title: Immobilised LSP BIO3

type: mixed

city: Kgs. Lyngby

first: Gitte

middle: M.

last: Knudsen

address: Søltofts Plads, Building 221

department: National Food Institute

country: Denmark

institute: Technical University of Denmark

hyb_protocol: Slides blocked with DCE according to protocol at PMID 11266573. Microarray Hybridisations  1. To 10 μl of dried down labelling reactions add:  a) 1.5 μl of 50 X Denhardts solution  b) 2.25 μl of 20 X SSC  c) 1.125 μl of E. coli tRNA (10 μg/μl) (Sigma, Cat: R8759)  d) 0.375 μl of 1M HEPES, pH 7.0.  NOTE: The hybridisation components can be mixed together as a stock solution, either from a fresh or frozen stock, and added to labelling reactions.  2. Add 0.375 μl of 10% SDS to the mixture  3. Incubate 100 °C for 2 min. Let stand on the bench for 5 - 10 min. Do not place the reaction on ice because the SDS will precipitate.  4. Spin in a microfuge at full speed for 5 min, transfer supernatant to clean tube and repeat spin.  5. Place a microarray slide in a metal hybridisation chamber (Photo 3).  6. Pipette the hybridisation solution towards one edge of an array.  7. Place edge of a clean (washed with 70% ethanol), dry, coverslip on the edge of the array.  8. Position fine nosed forceps (VWR, Cat: 406/0073/04) under the coverslip.  9. Using the forceps slowly lower the coverslip down onto the hybridisation solution and the array ensuring no air bubbles are present under the coverslip. If air bubbles are present it is sometimes possible to move them to the edge of the coverslip by pressing gently with the forceps.  10. Apply 20 μl of 3 X SSC around the 4 corners of the slide using a pipette (approximately 5 μl per corner). This will maintain the correct humidity in the hybridisation chamber.  11. Screw the lid onto the chamber and place at the bottom of a water bath at 63°C overnight.  12. Remove the slides from the hybridisation chambers and place them in a plastic slide rack (Photo 4). The slides are then placed into a 63°C wash solution containing 2 X SSC, 0.1% SDS and agitated for 5 mins. (We use a 2L beaker with a stirrer bar in  the bottom and a platform for the slide rack to sit on for this, Photos 5 & 6). It may be necessary to tap the slide rack on the platform to make sure that the cover slips are removed. We use a thermocouple attached to a hot-plate stirrer for this step. Repeat the washing step.  Note: SDS fluoresces so you should avoid carrying it over into the washes in steps 13 and 14.  13. Transfer slide rack to a solution of 1 X SSC at room temperature and agitate for 5 min. Repeat. (We use black tissue boxes often used for histology for this)  14. Wash in 0.2 X SSC for 5 min. Repeat.  15. Spin dry in a centrifuge at 1200 rpm for 5 min at room temp in an enclosed slide chamber (with blotting paper lining the bottom of the chamber).  16. Scan.  NB All solutions are filter sterilised (0.1 μm) and made up with Sigma water (molecular biology reagent, Sigma, Cat: W4502).

scan_protocol: Axon GenePix4000A 10 µm resolution scan and see http://www.ifr.ac.uk/safety/Microarrays/default.html#protocols

data_processing: Intensity ratios of channel 1 (cDNA) divided by channel 2 (reference gDNA) signals are calculated using BlueFuse 3.1 or 3.6 (BlueGnome, Cambridge, UK). Data centering was subsequently performed using the block median

organism: Salmonella enterica subsp. enterica serovar Typhimurium str. ST4/74

source_name: S. Typhimurium Immobilised LSP BIO3

label_protocol: Set up random priming reactions in 1.5 ml microfuge tubes. Add:  a) 10 μg of total RNA (this may require concentrating using a speed vac)  b) 5 μg of random hexamers (Invitrogen, Cat: 48190011)  c) In a total volume of 9.4 μl of Sigma ultra-pure water (mol. biol. reagent, Cat: W4502).  d) Incubate 70°C for 5 min protected from light then chill on ice for 10 min. Spin briefly in microfuge.  2) Using the Stratagene AffinityScript multi-temperature Reverse Transcriptase (Cat: 600109) Prepare RT reaction mix (sufficient for one labelling reaction)  a) 2.0 μl of 10 X RT buffer  b) 2.0 μl of 0.1 M DTT  c) 0.6 μl of 50 X dNTP's *  3) Add RT reaction mix to RNA (4.6 μl per reaction)  4) Finally add:  a) 2 μl of Cy3 or Cy5-dCTP (1mM stock, GE Healthcare Lifesciences, Cat: PA55321)  b) 4 μl of reverse transcriptase.  The total reaction volume is 20 μl.  5) Mix and incubate at 25oC for 10 mins.  6) Incubate overnight at 42°C.  7) Add 15 μl of freshly prepared 0.1M NaOH and hydrolyse the RNA at 70°C for 10 minutes. Add 15 μl of 0.1M HCl to neutralise the alkali.  8) If performing a type I experiment follow section ‘a’ or for a type II follow section ‘b’  a) If performing a type I experiment (RNA verses RNA) (DeRisi et al, Science, 1997, 278, 680-686), combine reactions and clean up using Qia-quick PCR purification kit (Qiagen, Cat: 28104) to remove unincorporated/quenched cy dyes. Elute twice using 50 μl Sigma water as final eluant to maximise recovery. Speed vac on medium setting and redissolve pellet in 10 μl Sigma water (mol. biol. reagent, Cat: W4502). Proceed to 'Microarray Hybridisations' protocol.  b) If performing a type II experiment (RNA verses genomic DNA), mix labelled cDNA with labelled genomic DNA (see 'Direct labelling of DNA' protocol). Clean up using Qia-quick PCR purification kit (Qiagen, Cat: 28104) to remove unincorporated/quenched cy dyes. Elute twice using 50 μl Sigma water as final eluant to maximise recovery. Speed vac on medium setting and redissolve pellet in  www.ifr.ac.uk/safety/molmicro  Molecular Microbiology Group, Institute of Food Research, Norwich Research Park,  Colney, Norwich NR4 7UA UK  10 μl Sigma water (mol. biol. reagent, Cat: W4502). Proceed to 'Microarray Hybridisations' protocol.  *To prepare 50 X dNTP's, mix 25 mM dATP, dGTP, dTTP and 10 mM dCTP, (25 μl of dA, dG, dT and 10 μl of dCTP from 100 mM stock dNTP kit (GE Healthcare Lifesciences, Cat: 27-2035-02). Add Sigma water to 100 μl.  NB: Sigma water (molecular biology reagent, Cat: W4502) was used to prepare all solutions

growth_protocol: Inoculum cultures  were obtained by inoculating a few colonies in 10 ml LB broth and incubated statically at 25°C for 24 hr. Then a second subculture was made by inoculation 100 µl of the first subculture in 10 ml LB broth, which was incubated under the same conditions. For immobilised and planktonic growth of Salmonella was the IFR Gel Cassette system used. For immobilised growth, a 24 hr standard inoculum culture was diluted 1000-fold in diluents, which was futher diluted 1000-fold in LB with 22% (w/w) Pluronic® to give 103 CFU g per gel. This mixture was loaded into sterile Gel Cassettes. For planktonic growth in the Gel Cassette system, metal holders with a metal mesh or acetal frames with 644 holes was used. Similar to the gel, the standard inocula were diluted and inoculated at 103 CFU per ml in LB broth. Immobilised and planktonic cultures were incubated at 25°C without aeration.

