title: 13757356 - cat2xcad2-ld-2 vs Col0-ld-2

description:
ch1: arabidopsis thaliana (columbia)  - harvest date: 07-07-08
ch2: arabidopsis thaliana (columbia)  mutant (cat2xcad2) - harvest date: 07-07-08
How does glutathione content or reduction state affect H2O2-induced changes in the transcriptome?

hyb_protocol: Col0-ld-2 Cy5 / cat2xcad2-ld-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756994 - Col0-ld-2 vs cat2-ld-2

status: Public on Jul 22 2010

type: RNA

city: evry

first: Soubigou-Taconnat

last: Ludivine

address: 2 rue gaston crémieux

department: URGV

country: France

institute: INRA

description:
ch1: arabidopsis thaliana (columbia)  mutant (cat2) - harvest date: 07-07-08
ch2: arabidopsis thaliana (columbia)  - harvest date: 07-07-08
How does glutathione content or reduction state affect H2O2-induced changes in the transcriptome?

hyb_protocol: cat2-ld-2 Cy5 / Col0-ld-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.

scan_protocol: GenePix Pro 3.0, Cy3:pmt voltage 532nm,460V,laser power 35%, Cy5:635nm,pmt voltage 460V,laser power 30%

data_processing: The raw data comprised the logarithm of median feature pixel intensity at wavelength 635 nm (red) and 532 nm (green). No background was subtracted. An array-by-array normalization was performed to remove systematic biases. First, we excluded spots that were considered badly formed features by the experimenter (Flags=-100). Then we performed a global intensity-dependent normalization using the loess procedure (see Yang et al., 2002) to correct the dye bias. Finally, on each block, the log-ratio median is subtracted from each value of the log-ratio of the block to correct a print-tip effect on each metablock.

molecule: total RNA

organism: Arabidopsis thaliana

label_protocol: labelling Cy3 and Cy5 direct, amplification=yes, aRNA 5 ug. Labelling protocol: 5 µg of aRNA (8µl) mixed with 2 µl of random nonamers 1µg/µl and 0.5 µl of RNAse Out 40 U/µl, denatured 10 min at 70 °C and chilled on ice. The followig components were added to the sample 4 µl of first strand buffer 5 X, 1 µl of 10 mM dNTP/2mM dCTP, 2 µl of 0.1 M DTT, 1.5 µl of Cy3-dCTP or Cy5-dCTP (Amersham Pharmacia Biotech, 25nmol tube, PA5502), 1 µl of SuperScript II RT 200U/µl. Then it was incubated at 42°C for 2.5 hours and chilled on ice. The sample was denaturated by adding 2 µl of NaOH 2.5M and incubated at 37 °C for exactly 15 min, then was added 10 µl of 2M MOPS and put on ice. The dyes were purified with the QIAquick PCR Purification Kit (QIAGEN).

growth_protocol: rosette - Media=soil hygrometry=70% Temperature=25°C  Light=200 µmol/m2/s long day=16h photoperiod.

treatment_protocol: no treatment

tag: genetic background

value: Columbia

tag: genotype

value: mutant (cat2) -

tag: harvest date

value: wild type


title: 14054284 - cat2xcytGR-ld-2 vs Col0-ld-2

description:
ch1: arabidopsis thaliana (columbia)  - harvest date: 07-07-08
ch2: arabidopsis thaliana (columbia)  mutant (cat2xcytgr) - harvest date: 07-07-08
How does glutathione content or reduction state affect H2O2-induced changes in the transcriptome?

hyb_protocol: Col0-ld-2 Cy5 / cat2xcytGR-ld-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.

scan_protocol: GenePix Pro 3.0, Cy3:pmt voltage 532nm,480V,laser power 35%, Cy5:635nm,pmt voltage 480V,laser power 30%

value: mutant (cat2xcytgr) -


title: 0.625-2 vs 2.5-1

status: Public on Jul 30 2010

city: Vicksburg

first: Kurt

middle: A

last: Gust

state: MS

address: 3909 Halls Ferry Rd.

department: Environmental Laboratory

country: USA

institute: US Army ERDC

description: Hybridizations conducted using Balanced, Interwoven Loop Design: 2 Dyes Swaps and 4 total technical replicates

hyb_protocol: Products of the cDNA synthesis reactions were combined according to dye-swap assignments and hybridized onto microarrays.  Two µL of blocking agent, locked nucleic acid (LNA) dT Blocker (Genisphere) was included in each hybridization.  RNA sample probes were hybridized at 55ºC overnight (~16h) then washed for 20 min in 60ºC 2x SSC including 0.2% SDS, 20 min in 2x SSC at room temperature, and 20 min in 0.2x SSC also at room temperature.  Slides were then spin dried and subsequently hybridized to A647 and Cy3 labeled secondary hybridization probes in 100 µL hybridization buffer with 1 µL of Anti-Fade Reagent for 4h at 60ºC.  Microarrays were washed as described above only at 15 minute wash intervals and spin dried.  Slides were stored for no more than 1 week in a darkened desiccation chamber prior to microarray scanning.

scan_protocol: Microarrays were scanned at 5μm using a VersArray Chipreader™ (Bio-Rad, Hercules, CA).  Grids were manually fit to microarray spots for each image file and both positive and negative controls were flagged prior to spot analysis.  Data for which the spot intensity was less than the background intensity plus 2x the background standard deviation for ≥1 signal channel were excluded from the analysis

data_processing: Summary data consisted of LOWESS-normalized Cy3:A647 ratio values calculated using background-normalized median-signal intensities.  The statistical analysis was conducted using Bayesian Analysis of Gene Expression Levels (BAGEL) software version 3.62 (Townsend and Hartl, 2002).  Non-overlapping 97.5% confidence intervals (C.I.) among treatments were established a priori as statistically significant differences in expression.

organism: Pimephales promelas

source_name: RDX 0.625, replicate 2

label_protocol: Microarray experiments were conducted using a balanced interwoven-loop design using Cyanine-3 (Cy3) and A647.  The experiment compared controls with 0.625, 2.5, 5 and 10 mg/L RDX-exposed fish with each condition including 3 biological replicates.  Hybridizations included four technical replicates and two dye swaps per biological replicate resulting in a total of 30 micorarrays assayed.  Probes for microarray hybridizations were prepared from 600 ng total RNA / sample following the Genisphere®, Array 900™ Alexa Fluor 647 and Cy3 secondary hybridization labeling protocol (Genisphere, Hatfield, PA) according to manufacturer’s instructions.

growth_protocol: Fish for each treatment were reared in a 25L aquarium where water quality parameters including temperature, dissolved oxygen, pH, ammonia concentration, hardness, and alkalinity were measured at the initiation and termination of each bioassay.  During the bioassays, temperature, dissolved oxygen, and pH were measured on days 1, 2 and then every other day until bioassay termination.

extract_protocol: RNeasy® Mini RNA extraction kits (Qiagen, Valencia, CA) were used for total RNA extractions.  RNA quality was assessed using an Agilent 2100 Bioanalyzer (Agilent Technologies, Waldbronn, Germany) with RNA 6000 Nano LabChips® RNA. Only samples with a 28s/18s ratio {greater than or equal to} 1.4 and RNA integrity number (RIN) {greater than or equal to} 6 were used for downstream applications.  The majority of RNA samples greatly exceeded these minimum requirements.    Total RNA was utilized for cDNA library construction, microarray analysis and RT-qPCR.

treatment_protocol: Fathead minnows were exposed to RDX in a 10d dose-series experiment (0.625, 1.25, 2.5, 5.0, or 10 mg/L) which included an acetone-spike control (1% acetone).  Each experimental treatment included 8 replicate fish.

