title: Jurkat +Dox rep6

description: Jurkat Dox+6

tag: protocol


title: cleavage_45min_mixed_rep2

status: Public on Dec 14 2010

type: RNA

city: Zagreb

first: Tomislav

last: Domazet-Loso

address: Bijenicka cesta 54

department: Division of Molecluar Biology

country: Croatia

institute: Ruder Boskovic Institute

description:
Agilent Feature Extraction file:
US84303580_251916110393_S01_GE1_105_Dec08_1_2.txt.
Gene expression in wild type zebrafish cleavage, 45min, mixed sex, biological replicate 2.

hyb_protocol: 1650 ng of Cy3-labelled cRNA (specific activity >9.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 55 ul  following the manufacturers instructions. On completion of the fragmentation reaction, 55 ul of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Zebrafish (V2) Gene Expression Microarray slides (4x44K) (G2519F) for 17 hours at 65°C in a rotating Agilent hybridization oven.  After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 34°C GE Wash buffer 2 (Agilent), then dried 1 minute at room temperature.

scan_protocol: Slides were scanned immediately after washing and drying on the Agilent High-Resolution Microarray Scanner (G2505C) using one color scan setting for 4x44k array slides (Scan resolution 5um, Dye channel is set to Green and Green PMT is set to 100%).

data_processing: The scanned images were analyzed with Feature Extraction 10.7 Image Analysis Software (Agilent) using default parameters (protocol GE1_105_Dec08 and Grid: 019161_D_F_20080627) to obtain background subtracted and spatially detrended Processed Signal intensities.

molecule: total RNA

organism: Danio rerio

source_name: cleavage, 45min, mixed sex, replicate 2

label_protocol: Cyanine-3 (Cy3) labeled cRNA was prepared from 400 ng RNA using the Agilent one-color Quick Amp Labeling Kit protocol according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA).  Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.

growth_protocol: Zebrafish (Danio rerio) were kept in 12L flow-through tanks at 26.5 °C (around 60 animals per tank). For accurate staging, fertilized eggs were collected within 15 min intervals and incubated in Petri dishes at 28.5°C with water changes every 2-6 hours. After hatching, larvae were transferred to 1L tanks and kept at 28.5 °C.

extract_protocol: Frozen tissues were homogenized in TRIZOL (Invitrogen) by shaking samples 4 min together with a 5mm steel bead in a bead mill (TissueLyser II, Qiagen) at 30 Hz (embryos and smaller larve) or by disruption (larger larvae, juveniles and adults) using rotor-stator homogenizer (Tissue Ruptor, Qiagen). Total RNA was isolated using TRIZOL plus protocol from Invitrogen and the quality and the yield was determined by BioAnalyzer 2100 (Agilent) and NanoDrop ND-1000 spectrophotometer (Thermo scientific)

treatment_protocol: Staging was done according to post-fertilization time. Embryos were additionally staged under the dissecting microscope according to ref. 1 to check for the consistency of post-fertilization timing and morphological development at standard temperature (28.5 °C). Only healthy animals that showed the expected morphological features for a given post-fertilization time were sampled. Each sample contained around 50 individuals until the 1 day and 3 hour embryo stage, 15 individuals until the 10 day larval stage, 10 individuals until the 18 day larval stage, 5 individuals until the 45 day juvenile stage, while in later juvenile and adult stages we sampled males and females separately and each sample contained 2 individuals. All samples were snap frozen in liquid nitrogen and stored at -80 °C until RNA extraction. To avoid severe biases due to the excess of unfertilized eggs, we squeezed eggs from adult females before freezing them in liquid nitrogen. 1. (Kimmel CB, Ballard WW, Kimmel SR, Ullmann B, Schilling TF. Stages of embryonic development of the zebrafish. Dev Dyn. 1995 Jul;203(3):253-310.)

tag: strain

value: wild type

tag: developmental stage

value: cleavage

tag: developmental timing

tag: gender

value: mixed

tag: number of individuals per sample


title: human erythroid progenitors scramble replicate 1

city: Ottawa

first: Theodore

last: Perkins

state: ON

address: 501 Smyth Rd.

department: Cellular and Molecular Medicine

country: Canada

institute: Ottawa Hospital Research Institute

description: pro-erythroblasts scr 1

hyb_protocol: Affymetrix standard protocol

scan_protocol: Affymetrix standard protocol

data_processing: The data were analyzed with RMA as background correction, summarization & normalization method.