extract_protocol: 1) Harvest 2.0 OD600 units of bacterial culture (for example 4 mls of a culture with an OD600 = 0.5)  2) Prepare a 50 ml Falcon tube containing 2/5 of the culture volume ice-cold 5% (v/v) phenol pH 4.3, 95% (v/v) ethanol (Phenol Sigma order code: P-4682). Add the appropriate volume of culture e.g. add 4 mls of culture to 1.6 mls of phenol/ethanol. Stand on ice for at least 30 min but no longer than 2 hours to stabilise the RNA and prevent degradation.  3) Centrifuge samples at 3220 X g 4°C for 10 min. Discard supernatant, resuspend bacterial pellets using residual liquid in tubes and transfer to 1.5 ml microcentrifuge tubes.  4) Spin tubes 60 secs at maximum speed in a microfuge and discard remaining liquid.  5) Optional: Freeze pellets at -80°C. Pellets can be kept for up to 1 month before continuing with RNA prep.  6) Resuspend pellets in 100 μl TE buffer containing 50 mg/ml lysozyme. Incubate at room temperature for 5 min.  7) Add 75 μl lysis reagent (Promega SV Total RNA Purification kit. Cat: Z3100) and mix by inversion several times.  8) Add 350 μl RNA dilution buffer from kit (Promega SV, Cat: Z3100). Mix well by inversion.  9) Heat samples at 70 °C for 3 min and then centrifuge for 10 min at full speed (13000 rpm).  10) Transfer supernatant to clean tubes supplied with the kit. Add 200 μl ethanol and mix by pipette. Transfer to spin columns (Cat: Z3100) and centrifuge columns for 30 secs at full speed. Discard eluate.  11) Wash columns with 600 μl wash buffer from the kit. Spin 30 secs at full speed.  12) Prepare DNase mix (all reagents supplied with the kit):  a) 5 μl 90 mM MnCl2  b) 40 μl DNase core buffer  c) 5 μl DNase  13) Apply 50 μl of DNase mix to column matrix and incubate at room temperature for 15 min.  14) Add 200 μl DNase stop mix (kit) and centrifuge 30 secs at full speed.  15) Wash columns with 600 μl wash buffer, by centrifugation for 30 secs at full speed. Discard eluate.  16) Wash columns with 250 μl wash buffer, by centrifugation for 30 secs at full speed. Discard eluate.  17) Transfer columns to sterile microcentrifuge tubes, add 100 μl RNAse-free distilled H2O and allow to stand for 1 min.  18) Centrifuge at 4500 X g for 2 min. Discard column.  19) Check the RNA concentration using a spectrophotometer. From exponential phase cultures, you should typically obtain 50-60 μg RNA. Stationary phase and near-stationary phase samples will yield less, but nevertheless plenty to use. Quality of RNA should also be checked by the Aligent Bioanalyzer or a similar method to ensure the RNA is not degraded.

tag: growth phase

value: late stationary phase LSP

tag: growth type

value: Immobilised

source_name: S. typhimurium genomic DNA

value: genomic DNA as reference


title: Immobilised LSP BIO5

status: Public on Nov 01 2011

molecule: total RNA

source_name: S. Typhimurium Immobilised LSP BIO5

tag: strain

molecule: genomic DNA

tag: sample type


title: Ovary light 0 min rep1

type: RNA

city: Onnna, Kunigami

first: Takeshi

last: Kawashima

state: Okinawa

country: Japan

institute: Okinawa Institute of Science and Technology

description: This sample is extracted from a same individual with 469376A01. This individual is mature adult of wild-type and was kept in fresh-sea water individually.

hyb_protocol: standard NimbleGen protocol

scan_protocol: standard NimbleGen protocol

data_processing: RMA and summalization were executed by NimbleScan.

organism: Ciona intestinalis

source_name: Ovary light 0 min

label_protocol: 1 color, 24 reaction kit, official instruction

extract_protocol: All samples were saturated in RNAlater. RNA extraction: Trizol Reagent and Rneasy micro. Total RNA samples from Neural complex were amplified by Ovation Pico SL.

tag: tissue

value: Ovary

tag: light condition

value: light

tag: time

value: 0 min


title: Immobilised LSP BIO4

source_name: S. Typhimurium Immobilised LSP BIO4


title: H5 D1-1 549 RNA Cy3 2

city: Seattle

first: Michael

last: Katze

state: WA

address: Rosen Building 960 Republican St.

department: Microbiology

country: USA

institute: University of Washington

description: Thirty-four cynomolgus macaques (M. fascicularus) (half of each sex), weighing 2.1–2.7 kg, were exported from the Huazheng laboratory animal breeding center, Zuo Village Chengjiao Town, Conghua, Guangzhou, China and received by Covance Research Products. Animals were quarantined for 6 weeks, prescreened, and certified as specific pathogen-free. Husbandry and procedures were approved by the Battelle Institutional Animal Care and Use Committee and by the Battelle Biosafety Committee. Influenza-infected animals were housed in an ABSL-3 enhanced facility

hyb_protocol: The Agilent One-Color Microarray-Based Gene Expression Analysis Protocol was followed for hybridization and array washing. Two hundred fifty ng of each RNA sample was hybridized to one Agilent 4X44K human HG (Design ID 014850) array.

scan_protocol: Dry slides were scanned on an Agilent DNA microarray scanner (Model G2505B) using the XDR setting.

data_processing: Raw images were analyzed using the Agilent Feature Extraction software (version 9.5.3.1) and the GE1-v5_95_Feb07 extraction protocol. All arrays were required to pass Agilent QC flags. Extracted raw data were background corrected using the norm-exp method and quantile normalized using Agi4x44PreProcess and RMA Bioconductor packages.

organism: Macaca fascicularis

source_name: RNA from lung tissue infected with VN1203, 1 dpi, animal 549

label_protocol: The Agilent One-Color Microarray-Based Gene Expression Analysis Protocol was followed for the Cy3-cDNA probe preparation.

extract_protocol: Tissues were collected at necropsy and immersed immediately in solution D. Tissue was homogenized immediately for 30 seconds with a Kinematica Polytron PT1200 instrument and a model PT-DA1212/2 generator (Kinematica, Lucerne, Switzerland) in 10-ml round-bottom polypropylene test tubes with 5 ml of solution D. In order to reduce the generation of aerosols during this process, the Polytron generator was passed through a hole in the test tube lid, which had been drilled in a manner that ensured a tight fit with the instrument. To further minimize possible contact with aerosols, a barrier shield was used in addition to positive-air-pressure respirators and full-protective personal protection equipment. The homogenized samples were then snap frozen on dry ice and stored at −70°C . Added to this were 650 μl of 2 M sodium acetate, 6.5 ml of water-saturated phenol, and 1.3 ml chloroform-isoamyl alcohol (49:1). Tubes were vortexed and gently rocked for 10 min at room temperature. The mixture was then centrifuged for 30 min at 8,500 rpm and 4°C by using a Beckman L8-80 M ultracentrifuge with a JA18 rotor. The resulting aqueous layer was transferred to a new tube and precipitated with an equal volume of cold isopropanol at −20°C overnight. Centrifugation was performed again as described above. The resulting pellet was rinsed in 75% ethanol and resuspended in 150 μl of RNase-free water. A Beckman Coulter DU 640B spectrophotometer was used to quantify total RNA. One hundred micrograms of total RNA was purified with an RNA-Easy column according to the manufacturer's specifications. A Hewlett-Packard Kayak XM600 bioanalyzer was used to check the purity of the RNA prior to amplification.

treatment_protocol: The H5N1 human influenza isolate A/Vietnam/1203/2004 was obtained from the World Health Organization (WHO) influenza collaborating laboratory at the Centers for Disease Control (CDC), Atlanta, GA. The reassortant A/Texas/36/1991 influenza viruses possessing the A/South Carolina/1/18 HA (GenBank AF117241) and A/Brevig Mission/1/1918 NA (GenBank AF250356) or the A/South Carolina/1/18 HA, A/Brevig Mission/1/1918 NA, and A/Brevig Mission/1/18 NS (GenBank AF333238) were generated as previously described (50, 51). Recombinant viruses were plaque-purified, and the sequences of the virus segments were confirmed by reverse transcription-PCR and sequencing.intratracheal, intranasal, tonsillar, and conjunctival routes with a total of 107 pfu of A/Vietnam/1203/2004 (H5N1) virus, A/Texas/36/91 (H1N1) virus or reassortants of this virus containing either 2 (HA, NA) or 3 (HA, NA, NS) genes from the 1918 virus. Two animals per group were scheduled for sacrifice on days 1, 2, 4, and 7. Two additional animals were used as uninfected control animals and killed on day 7. Blood was collected from all animals 2 weeks before infection and on days 2, 4, and 7 or until sacrifice, for hematology, immunophenotyping, and cytokine analysis. Blood was also collected on day 1 from all animals except those with a day 7 endpoint, due to limitations in the total permissible blood draw. At necropsy, the lungs, tracheobronchial and retropharyngeal lymph nodes, heart, liver, spleen, kidneys, brain, and gastrointestinal tract were examined and sampled either by snap-freezing for viral isolation, cytometric bead array, real-time PCR, or microarray analysis, or by fixation in 10% formalin for histology and immunohistochemistry.