tag: agent

value: RDX

tag: dose

tag: tissue type

value: Fathead minnow brain tissue

source_name: RDX 2.5, replicate 1


title: 13756990 - cytGR-ld-1 vs Col0-ld-1

description:
ch1: arabidopsis thaliana (columbia)  - harvest date: 07-07-08
ch2: arabidopsis thaliana (columbia)  mutant (cytgr) - harvest date: 07-07-08
How does glutathione content or reduction state affect H2O2-induced changes in the transcriptome?

hyb_protocol: Col0-ld-1 Cy5 / cytGR-ld-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.

value: mutant (cytgr) -


title: 13757354 - cat2xcad2-ld-1 vs Col0-ld-1

hyb_protocol: Col0-ld-1 Cy5 / cat2xcad2-ld-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.

value: mutant (cat2xcad2) -


title: 13757347 - Col0-ld-1 vs cat2-ld-1

hyb_protocol: cat2-ld-1 Cy5 / Col0-ld-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756992 - cytGR-ld-2 vs Col0-ld-2

hyb_protocol: Col0-ld-2 Cy5 / cytGR-ld-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13757353 - Col0-ld-1 vs cat2xcad2-ld-1

description:
ch1: arabidopsis thaliana (columbia)  mutant (cat2xcad2) - harvest date: 07-07-08
ch2: arabidopsis thaliana (columbia)  - harvest date: 07-07-08
How does glutathione content or reduction state affect H2O2-induced changes in the transcriptome?

hyb_protocol: cat2xcad2-ld-1 Cy5 / Col0-ld-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13757348 - Col0-ld-1 vs cad2-ld-1

description:
ch1: arabidopsis thaliana (columbia)  mutant (cad2) - harvest date: 07-07-08
ch2: arabidopsis thaliana (columbia)  - harvest date: 07-07-08
How does glutathione content or reduction state affect H2O2-induced changes in the transcriptome?

hyb_protocol: cad2-ld-1 Cy5 / Col0-ld-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.

value: mutant (cad2) -


title: 13756995 - cat2-ld-2 vs Col0-ld-2

description:
ch1: arabidopsis thaliana (columbia)  - harvest date: 07-07-08
ch2: arabidopsis thaliana (columbia)  mutant (cat2) - harvest date: 07-07-08
How does glutathione content or reduction state affect H2O2-induced changes in the transcriptome?

hyb_protocol: Col0-ld-2 Cy5 / cat2-ld-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 14054283 - Col0-ld-2 vs cat2xcytGR-ld-2

description:
ch1: arabidopsis thaliana (columbia)  mutant (cat2xcytgr) - harvest date: 07-07-08
ch2: arabidopsis thaliana (columbia)  - harvest date: 07-07-08
How does glutathione content or reduction state affect H2O2-induced changes in the transcriptome?

hyb_protocol: cat2xcytGR-ld-2 Cy5 / Col0-ld-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756941 - cytGR-sd-2 vs Col0-sd-2

hyb_protocol: Col0-sd-2 Cy5 / cytGR-sd-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.

growth_protocol: rosette - Media=soil hygrometry=70% Temperature=25°C  Light=200 µmol/m2/s short day=8h photoperiod.


title: 13756989 - Col0-ld-1 vs cytGR-ld-1

description:
ch1: arabidopsis thaliana (columbia)  mutant (cytgr) - harvest date: 07-07-08
ch2: arabidopsis thaliana (columbia)  - harvest date: 07-07-08
How does glutathione content or reduction state affect H2O2-induced changes in the transcriptome?

hyb_protocol: cytGR-ld-1 Cy5 / Col0-ld-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 14054281 - cat2xcytGR-ld-1 vs Col0-ld-1

hyb_protocol: Col0-ld-1 Cy5 / cat2xcytGR-ld-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13757350 - Col0-ld-2 vs cad2-ld-2

hyb_protocol: cad2-ld-2 Cy5 / Col0-ld-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13757349 - cad2-ld-1 vs Col0-ld-1

description:
ch1: arabidopsis thaliana (columbia)  - harvest date: 07-07-08
ch2: arabidopsis thaliana (columbia)  mutant (cad2) - harvest date: 07-07-08
How does glutathione content or reduction state affect H2O2-induced changes in the transcriptome?

hyb_protocol: Col0-ld-1 Cy5 / cad2-ld-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13757352 - cad2-ld-2 vs Col0-ld-2

hyb_protocol: Col0-ld-2 Cy5 / cad2-ld-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756988 - cat2xcad2-sd-2 vs Col0-sd-2

hyb_protocol: Col0-sd-2 Cy5 / cat2xcad2-sd-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756993 - cat2-ld-1 vs Col0-ld-1

hyb_protocol: Col0-ld-1 Cy5 / cat2-ld-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13757360 - Col0-sd-2 vs cytGR-sd-2

hyb_protocol: cytGR-sd-2 Cy5 / Col0-sd-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 14054280 - Col0-ld-1 vs cat2xcytGR-ld-1

hyb_protocol: cat2xcytGR-ld-1 Cy5 / Col0-ld-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756983 - cat2xcad2-sd-1 vs Col0-sd-1

hyb_protocol: Col0-sd-1 Cy5 / cat2xcad2-sd-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756982 - Col0-sd-1 vs cat2xcad2-sd-1

hyb_protocol: cat2xcad2-sd-1 Cy5 / Col0-sd-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: peripheral blood collected on day 3 from a pediatric non-responder to IVCS therapy SID_2588

status: Public on Aug 31 2010

city: Toronto

first: Boyko

last: Kabakchiev

state: Ontario

address: 60 Murray Street, Room L3-004

department: Samuel Lunenfeld Research Institute

country: Canada

institute: Mount Sinai Hospital

hyb_protocol: Samples were hybridized using Affymetrix GeneChip Hybridization, Wash and Stain Kit

scan_protocol: Affymetrix Gene ChIP Scanner 3000 7G

data_processing:
Probesets lacking annotation information were removed from further analysis. Raw data were background corrected, log transformed, and quantile normalized using a robust multi-array average (RMA) algorithm in R. All 40 samples were scaled using the R function “scale” (http://www.r-project.org/), and then correlated with a binary intravenous-corticosteroid therapy response variableusing the local-pooled error (LPE) method for detection of significance while controlling for batch effects.
probe group file: HuGene-1_0-st-v1.r4.pgf
meta-probeset file: HuGene-1_0-st-v1.r4.mps

label: biotin

organism: Homo sapiens

source_name: peripheral blood, 3 days on IVCS therapy

label_protocol: Samples were labeled using Affymetrix GeneChip Whole Transcript (WT) Sense Target Labeling Assay - 100 ng Total RNA Labeling Protocol

extract_protocol: Total RNA was extracted from all samples using the PAXgene Blood RNA Kit (QIAGEN).

treatment_protocol: Blood samples were collected in PAXgene tubes (PreAnalytiX, QIAGEN) and were incubated at room temperaturte between 2 and 24 hours priot to being frozen at -20°C. Samples were then stored at -80°C until extraction.

