title: Day 14 lens RNA, scan 2.2

status: Public on Aug 11 2007

description: The results were analysed using an image analysis program, ImaGene 5 (Biodiscovery). Using ImaGene, a virtual grid was placed over the image of the arrays assigning each spot a know grid reference; this allowed for the intensity of each spot to be calculated and also allowed the correct gene ID to be allocated to each spot (see Figure 3.2.). Once ImaGene had calculated the spot intensities, the data were then exported to Microsoft® Excel for further analysis. The data obtained from ImaGene produces a spot intensity and a background signal value for each individual spot. The background value for each spot was calculated as a mean of the intensity of a set number of pixels surrounding the spot (as shown in Figure 2.1.).

hyb_protocol: Panorama™ mouse apoptosis arrays (Sigma-Genosys) were used for this set of experiments. These arrays are nylon membranes containing 243 known apoptosis related genes. The complete list of genes present on the array is given in Appendix 2. Before the arrays were hybridised with the radiolabelled cDNA they were first pre-hybridised to prevent non-specific binding of DNA to the arrays. The arrays were washed in 50ml 2x SSPE (Sigma) at room temperature for 5 minutes. The arrays were then pre-hybridised in hybridisation solution (Sigma) containing salmon testes DNA (100µl Salmon testes DNA in 10ml hybridisation solution) at 65ºC for at least an hour before the addition of the radiolabelled cDNA. The unincorporated radiolabelled nucleotides had to be removed from the reaction mix before it was added to the arrays. This should result in a reduction in the background signal during the hybridisation reaction. To remove the unincorporated radiolabelled nucleotides a spin column containing sephadex beads (Sigma-Genosys) was used. The spin column was centrifuged to remove any excess buffer before the sample was added. Once the sample had been added to the centre of the spin column, the column was centrifuged at 1100 x g for 4 minutes. The eluate from the column contained the purified cDNA sample; this sample could then be added to the arrays. The purified labelled cDNA was then added to 2-3mls hybridisation solution (containing 100μl salmon testes DNA per 10ml hybridisation solution). The DNA was then denatured by heating at 95°C for 10 minutes. The hybridisation solution containing the labelled cDNA was then added to the arrays, which were hybridised overnight for 18 hours in the hybridisation oven at 65°C (UVP, HC-3000 Hybricycler). After the hybridisation reaction was complete, the arrays were then washed to remove any unbound DNA with increasing concentrations of salt solution. The hybridisation solution was decanted and discarded and 40-50ml of wash solution I (composition in Appendix 1) was added to the roller bottles. The arrays were washed by gently inverting the bottles by hand at room temperature for 2-3 minutes. This process was repeated twice more, with a new aliquot of wash solution I each time. 80-100ml of wash solution I (pre-warmed to 65°C) was then added to the roller bottles and the arrays were washed in the hybridisation oven at 65°C for 20 minutes, before the wash solution was discarded. This process was repeated once more. Wash solution II (pre-warmed to 65ºC: composition given in Appendix 1) was then added to the roller bottles (80-100ml) and washed for a further 20 minutes in the hybridisation oven. The wash solution was then discarded and the arrays were wrapped in clingfilm before placing them into a storage phosphor screen (Amersham Biosciences). The arrays were exposed to the phosphor screen for 5-7days.

scan_protocol: The phosphor screen was then scanned using the Typhoon scanner (Amersham Biosciences, Typhoon 9410 Variable Mode Imager) which extracted the energy from the screen and converted it into a computer image which could later be analysed by use of image analysis software. Once the phosphor screen had been scanned, the arrays were stripped removing any residual bound DNA. This ensured that each array could be reused. The arrays were added to 200ml of stripping solution (composition given in Appendix 1), which had been brought to the boil. The arrays were then added to the stripping solution, which was then allowed to cool. The arrays were then removed from the solution, the excess drained and the arrays wrapped in clingfilm. Once stripped, the arrays were stored at -20°C. Stripped arrays were exposed to the phosphor screen to check that the residual radiolabelled cDNA had been removed.

data_processing: The data produced were normalised using the mean expression of the housekeeping genes from the arrays; housekeeping genes are genes continuously expressed throughout development. The normalisation process allowed comparisons of gene expression to be made between the completed arrays; accounting for variations of the signal between arrays. The mean and standard deviation were calculated using the individual background values for each spot; these values were subsequently used to filter the data later on in the analysis process. The spots whose original signal was lower than the mean plus two standard deviations of the background values were removed from the rest of the results following normalisation. The individual background values were then subtracted from the corresponding spot intensity, to give a corrected intensity. Each corrected spot value was normalised before being filtered. The spots were normalised using the housekeeping genes printed on the apoptosis arrays. A mean value was calculated from the corrected spot intensities from each of these housekeeping genes as well as Gapdh (a standard housekeeping gene), which was also printed on the array. The array data were then normalised using the ratios of these mean values. One of the arrays was designated as standard. A ratio was calculated for each array by dividing the mean housekeeping value from the standard array by the mean housekeeping value from that array. The corrected spot intensities were all multiplied by this ratio. After normalisation, a mean value for each spot across the set of arrays was calculated using the corrected intensities. The mean values for each spot were then compared for P7 and P14 using an unpaired t-test. Spots with a 2-fold difference between these two time points and a p-value <0.05 were considered to have a significant change in gene expression.

label: 33P-dCTP (Amersham Biosciences)

source_name: 14 day old mouse lens, strain 129Sv/Ev

label_protocol: The labelling reactions are performed in two steps. The first step, the annealing reaction, shown in Table 2.2, involves the annealing of mouse apoptosis cDNA primers (Sigma-Genosys) to the RNA template. During the second step, radiolabelled nucleotides and reverse transcriptase (Sigma-Genosys) were added to initiate the cDNA synthesis reaction. Final Concentration Volume for One Reaction 2µg total RNA 2-11µl Mouse Apoptosis cDNA Labelling Primers 4µl Sterile distilled water to 15µl Table 2.2: Composition of the Annealing Reaction. Table taken from Panorama™ Mouse Apoptosis Gene Arrays Protocol. For the annealing step, the components from the table above were added to an eppendorf tube. The primers were annealed to the RNA template by heating at 90ºC for 2 minutes and then transferring to a heating block set at 42ºC for 20 minutes. The components of the annealing reaction were added to the components of the reverse transcriptase reaction, (shown in Table 2.3), mixed and then incubated at 42ºC for 2-3 hours on a heating block (Grant, QBT2). Final Concentration Stock Reagent Volume for One Reaction Components from table 2.1 15 µl 1x Reverse Transcriptase Buffer 5x 6 µl 333µM dATP 10 mM 1 µl 333µM dGTP 10 mM 1 µl 333µM dTTP 10 mM 1 µl 1.67µM dCTP 100 µM 0.5 µl 20µCi [α-33P] dCTP 10 µCi/µl 3 µl 20 U Ribonuclease Inhibitor 40 U/µl 0.5 µl 50 U AMV Reverse Transcriptase 25 u/µl 2 µl Sterile distilled water to a final volume of 30 µl Table 2.3: Composition of the Reverse Transcriptase Reaction. Table adapted from Panorama™ Mouse Apoptosis Gene Arrays Protocol provided by Sigma-Genosys.

growth_protocol: Mice were maintained and killed under Home Office licence in accordance with British law (comparable with U.S. Public Health Service Policy on Humane Care and Use of Laboratory Animals). Transgenic and control animals were maintained on RM3 diet (1.15% calcium, 0.82% phosphorus, 4088.65I.U./kg Vitamin D, Special Diet Services, Witham, Essex CM8 3AD, UK), ad libitum. The timing of plugs and births were noted, litters were sacrificed at 7 and 14 days

extract_protocol: Once collected, the lenses were immediately homogenised in TRIzol® reagent (Invitrogen, Paisley, UK) using a tissue grinder (Wheaton). RNA was then isolated from these lenses using the protocol provided by Invitrogen.

treatment_protocol: We used wild type 129Sv/Ev mice, which were maintained under normal conditions and not treated with anything (see growth protocol)

tag: 14 day old mouse lens, strain 129Sv/Ev, dissected out from eyes using forceps in PBS under a dissecting microscope


title: MCF7/XBP1 p.3

status: Public on Aug 13 2007

type: RNA

city: Washington

first: Rebecca

middle: B.

last: Riggins

state: DC

address: E412 Research Building, 3970 Reservoir Rd. NW

department: Oncology, and Lombardi Comprehensive Cancer Center

country: USA

institute: Georgetown University

hyb_protocol: Standard Protocol for Hybridization:

scan_protocol: Standard Protocol for Affymetrix GeneChip Scanner 3000:

data_processing:
For .CHP file: MAS5.
For .CEL file and manuscript: Robust Multiple-Array Average (RMA) method from the Bioconductor Project.

label: biotin

molecule: total RNA

organism: Homo sapiens

source_name: Breast Cancer Cell Line

label_protocol: Standard Protocol for One-Cycle Labeling:

growth_protocol: Cells were cultured in IMEM supplemented with 5% FBS and 200 ug/ml Geneticin

extract_protocol: Trizol Extraction (Invitrogen), followed by RNeasy Column Cleanup (Qiagen)

tag: MCF7 cells stably transfected with XBP1, passage 3


title: Day 14 lens RNA, scan 6.2

city: Cardiff

first: Fiona

middle: Catherine

last: Mansergh

state: Wales

address: Museum Avenue

department: Biosciences

country: United Kingdom

institute: Cardiff University

organism: Mus musculus


title: Day 14 lens RNA, scan 2.1


title: Day 7 lens RNA, scan 11.2

source_name: 7 day old mouse lens, strain 129Sv/Ev

tag: 7 day old mouse lens, strain 129Sv/Ev, dissected out from eyes using forceps in PBS under a dissecting microscope


title: Day 7 lens RNA, scan 5.1


title: MCF7/c p.2

tag: MCF7 cells stably transfected with pCDNA3.1, passage 2


title: Day 7 lens RNA, scan 11.1

description: The results were analysed using an image analysis program, ImaGene 5 (Biodiscovery).  Using ImaGene, a virtual grid was placed over the image of the arrays assigning each spot a know grid reference; this allowed for the intensity of each spot to be calculated and also allowed the correct gene ID to be allocated to each spot (see Figure 3.2.).  Once ImaGene had calculated the spot intensities, the data were then exported to Microsoft® Excel for further analysis.  The data obtained from ImaGene produces a spot intensity and a background signal value for each individual spot.  The background value for each spot was calculated as a mean of the intensity of a set number of pixels surrounding the spot (as shown in Figure 2.1.).