tag: time (post virus infection)

value: Day 1

tag: infection

value: lung


title: H5 D1-1 549 RNA Cy3 1


title: H5 D1-1 514 RNA Cy3 1

source_name: RNA from lung tissue infected with VN1203, 1 dpi, animal 514


title: Immobilised LSP BIO1

source_name: S. Typhimurium Immobilised LSP BIO1


title: Immobilised ESP BIO5

source_name: S. Typhimurium Immobilised ESP BIO5

value: early stationary phase ESP


title: shoot meristem of triploid hybrids showing growth abortion-replicate2

city: Kobe

first: Shigeo

last: Takumi

state: Hyogo

address: Rokkodai 1-1, Nada

department: Graduate School of Agricultural Science

institute: Kobe University

description:
shoots containig shoot apical meristem
Gene expression in 3-week-old seedling leaves

hyb_protocol: Hybridization of the Cy3-labeled cRNA to the microarrays and washing were performed using a Gene Expression Hybridization kit and Gene Expression Wash Pack (Agilent Technologies) according to the manufacturer's instructions.

scan_protocol: Signal intensities were detected by Feature Extraction software (Agilent Technologies). Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505B) using one color scan setting for 1x44k array slides (Scan Area 61x21.6 mm, Scan resolution 10um, Dye channel is set to Green and Green PMT is set to 100%).

data_processing: The scanned images were analyzed with Feature Extraction Software 9.5 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities.  Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.

organism: Aegilops tauschii x Triticum turgidum

source_name: triploid wheat hybrid

label_protocol: Cyanine-3 (Cy3) labeled cRNA was prepared from 0.5 ug RNA using the Quick Amp Labeling kit (Agilent) according to the manufacturer's instructions.  Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.

growth_protocol: Plants were grown at 23°C for 3 weeks.

extract_protocol: Total RNA was extracted from the crown tissues of WT and SGA lines , using an RNeasy Plant Mini kit (Qiagen). RNA quality was checked using an Agilent 2100 BioAnalyzer.

value: crown leaf

tag: age

value: 3 wk

tag: phenotype

value: growth abortion

tag: line

value: SGA


title: Immobilised ESP BIO3

source_name: S. Typhimurium Immobilised ESP BIO3


title: shoot meristem of triploid hybrids showing growth abortion-replicate1


title: 2000 mg/kg PMCol,  Day 29, biological rep 4

city: Menlo Park

first: Hanna

middle: Hongchin

last: Ng

state: CA

address: 333 Ravenswood Avenue

institute: SRI International

description: Gene expression data from 2000 mg/kg animal sacrificed on Day 29.

hyb_protocol: Hybridization to GeneChip Rat Gene 1.0 ST Arrays were performed as described in the Affymetrix GeneChip Whole Transcript (WT) Sense Target Labeling Assay, revision 5

scan_protocol: hydrization oven 640, GeneChip Fluidics Station 450, GeneChip Scanner 3000 7G, and GeneChip command console software.

data_processing: MAS5, In addition,  the probe logarithmic intensity error (PLIER) method was used to build upon the summarization algorithm provided in Affymetrix® Microarray Suite 5.0 (MAS 5) by taking into account the experimentally validated value of weighting feature intensities to determine an overall probe set summary. GeneSpring GX11.0 software (Agilent Technologies)

label: biotin

organism: Rattus norvegicus

source_name: 2000 mg/kg PMCol at Day 29

label_protocol: Total RNA (150 ng) from each liver sample was processed to double-stranded cDNA using the Ambion WT Expression kit (Applied Biosystems), followed by a 16-hr in vitro transcription reaction. Second-cycle cDNA was synthesized using 12 µg cRNA, and then 5.5 µg was fragmented and biotin-labeled using the GeneChip WT Terminal Labeling kit (Affymetrix, Santa Clara, CA).

growth_protocol: Male Crl:CD Sprague-Dawley Virus Antibody Free rats (Harlan, Livermore, CA) 6-7 weeks of age were maintained on Purina Certified Rodent Chow 5002 (Richmond, IN) and reverse osmosis purified tap water ad libitum under controlled lighting (12 h light-dark cycle).

extract_protocol: Total RNA was extracted and purified using the Ambion RiboPure RNA isolation kit (Applied Biosystems, Life Technologies, Carlsbad, CA) with 1-bromo-3-chloropropane (Sigma-Aldrich, St. Louis, MO).  Residual genomic DNA was removed using the Ambion TURBO DNA-free kit (Applied Biosystems).

treatment_protocol: Rats were given once daily with vehicle control (1% methy cellulose in steril water) or 200 mg/kg (low dose) or 2000 mg/kg (ligh dose) PMCol by oral gavage for either 7 days or 28 days, and eutahnized on Day 8 or Day 29. Liver left lobe was cut into ≤ 0.3 cm cubed sections, submerged in RNAlater. refrigerated overnight, and then stored frozen at ≤ -60°C until RNA isolation

value: Sprague-Dawley

value: liver

tag: treatment

value: 2000 mg/kg PMCol

value: day 29


title: 2000 mg/kg PMCol,  Day 29, biological rep 5


title: Immobilised ESP BIO4

source_name: S. Typhimurium Immobilised ESP BIO4


title: 2000 mg/kg PMCol,  Day 29, biological rep 3


title: CR Lung Pool RNA Cy3 1

source_name: Pooled equal masses of mRNA from all lung lobes of 7 reference cynomolgus macaques obtained through the tissue program of the University of Washington National Primate Research Center

value: Not infected

value: no infection


title: CR Lung Pool RNA Cy3 2


title: 2000 mg/kg PMCol,  Day 29, biological rep 2


title: shoot meristem of wild type triploid hybrids-replicate2

value: wild type


title: Immobilised LEP BIO4

source_name: S. Typhimurium Immobilised LEP BIO4

value: late exponential phase LEP

value: Immobilised

value: genomic DNA as reference


title: Immobilised LEP BIO5

source_name: S. Typhimurium Immobilised LEP BIO5


title: Cont D7-1 614 RNA Cy3 2

source_name: RNA from lung tissue infected control, 7dpi, animal 614

value: Day 7


title: H5 D1-1 514 RNA Cy3 2

value: lung


title: Cont D7-1 614 RNA Cy3 1


title: Cont D7-1 474 RNA Cy3 2

source_name: RNA from lung tissue infected control, 7dpi, animal 472


title: Cont D7-1 474 RNA Cy3 1


title: 2000 mg/kg PMCol,  Day 29, biological rep 1


title: Immobilised LEP BIO1

source_name: S. Typhimurium Immobilised LEP BIO1

source_name: S. typhimurium genomic DNA


title: Immobilised LEP BIO3

source_name: S. Typhimurium Immobilised LEP BIO3


title: 2000 mg/kg PMCol,  Day 8, biological rep 5

description: Gene expression data from 2000 mg/kg animal sacrificed on Day 8.

source_name: 2000 mg/kg PMCol at Day 8

value: day 8


title: 2000 mg/kg PMCol,  Day 8, biological rep 6


title: 2000 mg/kg PMCol,  Day 8, biological rep 4


title: 3/5 D7-1 581 RNA Cy3 2

source_name: RNA from lung tissue infected with 3:5 (1918:A/Texas/36/91), 7dpi, animal 581


title: Immobilised MEP BIO5

source_name: S. Typhimurium Immobilised MEP BIO5

value: mid-exponential phase MEP


title: Immobilised MEP BIO4

source_name: S. Typhimurium Immobilised MEP BIO4


title: 2000 mg/kg PMCol, Day 8, biological rep 3


title: 3/5 D7-1 581 RNA Cy3 1


title: Immobilised MEP BIO1

source_name: S. Typhimurium Immobilised MEP BIO1


title: 3/5 D7-1 461 RNA Cy3 1

source_name: RNA from lung tissue infected with 3:5 (1918:A/Texas/36/91), 7dpi, animal 461


title: Immobilised MEP BIO3

source_name: S. Typhimurium Immobilised MEP BIO3


title: shoot meristem of wild type triploid hybrids-replicate1

source_name: triploid wheat hybrid

value: crown leaf

value: 3 wk


title: 2000 mg/kg PMCol, Day 8, biological rep 2


title: 3/5 D4-1 635 RNA Cy3 1

source_name: RNA from lung tissue infected with 3:5 (1918:A/Texas/36/91), 4dpi, animal 635

value: Day 4


title: 3/5 D7-1 461 RNA Cy3 2


title: 200 mg/kg PMCol, Day 29, biological rep 5

description: Gene expression data from 200 mg/kg animal sacrificed on Day 29.