tag: tissue

value: peripheral blood

tag: age (in years)

value: No

tag: batch


title: 13756987 - Col0-sd-2 vs cat2xcad2-sd-2

hyb_protocol: cat2xcad2-sd-2 Cy5 / Col0-sd-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756979 - cad2-sd-1 vs Col0-sd-1

hyb_protocol: Col0-sd-1 Cy5 / cad2-sd-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756981 - cad2-sd-2 vs Col0-sd-2

hyb_protocol: Col0-sd-2 Cy5 / cad2-sd-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756974 - cat2xcytGR-sd-1 vs Col0-sd-1

hyb_protocol: Col0-sd-1 Cy5 / cat2xcytGR-sd-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756945 - cat2-sd-2 vs Col0-sd-2

hyb_protocol: Col0-sd-2 Cy5 / cat2-sd-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756942 - Col0-sd-1 vs cat2-sd-1

hyb_protocol: cat2-sd-1 Cy5 / Col0-sd-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756943 - cat2-sd-1 vs Col0-sd-1

hyb_protocol: Col0-sd-1 Cy5 / cat2-sd-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756975 - Col0-sd-1 vs cat2xcytGR-sd-1

hyb_protocol: cat2xcytGR-sd-1 Cy5 / Col0-sd-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756980 - Col0-sd-2 vs cad2-sd-2

hyb_protocol: cad2-sd-2 Cy5 / Col0-sd-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756977 - Col0-sd-2 vs cat2xcytGR-sd-2

hyb_protocol: cat2xcytGR-sd-2 Cy5 / Col0-sd-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756978 - Col0-sd-1 vs cad2-sd-1

hyb_protocol: cad2-sd-1 Cy5 / Col0-sd-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756938 - Col0-sd-1 vs cytGR-sd-1

hyb_protocol: cytGR-sd-1 Cy5 / Col0-sd-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756939 - cytGR-sd-1 vs Col0-sd-1

hyb_protocol: Col0-sd-1 Cy5 / cytGR-sd-1 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: 13756944 - Col0-sd-2 vs cat2-sd-2

hyb_protocol: cat2-sd-2 Cy5 / Col0-sd-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: Newman (ausA) late exponential replicate 1

status: Public on Jul 01 2010

city: Omaha

first: Paul

middle: M

last: Dunman

state: NE

address: 986495 Nebraska Medical Center

department: Pathology and Microbiology

institute: University of Nebraska Medical Center

hyb_protocol: 1.5 micrograms (ug) of labeled total RNA was applied to an S. aureus Affymetrix GeneChip hybridized @ 45 degrees Celsius (C) for 16 hr in an Affymetrix hybridization oven.  The sample was then washed 60 times in non-stringent buffer (Wash A; 6X SSPE, 0.01% Tween-20) at 45C followed by 60 washes in stringent buffer (Wash B; 1X MES, 25 mM NaCl, 0.01% Tween-20) at 45 C in an Affymetrix Fluidics station (450).  The sample was then subjected to the following stains/wash steps: 1. 10 min of Stain 1 (1X MES, 1.2 mg BSA, 6 ug Streptavidin) at 25C, 2. 40 washes with Wash A, 3. 10 min of Stain 2 (1X MES, 1.2 mg BSA, 60 ug Goat-IgG, and 3 ug biotinylated antistreptavidin), 4. 10 min of Stain 3 (1X MES, 1.2 mg BSA, 6 ug Streptavidin-Phycoerythrin), 5. 60 washes with Wash A at 25C.

scan_protocol: The GeneChip was scanned in an Affymetrix 7G scanner.

data_processing: The raw signal intensity value and present/absent determinations were calculated based on Affymetrix algorithms.

label: Biotin

organism: Staphylococcus aureus

source_name: Newman (ausA) late exponential

label_protocol: RNA was reverse transcribed, cDNA fragmented, 3’ biotinylated, and hybridized to S. aureus GeneChips following the manufacturer’s recommendations for antisense prokaryotic arrays (Affymetrix, Santa Clara, CA).

growth_protocol: S. aureus colonies from tryptic soy agar plates were used to inoculate tryptic soy broth (TSB; 1:100) and grown to late exponential phase (OD600 of 1.2).  RNA was extracted.

extract_protocol: Twenty ml of cells were added to 20 ml ice cold acetone:ethanol (1:1) and stored at -80oC overnight. For RNA isolation, -80oC suspensions were thawed on ice and centrifuged at 5,000 X g at 4oC for 10 min.  The supernatant was removed, the cell pellet was resuspended in 1 ml ice cold TE buffer (10 mM Tris, 1 mM EDTA, pH 8.0), and transferred to a pre-chilled lysing matrix B tube (Q-BIOgene, Irvine, CA).  Cells were lysed by shaking in an FP120 shaker (Q-BIOgene) two times at a setting of 4.5 for 20 s.  Suspensions were then centrifuged at 13,000 X g at 4oC for 10 min and the supernatant was used for RNA isolation with an RNeasy Mini column, following the manufacturer recommendations for on-the-column DNase treatment and RNA purification (Qiagen, Valencia, CA).

tag: strain

value: Newman

tag: genotype variation

value: aureusimine defective (ausA)


title: Newman early exponential replicate 2

source_name: Newman early exponential phase

growth_protocol: S. aureus colonies from tryptic soy agar plates were used to inoculate tryptic soy broth (TSB; 1:100) and grown to early exponential phase (OD600 of 0.3).  RNA was extracted.


title: Newman (ausA) early exponential replicate 2

source_name: Newman (ausA) early exponential


title: Newman late exponential replicate 2

source_name: Newman late exponential


title: Newman (ausA) early exponential replicate 1

source_name: Newman (ausA) early exponential phase


title: Newman (ausA) late exponential replicate 2


title: Newman late exponential replicate 1


title: Newman early exponential replicate 1

source_name: Newman Early Exponential  1


title: Untreated 3

status: Public on Jul 15 2010

city: Aurora

first: Tzu Lip

last: Phang

state: CO

address: 12700 East 19th Avenue

department: Medicine

institute: University of Colorado, Denver

description: K562 CML untreated

hyb_protocol: Following fragmentation, 10 ug of cRNA were hybridized for 16 hr at 45C on HGU133plus2 chip array. GeneChips were washed and stained in the Affymetrix Fluidics Station 400.

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

data_processing: Hybridization data was analyzed using GeneNet software (Systems Biosciences), which creates a report file in a format (.txt) compatible with common spreadsheet and statistical programs. The file lists the intensities of signal—which correspond to the abundance level—for each of the specific siRNA species in the library.

source_name: Chronic Myeloid Leukemia cell line

label_protocol: RNA was reverse-transcribed with M-MLV Reverse Transcriptase (Epicentre) followed by 2 rounds of PCR. During the second round of PCR with two nested primers (one primer has Biotin residues at the 5’-end), the amplified siRNA targets are labeled with biotin and can be used as hybridization targets with Affymetrix GeneChip® Arrays

growth_protocol: Cells were maintained in Iscove's Modified Dulbecco's Medium supplemented with 10% Fetal Bovine Serum, Pen Strep, and 2.5 µg/ml amphotericin B at 37°C in a 5% CO2 humidified atmosphere

extract_protocol: Total RNA was extracted using Trizol reagent followed by purification on a PureLink column according to the manufacturers instructions.

treatment_protocol: Cells were treated with imatinib mesylate at 1 µM for a period of 72 hr

tag: cell line

value: K562 Chronic Myeloid Leukemia

value: untreated


title: Imatinib 2

description: K562 CML imatinib-treated

value: imatinib


title: Imatinib 3

description: K562 CML imatinib-treated


title: Imatinib 1

description: K562 CML imatinib-treated


title: Untreated 1

description: K562 CML untreated


title: 13756976 - cat2xcytGR-sd-2 vs Col0-sd-2

hyb_protocol: Col0-sd-2 Cy5 / cat2xcytGR-sd-2 Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.


title: Untreated 2

description: K562 CML untreated


title: Tumor, T_310, Sty

type: genomic

city: Hanover

first: Brock

middle: Clarke

last: Christensen

state: NH

address: 7650 Remsen

institute: Dartmouth Medical School

description:
Hybridized to 250K_Sty.
T_310

hyb_protocol: As per standard manufacturer protocol (Affymetrix).

data_processing:
Data were collected at the Harvard Partners Microarray Core Facility. Inferred copy number states of each SNP were extracted and converted from .CEL files into signal intensities using GCOS and Affymetrix Copy Number Analysis Tool (CNAT) v4.0.1. We utilized the HMM-based algorithm with defaults for tuning parameters, Gaussian smoothing, transition decay, and median scaling.