hyb_protocol: Panorama™ mouse apoptosis arrays (Sigma-Genosys) were used for this set of experiments.  These arrays are nylon membranes containing 243 known apoptosis related genes.  The complete list of genes present on the array is given in Appendix 2.  Before the arrays were hybridised with the radiolabelled cDNA they were first pre-hybridised to prevent non-specific binding of DNA to the arrays.  The arrays were washed in 50ml 2x SSPE (Sigma) at room temperature for 5 minutes.  The arrays were then pre-hybridised in hybridisation solution (Sigma) containing salmon testes DNA (100µl Salmon testes DNA in 10ml hybridisation solution) at 65ºC for at least an hour before the addition of the radiolabelled cDNA. The unincorporated radiolabelled nucleotides had to be removed from the reaction mix before it was added to the arrays.  This should result in a reduction in the background signal during the hybridisation reaction.  To remove the unincorporated radiolabelled nucleotides a spin column containing sephadex beads (Sigma-Genosys) was used.  The spin column was centrifuged to remove any excess buffer before the sample was added.  Once the sample had been added to the centre of the spin column, the column was centrifuged at 1100 x g for 4 minutes.  The eluate from the column contained the purified cDNA sample; this sample could then be added to the arrays. The purified labelled cDNA was then added to 2-3mls hybridisation solution (containing 100μl salmon testes DNA per 10ml hybridisation solution).  The DNA was then denatured by heating at 95°C for 10 minutes.  The hybridisation solution containing the labelled cDNA was then added to the arrays, which were hybridised overnight for 18 hours in the hybridisation oven at 65°C (UVP, HC-3000 Hybricycler). After the hybridisation reaction was complete, the arrays were then washed to remove any unbound DNA with increasing concentrations of salt solution.  The hybridisation solution was decanted and discarded and 40-50ml of wash solution I (composition in Appendix 1) was added to the roller bottles.  The arrays were washed by gently inverting the bottles by hand at room temperature for 2-3 minutes.  This process was repeated twice more, with a new aliquot of wash solution I each time.  80-100ml of wash solution I (pre-warmed to 65°C) was then added to the roller bottles and the arrays were washed in the hybridisation oven at 65°C for 20 minutes, before the wash solution was discarded.  This process was repeated once more.  Wash solution II (pre-warmed to 65ºC: composition given in Appendix 1) was then added to the roller bottles (80-100ml) and washed for a further 20 minutes in the hybridisation oven.  The wash solution was then discarded and the arrays were wrapped in clingfilm before placing them into a storage phosphor screen (Amersham Biosciences).  The arrays were exposed to the phosphor screen for 5-7days.

scan_protocol: The phosphor screen was then scanned using the Typhoon scanner (Amersham Biosciences, Typhoon 9410 Variable Mode Imager) which extracted the energy from the screen and converted it into a computer image which could later be analysed by use of image analysis software. Once the phosphor screen had been scanned, the arrays were stripped removing any residual bound DNA.  This ensured that each array could be reused.  The arrays were added to 200ml of stripping solution (composition given in Appendix 1), which had been brought to the boil.  The arrays were then added to the stripping solution, which was then allowed to cool.  The arrays were then removed from the solution, the excess drained and the arrays wrapped in clingfilm.  Once stripped, the arrays were stored at -20°C.  Stripped arrays were exposed to the phosphor screen to check that the residual radiolabelled cDNA had been removed.

data_processing:
The data produced were normalised using the mean expression of the housekeeping genes from the arrays; housekeeping genes are genes continuously expressed throughout development.  The normalisation process allowed comparisons of gene expression to be made between the completed arrays; accounting for variations of the signal between arrays. The mean and standard deviation were calculated using the individual background values for each spot; these values were subsequently used to filter the data later on in the analysis process.  The spots whose original signal was lower than the mean plus two standard deviations of the background values were removed from the rest of the results following normalisation.
The individual background values were then subtracted from the corresponding spot intensity, to give a corrected intensity.  Each corrected spot value was normalised before being filtered.  The spots were normalised using the housekeeping genes printed on the apoptosis arrays.  A mean value was calculated from the corrected spot intensities from each of these housekeeping genes as well as Gapdh (a standard housekeeping gene), which was also printed on the array.  The array data were then normalised using the ratios of these mean values.
One of the arrays was designated as standard.  A ratio was calculated for each array by dividing the mean housekeeping value from the standard array by the mean housekeeping value from that array.  The corrected spot intensities were all multiplied by this ratio.
After normalisation, a mean value for each spot across the set of arrays was calculated using the corrected intensities.  The mean values for each spot were then compared for P7 and P14 using an unpaired t-test.  Spots with a 2-fold difference between these two time points and a p-value <0.05 were considered to have a significant change in gene expression.

label_protocol:
The labelling reactions are performed in two steps.  The first step, the annealing reaction, shown in Table 2.2, involves the annealing of mouse apoptosis cDNA primers (Sigma-Genosys) to the RNA template.  During the second step, radiolabelled nucleotides and reverse transcriptase (Sigma-Genosys) were added to initiate the cDNA synthesis reaction.
Final Concentration	Volume for One Reaction
2µg total RNA	2-11µl
Mouse Apoptosis cDNA Labelling Primers	4µl
Sterile distilled water to	15µl
Table 2.2: Composition of the Annealing Reaction.  Table taken from Panorama™ Mouse Apoptosis Gene Arrays Protocol.
For the annealing step, the components from the table above were added to an eppendorf tube.  The primers were annealed to the RNA template by heating at 90ºC for 2 minutes and then transferring to a heating block set at 42ºC for 20 minutes.
The components of the annealing reaction were added to the components of the reverse transcriptase reaction, (shown in Table 2.3), mixed and then incubated at 42ºC for 2-3 hours on a heating block (Grant, QBT2).
Final Concentration	Stock Reagent	Volume for One Reaction
Components from table 2.1		15 µl
1x Reverse Transcriptase Buffer	5x	6 µl
333µM dATP	10 mM	1 µl
333µM dGTP	10 mM	1 µl
333µM dTTP	10 mM	1 µl
1.67µM dCTP	100 µM	0.5 µl
20µCi [α-33P] dCTP	10 µCi/µl	3 µl
20 U Ribonuclease Inhibitor	40 U/µl	0.5 µl
50 U AMV Reverse Transcriptase	25 u/µl	2 µl
Sterile distilled water to a final volume of	30 µl
Table 2.3: Composition of the Reverse Transcriptase Reaction.  Table adapted from Panorama™ Mouse Apoptosis Gene Arrays Protocol provided by Sigma-Genosys.

extract_protocol: Once collected, the lenses were immediately homogenised in TRIzol® reagent (Invitrogen, Paisley, UK) using a tissue grinder (Wheaton).  RNA was then isolated from these lenses using the protocol provided by Invitrogen.


title: MCF7/XBP1 p.2

tag: MCF7 cells stably transfected with XBP1, passage 2


title: Day 7 lens RNA, scan 5.2


title: MCF7/XBP1 p.6

tag: MCF7 cells stably transfected with XBP1, passage 6


title: VA1.2

status: Public on Aug 14 2007

city: Roslin

first: Michael

middle: Bryan

last: Watson

state: Midlothian, EH25 9PS

address: Roslin Biocentre

department: ARK-Genomics

institute: The Roslin Institute

description: IAH/VLA sample VA1.2

hyb_protocol: Hybridised (2h) according to previously published protocol: 1. Burton, J.E., et al., Development of a multi-pathogen oligonucleotide microarray for detection of Bacillus anthracis. Mol Cell Probes, 2005. 19(5): p. 349-57.

scan_protocol: Slides were scanned using a Hewlett Packard ScanArray Lite

data_processing: Fluorescence was quantified using ScanArray Express 2.0.

organism: Foot-and-mouth disease virus O

source_name: Amplified RNA from Cell cultured FMDV type O

label_protocol: MultiScribe reverse transcriptase (Applied Biosystems) and random amplification followed by direct labelling with Cy3-dCTP. Primers used:GTT TCC CAG TCA CGA TCN NNN NNN NN and GTT TCC CAG TCA CGA TC

extract_protocol: Total RNA was extracted from FMD virus cultured in primary bovine thyroid cells (Rneasy Mini Kit, Qiagen)


title: Zebrafish-C

city: Wuhan

first: M

middle: X

last: Chang

address: Luojiaashan

country: China

institute: Institute of Hydrobiology, Chinese Academy of Sciences

description: Gene expression data from the uninjected embryos

hyb_protocol: Fifteen microgram aliquants of the fragmented cRNA were hybridized with the D. rerio Genome AFF-F0027 GeneChip® microarray (Affymetrix, CA) in hybridization cocktail. Hybridization was allowed to proceed overnight (16 h) at 45 °C

scan_protocol: The distribution of fluorescent material on the array was obtained using GeneChip® Scanner 3000 (Affymetrix, CA)

data_processing: The scanned images were firstly assessed by visual inspection then analyzed to generate raw data files saved as CEL files using the default setting of GCOS 1.4. A global scaling factor of 500 was used to normalize the different arrays. The differentially expressed genes were judged according to change p-value calculated by GCOS 1.4, and 2-fold change as an empirical criterion.

organism: Danio rerio

source_name: 24 hours post-fertilization embryos

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

growth_protocol: Zebrafish embryos were maintained at 28 ± 0.5 °C

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

treatment_protocol: Selected embryos were transferred to 2 mL Eppendorf tube, rinsed with DEPC-treated water. After discarding water, the Trizol solution (GibcoBRL) was added to Eppendorf tube containing embryos.

tag: the uninjected embryos


title: MCF7/c p.1

tag: MCF7 cells stably transfected with pCDNA3.1, passage 1


title: MCF7/c p.6

tag: MCF7 cells stably transfected with pCDNA3.1, passage 6


title: P3_AS_HeLa_sRW_C79_T1

status: Public on Aug 10 2007

city: Santa Clara

first: Thomas

middle: R

last: Gingeras

state: CA

address: 3380 Central Expressway

institute: Affymetrix, Inc.