source_name: 200 mg/kg PMCol at Day 29

value: Sprague-Dawley

value: 200 mg/kg PMCol


title: 3/5 D4-1 635 RNA Cy3 2


title: 200 mg/kg PMCol, Day 29, biological rep 3

value: liver


title: 200 mg/kg PMCol, Day 29, biological rep 6

value: day 29


title: 2000 mg/kg PMCol, Day 8, biological rep 1

value: 2000 mg/kg PMCol


title: 200 mg/kg PMCol, Day 29, biological rep 4


title: Planktonic LSP BIO5

source_name: S. Typhimurium Planktonic LSP BIO5

value: late stationary phase LSP

value: Planktonic


title: Planktonic LSP BIO3

source_name: S. Typhimurium Planktonic LSP BIO3


title: Planktonic LSP BIO4

source_name: S. Typhimurium Planktonic LSP BIO4


title: 3/5 D4-1 515 RNA Cy3 1

source_name: RNA from lung tissue infected with 3:5 (1918:A/Texas/36/91), 4dpi, animal 515


title: 3/5 D4-1 515 RNA Cy3 2


title: Planktonic LSP BIO2

source_name: S. Typhimurium Planktonic LSP BIO2


title: 200 mg/kg PMCol, Day 29, biological rep 2


title: Planktonic ESP BIO5

source_name: S. Typhimurium Planktonic ESP BIO5

value: early stationary phase ESP

source_name: S. typhimurium genomic DNA


title: 3/5 D2-1 595 RNA Cy3 2

source_name: RNA from lung tissue infected with 3:5 (1918:A/Texas/36/91), 2dpi, animal 595

value: Day 2


title: 3/5 D2-1 595 RNA Cy3 1


title: Planktonic ESP BIO3

source_name: S. Typhimurium Planktonic ESP BIO3

value: genomic DNA as reference


title: Control at Day 29, biological rep 3

description: Gene expression data from vehicle control animal sacrificed on Day 29.

source_name: Control at Day 29

value: control


title: 3/5 D1-1 603 RNA Cy3 1

source_name: RNA from lung tissue infected with 3:5 (1918:A/Texas/36/91), 1dpi, animal 603

value: lung


title: Control at Day 29, biological rep 5


title: 200 mg/kg PMCol, Day 29, biological rep 1


title: Planktonic ESP BIO4

source_name: S. Typhimurium Planktonic ESP BIO4


title: Control at Day 29, biological rep 4


title: 3/5 D1-1 603 RNA Cy3 2

value: Day 1


title: Control at Day 29, biological rep 6


title: Planktonic ESP BIO2

source_name: S. Typhimurium Planktonic ESP BIO2


title: 3/5 D2-1 538 RNA Cy3 2

source_name: RNA from lung tissue infected with 3:5 (1918:A/Texas/36/91), 2dpi, animal 538


title: 3/5 D1-1 472 RNA Cy3 2

source_name: RNA from lung tissue infected with 3:5 (1918:A/Texas/36/91), 1dpi, animal 472


title: Planktonic LEP BIO5

source_name: S. Typhimurium Planktonic LEP BIO5

value: late exponential phase LEP


title: 3/5 D2-1 538 RNA Cy3 1


title: Planktonic LEP BIO3

source_name: S. Typhimurium Planktonic LEP BIO3


title: 3/5 D1-1 472 RNA Cy3 1


title: Planktonic LEP BIO4

source_name: S. Typhimurium Planktonic LEP BIO4


title: 14227223 - tt7 24h 4 vs adj 24h 4

city: Evry

first: Eddy

last: Blondet

address: 2 rue Gaston crémieux

department: URGV

country: France

institute: INRA

description: Do the treatments modify gene expression in a way that can give us a hint as to their use in crop protection? What is the effect of the adjuvant on gene expression?

hyb_protocol: adj 24h 4 Cy5 / tt7 24h 4 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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%

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.

organism: Arabidopsis thaliana

source_name: adj 24h 4

label_protocol: labelling Cy3 and Cy5 direct, amplification=yes, aRNA 5 ug. (Labelling_protocol.txt) 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: jiffy (peat pellets)  hygrometry: 70-85%  Temperature: 18°C (night) / 21°C (day)  Light: 110-150 µE

extract_protocol: adj 24h 4:5ug. (Promega.pdf)

treatment_protocol: Name:adj - control buffer - elicitor,oligosaccharide:quantity 2.5g/l time 24hour . 6,5 week-old arabidopsis plants were spray-treated with a hand-sprayer with water, an adjuvant or 7 oligosaccharides at 2,5 g/L in the adjuvant

tag: ecotype

value: Columbia

tag: dev.stage (boyes et al. plant cell 2001)

extract_protocol: tt7 24h 4:5ug. (Promega.pdf)

treatment_protocol: Name:tt7 - environmental treatment - elicitor,oligosaccharide:quantity 2.5g/l time 24hour . 6,5 week-old arabidopsis plants were spray-treated with a hand-sprayer with water, an adjuvant or 7 oligosaccharides at 2,5 g/L in the adjuvant


title: 14227220 - tt7 24h 2 vs adj 24h 2

hyb_protocol: adj 24h 2 Cy5 / tt7 24h 2 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

source_name: adj 24h 2

extract_protocol: adj 24h 2:5ug. (Promega.pdf)

extract_protocol: tt7 24h 2:5ug. (Promega.pdf)


title: Control at Day 8, biological rep 6

description: Gene expression data from vehicle control animal sacrificed on Day 8.

source_name: Control at Day 8

value: day 8


title: Control at Day 8, biological rep 4


title: Control at Day 8, biological rep 5


title: 14227214 - tt6 24h 2 vs adj 24h 2

hyb_protocol: adj 24h 2 Cy5 / tt6 24h 2 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt6 24h 2:5ug. (Promega.pdf)

treatment_protocol: Name:tt6 - environmental treatment - elicitor,oligosaccharide:quantity 2.5g/l time 24hour . 6,5 week-old arabidopsis plants were spray-treated with a hand-sprayer with water, an adjuvant or 7 oligosaccharides at 2,5 g/L in the adjuvant


title: 14227215 - adj 24h 3 vs tt6 24h 3

hyb_protocol: tt6 24h 3 Cy5 / adj 24h 3 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt6 24h 3:5ug. (Promega.pdf)

source_name: adj 24h 3

extract_protocol: adj 24h 3:5ug. (Promega.pdf)


title: Control at Day 29, biological rep 2


title: Planktonic LEP BIO2

source_name: S. Typhimurium Planktonic LEP BIO2


title: 2/6 D7-1 586 RNA Cy3 1

source_name: RNA from lung tissue infected with 2:6 (1918:A/Texas/36/91), 7dpi, animal 586

value: Day 7


title: 14227222 - adj 24h 3 vs tt7 24h 3

hyb_protocol: tt7 24h 3 Cy5 / adj 24h 3 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt7 24h 3:5ug. (Promega.pdf)


title: Control at Day 8, biological rep 2

value: Sprague-Dawley


title: Planktonic MEP BIO5

source_name: S. Typhimurium Planktonic MEP BIO5

value: mid-exponential phase MEP

source_name: S. typhimurium genomic DNA


title: Control at Day 29, biological rep 1

source_name: Control at Day 29

value: liver


title: 14227217 - tt6 24h 4 vs adj 24h 4

hyb_protocol: adj 24h 4 Cy5 / tt6 24h 4 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt6 24h 4:5ug. (Promega.pdf)


title: 14227219 - adj 24h 1 vs tt7 24h 1

hyb_protocol: tt7 24h 1 Cy5 / adj 24h 1 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt7 24h 1:5ug. (Promega.pdf)

source_name: adj 24h 1

extract_protocol: adj 24h 1:5ug. (Promega.pdf)


title: Planktonic MEP BIO4

source_name: S. Typhimurium Planktonic MEP BIO4


title: Control at Day 8, biological rep 1


title: Planktonic MEP BIO3

source_name: S. Typhimurium Planktonic MEP BIO3


title: 14184031 - adj 24h 1 vs tt6 24h 1

hyb_protocol: tt6 24h 1 Cy5 / adj 24h 1 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt6 24h 1:5ug. (Promega.pdf)