The .CHP files contain the normalized, summarized signal intensities generated by the primary analysis software (CNAT). The 'GSE21057_tumor_copy_number.txt' file, which is linked to the Series GSE21057 record as a supplementary file, contains the (blood-adjusted) tumor copy number calls used in our final analyses.

molecule: genomic DNA

source_name: Mesothelioma

growth_protocol: Fresh-frozen human tissue.

extract_protocol: Qiagen Qiamp DNA extraction kit.

treatment_protocol: None.

value: pleural mesothelioma

tag: patient


title: Tumor, T_300, Sty

description:
Hybridized to 250K_Sty.
T_300

scan_protocol: As per standard manufacturer protocol (Affymetrix).


title: Tumor, T_264, Sty

description:
Hybridized to 250K_Sty.
T_264

label_protocol: As per standard manufacturer protocol (Affymetrix).


title: Tumor, T_294, Sty

description:
Hybridized to 250K_Sty.
T_294


title: Tumor, T_248, Sty

description:
Hybridized to 250K_Sty.
T_248


title: Tumor, T_247, Sty

description:
Hybridized to 250K_Sty.
T_247


title: Tumor, T_282, Sty

description:
Hybridized to 250K_Sty.
T_282


title: Tumor, T_310, Nsp

description:
Hybridized to 250K_Nsp.
T_310


title: Tumor, T_296, Sty

description:
Hybridized to 250K_Sty.
T_296


title: Tumor, T_284, Sty

description:
Hybridized to 250K_Sty.
T_284


title: Tumor, T_162, Sty

description:
Hybridized to 250K_Sty.
T_162


title: Tumor, T_236, Sty

description:
Hybridized to 250K_Sty.
T_236


title: Tumor, T_198, Sty

description:
Hybridized to 250K_Sty.
T_198


title: Tumor, T_296, Nsp

description:
Hybridized to 250K_Nsp.
T_296


title: Tumor, T_278, Sty

description:
Hybridized to 250K_Sty.
T_278


title: Tumor, T_225, Sty

description:
Hybridized to 250K_Sty.
T_225


title: Tumor, T_177, Sty

description:
Hybridized to 250K_Sty.
T_177


title: Tumor, T_290, Nsp

description:
Hybridized to 250K_Nsp.
T_290


title: Tumor, T_223, Sty

description:
Hybridized to 250K_Sty.
T_223


title: Tumor, T_224, Sty

description:
Hybridized to 250K_Sty.
T_224


title: Tumor, T_290, Sty

description:
Hybridized to 250K_Sty.
T_290


title: Tumor, T_159, Sty

description:
Hybridized to 250K_Sty.
T_159


title: Tumor, T_172, Sty

description:
Hybridized to 250K_Sty.
T_172


title: Tumor, T_294, Nsp

description:
Hybridized to 250K_Nsp.
T_294


title: Tumor, T_215, Sty

description:
Hybridized to 250K_Sty.
T_215


title: Tumor, T_284, Nsp

description:
Hybridized to 250K_Nsp.
T_284


title: Tumor, T_281, Sty

description:
Hybridized to 250K_Sty.
T_281


title: Tumor, T_023, Sty

description:
Hybridized to 250K_Sty.
T_023


title: Tumor, T_300, Nsp

description:
Hybridized to 250K_Nsp.
T_300


title: Matched blood, B_310, Sty

description:
Hybridized to 250K_Sty.
B_310

source_name: Blood


title: Matched blood, B_300, Sty

description:
Hybridized to 250K_Sty.
B_300


title: Tumor, T_282, Nsp

description:
Hybridized to 250K_Nsp.
T_282


title: Matched blood, B_296, Sty

description:
Hybridized to 250K_Sty.
B_296


title: Matched blood, B_278, Sty

description:
Hybridized to 250K_Sty.
B_278


title: Tumor, T_264, Nsp

description:
Hybridized to 250K_Nsp.
T_264


title: Tumor, T_278, Nsp

description:
Hybridized to 250K_Nsp.
T_278


title: Tumor, T_248, Nsp

description:
Hybridized to 250K_Nsp.
T_248


title: Matched blood, B_284, Sty

description:
Hybridized to 250K_Sty.
B_284


title: Matched blood, B_294, Sty

description:
Hybridized to 250K_Sty.
B_294


title: Matched blood, B_248, Sty

description:
Hybridized to 250K_Sty.
B_248


title: Matched blood, B_224, Sty

description:
Hybridized to 250K_Sty.
B_224


title: Matched blood, B_247, Sty

description:
Hybridized to 250K_Sty.
B_247


title: Tumor, T_236, Nsp

description:
Hybridized to 250K_Nsp.
T_236


title: Matched blood, B_281, Sty

description:
Hybridized to 250K_Sty.
B_281


title: Matched blood, B_282, Sty

description:
Hybridized to 250K_Sty.
B_282


title: Matched blood, B_162, Sty

description:
Hybridized to 250K_Sty.
B_162


title: Matched blood, B_225, Sty

description:
Hybridized to 250K_Sty.
B_225


title: Tumor, T_224, Nsp

description:
Hybridized to 250K_Nsp.
T_224


title: Tumor, T_247, Nsp

description:
Hybridized to 250K_Nsp.
T_247


title: Matched blood, B_290, Sty

description:
Hybridized to 250K_Sty.
B_290


title: Matched blood, B_223, Sty

description:
Hybridized to 250K_Sty.
B_223


title: Tumor, T_225, Nsp

description:
Hybridized to 250K_Nsp.
T_225


title: Matched blood, B_177, Sty

description:
Hybridized to 250K_Sty.
B_177


title: Matched blood, B_236, Sty

description:
Hybridized to 250K_Sty.
B_236


title: Matched blood, B_215, Sty

description:
Hybridized to 250K_Sty.
B_215


title: Tumor, T_215, Nsp

description:
Hybridized to 250K_Nsp.
T_215


title: Matched blood, B_264, Sty

description:
Hybridized to 250K_Sty.
B_264


title: Palate_Adamts9+/-:bt/bt_Rep3

city: Cleveland

first: Courtney

last: Nelson

state: OH

address: 9500 Euclid Avenue

department: Department of Biomedical Engineering

institute: Cleveland Clinic

hyb_protocol: cRNA was quantified using a nanodrop spectrophotometer and the cRNA quality (size distribution) was further analyzed on a 1% agarose gel. cRNA was hybridized to the Illumina MouseRef-8 v2 Expression BeadChip using standard protocols (provided by Illumina, San Diego, CA). Individual hybridizations were performed on each sample, and no pooling of samples took place.

scan_protocol: Scan and Imaging procedures adhered strictly to standard protocols provided by Illumina, San Diego, CA.

data_processing: The summarized data from the raw microarray data were log2 transformed and processed with force-positive background correction and quantile normalization.  Quality control analyses were applied to detect the outlier samples.