description: See publication for details

hyb_protocol: See publication for details

scan_protocol: See publication for details

data_processing: See publication for details

source_name: Whole cell RNA from HeLa cell line

label_protocol: See publication for details

extract_protocol: Whole-cell short RNA protocol - see publication for details

tag: See publication for details


title: P3_S_HepG2_sRW_C11_T1

description: See publication for details

source_name: Whole-cell short RNA from HepG2 cell line


title: P3_S_HepG2_sRW_C80_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C79_T1

description: See publication for details


title: P3_S_HeLa_sRW_C14_T1

description: See publication for details

source_name: Whole-cell short RNA from HeLa cell line


title: P3_AS_HepG2_sRW_C14_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C80_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C12_T1

description: See publication for details


title: P3_S_HepG2_sRW_C28_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C11_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C28_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C11_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C14_T1

description: See publication for details


title: P3_S_HeLa_sRW_C28_T1

description: See publication for details


title: P3_S_HeLa_sRW_C79_T1

description: See publication for details


title: P3_S_HeLa_sRW_C11_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C80_T1

description: See publication for details


title: P3_S_HeLa_sRW_C80_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C28_T1

description: See publication for details


title: P3_S_HepG2_sRW_C79_T1

description: See publication for details


title: P3_S_HepG2_sRW_C29_T1

description: See publication for details


title: P3_S_HepG2_sRW_C12_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C12_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C13_T1

description: See publication for details


title: P3_S_HepG2_sRW_C13_T1

description: See publication for details


title: P3_S_HeLa_sRW_C13_T1

description: See publication for details


title: P3_S_HepG2_sRW_C14_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C13_T1

description: See publication for details


title: P3_S_HeLa_sRW_C12_T1

description: See publication for details


title: P4

type: genomic

city: Heidelberg

first: Stefan

middle: M

last: Pfister

address: INF 580

department: Molecular Genetics

country: Germany

institute: German Cancer Research Center

description: Comparison of tumor DNA (Cy3-labeled) vs. normal DNA (Cy5-labeled). Whole-genome screening of DNA copy-number aberrations using array-CGH

hyb_protocol: Ten micrograms each of tumor and reference DNA were combined with 300µg of human Cot-1 DNA (Roche Diagnostics, Mannheim, Germany) and suspended in 125µl of UltrahybTM hybridization buffer (Ambion, Bad Soden, Germany). Denaturation of combined DNA was performed at 75°C for 10 min, followed by preannealing at 37°C for 30 min. Hybridization was carried out in a GeneTAC Hybridization Station (Genomic Solutions, Oberhaching, Germany) for 36 hr at 37°C. Slides were washed with 2× SSC/0.05% Tween-20 at 37°C for 1 cycle, 2× SSC/0.05% Tween-20 at 44°C for 2 cycles, 50% formamide/2× SSC/0.1% Tween-20 at 44°C for 3 cycles, 2× SSC/0.05% Tween-20 at 44°C for 2 cycles, and finally 1× PBS/0.05% Tween-20 at 25°C for 2 cycles. Slides were dried by centrifugation at 54 g for 1 min, followed by centrifugation at 216 g for 1 min in a clinical centrifuge.

scan_protocol: Arrays were scanned at a 5µm resolution with an Axon 4000B scanner (Axon Instruments, Burlingame, USA) and images analyzed using GenePix Pro 6.0 software (Axon Instruments). Results files containing all relevant raw data were processed by use of ChipYard microarray analysis platform software (developed in-house), the statistical programming language R (http://www.r-project.org), and packages of the Bioconductor project (http://bioconductor.org; Ihaka and Gentleman, 1996).

data_processing: Fluorescence intensities of all spots were filtered consistently (intensity/local background >3; mean/median intensity <1.3; standard deviation of clone log2 ratios <0.2) and globally normalized using the printtip-loess algorithm (Smyth and Speed, 2003).

molecule: genomic DNA

source_name: tumor DNA

label_protocol: Genomic DNA (300-500 ng) was labeled with Cy3-labeled dCTP (tumor) or Cy5-conjugated dCTP (control) by use of a BioPrime DNA Labeling Kit (Invitrogen, Karlsruhe, Germany). Unincorporated nucleotides, random primers, and dyes were removed by use of MicroCon YM30 Spin Columns (Millipore, Schwalbach, Germany). Incorporation of dyes into the labeled DNA was determined by UV spectrophotometry.

extract_protocol: Genomic DNA from tumor tissue was isolated using the Blood and Cell Culture Kit (Qiagen,Hilden, Germany) following the instructions of the suppliers.  Genomic DNA from peripheral blood mononuclear cells of healthy donors was isolated by use of the Quiagen DNA Blood Midi-Kit (Hilden, Germany).

tag: 3 years old female patient with stPNET of Mstage 0

source_name: Pooled lymphocyte DNA from 10 healthy individuals (age 25-40 years)

tag: pooled DNA from peripheral blood mononuclear cells of ten healthy donors


title: P3

tag: 7 years old female patient with stPNET of Mstage 0


title: P3_S_HeLa_sRW_C29_T1

description: See publication for details


title: P3_S_HeLa_sRW_C30_T1

description: See publication for details


title: M7

tag: 12 years old female patient with stPNET of Mstage N/A


title: P3_S_HepG2_sRW_C30_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C30_T1

description: See publication for details


title: M6

tag: <18 years old male patient with stPNET of Mstage N/A


title: P3_S_HepG2_sRW_C31_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C01_T1

description: See publication for details


title: P3_S_HepG2_sRW_C44_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C44_T1

description: See publication for details


title: P3_S_HeLa_sRW_C31_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C01_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C45_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C29_T1

description: See publication for details


title: 682

tag: 2 years old male patient with stPNET of Mstage 1


title: P3_AS_HeLa_sRW_C44_T1

description: See publication for details


title: 750

tag: 3 years old male patient with stPNET of Mstage 0


title: P3_S_HepG2_sRW_C01_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C45_T1

description: See publication for details


title: P3_S_HepG2_sRW_C46_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C31_T1

description: See publication for details


title: 584

tag: 6 years old male patient with stPNET of Mstage 1


title: P3_AS_HeLa_sRW_C30_T1

description: See publication for details


title: P3_S_HepG2_sRW_C08_T1

description: See publication for details


title: P3_S_HeLa_sRW_C45_T1

description: See publication for details


title: P3_S_HeLa_sRW_C08_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C46_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C31_T1

description: See publication for details


title: P3_S_HeLa_sRW_C46_T1

description: See publication for details


title: 254

tag: 1 years old female patient with stPNET of Mstage 0


title: P3_S_HeLa_sRW_C01_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C46_T1

description: See publication for details


title: 446

tag: 0 years old male patient with stPNET of Mstage 0


title: P3_AS_HepG2_sRW_C08_T1

description: See publication for details


title: P3_S_HepG2_sRW_C45_T1

description: See publication for details


title: 563

tag: 11 years old female patient with stPNET of Mstage 1


title: eugenol_45min

city: Changchun

first: Xuming

last: Deng

address: Xi'an Street 5333#

institute: College of Animal Science and Veterinary Medicine, Jilin University

description: eugenol treatment

hyb_protocol: Standard Affymetrix procedures

scan_protocol: Standard Affymetrix procedures

data_processing: Affymetrix GeneChip Opererating Software (GCOS) version 1.0

label: Biotin

organism: Staphylococcus aureus

source_name: Staphylococcus aureus_eugenol

label_protocol: Conversion to cDNA and labeling was performed as described in Beenken et al., Journal of Bacteriology, 2004 July, 186(14):4665-4684

growth_protocol: (i) S. aureus strain ATCC25923 was grown overnight at 200 rpm in a rotary shaker at 37°C in 10 ml of MHB II, (ii) the culture were inoculated to an initial OD600 of 0.05 in 100 ml of MHB II in 250-ml Erlenmeyer flasks, the bacteria were grown at 37°C at 200 rpm to an OD600 of 0.3. (iii)  eugenol , prepared in dimethyl sulfoxide (DMSO), was added to the cultures, giving a final concentration of 1/2× MIC (250 µg/ml), and the final concentration of DMSO was 1% (vol/vol). Then, the bacterial suspensions were further incubated for 45 min at 37°C

extract_protocol: Total bacterial RNA was isolated as described by Beenken et al., 2004 July, 186(14):4665-4684

treatment_protocol: eugenol (250 µg/ml)

tag: Standard strain ATCC25923


title: 24h +TGF-beta pulp tissu

city: Oulu

first: Virve

last: Pääkkönen

address: Aapistie 3

department: Institute of Dentistry

country: Finland

institute: University of Oulu

hyb_protocol: Experimental procedures for GeneChip (Affymetrix, Santa Clara, CA, USA) were performed according to the Affymetrix GeneChip Expression Analysis Technical Manual.

scan_protocol: HP GeneArray Scanner was used.

data_processing: The expression data was analyzed using Affymetrix GeneChip Operating Software version 1.1.1. Signal intensities of all probe sets were scaled to the target value of 500.

source_name: Pulp tissue, tissue cultured for 24h with TGF-beta

label_protocol: Double-stranded DNA was synthesized by means of the Superscript Choice System (Gibco BRL Life Technologies, Rockville, MD, USA) and T7-(dT)24 primer. The DNA was purified using GeneChip Sample Cleanup Module (Qiagen, Valencia, CA, USA). In vitro transcription was performed to produce biotin labeled cRNA using a BioArray HighYield RNA Transcription Labeling Kit (Enzo Diagnostics, Farmingdale, NY, USA) according to the manufacturer’s instructions. Biotinylated cRNA was cleaned with an GeneChip Sample Cleanup Module (Qiagen), fragmented to 35 to 200 nt, and hybridized to Affymetrix HGU133A . After being washed, the array was stained with streptavidin-phycoerythrin (Molecular Probes, Eugene, OR, USA). Staining signal was amplified by biotinylated anti-streptavidin (Vector Laboratories, Burlingame, CA, USA) and second staining with streptavidin-phycoerythrin.

extract_protocol: Total RNA was isolated using Trizol-protocol and further purified by Qiagen RNeasy Mini Kit

treatment_protocol: Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The pulps were cultured on 24-well culture plates in OPTIMEM-medium for 24h with TGF-beta.

tag: Pulp tissue obtained from third molars of young patients and tissue cultured for 24h with TGF-beta


title: 48h +TGF-beta pulp tissue

source_name: Pulp tissue, tissue cultured for 48h with TGF-beta

treatment_protocol: Freshly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The pulps were cultured on 24-well culture plates in OPTIMEM-medium for 48h with TGF-beta.

tag: Pulp tissue obtained from third molars of young patients and tissue cultured for 48h with TGF-beta


title: P3_S_HeLa_sRW_C44_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C29_T1

description: See publication for details


title: P3_S_HepG2_sRW_C47_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C08_T1

description: See publication for details


title: P3_S_HepG2_sRW_C10_T1

description: See publication for details


title: P3_S_HeLa_sRW_C47_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C47_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C47_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C10_T1

description: See publication for details


title: P3_S_HeLa_sRW_C48_T1

description: See publication for details


title: P3_S_HepG2_sRW_C03_T1

description: See publication for details


title: P3_S_HeLa_sRW_C03_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C48_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C03_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C15_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C10_T1

description: See publication for details


title: P3_S_HepG2_sRW_C15_T1

description: See publication for details


title: P3_S_HepG2_sRW_C61_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C03_T1

description: See publication for details


title: 48h -TGF-beta pulp tissue

source_name: Pulp tissue, tissue cultured for 48h without TGF-beta

treatment_protocol: Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The pulps were cultured on 24-well culture plates in OPTIMEM-medium for 48h without TGF-beta.

tag: Pulp tissue obtained from third molars of young patients and tissue cultured for 48h without TGF-beta


title: P3_S_HepG2_sRW_C04_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C15_T1

description: See publication for details


title: P3_S_HepG2_sRW_C17_T1

description: See publication for details


title: 5h +TGF-beta pulp tissue

source_name: Pulp tissue, tissue cultured for 5h with TGF-beta

treatment_protocol: Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The pulps were cultured on 24-well culture plates in OPTIMEM-medium for 5h with TGF-beta.

tag: Pulp tissue obtained from third molars of young patients and tissue cultured for 5h with TGF-beta


title: 24h -TGF-beta pulp tissue

source_name: Pulp tissue, tissue cultured for 24h without TGF-beta

treatment_protocol: Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The pulps were cultured on 24-well culture plates in OPTIMEM-medium for 24h without TGF-beta.