title: Planktonic MEP BIO2

source_name: S. Typhimurium Planktonic MEP BIO2

extract_protocol: 1) Harvest 2.0 OD600 units of bacterial culture (for example 4 mls of a culture with an OD600 = 0.5)  2) Prepare a 50 ml Falcon tube containing 2/5 of the culture volume ice-cold 5% (v/v) phenol pH 4.3, 95% (v/v) ethanol (Phenol Sigma order code: P-4682). Add the appropriate volume of culture e.g. add 4 mls of culture to 1.6 mls of phenol/ethanol. Stand on ice for at least 30 min but no longer than 2 hours to stabilise the RNA and prevent degradation.  3) Centrifuge samples at 3220 X g 4°C for 10 min. Discard supernatant, resuspend bacterial pellets using residual liquid in tubes and transfer to 1.5 ml microcentrifuge tubes.  4) Spin tubes 60 secs at maximum speed in a microfuge and discard remaining liquid.  5) Optional: Freeze pellets at -80°C. Pellets can be kept for up to 1 month before continuing with RNA prep.  6) Resuspend pellets in 100 μl TE buffer containing 50 mg/ml lysozyme. Incubate at room temperature for 5 min.  7) Add 75 μl lysis reagent (Promega SV Total RNA Purification kit. Cat: Z3100) and mix by inversion several times.  8) Add 350 μl RNA dilution buffer from kit (Promega SV, Cat: Z3100). Mix well by inversion.  9) Heat samples at 70 °C for 3 min and then centrifuge for 10 min at full speed (13000 rpm).  10) Transfer supernatant to clean tubes supplied with the kit. Add 200 μl ethanol and mix by pipette. Transfer to spin columns (Cat: Z3100) and centrifuge columns for 30 secs at full speed. Discard eluate.  11) Wash columns with 600 μl wash buffer from the kit. Spin 30 secs at full speed.  12) Prepare DNase mix (all reagents supplied with the kit):  a) 5 μl 90 mM MnCl2  b) 40 μl DNase core buffer  c) 5 μl DNase  13) Apply 50 μl of DNase mix to column matrix and incubate at room temperature for 15 min.  14) Add 200 μl DNase stop mix (kit) and centrifuge 30 secs at full speed.  15) Wash columns with 600 μl wash buffer, by centrifugation for 30 secs at full speed. Discard eluate.  16) Wash columns with 250 μl wash buffer, by centrifugation for 30 secs at full speed. Discard eluate.  17) Transfer columns to sterile microcentrifuge tubes, add 100 μl RNAse-free distilled H2O and allow to stand for 1 min.  18) Centrifuge at 4500 X g for 2 min. Discard column.  19) Check the RNA concentration using a spectrophotometer. From exponential phase cultures, you should typically obtain 50-60 μg RNA. Stationary phase and near-stationary phase samples will yield less, but nevertheless plenty to use. Quality of RNA should also be checked by the Aligent Bioanalyzer or a similar method to ensure the RNA is not degraded.


title: Inoculum B BIO5

source_name: S. Typhimurium Inoculum B BIO5

value: Inoculum


title: 14184030 - tt5 24h 4 vs adj 24h 4

hyb_protocol: adj 24h 4 Cy5 / tt5 24h 4 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt5 24h 4:5ug. (Promega.pdf)

treatment_protocol: Name:tt5 - environmental treatment - elicitor,oligosaccharide:quantity 2.5g/l time 24hour . 6,5 week-old arabidopsis plants were spray-treated with a hand-sprayer with water, an adjuvant or 7 oligosaccharides at 2,5 g/L in the adjuvant


title: Control at Day 8, biological rep 3


title: 2/6 D7-1 586 RNA Cy3 2

extract_protocol: Tissues were collected at necropsy and immersed immediately in solution D. Tissue was homogenized immediately for 30 seconds with a Kinematica Polytron PT1200 instrument and a model PT-DA1212/2 generator (Kinematica, Lucerne, Switzerland) in 10-ml round-bottom polypropylene test tubes with 5 ml of solution D. In order to reduce the generation of aerosols during this process, the Polytron generator was passed through a hole in the test tube lid, which had been drilled in a manner that ensured a tight fit with the instrument. To further minimize possible contact with aerosols, a barrier shield was used in addition to positive-air-pressure respirators and full-protective personal protection equipment. The homogenized samples were then snap frozen on dry ice and stored at −70°C . Added to this were 650 μl of 2 M sodium acetate, 6.5 ml of water-saturated phenol, and 1.3 ml chloroform-isoamyl alcohol (49:1). Tubes were vortexed and gently rocked for 10 min at room temperature. The mixture was then centrifuged for 30 min at 8,500 rpm and 4°C by using a Beckman L8-80 M ultracentrifuge with a JA18 rotor. The resulting aqueous layer was transferred to a new tube and precipitated with an equal volume of cold isopropanol at −20°C overnight. Centrifugation was performed again as described above. The resulting pellet was rinsed in 75% ethanol and resuspended in 150 μl of RNase-free water. A Beckman Coulter DU 640B spectrophotometer was used to quantify total RNA. One hundred micrograms of total RNA was purified with an RNA-Easy column according to the manufacturer's specifications. A Hewlett-Packard Kayak XM600 bioanalyzer was used to check the purity of the RNA prior to amplification.


title: 14184029 - adj 24h 3 vs tt5 24h 3

hyb_protocol: tt5 24h 3 Cy5 / adj 24h 3 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt5 24h 3:5ug. (Promega.pdf)


title: Inoculum B BIO4

source_name: S. Typhimurium Inoculum B BIO4


title: 2/6 D4-1 574 RNA Cy3 1

source_name: RNA from lung tissue infected with 2:6 (1918:A/Texas/36/91), 4dpi, animal 574

value: Day 4


title: 2/6 D7-1 479 RNA Cy3 1

source_name: RNA from lung tissue infected with 2:6 (1918:A/Texas/36/91), 7dpi, animal 479


title: Inoculum B BIO2

source_name: S. Typhimurium Inoculum B BIO2


title: MPNST biological rep3

city: Dallas

first: James

middle: F

last: Amatruda

state: TX

address: 5323 Harry Hines Boulevard

department: Pediatrics and Molecular Biology

institute: University of Texas Southwestern Medical Center

description: Malignant peripheral nerve sheath tumor arising in p53M214K/M214K zebrafish

hyb_protocol: Hybridization was carried out according to Affymetrix protocol 701027 rev 5, using Affymetrix Fluidics Station 450.

scan_protocol: Arrays were scanned with the Affymetrix GeneChip Scanner 3000 using default parameters

data_processing: The probe-level data were background adjusted, normalized and summarized by a Robust Multi-array Average (RMA) algorithm implemented in the software package affy of the Bioconductor project release 1.7, running in the R statistical environment version 2.2.0 using default parameters.

organism: Danio rerio

source_name: Malignant peripheral nerve sheath tumor

label_protocol: Total RNA (2.5ug) from tumors was reverse-transcribed using a T7-Oligo(dT) Promoter Primer, followed by RNAseH-mediated second-strand cDNA synthesis.  Biotinylated cRNA probe was prepared with MessageAmp II (Ambion).

growth_protocol: Wild-type or p53(M214K) homozygous zebrafish embryos were injected at the one cell stage with a transposon in which the Heat Shock Protein 70 (HSP70) promoter drove expression of a human EWS-FLI1 fusion oncoprotein [Tg(HSP:EWS-FLI1)].

extract_protocol: Total RNA was extracted from tumors using Trizol reagent according to the manufacturer's instructions.

treatment_protocol: Fish were grown under standard conditions and monitored for tumor development.  Fish developing tumors were euthanized and the tumor tissue removed for histology and RNA preparation.

value: tumor

tag: genotype/variation


title: 14184028 - tt5 24h 2 vs adj 24h 2

hyb_protocol: adj 24h 2 Cy5 / tt5 24h 2 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

source_name: adj 24h 2

extract_protocol: tt5 24h 2:5ug. (Promega.pdf)


title: 2/6 D4-1 574 RNA Cy3 2


title: MPNST biological rep1


title: 2/6 D7-1 479 RNA Cy3 2


title: 2/6 D4-1 508 RNA Cy3 2

source_name: RNA from lung tissue infected with 2:6 (1918:A/Texas/36/91), 4dpi, animal 508