organism: Mus musculus

source_name: Secondary mouse palatal shelf tissue at embryonic stage E13.75

label_protocol: 250 ng of RNA was reverse transcribed into cRNA and biotin-UTP labeled using the Illumina TotalPrep RNA Amplification Kit (Ambion, Austin, TX).

growth_protocol: No growth protocol

extract_protocol: RNA was extracted from tissue samples using the RNeasy kit from Qiagen

treatment_protocol: No treatment of the tissue. They were directly taken for RNA after dissection

value: Murine palatal shelf tissue

tag: genotype/variation

tag: age


title: Palate_Adamts9+/-:bt/bt_Rep2

data_processing: The summarized data from the raw microarray data were log2 transformed and processed with force-positive background correction and quantile normalization. Quality control analyses were applied to detect the outlier samples.

extract_protocol: RNA was extracted from the tissue samples using the RNeasy kit from Qiagen


title: Matched blood, B_023, Sty

description:
Hybridized to 250K_Sty.
B_023


title: Palate_WT_Rep3

extract_protocol: RNA was extracted from the tissue samples using the RNeasy kit from Qiagen.

treatment_protocol: No treatment of the tissue. They were directly taken for RNA after dissection.


title: Palate_Adamts9+/-:bt/bt_Rep1


title: Palate_WT_Rep2


title: Matched blood, B_198, Sty

description:
Hybridized to 250K_Sty.
B_198


title: Matched blood, B_172, Sty

description:
Hybridized to 250K_Sty.
B_172


title: Matched blood, B_159, Sty

description:
Hybridized to 250K_Sty.
B_159


title: Palate_Adamts9+/-:bt/bt_Rep4


title: Palate_WT_Rep1


title: Tumor, T_177, Nsp

description:
Hybridized to 250K_Nsp.
T_177


title: Tumor, T_172, Nsp

description:
Hybridized to 250K_Nsp.
T_172


title: Tumor, T_281, Nsp

description:
Hybridized to 250K_Nsp.
T_281


title: Tumor, T_023, Nsp

description:
Hybridized to 250K_Nsp.
T_023


title: Tumor, T_162, Nsp

description:
Hybridized to 250K_Nsp.
T_162


title: Tumor, T_159, Nsp

description:
Hybridized to 250K_Nsp.
T_159


title: xenograft spheroid, biological rep 3

city: San Diego

first: Douglas Fang

last: Fang

state: CA

address: 10777 Science Center Drive, CB5

department: Oncology Research Unit

institute: Pfizer Inc.

hyb_protocol: Fifteen micrograms of labeled, fragmented cRNA was hybridized to Affymetrix HGU133 Plus 2.0 array (Affymetrix, Santa Clara, CA).

scan_protocol: GeneChips were scanned using Affymetrix GeneChip Scanner 3000.

data_processing: The data were RMA normalized using Bioconductor Affy package.

source_name: xenografted spheroid cells

label_protocol: For microarray sample processing, 2 µg of total RNA was used to synthesize cRNA using Affymetrix IVT one-cycle labeling kit per manufacturer's protocol.

growth_protocol: A non-small cell lung cancer cell line NCI-NCI-H1299, purchased from the ATCC (Manassas, VA), was cultured at 37  C, 5% CO2 in humidified atmosphere

extract_protocol: Total RNA was isolated using RNeasy kit (Qiagen, Germantown, MD) according to manufacturer's protocol.

treatment_protocol: To generate spheroid cultures, exponentially growing NCI-H1299 monolayer cultures were changed into the CSC medium. After 7-10 days of continual cultivation with addition of fresh culture medium every other day, the cells in suspension were collected and plated in ultra low attachment flasks (Corning, Corning, NY).

tag: medium

value: stem-cell medium

tag: condition

value: xenografted


title: xenograft spheroid, biological rep 1


title: xenograft parental, biological rep 3

source_name: xenografted parental cells

value: serum-containing medium


title: Matched blood, B_296, Nsp

description:
Hybridized to 250K_Nsp.
B_296


title: xenograft spheroid, biological rep 2


title: Tumor, T_223, Nsp

description:
Hybridized to 250K_Nsp.
T_223


title: Matched blood, B_278, Nsp

description:
Hybridized to 250K_Nsp.
B_278


title: Matched blood, B_281, Nsp

description:
Hybridized to 250K_Nsp.
B_281


title: Matched blood, B_310, Nsp

description:
Hybridized to 250K_Nsp.
B_310


title: Matched blood, B_300, Nsp

description:
Hybridized to 250K_Nsp.
B_300


title: Matched blood, B_282, Nsp

description:
Hybridized to 250K_Nsp.
B_282


title: Matched blood, B_264, Nsp

description:
Hybridized to 250K_Nsp.
B_264


title: cultured spheroid, biological rep 2

source_name: cultured spheroid cells

value: cultured


title: cultured spheroid, biological rep 1


title: Matched blood, B_248, Nsp

description:
Hybridized to 250K_Nsp.
B_248


title: xenograft parental, biological rep 1


title: Matched blood, B_236, Nsp

description:
Hybridized to 250K_Nsp.
B_236


title: Matched blood, B_224, Nsp

description:
Hybridized to 250K_Nsp.
B_224


title: Matched blood, B_290, Nsp

description:
Hybridized to 250K_Nsp.
B_290


title: Matched blood, B_247, Nsp

description:
Hybridized to 250K_Nsp.
B_247


title: Matched blood, B_223, Nsp

description:
Hybridized to 250K_Nsp.
B_223


title: Matched blood, B_284, Nsp

description:
Hybridized to 250K_Nsp.
B_284


title: cultured parental, biological rep 2

source_name: cultured parental cells


title: cultured spheroid, biological rep 3


title: Matched blood, B_162, Nsp

description:
Hybridized to 250K_Nsp.
B_162


title: xenograft parental, biological rep 2


title: Matched blood, B_294, Nsp

description:
Hybridized to 250K_Nsp.
B_294


title: Matched blood, B_215, Nsp

description:
Hybridized to 250K_Nsp.
B_215


title: Matched blood, B_198, Nsp

description:
Hybridized to 250K_Nsp.
B_198


title: Matched blood, B_023, Nsp

description:
Hybridized to 250K_Nsp.
B_023


title: Matched blood, B_172, Nsp

description:
Hybridized to 250K_Nsp.
B_172


title: Matched blood, B_177, Nsp

description:
Hybridized to 250K_Nsp.
B_177


title: cultured parental, biological rep 3


title: Matched blood, B_159, Nsp

description:
Hybridized to 250K_Nsp.
B_159


title: Matched blood, B_225, Nsp

description:
Hybridized to 250K_Nsp.
B_225


title: Tumor, T_198, Nsp

description:
Hybridized to 250K_Nsp.
T_198


title: cultured parental, biological rep 1


title: CS line, 2 d after Fg inoculation, rep 2

status: Public on Jun 15 2010

city: Ottawa

first: Thérèse

last: Ouellet

state: ON

address: 960 Carling Ave, room 2091

department: Eastern Cereal and Oilseed Research Centre

institute: Agriculture and Agri-Food Canada

description: Gene expression data from Fusarium graminearum-inoculated wheat spikelets from parental line CS, 2 DAI

hyb_protocol: According to Affymetrix instructions for the GeneChip® HT One-Cycle Target Labeling and Controls Kit. All samples were processed in one experimental batch.

scan_protocol: According to Affymetrix instructions for the GeneChip® HT One-Cycle Target Labeling and Controls Kit, using the Affymetrix GeneChip® 3000 Scanner.

data_processing: The probe-level data for the whole profiling experiment were assembled into one dataset using the Robust Multichip Analysis (Irizarry et al, 2002) implemented in the software Acuity 4.0 (Molecular Devices, CA, USA), with quantile normalization and summarization by median polish.