tag: Pulp tissue obtained from third molars of young patients and tissue cultured for 24h without TGF-beta


title: 5h -TGF-beta pulp tissue

source_name: Pulp tissue, tissue cultured for 5h without TGF-beta

treatment_protocol: Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The pulps were cultured on 24-well culture plates in OPTIMEM-medium for 5h without TGF-beta.

tag: Pulp tissue obtained from third molars of young adults, and cultured for 5h without TGF-beta


title: P3_S_HeLa_sRW_C04_T1

description: See publication for details

tag: See publication for details


title: 1h -TGF-beta pulp tissue

source_name: Pulp tissue, tissue cultured for 1h without TGF-beta

treatment_protocol: Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The pulps were cultured on 24-well culture plates in OPTIMEM-medium for 1h without TGF-beta.

tag: Pulp tissue obtained from third molars of young patients, and tissue cultured for 1h without TGF-beta


title: P3_S_HeLa_sRW_C61_T1

description: See publication for details


title: 1h +TGF-beta pulp tissue

source_name: Pulp tissue, tissue cultured for 1h with TGF-beta

treatment_protocol: Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The pulps were cultured on 24-well culture plates in OPTIMEM-medium for 1h with TGF-beta.

tag: Pulp tissue obtained from third molars of young patients and tissue cultured for 1h with TGF-beta


title: P3_AS_HepG2_sRW_C04_T1

description: See publication for details


title: P3_S_HeLa_sRW_C10_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C17_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C48_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C61_T1

description: See publication for details


title: P3_S_HepG2_sRW_C62_T1

description: See publication for details


title: P3_S_HeLa_sRW_C05_T1

description: See publication for details


title: P3_S_HepG2_sRW_C05_T1

description: See publication for details


title: P3_S_HeLa_sRW_C17_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C62_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C61_T1

description: See publication for details


title: P3_S_HepG2_sRW_C24_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C62_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C17_T1

description: See publication for details


title: P3_S_HepG2_sRW_C63_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C04_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C05_T1

description: See publication for details


title: P3_S_HeLa_sRW_C63_T1

description: See publication for details


title: P3_S_HeLa_sRW_C62_T1

description: See publication for details


title: P3_S_HeLa_sRW_C02_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C24_T1

description: See publication for details


title: P3_S_HepG2_sRW_C26_T1

description: See publication for details


title: P3_S_HepG2_sRW_C48_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C24_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C05_T1

description: See publication for details


title: P3_S_HeLa_sRW_C26_T1

description: See publication for details


title: P3_S_HeLa_sRW_C06_T1

description: See publication for details


title: P3_S_HeLa_sRW_C15_T1

description: See publication for details


title: P3_S_HeLa_sRW_C24_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C63_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C06_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C06_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C63_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C26_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C02_T1

description: See publication for details


title: P3_S_HepG2_sRW_C02_T1

description: See publication for details


title: P3_S_HepG2_sRW_C64_T1

description: See publication for details


title: P3_S_HepG2_sRW_C19_T1

description: See publication for details


title: P3_S_HepG2_sRW_C35_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C02_T1

description: See publication for details


title: P3_S_HepG2_sRW_C07_T1

description: See publication for details


title: P3_S_HeLa_sRW_C07_T1

description: See publication for details


title: P3_S_HeLa_sRW_C64_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C64_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C64_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C07_T1

description: See publication for details


title: P3_S_HeLa_sRW_C19_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C35_T1

description: See publication for details


title: P3_S_HeLa_sRW_C35_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C26_T1

description: See publication for details


title: P3_S_HepG2_sRW_C78_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C35_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C07_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C19_T1

description: See publication for details


title: P3_S_HeLa_sRW_C78_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C19_T1

description: See publication for details


title: P3_S_HepG2_sRW_C40_T1

description: See publication for details


title: P3_S_HepG2_sRW_C06_T1

description: See publication for details


title: P3_S_HepG2_sRW_C09_T1

description: See publication for details


title: P3_S_HeLa_sRW_C20_T1

description: See publication for details


title: P3_S_HepG2_sRW_C20_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C78_T1

description: See publication for details


title: P3_S_HepG2_sRW_C81_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C40_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C78_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C40_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C20_T1

description: See publication for details


title: P3_S_HeLa_sRW_C09_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C09_T1

description: See publication for details


title: P3_S_HeLa_sRW_C21_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C09_T1

description: See publication for details


title: P3_S_HepG2_sRW_C21_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C21_T1

description: See publication for details


title: P3_S_HeLa_sRW_C40_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C20_T1

description: See publication for details


title: P3_S_HeLa_sRW_C81_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C21_T1

description: See publication for details


title: P3_S_HepG2_sRW_C42_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C16_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C16_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C42_T1

description: See publication for details


title: 1017

tag: 10 years old male patient with astrocytoma WHO °2; localization:NA

tag: pooled DNA from peripheral blood mononuclear cells of ten healthy donors


title: P3_S_HeLa_sRW_C16_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C42_T1

description: See publication for details


title: P3_S_HeLa_sRW_C42_T1

description: See publication for details


title: P3_S_HepG2_sRW_C22_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C81_T1

description: See publication for details


title: 1003

tag: 7 years old male patient with astrocytoma WHO °1; localization:fourth ventricle


title: 1002

tag: 1 year old female patient with astrocytoma WHO °1; localization:cerebellum


title: P3_AS_HepG2_sRW_C81_T1

description: See publication for details


title: P3_S_HepG2_sRW_C18_T1

description: See publication for details


title: P3_S_HepG2_sRW_C49_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C18_T1

description: See publication for details


title: 1001

tag: 6 years old female patient with astrocytoma WHO °1; localization:cerebellum


title: P3_S_HeLa_sRW_C22_T1

description: See publication for details


title: P3_S_HepG2_sRW_C25_T1

description: See publication for details


title: 986

tag: 5 years old male patient with astrocytoma WHO °1; localization:cerebellum


title: P3_S_HepG2_sRW_C16_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C22_T1

description: See publication for details


title: P3_S_HeLa_sRW_C25_T1

description: See publication for details


title: P3_S_HeLa_sRW_C49_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C49_T1

description: See publication for details


title: P3_S_HepG2_sRW_C51_T1

description: See publication for details


title: P3_S_HeLa_sRW_C18_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C18_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C23_T1

description: See publication for details


title: 984

tag: 3 years old male patient with astrocytoma WHO °1; localization:cerebellum


title: P3_S_HepG2_sRW_C23_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C49_T1

description: See publication for details


title: 966

tag: 11 years old male patient with astrocytoma WHO °2; localization:NA


title: 963

tag: 10 years old male patient with astrocytoma WHO °1; localization:NA


title: P3_AS_HepG2_sRW_C25_T1

description: See publication for details


title: 960


title: P3_AS_HepG2_sRW_C22_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C51_T1

description: See publication for details


title: 957

tag: 3 years old female patient with astrocytoma WHO °1; localization:cerebellum


title: P3_AS_HeLa_sRW_C23_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C51_T1

description: See publication for details


title: P3_S_HepG2_sRW_C36_T1

description: See publication for details


title: P3_S_HeLa_sRW_C51_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C25_T1

description: See publication for details


title: 922

tag: 2 years old male patient with astrocytoma WHO °1; localization:chiasma & diencephalon


title: P3_S_HepG2_sRW_C27_T1

description: See publication for details


title: P3_S_HepG2_sRW_C58_T1

description: See publication for details


title: P3_S_HeLa_sRW_C27_T1

description: See publication for details


title: 899

tag: 2 years old male patient with astrocytoma WHO °1; localization:chiasma opticum


title: 816

tag: 15 years old male patient with astrocytoma WHO °2; localization:NA


title: P3_S_HeLa_sRW_C23_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C58_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C36_T1

description: See publication for details


title: 813

tag: 3 years old female patient with astrocytoma WHO °1; localization:NA


title: P3_S_HeLa_sRW_C36_T1

description: See publication for details

tag: See publication for details


title: P3_S_HepG2_sRW_C60_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C36_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C27_T1

description: See publication for details


title: P3_S_HepG2_sRW_C32_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C58_T1

description: See publication for details


title: 740


title: 651

tag: 1 year old male patient with astrocytoma WHO °1; localization:third ventricle


title: P3_AS_HeLa_sRW_C27_T1

description: See publication for details


title: P3_S_HeLa_sRW_C37_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C32_T1

description: See publication for details


title: P3_S_HepG2_sRW_C37_T1

description: See publication for details


title: P3_S_HeLa_sRW_C58_T1

description: See publication for details


title: P3_S_HeLa_sRW_C32_T1

description: See publication for details


title: P3_S_HeLa_sRW_C60_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C60_T1

description: See publication for details


title: 623


title: P3_AS_HeLa_sRW_C32_T1

description: See publication for details


title: P3_S_HepG2_sRW_C33_T1

description: See publication for details


title: P3_S_HepG2_sRW_C34_T1

description: See publication for details


title: 581

tag: 14 years old male patient with astrocytoma WHO °1; localization:NA


title: 593

tag: <18 years old male patient with astrocytoma WHO °2; localization:intramedullary


title: P3_S_HepG2_sRW_C38_T1

description: See publication for details


title: 463

tag: <18 years old female patient with astrocytoma WHO °1; localization:NA

tag: pooled DNA from peripheral blood mononuclear cells of ten healthy donors


title: P3_AS_HeLa_sRW_C60_T1

description: See publication for details


title: P3_S_HeLa_sRW_C34_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C38_T1

description: See publication for details


title: 458


title: P3_S_HeLa_sRW_C38_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C37_T1

description: See publication for details


title: P3_S_HeLa_sRW_C33_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C33_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C34_T1

description: See publication for details


title: P3_S_HepG2_sRW_C65_T1

description: See publication for details


title: P3_S_HepG2_sRW_C39_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C65_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C33_T1

description: See publication for details


title: 397

tag: 6 years old female patient with astrocytoma WHO °1; localization:brain stem


title: P3_S_HepG2_sRW_C41_T1

description: See publication for details


title: P3_S_HeLa_sRW_C41_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C38_T1

description: See publication for details


title: 440

tag: 11 years old male patient with astrocytoma WHO °1; localization:occipital lobe


title: 391

tag: 13 years old male patient with astrocytoma WHO °2; localization:NA


title: P3_S_HeLa_sRW_C39_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C39_T1

description: See publication for details


title: P3_S_HepG2_sRW_C53_T1

description: See publication for details


title: 396

tag: 6 years old female patient with astrocytoma WHO °2; localization:NA


title: P3_AS_HeLa_sRW_C41_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C41_T1

description: See publication for details


title: 332

tag: <18 years old male patient with astrocytoma WHO °1; localization:NA


title: P3_AS_HepG2_sRW_C37_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C65_T1

description: See publication for details


title: P3_S_HepG2_sRW_C67_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C53_T1

description: See publication for details


title: P3_S_HeLa_sRW_C43_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C67_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C43_T1