title: 2/6 D4-1 508 RNA Cy3 1


title: MPNST biological rep2


title: Inoculum A BIO4

hyb_protocol: Slides blocked with DCE according to protocol at PMID 11266573. Microarray Hybridisations  1. To 10 μl of dried down labelling reactions add:  a) 1.5 μl of 50 X Denhardts solution  b) 2.25 μl of 20 X SSC  c) 1.125 μl of E. coli tRNA (10 μg/μl) (Sigma, Cat: R8759)  d) 0.375 μl of 1M HEPES, pH 7.0.  NOTE: The hybridisation components can be mixed together as a stock solution, either from a fresh or frozen stock, and added to labelling reactions.  2. Add 0.375 μl of 10% SDS to the mixture  3. Incubate 100 °C for 2 min. Let stand on the bench for 5 - 10 min. Do not place the reaction on ice because the SDS will precipitate.  4. Spin in a microfuge at full speed for 5 min, transfer supernatant to clean tube and repeat spin.  5. Place a microarray slide in a metal hybridisation chamber (Photo 3).  6. Pipette the hybridisation solution towards one edge of an array.  7. Place edge of a clean (washed with 70% ethanol), dry, coverslip on the edge of the array.  8. Position fine nosed forceps (VWR, Cat: 406/0073/04) under the coverslip.  9. Using the forceps slowly lower the coverslip down onto the hybridisation solution and the array ensuring no air bubbles are present under the coverslip. If air bubbles are present it is sometimes possible to move them to the edge of the coverslip by pressing gently with the forceps.  10. Apply 20 μl of 3 X SSC around the 4 corners of the slide using a pipette (approximately 5 μl per corner). This will maintain the correct humidity in the hybridisation chamber.  11. Screw the lid onto the chamber and place at the bottom of a water bath at 63°C overnight.  12. Remove the slides from the hybridisation chambers and place them in a plastic slide rack (Photo 4). The slides are then placed into a 63°C wash solution containing 2 X SSC, 0.1% SDS and agitated for 5 mins. (We use a 2L beaker with a stirrer bar in  the bottom and a platform for the slide rack to sit on for this, Photos 5 & 6). It may be necessary to tap the slide rack on the platform to make sure that the cover slips are removed. We use a thermocouple attached to a hot-plate stirrer for this step. Repeat the washing step.  Note: SDS fluoresces so you should avoid carrying it over into the washes in steps 13 and 14.  13. Transfer slide rack to a solution of 1 X SSC at room temperature and agitate for 5 min. Repeat. (We use black tissue boxes often used for histology for this)  14. Wash in 0.2 X SSC for 5 min. Repeat.  15. Spin dry in a centrifuge at 1200 rpm for 5 min at room temp in an enclosed slide chamber (with blotting paper lining the bottom of the chamber).  16. Scan.  NB All solutions are filter sterilised (0.1 μm) and made up with Sigma water (molecular biology reagent, Sigma, Cat: W4502).

source_name: S. Typhimurium Inoculum A BIO4


title: 14184024 - adj 24h 3 vs tt4 24h 3

hyb_protocol: tt4 24h 3 Cy5 / adj 24h 3 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt4 24h 3:5ug. (Promega.pdf)

treatment_protocol: Name:tt4 - environmental treatment - elicitor,oligosaccharide:quantity 2.5g/l time 24hour . 6,5 week-old arabidopsis plants were spray-treated with a hand-sprayer with water, an adjuvant or 7 oligosaccharides at 2,5 g/L in the adjuvant

source_name: adj 24h 3


title: Inoculum A BIO2

source_name: S. Typhimurium Inoculum A BIO2


title: 2/6 D2-1 550 RNA Cy3 1

hyb_protocol: The Agilent One-Color Microarray-Based Gene Expression Analysis Protocol was followed for hybridization and array washing. Two hundred fifty ng of each RNA sample was hybridized to one Agilent 4X44K human HG (Design ID 014850) array.

source_name: RNA from lung tissue infected with 2:6 (1918:A/Texas/36/91), 2dpi, animal 550


title: 14184023 - tt4 24h 2 vs adj 24h 2

hyb_protocol: adj 24h 2 Cy5 / tt4 24h 2 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: adj 24h 2:5ug. (Promega.pdf)

extract_protocol: tt4 24h 2:5ug. (Promega.pdf)


title: 2/6 D2-1 550 RNA Cy3 2


title: 14184025 - tt4 24h 4 vs adj 24h 4

hyb_protocol: adj 24h 4 Cy5 / tt4 24h 4 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

source_name: adj 24h 4

extract_protocol: adj 24h 4:5ug. (Promega.pdf)

extract_protocol: tt4 24h 4:5ug. (Promega.pdf)


title: SRBCT biological rep1

description: Ewing's Sarcoma-like small round blue cell tumor arising in Tg(HSP:EWS-FLI1); p53M214K/M214K zebrafish

source_name: Small Round Blue Cell Tumor

value: Tg(HSP:EWS-FLI1); p53M214K/M214K


title: 14184027 - adj 24h 1 vs tt5 24h 1

hyb_protocol: tt5 24h 1 Cy5 / adj 24h 1 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt5 24h 1:5ug. (Promega.pdf)

source_name: adj 24h 1


title: Inoculum A BIO5

source_name: S. Typhimurium Inoculum A BIO5

source_name: S. typhimurium genomic DNA


title: Tg(HSP:EWS-FLI1) embryos biological rep3

source_name: g(HSP:EWS-FLI1) embryos, heat-shocked 39 degrees C for 30 mins at 50% epiboly stage

label_protocol: Total RNA (2.5ug) from embryos was reverse-transcribed using a T7-Oligo(dT) Promoter Primer, followed by RNAseH-mediated second-strand cDNA synthesis.  Biotinylated cRNA probe was prepared with MessageAmp II (Ambion).

growth_protocol: Wild-type AB strain embryos and Tg(HSP:EWS-FLI1) embryos were derived from crosses under standard conditions and raised until the 50% epiboly stage.

extract_protocol: Total RNA was extracted from groups of 40 embryos using Trizol reagent according to the manufacturer's instructions.

treatment_protocol: Embryos were heat shocked for 30 minutes at 39 degrees C., then incubated 2 hours at 28.5 degrees C. prior to RNA collection.

value: embryo


title: SRBCT biological rep2


title: SRBCT biological rep3


title: 2/6 D2-1 503 RNA Cy3 1

source_name: RNA from lung tissue infected with 2:6 (1918:A/Texas/36/91), 2dpi, animal  503

value: Day 2


title: 14184022 - adj 24h 1 vs tt4 24h 1

hyb_protocol: tt4 24h 1 Cy5 / adj 24h 1 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt4 24h 1:5ug. (Promega.pdf)


title: 2/6 D1-1 599 RNA Cy3 2

source_name: RNA from lung tissue infected with 2:6 (1918:A/Texas/36/91), 1dpi, animal 599


title: 2/6 D2-1 503 RNA Cy3 2

source_name: RNA from lung tissue infected with 2:6 (1918:A/Texas/36/91), 2dpi, animal 503


title: 2/6 D1-1 599 RNA Cy3 1


title: Duchenne Muscular Dystrophy sample 3 Replicate 2

city: Nantes

first: Daniel

last: BARON

state: Loire-Atlantique (44)

address: 30 Bd Jean Monnet

department: UMR1064 - Centre de Recherche en Transplantation & Immunologie

institute: Institut national de la santé et de la recherche médicale-INSERM

description: DMD, Biological replicate 3 of 4, Technical replicate 2 of 2.

hyb_protocol: Microarrays were prehybridized for 1 hour at 42° C (in 10% BSA, 20XSSC, 10% SDS). 0.5 µg of labeled sample cDNA was mixed with 0.5 µg of labeled reference cDNA in 40 µl of hybridization mix (50X denhardt, 20XSSC, 10 µg/µl PolyA RNA, 10 µg/µl Yeast tRNA, 10% SDS, 50% formamide) and hybridized to the microarrays. Microarray hybridization was carried out overnight at 42° C in a humidified chamber. Finally, slides were washed and dried by centrifugation.

scan_protocol: Array scanning was carried out using a GSI lumonics LITE dual confocal laser scanner with ScanArray Microarray Analysis Software. Raw scanner images were analyzed with Genepix Software (AXON LABORATORY).

data_processing: Data were normalized (LOWESS) and analyzed (data filtering) using the Micro-Array Data Suite of Computed Analysis (MADSCAN)

organism: Homo sapiens

source_name: DMD

label_protocol: Cy3- and Cy5-labeled cDNA was prepared using the CyScribe cDNA post labeling Kit (Amersham Pharmacia Biotech). Samples from DMD and AGING patients were each labeled individually with Cy5. Reference patients were pooled and then labeled with Cy3.

extract_protocol: Normal and patient total RNA was purified using Trizol Reagent (Life technologies). Poly (A)+ RNA was purified from total RNA using oligo-dt linked oligotex resin (QIAGEN).

tag: gender

value: Male

tag: age (years)

tag: muscle

value: Fascia lata tensor

source_name: CONTROL

tag: description

value: REFERENCE:pool of total RNA from 7 younger CONTROL muscle biopsies

value: 5 Females / 2 Males

value: Paravertebral


title: Duchenne Muscular Dystrophy sample 4 Replicate 1

description: DMD, Biological replicate 4 of 4, Technical replicate 1 of 2.


title: 14184021 - tt3 24h 4 vs adj 24h 4

hyb_protocol: adj 24h 4 Cy5 / tt3 24h 4 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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: Columbia

extract_protocol: tt3 24h 4:5ug. (Promega.pdf)

treatment_protocol: Name:tt3 - environmental treatment - elicitor,oligosaccharide:quantity 2.5g/l time 24hour . 6,5 week-old arabidopsis plants were spray-treated with a hand-sprayer with water, an adjuvant or 7 oligosaccharides at 2,5 g/L in the adjuvant


title: Duchenne Muscular Dystrophy sample 3 Replicate 1

description: DMD, Biological replicate 3 of 4, Technical replicate 1 of 2.