organism: Triticum aestivum

growth_protocol: Plants were grown in growth cabinets at 20 C/16 hrs daylight; 16 C/8 hrs night until heads reach mid anthesis; then they were moved to the misting room.

extract_protocol: Total RNA from each sample was isolated using a modified form of the guanidine isothiocyanate-cesium chloride method of Chirgwin et al (Biochem. 18: 5294-5299, 1979), as outlined in Ohan and Heikkila (Dev. Genet. 17: 176-184, 1995).  The RNA samples were treated with an RNase-free DNase (Qiagen, Mississauga, Canada) and cleaned up using the RNeasy Mini Kit (Qiagen) according to manufacturer’s instructions.

treatment_protocol: Wheat heads at mid-anthesis were inoculated as follow: 10 µl of a 100,000 spores/ml suspension of the isolate Fusarium graminearum strain DAOM 180378 was point-inoculated into each of two basal florets of each developed spikelet of the wheat heads.  Water was used as a mock-inoculation treatment.  The inoculated plants were mist-irrigated for 10 second intervals every 15 minutes for 48 hours to maintain high humidity at a temperature of 25 oC. Five treated heads were used for each biological replicate, and inoculated spikelets were sampled at 0, 2 and 4 days post-inoculation (DPI).  The sampled tissues were immediately frozen in liquid nitrogen and stored at -80 C until used.

value: Chinese Spring wheat

value: inoculated spikelets

value: heads after mid-anthesis

value: Fusarium graminearum

tag: days after inoculation


title: CS line, 4 d after water inoculation, rep 2

description: Gene expression data from mock-inoculated wheat spikelets from parental line CS, 4 DAI

source_name: Mock-inoculated wheat spikelets from parental line CS, 4 DAI

label_protocol: According to Affymetrix instructions for the GeneChip® HT One-Cycle Target Labeling and Controls Kit. All samples were processed in one experimental batch.

tag: genotype

value: Water


title: MOF males_rep3

status: Public on Jun 26 2010

city: Munich

first: Christian

last: Feller

address: Schillerstraße 44

department: Molecular Biology

country: Germany

institute: Adolf-Butenandt Institute at the LMU University Munich

hyb_protocol: performed by imaGenes (www.imaGenes-bio.de) using standard NimbleGen protocols.

data_processing: raw signals of all channels in all replicate arrays were normalized and log2 transformed using the 'vsn' package in R/Bioconductor.

organism: Drosophila melanogaster

source_name: MOF ChIP DNA from males

extract_protocol: Chromatin was prepared from sex-sorted adult flies. The protocol from Negre et al. (Negre et al., 2006) was applied with some modification: 300 mg of 3-4 days old flies were crushed in 5 ml A1 (60 mM KCl, 15 mM NaCl, 4 mM MgCl2, 15 mM HEPES (pH 7.6), 0.5% Triton X-100, 0.5 mM DTT, 10 mM sodium butyrate, protease inhibitor cocktail (Roche, Cat No. 04693132011)) with a final concentration of 2.35% formaldehyde at 18°C for 15 min. Fixation was stopped with a final concentration of 250 mM glycine. After 3 washes with A1 the pellet was equilibrated in lysis buffer (140 mM NaCl, 15 mM HEPES (pH 7.6), 1 mM EDTA, 0.5 mM EGTA, 1% Triton X- 100, 0.5% sodium deoxycholate, protease inhibitor cocktail, 0.5mM DTT). The pellet was suspended in lysis buffer with 0.1% SDS and 0.5% N-lauroylsarcosine and rotated for 10 min at 4°C. Chromatin was sonicated (Branson (microtip), 4 times at 21 watt) in the presence of glass beads (212-300 μm). Cell debris was removed by centrifugation and the chromatin containing supernatant was stored in aliquots at -80°C. For chromatin immunoprecipitation (ChIP) the DNA concentration of the chromatin was determined and 7.5 μg of DNA was used per IP in 500 μl lysis buffer plus 250 μl PBS (1xPBS, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, protease inhibitor cocktail. Chromatin was pre-cleared with protein-A and -G beads at 4°C for 30 min. After pre-clearing an aliquot was saved for input DNA purification. Immunoprecipitation was carried out at 4°C overnight. The amount of antibody used per ChIP was adjusted individually. For the pull down 30 μl protein A and G beads were used per ChIP at 4°C for 2 h. Beads were washed 5 times in PBS, rinsed once in EB buffer (10 mM Tris-Cl, pH 8.5). De-crosslinking was performed in 100 μl EB with 0.6% SDS and a final concentration of 0.5 μg proteinase K and RNAse A. Digest was carried out at 55°C for 3 h and de-crosslinking at 65°C for 6 h. ChIP and input DNA were purified with phenol/chloroform using MaXtract High Density tubes (Qiagen, Cat No. 129056) and subsequent ethanol precipitation. Chromatin preparation of SL2 cells were performed as previously described in Straub et al. (2008). Briefly, cells were crosslinked in growth medium using 1% formaldehyde for 60 minutes in icewater. Fixation was quenched by addition of glycine to a final concentration of 125 mM. After washing, cells were resuspended in RIPA buffer and sonciated using the Bioruptor (Diagenode, Belgium) 8 times 30 seconds using the ‘high’ setting. Fragment size of the obtained chromatin was checked to be between 300 bp and 700 bp. Chromatin was precleared using a protein A/protein G-sepharose mixture for 1 hr at 4°C. 200 µl chromatin was incubated with appropriate amounts of antibodies in a total volume of 500 µl RIPA buffer at 4°C over night. After washing and crosslink revearsal, immunprecipitated nucleic acids were purified on GFX columns (GE Healthcare). IP DNA was amplified using the WGA kit (Sigma). For input DNA, chromatin was not subjected to an IP but otherwise processed identically to the ChIP material.

value: whole flies

tag: developmental stage

value: adult

tag: gender

value: males

tag: replicate

value: replicate 3

tag: chip antibody

value: MOF

value: none, input DNA


title: CS line, 2 d after Fg inoculation, rep 1

source_name: Fusarium graminearum-inoculated wheat spikelets from parental line CS, 2 DAI

tag: tissue

tag: agent


title: CS line, 4 d after Fg inoculation, rep 2

description: Gene expression data from Fusarium graminearum-inoculated wheat spikelets from parental line CS, 4 DAI


title: MSL1 males_rep4

scan_protocol: performed by imaGenes (www.imaGenes-bio.de) using standard NimbleGen protocols.

source_name: MSL1 ChIP DNA from males

value: replicate 4


title: CS line, 4 d after water inoculation, rep 1


title: CS-7EL line, 2 d after Fg inoculation, rep 2

description: Gene expression data from Fusarium graminearum-inoculated wheat spikelets from addition line CS-7EL, 2 DAI

value: Chinese Spring with Thinopyrum elongatum chromosome arm 7EL


title: CS line, 4 d after Fg inoculation, rep 1

source_name: Fusarium graminearum-inoculated wheat spikelets from parental line CS, 4 DAI


title: MSL1 males_rep2

label_protocol: performed by imaGenes (www.imaGenes-bio.de) using standard NimbleGen protocols.