description: See publication for details


title: 330

tag: 2 years old female patient with astrocytoma WHO °1; localization:hypothalamus


title: P3_S_HeLa_sRW_C53_T1

description: See publication for details


title: 293

tag: 7 years old female patient with astrocytoma WHO °1; localization:NA


title: P3_AS_HepG2_sRW_C39_T1

description: See publication for details


title: 292

tag: 3 years old female patient with astrocytoma WHO °1; localization:vermis cerebelli


title: P3_S_HeLa_sRW_C91_T1

description: See publication for details


title: P3_S_HeLa_sRW_C50_T1

description: See publication for details


title: P3_S_HepG2_sRW_C43_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C43_T1

description: See publication for details


title: P3_S_HepG2_sRW_C69_T1

description: See publication for details


title: 291

tag: 2 years old male patient with astrocytoma WHO °1; localization:optic nerve


title: P3_S_HeLa_sRW_C69_T1

description: See publication for details


title: P3_S_HepG2_sRW_C50_T1

description: See publication for details


title: 289

tag: 8 years old female patient with astrocytoma WHO °2; localization:NA


title: P3_AS_HepG2_sRW_C34_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C91_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C67_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C69_T1

description: See publication for details


title: 286

tag: 17 years old female patient with astrocytoma WHO °2; localization:NA


title: P3_AS_HepG2_sRW_C50_T1

description: See publication for details

tag: See publication for details


title: P3_S_HeLa_sRW_C67_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C53_T1

description: See publication for details


title: P3_S_HeLa_sRW_C71_T1

description: See publication for details


title: P3_S_HepG2_sRW_C72_T1

description: See publication for details


title: 283

tag: 10 years old female patient with astrocytoma WHO °1; localization:NA


title: 282

tag: 14 years old male patient with astrocytoma WHO °1; localization:NA


title: P3_S_HepG2_sRW_C71_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C69_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C50_T1

description: See publication for details


title: P3_S_HeLa_sRW_C65_T1

description: See publication for details


title: P3_S_HepG2_sRW_C74_T1

description: See publication for details


title: 270

tag: 11 years old male patient with astrocytoma WHO °1; localization:NA


title: P3_AS_HepG2_sRW_C72_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C71_T1

description: See publication for details


title: 271

tag: 2 years old female patient with astrocytoma WHO °2; localization:postcentral right hemisphere


title: P3_S_HepG2_sRW_C91_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C74_T1

description: See publication for details


title: P3_S_HeLa_sRW_C72_T1

description: See publication for details


title: 269

tag: 11 years old female patient with astrocytoma WHO °1; localization:brain stem

tag: pooled DNA from peripheral blood mononuclear cells of ten healthy donors


title: P3_S_HeLa_sRW_C74_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C91_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C72_T1

description: See publication for details


title: 264

tag: 1 year old female patient with astrocytoma WHO °2; localization:fourth ventricle


title: P3_AS_HeLa_sRW_C71_T1

description: See publication for details


title: 260

tag: 9/12 years old male patient with astrocytoma WHO °1; localization:thalamus


title: P3_S_HepG2_sRW_C52_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C74_T1

description: See publication for details


title: P3_S_HepG2_sRW_C54_T1

description: See publication for details


title: 249

tag: 4 years old male patient with astrocytoma WHO °2; localization:infratentorial


title: P3_S_HeLa_sRW_C85_T1

description: See publication for details


title: P3_S_HepG2_sRW_C85_T1

description: See publication for details


title: P3_S_HeLa_sRW_C52_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C52_T1

description: See publication for details


title: P3_S_HeLa_sRW_C54_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C54_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C85_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C54_T1

description: See publication for details


title: 243

tag: 4 years old male patient with astrocytoma WHO °2; localization:fourth ventricle


title: 34

tag: 2 years old female patient with astrocytoma WHO °1; localization:chiasma & diencephalon


title: P3_S_HeLa_sRW_C55_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C52_T1

description: See publication for details


title: 35

tag: 5 years old female patient with astrocytoma WHO °1; localization:cerebellum


title: P3_S_HepG2_sRW_C57_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C57_T1

description: See publication for details


title: P3_S_HepG2_sRW_C55_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C85_T1

description: See publication for details


title: P3_S_HepG2_sRW_C76_T1

description: See publication for details


title: P3_S_HeLa_sRW_C57_T1

description: See publication for details


title: 33


title: 32

tag: 5 years old male patient with astrocytoma WHO °1; localization:cerebellum


title: P3_AS_HepG2_sRW_C55_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C57_T1

description: See publication for details


title: P3_S_HepG2_sRW_C59_T1

description: See publication for details


title: P3_S_HeLa_sRW_C56_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C76_T1

description: See publication for details


title: P3_S_HepG2_sRW_C56_T1

description: See publication for details


title: P3_S_HeLa_sRW_C83_T1

description: See publication for details


title: P3_S_HepG2_sRW_C83_T1

description: See publication for details


title: P3_S_HeLa_sRW_C59_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C55_T1

description: See publication for details


title: 29

tag: 8 years old male patient with astrocytoma WHO °1; localization:cerebellum


title: gibberellin plus Yariv-reagent treatment rep2

city: Tokyo

first: kiyoshi

last: mashiguchi

address: 1-1-1 Yayoi, Bunkyo-ku

country: Japan

institute: University of Tokyo

description: Microarray analysis was performed to know how many gibberellin (GA)-responsive genes are inhibited by beta-Yariv reagent, a specific binder of plant arabinogalactan-proteins

data_processing: GCOS

molecule: polyA RNA

organism: Hordeum vulgare

source_name: gibberellin plus Yariv-reagent treated barley aleurone protoplasts

label_protocol: 0.3 microgram of mRNA was processed to produce biotinylated cRNA targets.

extract_protocol: Total RNA was extracted from protoplasts in 1:1 (v/v) mixture of TE-saturated phenol and 0.1 M Tris-HCl buffer (pH 8.0) containing 0.1 M LiCl, 10 mM EDTA and 1 % SDS, and precipitated in 2.5 M LiCl according to Shirzadegan et al. (1991) except that extracted samples were not incubated in 80°C.  Contaminated DNA was digested with DNase I (Takara, Otsu, Shiga, Japan).  mRNA was purified from total RNA using Dynabeads Oligo (dT)25 (DYNAL, Dynal Biotech, Oslo, Norway) after DNase I treatment.

tag: Protoplasts from Hordeum vulgare L. cv. Himalaya


title: P3_AS_HepG2_sRW_C76_T1

description: See publication for details


title: P3_S_HeLa_sRW_C76_T1

description: See publication for details


title: 27

tag: 1 year old female patient with astrocytoma WHO °1; localization:cerebellum


title: gibberellin treatment rep2

source_name: gibberellin treated barley aleurone protoplasts


title: P3_AS_HepG2_sRW_C56_T1

description: See publication for details

tag: See publication for details


title: 26

tag: 16 years old female patient with astrocytoma WHO °1; localization:parietal lobe


title: P3_AS_HeLa_sRW_C56_T1

description: See publication for details


title: 24


title: P3_AS_HepG2_sRW_C59_T1

description: See publication for details


title: Control (DMSO) treatment rep2

source_name: DMSO treated barley aleurone protoplasts


title: 22

tag: 4 years old male patient with astrocytoma WHO °1; localization:frontal lobe


title: 31

tag: 8 years old female patient with astrocytoma WHO °1; localization:frontal lobe


title: gibberellin plus Yariv-reagent treatment rep1


title: 21

tag: 9 years old male patient with astrocytoma WHO °1; localization:cerebellum


title: gibberellin treatment rep1


title: Control (DMSO) treatment rep1


title: P3_AS_HeLa_sRW_C59_T1

description: See publication for details

source_name: Whole cell RNA from HeLa cell line


title: P3_S_HepG2_sRW_C70_T1

description: See publication for details

source_name: Whole-cell short RNA from HepG2 cell line


title: P3_S_HepG2_sRW_C66_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C83_T1

description: See publication for details


title: 20


title: 19

tag: 1 year old male patient with astrocytoma WHO °1; localization:diencephalon


title: P3_S_HepG2_sRW_C90_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C66_T1

description: See publication for details


title: P3_S_HeLa_sRW_C70_T1

description: See publication for details

source_name: Whole-cell short RNA from HeLa cell line


title: 18

source_name: tumor DNA

tag: 1 year old female patient with astrocytoma WHO °1; localization:fourth ventricle

tag: pooled DNA from peripheral blood mononuclear cells of ten healthy donors


title: P3_AS_HepG2_sRW_C83_T1

description: See publication for details


title: P3_S_HeLa_sRW_C66_T1

description: See publication for details


title: 16

tag: 11 years old female patient with astrocytoma WHO °1; localization:cerebellum

source_name: Pooled lymphocyte DNA from 10 healthy individuals (age 25-40 years)


title: P3_AS_HepG2_sRW_C66_T1

description: See publication for details


title: 15


title: P3_AS_HepG2_sRW_C70_T1

description: See publication for details


title: P3_S_HeLa_sRW_C90_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C70_T1

description: See publication for details


title: P3_S_HepG2_sRW_C68_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C90_T1

description: See publication for details


title: 12

tag: 2 years old male patient with astrocytoma WHO °2; localization:temporal lobe


title: P3_AS_HepG2_sRW_C90_T1

description: See publication for details


title: P3_S_HepG2_sRW_C86_T1

description: See publication for details


title: 13

tag: 14 years old female patient with astrocytoma WHO °1; localization:cerebellum


title: P3_S_HepG2_sRW_C87_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C68_T1

description: See publication for details


title: 10

tag: 10 years old male patient with astrocytoma WHO °1; localization:occipital lobe


title: P3_AS_HeLa_sRW_C68_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C86_T1

description: See publication for details


title: P3_S_HeLa_sRW_C68_T1

description: See publication for details


title: P3_S_HeLa_sRW_C87_T1

description: See publication for details


title: 11

tag: 6 years old male patient with astrocytoma WHO °1; localization:cerebellum


title: 8

tag: 14 years old male patient with astrocytoma WHO °1; localization:cerebellum


title: P3_S_HeLa_sRW_C86_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C75_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C86_T1

description: See publication for details


title: 6

tag: 9/12 years old male patient with astrocytoma WHO °1; localization:chiasma & diencephalon


title: P3_S_HeLa_sRW_C75_T1

description: See publication for details


title: 9

tag: 8 years old female patient with astrocytoma WHO °1; localization:cerebellum


title: P3_S_HepG2_sRW_C88_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C87_T1

description: See publication for details


title: P3_AS_HeLa_sRW_C87_T1

description: See publication for details


title: 3


title: 2


title: P3_AS_HepG2_sRW_C75_T1

description: See publication for details


title: transfected HeLa cells; siGFP versus mock 2a

status: Public on Aug 09 2007

first: Cordula

last: Tschuch

address: Neuenheimer Feld 580

institute: DKFZ

description: Experiment 2a

hyb_protocol: Purified, dye-labeled cDAN was mixed with Ultra-Hyb buffer (Ambion), agitated for 60min at 60°, then for 10min at 70°C on a thermo mixer and subsequently applied to pre-heated microarrays mounted in a GeneTAC Hybridisation Station. After hybridization slides were washed sequential.