title: 2/6 D1-1 480 RNA Cy3 1

source_name: RNA from lung tissue infected with 2:6 (1918:A/Texas/36/91), 1dpi, animal 480

value: lung


title: Duchenne Muscular Dystrophy sample 4 Replicate 2

description: DMD, Biological replicate 4 of 4, Technical replicate 2 of 2.


title: AB control embryos biological rep3

source_name: control embryos, heat-shocked 39 degrees C for 30 mins at 50% epiboly stage

value: wildtype AB strain


title: Tg(HSP:EWS-FLI1) embryos biological rep1


title: 2/6 D1-1 480 RNA Cy3 2


title: 14184026 - adj 24h 1 vs tt3 24h 1

hyb_protocol: tt3 24h 1 Cy5 / adj 24h 1 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt3 24h 1:5ug. (Promega.pdf)

extract_protocol: adj 24h 1:5ug. (Promega.pdf)


title: AB control embryos biological rep2


title: AB control embryos biological rep1


title: Tg(HSP:EWS-FLI1) embryos biological rep2


title: 14184020 - adj 24h 3 vs tt3 24h 3

hyb_protocol: tt3 24h 3 Cy5 / adj 24h 3 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt3 24h 3:5ug. (Promega.pdf)

extract_protocol: adj 24h 3:5ug. (Promega.pdf)


title: 14184019 - tt3 24h 2 vs adj 24h 2

hyb_protocol: adj 24h 2 Cy5 / tt3 24h 2 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

source_name: adj 24h 2

extract_protocol: tt3 24h 2:5ug. (Promega.pdf)


title: 14184017 - tt2 24h 4 vs adj 24h 4

hyb_protocol: adj 24h 4 Cy5 / tt2 24h 4 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt2 24h 4:5ug. (Promega.pdf)

treatment_protocol: Name:tt2 - environmental treatment - elicitor,oligosaccharide:quantity 2.5g/l time 24hour . 6,5 week-old arabidopsis plants were spray-treated with a hand-sprayer with water, an adjuvant or 7 oligosaccharides at 2,5 g/L in the adjuvant


title: Duchenne Muscular Dystrophy sample 1 Replicate 2

description: DMD, Biological replicate 1 of 4, Technical replicate 2 of 2.


title: 14184016 - adj 24h 3 vs tt2 24h 3

hyb_protocol: tt2 24h 3 Cy5 / adj 24h 3 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt2 24h 3:5ug. (Promega.pdf)

source_name: adj 24h 3


title: 14184014 - tt2 24h 2 vs adj 24h 2

hyb_protocol: adj 24h 2 Cy5 / tt2 24h 2 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt2 24h 2:5ug. (Promega.pdf)


title: Duchenne Muscular Dystrophy sample 2 Replicate 2

description: DMD, Biological replicate 2 of 4, Technical replicate 2 of 2.


title: Duchenne Muscular Dystrophy sample 1 Replicate 1

description: DMD, Biological replicate 1 of 4, Technical replicate 1 of 2.


title: Aging muscular sample 4 Replicate 2

description: AGE, Biological replicate 4 of 4, Technical replicate 2 of 2.

source_name: AGE

value: Female


title: Duchenne Muscular Dystrophy sample 2 Replicate 1

description: DMD, Biological replicate 2 of 4, Technical replicate 1 of 2.


title: root control conditions at 12h, biological rep2

city: Santiago

first: Andrea

last: Vega

address: Vicuña Mackenna 4860

department: Ciencias Vegetales

country: Chile

institute: Pontificia Universidad Catolica de Chile

description: Gene expression data from root in control treatment at 12 hours

hyb_protocol: Following fragmentation, 10 ug of cRNA were hybridized for 16 hr at 45C on GeneChip® Arabidopsis ATH1 Genome Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 450.

scan_protocol: Array scanning was carried out with the GeneChip® scanner 300 and image analysis was performed using the GeneChip® Operating Software.

data_processing: The data were processed and normalized by RMA using the R package known as “affy”.

source_name: Root in control treatment at 12 hours

label_protocol: Biotinylated aRNA were prepared according to the standard Affymetrix protocol from 0.5 ug total RNA (GeneChip® 3’ IVT Express Kit User Manuall, 2008, Affymetrix).

growth_protocol: Arabidopsis seeds were surface-sterilized and germinated on an agar-solidified nutrient medium in Petri dishes. The nutrient medium was based on half Murashige-Skoog salts (MS) and the final pH was adjusted to 5,7. The seeds were vernalized at 4ºC for 2 d. Petri dishes were placed into a growth chamber, positioned vertically and kept under controlled environmental conditions at 22ºC and a 16/8 h day/night regime. After five days, seedlings were transferred to 1/2 MS plates containing BA (H3BO3, 5mM)

extract_protocol: Trizol extraction of total RNA was performed according to the manufacturer's instructions.

value: control

value: root

value: 5-days-old plant


title: root control conditions at 12h, biological rep3


title: Aging muscular sample 3 Replicate 1

description: AGE, Biological replicate 3 of 4, Technical replicate 1 of 2.


title: root BA treatment at 12h, biological rep3

description: Gene expression data from root treated with 5mM BA at 12 hours

source_name: Root in boron treatment (5mM) at 12 hours

value: boron (5mM)


title: DvH 0min Wildtype control

city: Berkeley

first: Adam

middle: P.

last: Arkin

address: 1 Cyclotron Road

department: Physical Biosciences Division

institute: Virtual Institute for Microbial Stress and Survival

description: This sample is one of a series of num_samples used to investigate the stress response pathways of Desulfovibrio vulgaris Hildenborough treated with Mutant. This sample was treated with hbaR (DVU2547)  Mutant.  Treatment time point = 0 min

hyb_protocol: He, Z., Wu, L., Li, X., Fields, M., and Zhou, J. 2005.  Empirical Establishment of Oligonucleotide Probe Design Criteria. Appl. Environ. Microbiol. 71:3753-3760.

scan_protocol: ImaGene software

data_processing: See Methods in: S. R. Chhabra, Q. He, K. H. Huang, S. P. Gaucher, E. J. Alm, Z. He, M. Z. Hadi, T. C. Hazen, J. D. Wall, J. Zhou, A.  . Arkin, and A. K. Singh, Global Analysis of Heat Shock Response in Desulfovibrio vulgaris Hildenborough, J Bacteriol. 2006 March; 188(5): 1817-1828. http://www.pubmedcentral.nih.gov/articlerender.fcgi?tool=pubmed&pubmedid=16484192

organism: Nitratidesulfovibrio vulgaris str. Hildenborough

source_name: Desulfovibrio vulgaris subsp. vulgaris str. Hildenborough, 0min Wildtype Control

growth_protocol: Inoculate 1 ml of log-phase inoculum culture to 100 ml of defined PIPES buffered medium LS4D (13, 27) in glass bottles. Grow culture at 37�C till mid-log phase (OD of ~0.4). There are three bottles for wild type D. vulgaris and each crp/fnr mutant.  Harvest the mid-log phase cells via centrifugation at 4oC. Remove the residual liquid by pipetting. Cell pellets were immediately frozen in liquid N2 and stored at -80�C until RNA isolation.

treatment_protocol: 0min Wildtype Control.

tag: strain

value: Hildenborough

tag: genotype

value: wild type


title: Aging muscular sample 2 Replicate 2

description: AGE, Biological replicate 2 of 4, Technical replicate 2 of 2.


title: Aging muscular sample 2 Replicate 1

description: AGE, Biological replicate 2 of 4, Technical replicate 1 of 2.


title: root BA treatment at 12h, biological rep2

tag: tissue


title: root control conditions at 12h, biological rep1


title: Aging muscular sample 4 Replicate 1

description: AGE, Biological replicate 4 of 4, Technical replicate 1 of 2.


title: 14184010 - tt1 24h 2 vs adj 24h 2

description: Do the treatments modify gene expression in a way that can give us a hint as to their use in crop protection? What is the effect of the adjuvant on gene expression?

