ul 5x Superscript II reaction buffer (Invitrogen)
3ul DTT (Invitrogen)
0.6ul 50x low-C dNTP (see below)
0.5ul RNase OUT (Invitrogen)
1.5ul Superscript II reverse transcriptase enzyme (Invitrogen)
50x low-C dNTP
25ul dATP (100mM)
25ul dGTP (100mM)
25ul dTTP (100mM)
10ul dCTP (100mM)
15ul NFW
2ul of test/control Lucidea Universal Scorecard (Amersham) spike in RNA was added to 14.9ul 4ug/ul RNA sample. 1.5ul oligo dT (Invitrogen) was added to the sample and then incubated at 70°C for 10 minutes.
Samples were immeadiately transferred to ice and cooled for at least one minute.
11.6ul master mix was added to each sample, followed by 3ul Cy3-dCTP (Amersham), which was then incubated at 42°C for 2 hours.
1.5ul 1M NaOH was added to the sample, which was then incubated at 70°C for 10 minutes.
1.5ul 1M HCl was added to the sample and then the reaction mixture was concentrated to under 20ul (required for DyeEx columns). The sample was concentrated by incubation at 80°C for 15-20 minutes, covered by aluminium foil.
Unincorporated Cy-dye was removed using the DyeEx 2.0 kit (QIAGEN), according to the manufacturer's protocol.

growth_protocol: D. melanogaster y; cn bw sp strain adults were allowed to lay eggs for a 6 hour period on petri dishes containing standard fly food with 2x agar. Petri dishes were then collected and incubated at 25°C. Larvae were allowed to develop on the original petri dishes until they were to be exposed, so that all larvae would be 4 1/2 days old when frozen after exposure.

extract_protocol:
RNA extraction
RNA was extracted using trizol reagent (Invitrogen). Eight samples (50 individuals per sample) were used. Frozen samples were crushed on liquid nitrogen and then resuspended in 1ml of trizol reagent. RNA extraction was then performed according to the manufacturer's instructions and the pellet was resuspended in 50ul of nuclease free water (NFW).
RNA clean-up
Samples were pooled to generate four 100ul samples. RNA was cleaned up using the column based RNeasy kit (QIAGEN). RNA clean up was performed according to the manufacturer�s instructions with the RNA being eluted in 80ul NFW. All four samples were pooled, with 5ul set aside for concentration determination using spectrophotometer analysis and an additional 5ul was set aside and run on a 1% agarose gel to assess RNA quality.
RNA concentration
23ul 3M NaOAc (pH 5.2) and 600ul ethanol was added to the RNA sample. This was vortexed and then precipitated for an hour at -20°C. After precipitation, the samples were centrifuged at maximum speed for 15 minutes at 4°C. The pellet was aspirated and 500ul 75% ethanol was added. The sample was vortexed and then centrifuged at maximum speed for 15 minutes at 4°C. The pellet was aspirated and allowed to air dry for 10 minutes in a fume hood. The pellet was then resuspended in NFW to give a final RNA concentration of 4ug/ul.

treatment_protocol: Fresh petri dishes containing standard fly food with 2x agar were prepared for exposure, containing either the compound or the appropriate control. A final concentration of 1.5mg/ml Caffeine (Sigma) was used. 50 larvae were transferred to each petri dish. After the appropriate 4 hours, the larvae were frozen in liquid nitrogen and stored at -80°C until RNA extraction.

value: Four and a half days post laying, 3rd instar feeding larvae

label_protocol:
Labeling master mix
6ul 5x Superscript II reaction buffer (Invitrogen)
3ul DTT (Invitrogen)
0.6ul 50x low-C dNTP (see below)
0.5ul RNase OUT (Invitrogen)
1.5ul Superscript II reverse transcriptase enzyme (Invitrogen)
50x low-C dNTP
25ul dATP (100mM)
25ul dGTP (100mM)
25ul dTTP (100mM)
10ul dCTP (100mM)
15ul NFW
2ul of test/control Lucidea Universal Scorecard (Amersham) spike in RNA was added to 14.9ul 4ug/ul RNA sample. 1.5ul oligo dT (Invitrogen) was added to the sample and then incubated at 70°C for 10 minutes.
Samples were immeadiately transferred to ice and cooled for at least one minute.
11.6ul master mix was added to each sample, followed by 3ul Cy5-dCTP (Amersham), which was then incubated at 42°C for 2 hours.
1.5ul 1M NaOH was added to the sample, which was then incubated at 70°C for 10 minutes.
1.5ul 1M HCl was added to the sample and then the reaction mixture was concentrated to under 20ul (required for DyeEx columns). The sample was concentrated by incubation at 80°C for 15-20 minutes, covered by aluminium foil.
Unincorporated Cy-dye was removed using the DyeEx 2.0 kit (QIAGEN), according to the manufacturer's protocol.

value: unexposed control


title: Caffeine treated 3rd instar larvae - Array 3

treatment_protocol: Fresh petri dishes containing standard fly food with 2x agar were prepared for exposure, containing either the compound or the appropriate control. A final concentration of 1.5mg/ml Caffeine (Sigma) was used. 50 larvae were transferred to each petri dish. After the appropriate 4 hours, the larvae were frozen in liquid nitrogen and stored at -80degC until RNA extraction.

value: exposed to caffeine