scan_protocol: Microarrays were scanned at 5 mm resolution and variable PMT voltage to abtain maximal signal intensities with <0.1% probe saturation, a count ratio of 0.8-1.2 (Cy5/Cy3) and maximal congruence of histogram curves, using a GenePix 4000B microarray scanner.

data_processing: Dta summaries on the spot intensities are provided by GenePix Pro, Version 5.1, using local background correction. This includes the mean and the median pixel intensities at each scan wavelength for both feature and background pixels.

source_name: Total RNA from siGFP transfected HeLa cells labeled with Cyanine-3

label_protocol: Samples were labeled with fluorochrome by the Klenow fragment using the Bioprime Kit (Invitrogen, Karlsruhe, Germany).

growth_protocol: Cells were grown in an incubator with 95% relative air humidity and 5% CO2 in the adequate medium supplemented with 10% FCS, L-glutamine and antibiotics.

extract_protocol: Total RNA extracted using Trizol following manufacturer's instructions

treatment_protocol: Cells were transfected using the transfection reagent Effentene according to the manufacturer's protocoll.

tag: HeLa cell line (CCL-2)

source_name: Total RNA from mock transfected HeLa cells labeled with Cyanine-5


title: transfected HeLa cells; siGFP versus mock 2

description: Experiment 2


title: transfected HEK cells; mock versus siGFP 1a

description: Colour-switch 1a

source_name: Total RNA from mock transfected HEK cells labeled with Cyanine-3

tag: HEK cell line (CRL-1573)

source_name: Total RNA from siGFP transfected HEK cells labeled with Cyanine-5


title: transfected HEK cells; mock versus siGFP 1

description: Colour-switch 1


title: P3_S_HeLa_sRW_C88_T1

description: See publication for details

tag: See publication for details


title: P3_S_HepG2_sRW_C75_T1

description: See publication for details


title: transfected HeLa cells; mock versus siGFP 2a

description: Colour-switch 2a

source_name: Total RNA from mock transfected HeLa cells labeled with Cyanine-3

source_name: Total RNA from siGFP transfected HeLa cells labeled with Cyanine-5


title: P3_S_HeLa_sRW_C77_T1

description: See publication for details


title: transfected HeLa cells; mock versus siGFP 2

description: Colour-switch 2


title: Wild type male, E15 bladder (WTM_525)

city: Columbus

first: Kirk

last: McHugh

state: OH

address: 700 Children's Drive

department: Cell & Developmental Biology

institute: Columbus Children's Hospital

hyb_protocol: For expression profiling, 25 ng total RNA per sample was processed using isothermal amplification SPIA Biotin System (NuGEN Technologies, Inc.) and 2.2 µg of cDNA was hybridized per microarray. The microarrays utilized were both obtained from Affymetrix (Santa Clara, CA) and included either the Mouse Genome 430 2.0 GeneChip or the Custom GeneChip Chr11/16. After 16 hours of hybridization at 45ºC, washing and staining of microarrays was performed using a Fluidics Station 450 (Affymetrix, Santa Clara, CA). GeneChips were scanned in a GeneChip Scanner 3000 (Affymetrix, Santa Clara, CA). All steps of sample and microarray processing were performed according to manufacturer's recommendations (Affymetrix, Santa Clara, CA).

scan_protocol: standard Affymetrix protocol

data_processing: Probe set expression estimates were calculated using WPP algorithm (Auer et al, 2007).

label_protocol: standard Affymetrix protocol

extract_protocol: Whole embryos were harvested from timed pregnant females at embryonic day 15 (E15). Embryonic bladders were micro-dissected, snap frozen in liquid nitrogen, and stored at -80C.  Bladder tissues were homogenized using a Kontes-pellet pestle (Fisher). High quality total cellular RNA was isolated using the Absolutely RNA Miniprep kit according to the manufacture’s protocol (Stratagene, La Jolla, CA). Sample concentration and purity was determined by optical density 260/280 nm ratios using standard procedures. RNA samples were stored at -80C until further use. Total cellular RNA was extracted from whole embryos by homogenizing samples in TRIzol Reagent as outlined by the supplier (Invitrogen, Carlsbad, CA).

tag: wild type male, E15 bladder


title: Wild type male, E15 bladder (WTM_465)


title: P3_S_HepG2_sRW_C77_T1

description: See publication for details


title: transfected HEK cells; siGFP versus mock 1

description: Experiment 1

source_name: Total RNA from siGFP transfected HEK cells labeled with Cyanine-3

source_name: Total RNA from mock transfected HEK cells labeled with Cyanine-5


title: P3_AS_HeLa_sRW_C88_T1

description: See publication for details


title: Wild type male, E15 bladder (WTM_437)


title: Wild type female, E15 bladder (WTF_528)

tag: wild type female, E15 bladder


title: Wild type female, E15 bladder (WTF_457)


title: mgb +/- male, E15 bladder (HTM_532)

tag: mgb +/- male, E15 bladder


title: P3_AS_HepG2_sRW_C77_T1

description: See publication for details


title: mgb +/- male, E15 bladder (HTM_459)


title: Wild type female, E15 bladder (WTF_462)


title: mgb +/- male, E15 bladder (HTM_441)


title: P3_AS_HepG2_sRW_C88_T1

description: See publication for details


title: mgb +/- male, E15 bladder (HTM_439)


title: mgb +/- female, E15 bladder (HTF_523)

tag: mgb +/- female, E15 bladder


title: mgb +/- female, E15 bladder (HTF_464)


title: P3_S_HepG2_sRW_C89_T1

description: See publication for details


title: mgb +/- female, E15 bladder (HTF_458)


title: mgb -/- male, E15 bladder (HMM_524)

tag: mgb -/- male, E15 bladder


title: mgb -/- male, E15 bladder (HMM_533)


title: mgb -/- male, E15 bladder (HMM_434)


title: mgb -/- female, E15 bladder (HMF_535)

tag: mgb -/- female, E15 bladder


title: Bee_worker/drone_antennae_rep4

city: Urbana

first: Hugh

last: Robertson

state: IL

address: 505 S. Goodwin Ave.

department: Entomology

institute: University of Illinois at Urbana-Champaign

hyb_protocol: Microarray slides were prepared for prehybridization by passing them array-side down twice over steam from a beaker of boiling water and dried briefly on a warm adjustable-temperature block. The oligonucleotides were cross-linked to the array in a Stratalinker (Stratagene) set at 6000 ujoules (x 1000).  Excess oligonucleotides were removed by washing the microarray slide quickly in 0.2% SDS and agitating vigorously for 2 minutes, then washing 2X in molecular biology-grade water (Eppendorf). Slides were transferred to Coplin jars containing prehybridization buffer (20% formamide, 5X Denhardt’s, 6X SSC, 0.1%SDS, 25ug/mL tRNA) in a 42°C waterbath for 1 hour with occasional shaking.  Slides were washed in molecular biology-grade water 5X, followed by one wash in isopropanol, and spun dry in a ChipMate (Tomy).  Slides were placed in Corning hybridization chambers with a LifterSlip (Eerie Scientific) positioned over the area of the spotted oligonucleotides. For each pair of Cy5/Cy3 probes to be hybridized together, one dried probe was suspended in 15 uL nuclease-free water and then transferred to the second probe for suspension.  The probes were heated to 95°C for 2-3 min, chilled on ice for 5 min and kept at room temperature.  To each labeled probe pair, 15 uL of pre-warmed (50°C) 2X hybridization buffer (50% formamide, 10X SSC, 0.2% SDS) was added and the entire volume pipetted under the Lifterslip edge.  A drop of hybridization solution was placed in each chamber well for humidity.  Chambers were incubated in a light-proof hybridization oven at 42°C overnight. Microarray slides were transferred to a Coplin jar containing Wash 1 (1X SSC, 0.2% SDS) at 42°C for 8 minutes with agitation.  The slides were agitated in Wash 2 (0.1X SSC, 0.2% SDS) for 8 min at room temperature followed by two successive washes in Wash 3 (0.1X SSC) for 8 min at room temperature. The slides were spun dry and stored in a dark container until scanning.

scan_protocol: Microarray slides were scanned on an Axon 4000B Scanner using GenePix v6.0 software.  Arrays were scanned using 33% power and manually-adjusted PMT values to achieve an overall count ratio of 1.0 from both channels.

data_processing: Analysis was performed using SAS software v.9.1 (SAS Institute Inc., Cary, North Carolina).  Spots that were flagged “bad” or “not found” in GenePix, as well as buffer-only and blank spots were removed.  Median background signal intensity values were subtracted from median foreground signal intensity values, negative values were set to 1.0.  A low-intensity filter was applied that removed spots where the background-subtracted intensity in both channels was less than the overall average array background intensity for each channel.  Additionally, any spot saturated (>65,535 pixels) in both channels was also excluded.  Background-subtracted intensity values were log2-transformed and then normalized within array using loess (local regression) transformation, a correction for intensity-dependent effects.  Normalization for common mean and variance between microarrays was achieved by scaling and centering the loess-transformed values.  Outliers were removed by excluding spots that were +/- 3 SD from the dataset mean. Using a mixed model ANOVA, gene-specific normalized observations were modeled as the fixed effect of sex and the random effects of biological replicate nested in sex and technical replicate nested in biological replicate.

organism: Apis mellifera

label_protocol: For each reverse transcription reaction, 20 ug of total RNA was suspended in 18.5 uL nuclease-free water containing 2 ug oligo-dT16 primer (Operon) and denatured for 10 min at 70°C. A cocktail containing 6.0 uL 5X first strand buffer (Invitrogen), 3.0 uL 0.1M DTT, 0.6 uL 50X amino-allyl dNTP mix (25mM each dATP, dCTP, dGTP, 15mM dTTP, 10mM amino-allyl-dUTP (Sigma)), 2.0 uL Superscript III (Invitrogen, 200U/uL), and 0.5 uL RNase inhibitor (Fisher, 40U/uL) was added to each reaction and incubated overnight at 46°C in a hybridization oven.  RNA was hydrolyzed by adding 10 uL 1M NaOH and 10 uL 0.5M EDTA with a 15 min incubation at 65°C followed by neutralized with 10 uL 1M HCl.  The 1st cDNA was purified using the Qiagen Qiaquick PCR purification kit, following manufacturer’s instructions with substitutions for the wash buffer (phosphate wash buffer: 5mM KPO4, pH 8.0, 80% EtOH) and elution buffer (phosphate elution buffer: 4mM KPO4, pH8.5).  1st cDNA was eluted from the column twice using 30 ul phosphate elution buffer and dried completely in a SpeedVac. Single-use CyDye label (Cy3, GE Healthcare) was suspended in 9 uL fresh 0.1M Na2CO3 pH 9.0 and transferred to the cDNA pellet for suspension. Labeling reactions were incubated 1 hour in complete darkness.  The pH of the labeling reaction was adjusted with 35 uL 3M NaOAc pH5.2.  The labeled-cDNA was purified using the Qiagen Qiaquick PCR purification kit following manufacturer’s instructions and eluted twice using 40ul elution buffer.  The entire sample was analyzed on a spectrophotometer at 260 nm, 320 nm, 550 nm, 650 nm, and 750 nm to calculate pmoles of cDNA synthesized and label incorporation.  Labeled probes were then dried to completion in a SpeedVac and stored at -20°C until hybridization.