value: replicate 2


title: CS line, 4 d after Fg inoculation, rep 3


title: MOF males_rep2


title: CS-7EL line, 4 d after water inoculation, rep 2

description: Gene expression data from mock-inoculated wheat spikelets from addition line CS-7EL, 4 DAI

source_name: Mock-inoculated wheat spikelets from addition line CS-7EL, 4 DAI


title: CS-7EL line, 4 d after Fg inoculation, rep 2

description: Gene expression data from Fusarium graminearum-inoculated wheat spikelets from addition line CS-7EL, 4 DAI


title: MSL1 males_rep1

value: replicate 1


title: CS-7EL line, 4 d after Fg inoculation, rep 1

source_name: Fusarium graminearum-inoculated wheat spikelets from addition line CS-7EL, 4 DAI


title: MBD-R2 males_rep1

source_name: MBD-R2 ChIP DNA from males


title: MBD-R2 females_rep2

source_name: MBD-R2 ChIP DNA from females

value: females


title: CS-7E line, 2 d after Fg inoculation, rep 2

description: Gene expression data from Fusarium graminearum-inoculated wheat spikelets from addition line CS-7E, 2 DAI

value: Chinese Spring with Thinopyrum elongatum chromosome 7E


title: MBD-R2 females_rep5

value: replicate 5


title: MSL1 males_rep3


title: OB06a

status: Public on Jun 01 2010

city: Minneapolis

first: Anthony

middle: James

last: Smith

state: MN

address: 420 Delaware Street S.E.

department: Microbiology

institute: University of Minnesota

description: lymph node, biological replicate 1

hyb_protocol: Fifteen micrograms of fragmented cRNA was hybridized to an Affymetrix Human Genome U133 Plus 2.0 array.

scan_protocol: After hybridization, chips were washed, stained with streptavidin-phycoerythrin, and scanned with GeneChip Operating Software at the Biomedical Genomics Center at the University of Minnesota.  The experiments from each RNA sample were duplicated in the preparation of each cRNA probe and microarray hybridization.

data_processing: The .cel files produced by the Affymetrix data analysis platform were uploaded into the Expressionist program (Genedata, Pro version 4.5) and the expression level for each of the ~56,000 probe sets on the arrays was quantified using the RMA algorithm.  The mean expression level from duplicate chips from the same individual’s RNA was computed and used in the subsequent analysis.  The RMA algorithm produces a summary of gene expression that is of log scale; we utilized this log scale for all analyses. To examine the relationship between gene expression and viral load, a linear regression model was fit with gene expression as the response variable and disease stage and the log of viral load as predictor variables.  We controlled for disease stage when examining the relationship between viral load and gene expression because we previously found an association between gene expression and disease stage for HIV-1 infection [9] (additionally, there is an association between disease stage and viral load for HIV-1-infected subjects naïve to HAART).  The p-values for the null hypothesis (testing no association between viral load and gene expression) were then transformed to q-values using the q-value package in the statistical software program R.  By varying the threshold for the q-values, one can obtain a set of genes that are found to be associated with viral load at a given false discovery rate. As the number of genes found to be associated with viral load increases dramatically as this threshold rises above ~7%, we used the conservative cut-off value of 7% for the false discovery rate.  To summarize the association between log of viral load and gene expression, we computed the partial correlation between these two quantities for all genes while controlling for the effect of disease stage.

source_name: Inguinal lymph node from HIV-1-infected individual

label_protocol: Double stranded cDNA and biotin-labeled cRNA probes were synthesized from 5 micrograms of total RNA with a MessageAmp II aRNA kit (Ambion). The cRNA probes were column purified and fragmented with a Fragmentation kit (Ambion).

extract_protocol: Inguinal lymph node biopsies from 22 HIV-1-infected subjects at different clinical stages were obtained for this University of Minnesota institutional review board–approved microarray study. Each lymph node biopsy was placed into a Falcon tube and snap-frozen by dropping it into liquid nitrogen. Frozen lymph nodes were homogenized with a power homogenizer (Heat Systems Ultrasonic) in TRIzol (Invitrogen) without thawing.  Total RNA was isolated following the manufacturer’s protocol and further purified with an RNeasy Mini Kit (Qiagen).

value: Inguinal lymph node

value: Male

tag: race

value: African American

tag: peripheral blood cd4+ t cells (cells/ml)

tag: plasma hiv-1 rna levels (copies/ml)


title: MBD-R2 females_rep3


title: MOF females_rep3

source_name: MOF ChIP DNA from females


title: CS-7E line, 4 d after Fg inoculation, rep 1

description: Gene expression data from Fusarium graminearum-inoculated wheat spikelets from addition line CS-7E, 4 DAI

value: inoculated spikelets

value: heads after mid-anthesis

value: Fusarium graminearum


title: CS-7EL line, 2 d after Fg inoculation, rep 1

source_name: Fusarium graminearum-inoculated wheat spikelets from addition line CS-7EL, 2 DAI


title: OB06b

description: lymph node, biological replicate 2


title: CS-7E line, 4 d after Fg inoculation, rep 2

source_name: Fusarium graminearum-inoculated wheat spikelets from addition line CS-7E, 4 DAI


title: CS-7E line, 4 d after Fg inoculation, rep 3


title: MOF females_rep1

value: whole flies

value: adult


title: MBD-R2 SL2_replicate2

source_name: MBD-R2 ChIP DNA from SL2

tag: cell line


title: CS-7EL line, 4 d after water inoculation, rep 1

value: Water


title: MOF females_rep2

value: MOF

value: none, input DNA


title: CS-7E line, 2 d after Fg inoculation, rep 3

source_name: Fusarium graminearum-inoculated wheat spikelets from addition line CS-7E, 2 DAI


title: MOF males_rep1

value: males


title: GL38b

description: lymph node, biological replicate 2

value: Caucasian


title: VM27a

description: lymph node, biological replicate 1

value: Female


title: TK62b

description: lymph node, biological replicate 2


title: MBD-R2 males_rep2

value: replicate 2


title: MBD-R2 SL2_replicate1

value: replicate 1


title: MBD-R2 males_rep4

value: replicate 4


title: TK62a

description: lymph node, biological replicate 1

value: Inguinal lymph node


title: SU36a

description: lymph node, biological replicate 1

value: Male


title: TK07a

description: lymph node, biological replicate 1


title: WU59b

description: lymph node, biological replicate 2


title: KS64a

description: lymph node, biological replicate 1


title: VM27b

description: lymph node, biological replicate 2

value: African American


title: MBD-R2 males_rep3

value: replicate 3


title: DS35b

description: lymph node, biological replicate 2


title: DS35a

description: lymph node, biological replicate 1


title: WU59a

description: lymph node, biological replicate 1


title: RJ56b

description: lymph node, biological replicate 2


title: RJ56a

description: lymph node, biological replicate 1


title: RM08a

description: lymph node, biological replicate 1


title: MBD-R2 females_rep4

value: females


title: MBD-R2 females_rep1


title: GL38a

description: lymph node, biological replicate 1


title: SU36b

description: lymph node, biological replicate 2


title: RM08b

description: lymph node, biological replicate 2


title: JR31a

description: lymph node, biological replicate 1


title: CF63a

description: lymph node, biological replicate 1


title: CF63b

description: lymph node, biological replicate 2


title: TS35a

description: lymph node, biological replicate 1


title: KS64b

description: lymph node, biological replicate 2


title: PH29a

description: lymph node, biological replicate 1


title: TH49a

description: lymph node, biological replicate 1


title: PH29b

description: lymph node, biological replicate 2


title: JR31b

description: lymph node, biological replicate 2


title: WB91a

description: lymph node, biological replicate 1


title: WB55b

description: lymph node, biological replicate 2


title: PR89b

description: lymph node, biological replicate 2


title: WB55a

description: lymph node, biological replicate 1


title: MH58a

description: lymph node, biological replicate 1


title: MH58b

description: lymph node, biological replicate 2


title: DP24b

description: lymph node, biological replicate 2


title: JS30a

description: lymph node, biological replicate 1


title: TH49b

description: lymph node, biological replicate 2


title: Elk/Sapko3.30CD3

city: London

first: Robert

middle: Harris

last: Nicolas

address: 44 Lincoln's Inn feilds

department: Transcription

country: United Kingdom

institute: LRI Cancer research UK

description: cDNA from Elk/Sap KO Tcell DPs mouse 3 treated with anti CD3 for 30 min

hyb_protocol: PICR 11uM feature

scan_protocol: Genechips were scanned using the Affymetrix Genechip Scanner 3000G

data_processing: Data was analysed within Bioconductor. Probe set expression measures were calculated using Robust Multichip Average default method from the Affy package.