hyb_protocol: adj 24h 2 Cy5 / tt1 24h 2 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: adj 24h 2:5ug. (Promega.pdf)

extract_protocol: tt1 24h 2:5ug. (Promega.pdf)

treatment_protocol: Name:tt1 - environmental treatment - elicitor,oligosaccharide:quantity 2.5g/l time 24hour . 6,5 week-old arabidopsis plants were spray-treated with a hand-sprayer with water, an adjuvant or 7 oligosaccharides at 2,5 g/L in the adjuvant


title: DvH 0min DVU2547 Mutant set4255

source_name: Desulfovibrio vulgaris subsp. vulgaris str. Hildenborough, 0min DVU2547  Mutant

label_protocol: He, Z., Wu, L., Li, X., Fields, M., and Zhou, J.  2005. Empirical Establishment of Oligonucleotide Probe Design Criteria. Appl. Environ. Microbiol. 71:3753-3760.

treatment_protocol: 0min hbaR (DVU2547)  Mutant.

value: DVU2547 Mutant


title: DvH 0min DVU3111 Mutant set4256

description: This sample is one of a series of num_samples used to investigate the stress response pathways of Desulfovibrio vulgaris Hildenborough treated with Mutant. This sample was treated with cooA-2 (DVU3111)  Mutant.  Treatment time point = 0 min

source_name: Desulfovibrio vulgaris subsp. vulgaris str. Hildenborough, 0min DVU3111  Mutant

extract_protocol: He, Z., Wu, L., Li, X., Fields, M., and Zhou, J.  2005. Empirical Establishment of Oligonucleotide Probe Design Criteria. Appl. Environ. Microbiol. 71:3753-3760.

treatment_protocol: 0min DVU3111 Mutant.

value: DVU3111 Mutant


title: DvH 0min DVU0379 Mutant set4253

description: This sample is one of a series of num_samples used to investigate the stress response pathways of Desulfovibrio vulgaris Hildenborough treated with Mutant. This sample was treated with DVU0379  Mutant.  Treatment time point = 0 min

source_name: Desulfovibrio vulgaris subsp. vulgaris str. Hildenborough, 0min DVU0379  Mutant

treatment_protocol: 0min DVU0379  Mutant.

value: DVU0379 Mutant


title: root BA treatment at 12h, biological rep1


title: 14184009 - adj 24h 1 vs tt1 24h 1

hyb_protocol: tt1 24h 1 Cy5 / adj 24h 1 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt1 24h 1:5ug. (Promega.pdf)

source_name: adj 24h 1


title: Aging muscular sample 3 Replicate 2

description: AGE, Biological replicate 3 of 4, Technical replicate 2 of 2.


title: Aging muscular sample 1 Replicate 1

description: AGE, Biological replicate 1 of 4, Technical replicate 1 of 2.

value: Male


title: 14184013 - adj 24h 1 vs tt2 24h 1

hyb_protocol: tt2 24h 1 Cy5 / adj 24h 1 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt2 24h 1:5ug. (Promega.pdf)


title: DvH 0min DVU2097 Mutant set4254

description: This sample is one of a series of num_samples used to investigate the stress response pathways of Desulfovibrio vulgaris Hildenborough treated with Mutant. This sample was treated with DVU209)  Mutant.  Treatment time point = 0 min

source_name: Desulfovibrio vulgaris subsp. vulgaris str. Hildenborough, 0min DVU2097 Mutant

treatment_protocol: 0min DVU209)  Mutant.

value: DVU2097 Mutant


title: 14162230 - adj 24h 4 vs H20 24h 4

hyb_protocol: H20 24h 4 Cy5 / adj 24h 4 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: H20 24h 4:5ug. (Promega.pdf)

treatment_protocol: Name:H20 - control - elicitor,oligosaccharide:time 24hour . 6,5 week-old arabidopsis plants were spray-treated with a hand-sprayer with water, an adjuvant or 7 oligosaccharides at 2,5 g/L in the adjuvant

source_name: adj 24h 4

extract_protocol: adj 24h 4:5ug. (Promega.pdf)


title: 14184012 - tt1 24h 4 vs adj 24h 4

hyb_protocol: adj 24h 4 Cy5 / tt1 24h 4 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt1 24h 4:5ug. (Promega.pdf)


title: 14184011 - adj 24h 3 vs tt1 24h 3

hyb_protocol: tt1 24h 3 Cy5 / adj 24h 3 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: tt1 24h 3:5ug. (Promega.pdf)


title: 14162229 - H20 24h 3 vs adj 24h 3

hyb_protocol: adj 24h 3 Cy5 / H20 24h 3 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: H20 24h 3:5ug. (Promega.pdf)


title: 14162228 - adj 24h 2 vs H20 24h 2

hyb_protocol: H20 24h 2 Cy5 / adj 24h 2 Cy3 : 30pmol. (Hybridization_Protocol.txt) 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.

extract_protocol: H20 24h 2:5ug. (Promega.pdf)


title: TE/Cdx2ER wt, rep2

status: Public on Oct 31 2011

city: Baltimore

first: Minoru

last: Ko

state: MD

address: 251 Bayview Blvd, Suite 100, 10C

department: National Institute on Aging

institute: NIH

description: TE Cdx2_ER Wildtype

hyb_protocol: Slides were hybridized according to the manufacturer's protocol (Two-Color Microarray-Based Gene Expression Analysis Protocol, Product # G4140-90050, Version 5.0.1, August 2006).

scan_protocol: Slides were scanned with an Agilent DNA Microarray Scanner (model G2505-64120) at 100% and 10% PMT in both channels, with a scan resolution of 5um.

data_processing:
Data were extracted with Agilent Feature Extraction Software. The data were further processed with NIA ANOVA tool utilities. See http://lgsun.grc.nia.nih.gov/ANOVA for details.

The data were normalized using the following method:
(1) Take values from the columns gDyeNormSignal, rDyeNormSignal in the raw files and log-transform them using: log10(max(x,1)). These values will be referenced below as Xgi and Xri, where i is array number.
(2) Take average of Xri's for the oligo among 8 arrays in the series: AverXr = average(Xri).
(3) For each array, estimate Yi = Xgi-Xri+AverXr
(4) Round Yi to 4 decimal digits. This is used as the normalized VALUE.

More detailed information with error correction can be found at http://lgsun.grc.nia.nih.gov/ANOVA/help.html#arrayjoin

organism: Mus musculus

source_name: TE Cdx2_ER Wildtype

label_protocol: Total RNA was labeled using the Agilent Low RNA Input Fluorescent Linear Amplification kit, according to manufacturer's instructions.

extract_protocol: Total RNA was extracted by TRizol Reagent (Invitrogen, 15596-018) according to the manufacturer's recommendation. Briefly, cells were disrupted by adding 1 ml of TRizol Reagent to a well of a 6-well plate and passing several times through a pipette. The cell lysate was transferred to a nuclease-free eppendorf tube and incubated for 5 minutes at room temperature. To separate phases, the lysate was mixed with 0.2 ml of chloroform, incubated at room temperature for 3 minutes and centrifuged at 12,000g for 15 minutes at 4C.  Following centrifugation, the aqueous phase were transferred to fresh tubes.  The RNA was precipitated from the aqueous phase by mixing with 0.5ml isopropyl alcohol, incubating at room temperature for 10 minutes and centrifuging at 12,000g for 10 minutes at 4C.  The RNA pellet was washed with 1ml of 75% ethanol and dried for less than 5 minutes.  RNA was dissolved in RNase-free water by passing the solution a few times through a pipette tip and incubating for 10 minutes at 60C. Dissolved RNA was stored at -70C.

treatment_protocol: To induce differentiation toward TE cells, the established ESC lines expressing tamoxifen-inducible Cdx2 cultured in the absence of tamoxifen were split at 1.2 x 104 cells/cm2 into a culture dish containing 30% TS medium, 70% FCM, 100 nM FGF4, 1 ug/ml Heparin and 1 ug/ml 4-hyroxy tamoxifen (Tx; Sigma).  The cells were cultured under this TS condition for 2 days without changing media.  TE cells were differentiated from the TS state cells by culturing in the 100% TS medium for additional 6 days, changing media every 2 days.

tag: cell type

value: trophectoderm cell (TE)

value: wildtype

value: male

tag: development stage

value: embryo

tag: genetic modification

value: gene knockout

tag: individual identifier

source_name: Universal Mouse Reference 129ES Cell

extract_protocol: Total RNA was extracted from cultured cells using a Stratagene Absolutely RNA kit. DNase-treated total RNA samples from 11 cultured cell lines derived from embryo, embryo fibroblast, kidney, liver/hepatocyte, lung/alveolar macrophage, B-lymphocyte, T-lymphocyte (thymus), mammary gland, muscle myoblast, skin, and testis were pooled in equal parts. The resulting URM RNA was mixed with 129ES cell RNA (Lif+) at 2:1 ratio and 2.5ug of mixed RNA was used for labeling in each tube.

tag: sample type

value: reference