extract_protocol: Total RNA was isolated using an RNeasy Mini Kit (Qiagen, Valencia, CA).

treatment_protocol: Whole antennae from drone bees collected at the hive entrance were dissected in pools of 50 sets of antennae.

tag: Bee_drone(male) antennae

label_protocol: For each reverse transcription reaction, 20 ug of total RNA was suspended in 18.5 uL nuclease-free water containing 2 ug oligo-dT16 primer (Operon) and denatured for 10 min at 70°C. A cocktail containing 6.0 uL 5X first strand buffer (Invitrogen), 3.0 uL 0.1M DTT, 0.6 uL 50X amino-allyl dNTP mix (25mM each dATP, dCTP, dGTP, 15mM dTTP, 10mM amino-allyl-dUTP (Sigma)), 2.0 uL Superscript III (Invitrogen, 200U/uL), and 0.5 uL RNase inhibitor (Fisher, 40U/uL) was added to each reaction and incubated overnight at 46°C in a hybridization oven.  RNA was hydrolyzed by adding 10 uL 1M NaOH and 10 uL 0.5M EDTA with a 15 min incubation at 65°C followed by neutralized with 10 uL 1M HCl.  The 1st cDNA was purified using the Qiagen Qiaquick PCR purification kit, following manufacturer’s instructions with substitutions for the wash buffer (phosphate wash buffer: 5mM KPO4, pH 8.0, 80% EtOH) and elution buffer (phosphate elution buffer: 4mM KPO4, pH8.5).  1st cDNA was eluted from the column twice using 30 ul phosphate elution buffer and dried completely in a SpeedVac. Single-use CyDye label (Cy5, GE Healthcare) was suspended in 9 uL fresh 0.1M Na2CO3 pH 9.0 and transferred to the cDNA pellet for suspension. Labeling reactions were incubated 1 hour in complete darkness.  The pH of the labeling reaction was adjusted with 35 uL 3M NaOAc pH5.2.  The labeled-cDNA was purified using the Qiagen Qiaquick PCR purification kit following manufacturer’s instructions and eluted twice using 40ul elution buffer.  The entire sample was analyzed on a spectrophotometer at 260 nm, 320 nm, 550 nm, 650 nm, and 750 nm to calculate pmoles of cDNA synthesized and label incorporation.  Labeled probes were then dried to completion in a SpeedVac and stored at -20°C until hybridization.

treatment_protocol: Whole antennae from worker bees collected at the hive entrance were dissected in pools of 50 sets of antennae.

tag: Bee_worker(female) antennae


title: mgb -/- female, E15 bladder (HMF_529)


title: mgb -/- male, E15 bladder (HMM_521)


title: P3_S_HeLa_sRW_C89_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C82_T1

description: See publication for details


title: mgb -/- female, E15 bladder (HMF_467)


title: mgb -/- female, E15 bladder (HMF_456)


title: P3_AS_HeLa_sRW_C89_T1

description: See publication for details


title: P3_AS_HepG2_sRW_C89_T1

description: See publication for details


title: mgb -/- female, E15 bladder (HMF_440)


title: P3_S_HeLa_sRW_C82_T1

description: See publication for details


title: mgb -/- female, E15 bladder (HMF_438)


title: Bee_worker/drone_antennae_rep3


title: P3_AS_HeLa_sRW_C77_T1

description: See publication for details


title: mgb -/- male, whole E15 embryo (HM407)

tag: mgb -/- male, whole E15 embryo


title: Wild type male, whole E15 embryo (WT410)

tag: wild type male, whole E15 embryo


title: P3_S_HepG2_sRW_C82_T1

description: See publication for details


title: mgb +/- female, whole E15 embryo (F382)

tag: mgb +/- female, whole E15 embryo


title: mgb -/- female, whole E15 embryo (F379)

tag: mgb -/- female, whole E15 embryo


title: P3_AS_HeLa_sRW_C82_T1

description: See publication for details


title: P3_S_HepG2_sRW_C73_T1

description: See publication for details


title: mgb -/- female, E15 bladder (HMF_436)


title: mgb +/- male, whole E15 embryo (HT409)

tag: mgb +/- male, whole E15 embryo


title: WT_vs_WT+perox

city: Wageningen

first: L.

middle: Mariela

last: Serrano

address: Nieuwe Kanaal 9A

country: Netherlands

institute: TI Food and Nutrition

hyb_protocol: Hybridization version 4.1 Agilent hybridization  protocol v 4.1

scan_protocol: Scanning protocol" Using the Scan Array  Express microarray  scanner. 1.Turn on the scanner and  the computer (in this order) 2.Login: scanner 3.Double click the  'ScanArray Express' icon 4.Switch on lasers 1 and 3  at least 15 minutes prior to  scanning arrays 5.Click the 'File' button This is to prevent you from  accidentally turning off the  laser after the 15 minutes  warming-up phase 6.Insert the slide with the  array-side up and the label  towards the outside. On  Agilent slides the array side  is the side  with the label that has the  word "Agilent" on it 7.Press 'Scan | Prescan' 8.Put resolution at 50  m,  select both labels used,  and set PMT values for  both channels at for  instance 40% (Agilent  arrays) 9.Press 'Start' 10.Press 'Palette,' 'Green'  as soon as projection of  the image has started (first  dye), select the red colour  as soon as  scanning of the second  layer (second dye) has  started 11.Adjust PMT values to  balance signals obtained  for both channels. If  necessary do a few  low-resolution scans to  obtain a  optimal balance 12.Click the 'Scan' button  again 13.Select frame for high  resolution scan, adjust  resolution (to 10  m) and  scan the array for both  dyes 14.Press 'File | Save' to  save the files, 'Save all'  saves both dye layers "Create a new folder for  each array "'Save all' saves the  signals from both layers in  individual files 15.Switch off the lasers

data_processing:
Data Processing 1.- Background correction  2.-Normalization lowess.
VALUE column reports log2 (test/ref) ratios.

organism: Lactiplantibacillus plantarum

label_protocol: cDNA synthesis and , Labeling Protocol 1.4  Annealing Annealing Mix 	x µl	 RNA preparation (=10  µg) 	y µl	 nuclease-free water 	1 µl	 Random Nonamers 	11 µl	 TOTAL  1.	mix Annealing  Mix in a 200 µl PCR  tube gently by pipetting  up and down 2.	incubate for  5’ 70°C 3.	cool at room  temperature for 10’  (annealing) 4.	spin down the  mixture to the bottom of  the tube 5.	place on ice  Reverse transcription  Reverse Transcription  Mix 	4 µl	5x  buffer 	2 µl	0.1  M DTT 	1 µl	 dNTP-mix 	1 µl	 AA-dUTP 	1 µl  	 TMIII (keep only briefly  outside  20°C) 	20 µl	 TOTAL  1.	Mix by very  gently by pipetting up  and down (vigorous  pipetting will denature  the enzyme!) 2.	Incubate for 3  hrs at 42°C (in  PCR-machine) 3.	Cool on ice  for immediate  purification or place at   20°C for storage  Degradation of mRNA 1.	Add 2 µl 2,5  M NaOH 2.	Mix (vortex)  and spin down 3.	Incubate for  15 min. at 37°C 4.	Add 10 µl 2 M  HEPES free acid 5.	Mix (vortex)  and spin down 6.	Ready for  purification or store at   20°C    Labelling of cDNA with  cyanine dyes  Purification of amino  allyl-modified cDNA Followed Ge Healthcare  kit "Amersham CyDye  (Cy3/Cy5)Reactive Dye  Protocols for Post  Labelling  Aminoallyl-cDNA and  Aminoallyl-aRNA" code  RPN5661  Labelling of amino  allyl-modified cDNA with  CyDye 1.	Add the  amino allyl modified  cDNA (in 0.1 M sodium  bicarbonate) directly into  one aliquot of CyDye  NHS Ester.  Resuspend the ester  completely by pipetting  several times and  transfer the solution to  an amber 1.5 ml tube 2.	Incubate at  room temperature, in the  dark for 60 to 90  minutes 3.	Add 15 µl 4 M  Hydroxylamine to each  coupling reaction 4.	Mix by stirring  and incubate at room  temperature, in the dark,  for 15 minutes 5.	Proceed  directly to purification of  CyDye-labelled cDNA  Followed Ge Healthcare kit "Amersham CyDye (Cy3/Cy5)Reactive Dye Protocols for Post Labelling Aminoallyl-cDNA and Aminoallyl-aRNA" code RPN5661

extract_protocol: Extraction RNA version ,1.3, Collecting cells Quenching Buffer 66.7 mM	HEPES (pH  6.5) 60%Methanol Extraction Mixture -500 µll  phenol/Chloroform -30 µl10% SDS -30 µl3 M NaAc (pH 5.2) -500 mg glass-beads  (75-150 µm) -400 µlTE buffer  Prepare in a screw-cap  tube  1.For each sample  prepare a tube with  Extraction Mixture 2.Harvest cells and  quench them  immediately in  Quenching Buffer. Add  4 volumes of Quenching  Buffer ( 40°C) to 1  volume of  cell culture 3.Mix and store the  sample at -40°C 4.Centrifuge cells at  -20  C at 9000 rpm for 10  minutes (Centrifuge  RC5B) 5.Discard the  supernatant; keep the  cell pellet cooled and  work quickly to prevent  condensation of water.  Transfer the pellet  with a pre-cooled  spatula to a screw-crap  tube with Extraction  Mixture 6.Mix the cell pellet  thoroughly with  Extraction Mixture by  shaking by hand 7.Freeze the tube  immediately in liquid  nitrogen 8.Break cells in the  Savant FastPrep  FP120. Three times 40  seconds at speed 4.0  (cool cells between  steps if necessary) 9.Centrifuge for 5  minutes at 4°C in an  Eppendorf centrifuge at  10.000 rpm 10.Transfer 500 µl of  the supernatant to a  new tube. Add 400 µl  chloroform (chloroform  should be cooled at  4°C) 11.Centrifuge for 2  minutes in an Eppendorf  centrifuge at full speed  (4°C) 5.Continue with the  "High Pure RNA  Isolation Kit" from  Roche (order # 1 828  665) 6.Incubate for 60  minutes with DnaseI,  elute with 50 µl elution  buffer 7.Store the RNA in at  least 2 aliquots, one of  20 µl (for concentration  and quality analysis and  labelling) and one  back-up  of 30 µl at  80°C