source_name: Elk/Sap KO/anti CD3 for 30 min

label_protocol: PICR 1 cycle Taget labelling V 1, cocktail V2 (DMSO)

growth_protocol: DP Tcells were purified on the cell sorter using anti-CD4 and anti-CD8 the Cells were then rested for 2 hours  in RPMI with 10% FCS at 37°C.

extract_protocol: Total RNA was purified further with the GenElute kit (Sigma) according to the manufacturers instructions

treatment_protocol: The DPs were spun onto anti-CD3 coated or non-coated plates and incubated 30min

tag: strain

value: Black 6J

value: male

value: 8 weeks

value: thymus

tag: cell type

value: double positive T cells (DPs)

value: Elk/Sap KO

tag: treatment

value: anti CD3 for 30 min


title: Elkko3.0

description: cDNA from Elk KO Tcell DPs mouse 3

source_name: Elk KO/Untreated

tag: age

tag: genotype/variation

value: Elk KO

value: Untreated


title: DP24a

description: lymph node, biological replicate 1

value: Caucasian


title: Elkko1.30CD3

description: cDNA from Elk KO Tcell DPs mouse 1 treated with anti CD3 for 30 min

source_name: Elk KO/anti CD3 for 30 min


title: Elk/Sapko1.30CD3

description: cDNA from Elk/Sap KO Tcell DPs mouse 1 treated with anti CD3 for 30 min


title: Elk/Sapko3.0

description: cDNA from Elk/Sap KO Tcell DPs mouse 3

source_name: Elk/Sap KO/Untreated


title: Elk/Sapko1.0

description: cDNA from Elk/Sap KO Tcell DPs mouse 1


title: Elkko2.0

description: cDNA from Elk KO Tcell DPs mouse 2


title: Elkko1.0

description: cDNA from Elk KO Tcell DPs mouse 1


title: Elk/Sapko2.30CD3

description: cDNA from Elk/Sap KO Tcell DPs mouse 2 treated with anti CD3 for 30 min


title: Elkko3.30CD3

description: cDNA from Elk KO Tcell DPs mouse 3 treated with anti CD3 for 30 min


title: Sapko2.30CD3

description: cDNA from Sap KO Tcell DPs mouse 2 treated with anti CD3 for 30 min

source_name: Sap KO/anti CD3 for 30 min

value: Sap KO


title: JS30b

description: lymph node, biological replicate 2


title: Elk/Sapko2.0

description: cDNA from Elk/Sap KO Tcell DPs mouse 2


title: Elkko2.30CD3

description: cDNA from Elk KO Tcell DPs mouse 2 treated with anti CD3 for 30 min


title: TK07b

description: lymph node, biological replicate 2


title: Sapko1.30CD3

description: cDNA from Sap KO Tcell DPs mouse 1 treated with anti CD3 for 30 min


title: Sapko2.0

description: cDNA from Sap KO Tcell DPs mouse 2

source_name: Sap KO/Untreated


title: WT1.0

description: cDNA from WT Tcell DPs mouse 1

source_name: wild-type/Untreated

value: wild-type


title: Sapko3.30CD3

description: cDNA from Sap KO Tcell DPs mouse 3 treated with anti CD3 for 30 min


title: WT1.30CD3

description: cDNA from WT Tcell DPs mouse 1 treated with anti CD3 for 30 min

source_name: wild-type/anti CD3 for 30 min


title: WT3.30CD3

description: cDNA from WT Tcell DPs mouse 3 treated with anti CD3 for 30 min


title: WT2.30CD3

description: cDNA from WT Tcell DPs mouse 2 treated with anti CD3 for 30 min


title: Sapko3.0

description: cDNA from Sap KO Tcell DPs mouse 3


title: WT3.0

description: cDNA from WT Tcell DPs mouse 3


title: WT2.0

description: cDNA from WT Tcell DPs mouse 2


title: Stroma_treatment HPHF_S 21

city: Corvallis

first: Massimo

last: Bionaz

state: OR

address: 316A Withycombe Hall

department: Animal and Rangeland Sciences

institute: Oregon State University

description:
A mixture of total RNA from 5 bovine tissues (mammary, liver, muscle, adipose, and placenta) were used as reference (Cy5 in Ch1 and Cy3 in Ch2).
S 21

hyb_protocol: Labelled cDNA hybridised using 80 μL of the hybridization buffer #1 (Ambion).  Slides placed in humidified slide chambers (Corning) in water bath for 40 h at 42 °C. Prior to scanning, the microarray were washed for 5 min by agitation 3 times with wash buffers in the following order: 1×SSC and 0.2% SDS solution preheated at 42°C, 0.1×SSC and 0.2% SDS solution, and 0.1×SSC solution. Lastly, slides were inserted into a 50 mL tube, spin-dried and gassed with Argon to preserve dye from bleaching.

scan_protocol: Arrays were scanned with a ScanArray 4000 (GSI-Lumonics) dual-laser confocal scanner and images were processed and edited using GenePix 6.0 (Axon Instruments).

data_processing: Array quality was assessed using an in-house parser written in Perl language. Spots that received a -100 flag by GenePix 6.0 were removed from further analysis and background intensity was subtracted from the foreground intensity. Spots on the slide were considered “good” if the median intensity was ≥3 standard deviation above median background for each channel (i.e., dye). Spots were flagged “present” when both dyes passed the criteria, “marginal” if only one dye passed the criteria, or “absent” when both dyes failed to pass the criteria. Statistical analysis was conducted on oligos that were flagged as “present” and “marginal”. Data from a total of 82 microarrays were normalized for dye and array effects (i.e., Lowess normalization and array centering and scaling) before statistical analysis.

organism: Bos taurus

source_name: Stromal tissue, HPHF diet

label_protocol: Labelled with Cy3 and Cy5 during cDNA synthesis from total RNA using Superscript III Kit (Invitrogen) as per manufacturer's instructions.

extract_protocol: Calves were killed and their mammary glands were removed. A sample of parenchyma and fat pad was dissected and snap-frozen in liquid nitrogen. Tissue was weighed (~0.5 g) and RNA extracted using ice-cold TRIZol. Genomic DNA was removed from RNA with DNAse (Qiagen) using RNeasy Mini Kit columns (Qiagen).

treatment_protocol: Holstein calves (1-3 days old) were fed 1 of 4 different milk replacers until the age of 65 days old.

tag: breed

value: Holstein

value: 65 days

tag: mammary tissue

value: stroma (fat pad)

tag: diet

value: HPHF

source_name: Reference

extract_protocol: Calves were killed, and tissue samples were dissected and snap-frozen in liquid nitrogen. Tissue was weighed (~0.5 g) and RNA extracted using ice-cold TRIZol. Genomic DNA was removed from RNA with DNAse (Qiagen) using RNeasy Mini Kit columns (Qiagen).

value: mammary, liver, muscle, adipose, placenta