tag: Lactobacillus plantarum WCFS1

tag: Lactobacillus plantarum WCFS1+ 3.5 mM hydrogen peroxide


title: mutTRXB1+perox _vs_WT+perox

description: mutTRXB1+perox _vs_WT+perox

scan_protocol: Scanning protocol Using the Scan Array  Express microarray  scanner. 1.Turn on the scanner and  the computer (in this order) 2.Login: scanner 3.Double click the  'ScanArray Express' icon 4.Switch on lasers 1 and 3  at least 15 minutes prior to  scanning arrays 5.Click the 'File' button This is to prevent you from  accidentally turning off the  laser after the 15 minutes  warming-up phase 6.Insert the slide with the  array-side up and the label  towards the outside. On  Agilent slides the array side  is the side  with the label that has the  word "Agilent" on it 7.Press 'Scan | Prescan' 8.Put resolution at 50  m,  select both labels used,  and set PMT values for  both channels at for  instance 40% (Agilent  arrays) 9.Press 'Start' 10.Press 'Palette,' 'Green'  as soon as projection of  the image has started (first  dye), select the red colour  as soon as  scanning of the second  layer (second dye) has  started 11.Adjust PMT values to  balance signals obtained  for both channels. If  necessary do a few  low-resolution scans to  obtain a  optimal balance 12.Click the 'Scan' button  again 13.Select frame for high  resolution scan, adjust  resolution (to 10  m) and  scan the array for both  dyes 14.Press 'File | Save' to  save the files, 'Save all'  saves both dye layers "Create a new folder for  each array "'Save all' saves the  signals from both layers in  individual files 15.Switch off the lasers  and the scanner after us

tag: Lactobacillus plantarum NZ7608 + 3.5 mM hydrogen peroxide


title: mutTRXB1_vs_WT

hyb_protocol: Hybridization version 4.1 Agilent hybridization  protocol v 4.

scan_protocol: Scanning protocol" Using the Scan Array  Express microarray  scanner. 1.Turn on the scanner and  the computer (in this order) 2.Login: scanner 3.Double click the  'ScanArray Express' icon 4.Switch on lasers 1 and 3  at least 15 minutes prior to  scanning arrays 5.Click the 'File' button This is to prevent you from  accidentally turning off the  laser after the 15 minutes  warming-up phase 6.Insert the slide with the  array-side up and the label  towards the outside. On  Agilent slides the array side  is the side  with the label that has the  word "Agilent" on it 7.Press 'Scan | Prescan' 8.Put resolution at 50  m,  select both labels used,  and set PMT values for  both channels at for  instance 40% (Agilent  arrays) 9.Press 'Start' 10.Press 'Palette,' 'Green'  as soon as projection of  the image has started (first  dye), select the red colour  as soon as  scanning of the second  layer (second dye) has  started 11.Adjust PMT values to  balance signals obtained  for both channels. If  necessary do a few  low-resolution scans to  obtain a  optimal balance 12.Click the 'Scan' button  again 13.Select frame for high  resolution scan, adjust  resolution (to 10  m) and  scan the array for both  dyes 14.Press 'File | Save' to  save the files, 'Save all'  saves both dye layers "Create a new folder for  each array "'Save all' saves the  signals from both layers in  individual files 15.Switch off the lasers  and the scanner after us

label_protocol: cDNA synthesis and , Labeling Protocol 1.4"  Annealing Annealing Mix 	x µl	 RNA preparation (=10  µg) 	y µl	 nuclease-free water 	1 µl	 Random Nonamers 	11 µl	 TOTAL  1.	mix Annealing  Mix in a 200 µl PCR  tube gently by pipetting  up and down 2.	incubate for  5’ 70°C 3.	cool at room  temperature for 10’  (annealing) 4.	spin down the  mixture to the bottom of  the tube 5.	place on ice  Reverse transcription  Reverse Transcription  Mix 	4 µl	5x  buffer 	2 µl	0.1  M DTT 	1 µl	 dNTP-mix 	1 µl	 AA-dUTP 	1 µl  	 TMIII (keep only briefly  outside  20°C) 	20 µl	 TOTAL  1.	Mix by very  gently by pipetting up  and down (vigorous  pipetting will denature  the enzyme!) 2.	Incubate for 3  hrs at 42°C (in  PCR-machine) 3.	Cool on ice  for immediate  purification or place at   20°C for storage  Degradation of mRNA 1.	Add 2 µl 2,5  M NaOH 2.	Mix (vortex)  and spin down 3.	Incubate for  15 min. at 37°C 4.	Add 10 µl 2 M  HEPES free acid 5.	Mix (vortex)  and spin down 6.	Ready for  purification or store at   20°C    Labelling of cDNA with  cyanine dyes  Purification of amino  allyl-modified cDNA Followed Ge Healthcare  kit "Amersham CyDye  (Cy3/Cy5)Reactive Dye  Protocols for Post  Labelling  Aminoallyl-cDNA and  Aminoallyl-aRNA" code  RPN5661  Labelling of amino  allyl-modified cDNA with  CyDye 1.	Add the  amino allyl modified  cDNA (in 0.1 M sodium  bicarbonate) directly into  one aliquot of CyDye  NHS Ester.  Resuspend the ester  completely by pipetting  several times and  transfer the solution to  an amber 1.5 ml tube 2.	Incubate at  room temperature, in the  dark for 60 to 90  minutes 3.	Add 15 µl 4 M  Hydroxylamine to each  coupling reaction 4.	Mix by stirring  and incubate at room  temperature, in the dark,  for 15 minutes 5.	Proceed  directly to purification of  CyDye-labelled cDNA  Followed Ge Healthcare kit "Amersham CyDye (Cy3/Cy5)Reactive Dye Protocols for Post Labelling Aminoallyl-cDNA and Aminoallyl-aRNA" code RPN5661

extract_protocol: Extraction RNA version ,1.3, Collecting cells" Quenching Buffer 66.7 mM	HEPES (pH  6.5) 60%Methanol Extraction Mixture -500 µll  phenol/Chloroform -30 µl10% SDS -30 µl3 M NaAc (pH 5.2) -500 mg glass-beads  (75-150 µm) -400 µlTE buffer  Prepare in a screw-cap  tube  1.For each sample  prepare a tube with  Extraction Mixture 2.Harvest cells and  quench them  immediately in  Quenching Buffer. Add  4 volumes of Quenching  Buffer ( 40°C) to 1  volume of  cell culture 3.Mix and store the  sample at -40°C 4.Centrifuge cells at  -20  C at 9000 rpm for 10  minutes (Centrifuge  RC5B) 5.Discard the  supernatant; keep the  cell pellet cooled and  work quickly to prevent  condensation of water.  Transfer the pellet  with a pre-cooled  spatula to a screw-crap  tube with Extraction  Mixture 6.Mix the cell pellet  thoroughly with  Extraction Mixture by  shaking by hand 7.Freeze the tube  immediately in liquid  nitrogen 8.Break cells in the  Savant FastPrep  FP120. Three times 40  seconds at speed 4.0  (cool cells between  steps if necessary) 9.Centrifuge for 5  minutes at 4°C in an  Eppendorf centrifuge at  10.000 rpm 10.Transfer 500 µl of  the supernatant to a  new tube. Add 400 µl  chloroform (chloroform  should be cooled at  4°C) 11.Centrifuge for 2  minutes in an Eppendorf  centrifuge at full speed  (4°C) 5.Continue with the  "High Pure RNA  Isolation Kit" from  Roche (order # 1 828  665) 6.Incubate for 60  minutes with DnaseI,  elute with 50 µl elution  buffer 7.Store the RNA in at  least 2 aliquots, one of  20 µl (for concentration  and quality analysis and  labelling) and one  back-up  of 30 µl at  80°C

tag: Lactobacillus plantarum NZ7608


title: P3_AS_HepG2_sRW_C84_T1

description: See publication for details

tag: See publication for details


title: WT+perox_vs_mutTRXB1+perox

hyb_protocol: Hybridization version 4.1" Agilent hybridization  protocol v 4.


title: 13281282 - B16-24h-mutant vs B11-24h-WT

city: Evry

first: Frederique

last: Bitton

address: 2 rue Gaston Cremieux

country: France

institute: INRA

description: Identification of Arabidopsis genes regulataed by Erwinia amylovora and of a subset of Arabiddopsis genes regulated by the type three secretion system of Erwinia amylovora

hyb_protocol: B11-24h-WT Cy5 / B16-24h-mutant Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.

scan_protocol: GenePix Pro 3.0, Cy3:pmt voltage 532nm,650V,laser power 100%, Cy5:635nm,pmt voltage 620V,laser power 100%

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

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

growth_protocol: leaf - media	soil	 hygrometry	70%	 temperature	21degreeC	 light	8h light(900 lux)/16h dark

treatment_protocol: Name:Ea-WT-24h - biotic stress - biotic stress,erwinia amylovora:time 24hour . infiltration of Arabidopsis leaves with water; harvest 24 hours post infiltration this is the water control : effect of infiltration

tag: Arabidopsis thaliana (columbia)  - dev.stage (Boyes et al. Plant Cell 2001):boyes

treatment_protocol: Name:Ea-mutant-24h - biotic stress - biotic stress,erwinia amylovora:time 24hour . infiltration of Arabidopsis leaves with Erwinia amylovora TTSS mutant CFBP6023; harvest 24 hours post infiltration


title: 13281281 - B06-24h-water vs B11-24h-WT

hyb_protocol: B11-24h-WT Cy5 / B06-24h-water Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.

treatment_protocol: Name:Water-24h - abiotic stress - abiotic stress,water:time 6hour . infiltration of Arabidopsis leaves with water; harvest 24 hours post infiltration this is the water control : effect of infiltration


title: Wild type female, whole E15 embryo (F384)

tag: wild type female, whole E15 embryo


title: 13281280 - B11-24h-WT vs B06-24h-water

hyb_protocol: B06-24h-water Cy5 / B11-24h-WT Cy3 : 30pmol. Hybridization Protocol: CATMA slides (Corning Microarray Technology, CORNING) are pretreated in the prehybridisation solution (1 % BSA, 0.1 SDS, 5X SSC ) at 42°C for 60 min. They are dipped a couple of times in distilled water at room temperature, then in isopropanol and dried immediately by compressed nitrogen stream. Slides were placed in Corning hybridization chambers with a 25x60 lifterslip and 10ul of distilled water for each groove. The target was diluted to a final volume of 60 µL as follows 15µl of purified, labeled cDNA, 15 µl of 4X Hybridization Buffer ( 20X SDS, 0.4 % SDS), 30 µL formamide. The target mixture is heated for 3 min at 95°C, put on ice for 30 sec and centrifuged to remove dust for 1 min. The target mixture was put on the chip as quickly as possible. The microarray is sealed in a chamber and submerged in a 42°C water bath for approximately 16 h. The microarray is washed for 4 min in 1xSSC, 0.2% SDS (42°C); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC, 0.2% SDS (RT); 4 min in 0.1x SSC (RT); dipped a few times in distilled water and dried immediately by compressed nitrogen stream.