label: biotin

organism: Homo sapiens

source_name: peripheral blood, human erythroid progenitors scramble

label_protocol: Whole transcript sense target labeling assay

growth_protocol: CD34+ cells were cultured in a modified serum-free medium (M-C Giarratana et al., Nature Biotechnology, 2005) with the modification of using 20% BIT (StemCell Technologies, Canada) which contains pre-tested batches of 50 mg/ml bovine serum albumin, 50 mg/ml rh insulin and 1 mg/ml human transferrin (iron-saturated) in s MDM instead of separate components. The amplification procedure was based on a four-step expansion culture of CD34+ cells, in IMDM (Invitrogen) media supplemented with 20% BIT, 4 mM L-glutamine (Gibco), 1% (v/v) penicilin/streptomycin, 40 mg/ml inositol, 10 mg/ml folic acid, 0.16 mM monothioglycerol, 900 ng/ml ferrous sulfate and 90 ng/ml ferric nitrate, by sequential supply of the culture with hydrocortisone (all from Sigma-Aldrich) and specific combinations of cytokines. In the first step (days 0-8), 5 x 104/ml CD34+ cells were cultured in the presence of 10-6 M hydrocortisone, 100 ng/ml stem cell factor (SCF), 3 IU/ml erythropoietin (EPO) and 5 ng/ml interleukin-3 (IL-3) (R&D Systems, Cedarlane Laboratories, Hornby, Ontario, Canada). On day 4, cells were diluted at 4 x 104/ml in fresh medium containing hydrocortisone and the same cytokines. On day 8, cells were washed and resuspended at a final concentration of 2x105 cells/ml in fresh medium containing only SCF and EPO. In the second step (days 11-15) cells were harvested, washed to remove growth factors and metabolites and resuspended at a final concentration of 2x105 cells/ml and co-cultured on mouse, bone marrow stromal cells (MS-5) (HyperCLDB, depositor : Prof. Dr. W. Ostertag, Heinrich-Pette-Institut, Hamburg, Germany) in fresh medium supplemented with erythropoietin. In the third step (day 15-18) all non adherent cells were harvested, washed to remove the residual Epo and cultured on MS-5 stromal layer in fresh medium without cytokines. In the fourth step (day 18-26) cells were resuspended at 5-7x106/ ml in a fresh media supplemented with 10% FBS and transferred in a new stromal layer to allow a better preservation of the cultured of red blood cells.

extract_protocol: RNeasy Mini Kit (Qiagen)

value: control [scramble (scr) shRNA]


title: human erythroid progenitors TAL1 knock down replicate 1

description: TAL1 KD 1

source_name: peripheral blood, human erythroid progenitors TAL1 knock down

value: TAL1 knockdown


title: egg_0min_mixed_rep2

description:
Agilent Feature Extraction file:
US84303580_251916110409_S01_GE1_105_Dec08_1_4.txt.
Gene expression in wild type zebrafish unfertilised egg, 0min, mixed sex, biological replicate 2.

source_name: unfertilised egg, 0min, mixed sex, replicate 2

value: unfertilised egg


title: Silver_ion_treatment_1a_1/2h

city: Bangalore

first: Genotypic

last: technology

state: Karnataka

address: 259, Apoorva 4th cross,80 feet Road,RMV 2ND STAGE

country: India

institute: Genotypic Technology

description: Gene expression of B. cereus with silver ion treatment 30 mins

hyb_protocol: 600 ng of cy3 labeled samples were fragmented and hybridized.  Fragmentation of labeled cRNA and hybridization were done using the Gene Expression Hybridization kit of Agilent (Part Number 5188-5242). Hybridization was carried out in Agilent’s Surehyb Chambers at 65º C for 16 hours. The hybridized slides were washed using Agilent Gene Expression wash buffers (Part No: 5188-5327)

scan_protocol: Scanned using the Agilent Microarray Scanner G Model G2565BA at 5 micron resolution.

data_processing: The normalization was done using GeneSpring GX 11 Software. Recommended Percentile Shift Normalization.

organism: Bacillus cereus

label_protocol: Poly (A)-tails were added to the 3’-end of RNA by using A-plus Poly (A) polymerase tailing kit (Epicentre Biotechnologies).Then  the samples  were labeled using Agilent Quick Amp Kit PLUS (Part number: 5190-0442).  Five hundred nanograms each of the samples were incubated with reverse trancription mix at 42°C and converted to double stranded cDNA primed by oligodT with a T7 polymerase promoter. The cleaned up double stranded cDNA were used as template for aRNA generation. cRNA was generated by in vitro transcription and the dye Cy3 CTP(Agilent) was incorporated during this step. The cDNA synthesis and in vitro transcription steps were carried out at 40°C. Labeled cRNA was cleaned up and quality assessed for yields and specific activity

growth_protocol: Overnight culture was innoculated into the LB broth and incubated it 37°C with agitation of 200 rpm until the O.D reach upto 0.5

extract_protocol: RNA was prepared using (QIAGEN MINI Rneays kit) followed according to the manufacturer's instructions

treatment_protocol: Exponentially grown bacterial cells were treated with 1 mM silver nitrate and incubated it for two different time points 30 and 60 mins at 37°C with agitation of 200 rpm.

tag: class

value: Gram-positive bacteria


title: Jurkat +Dox rep4

description: Jurkat Dox+4

source_name: Jurkat lines expressing a Dox-inducible shRNA against TAL1 with added Dox

growth_protocol: Growth medium: RPMI 1640 supplemented with 10% (v/v) fetal bovine serum, 100 units/ml of penicillin and 100 mg/ml of streptomycin, blasticidin and G418 at 1 mg/ml.


title: Uveal Melanoma MU3

city: Paris Cedex 05

first: Cecile

last: Laurent

address: 26 rue d'ulm

country: France

institute: Institut Curie

description: Gene expression data from Uveal Melanoma

hyb_protocol: Following fragmentation, 10 ug of labelled cRNA were hybridized for 16 hr on GeneChip Human Genome Array. GeneChips were washed and stained in the Affymetrix Fluidics Station FS450.

scan_protocol: GeneChips were scanned using Affymetrix GeneChip GCS3000 scanner.

data_processing: Raw data were controlled using Genechip Operating Software and Affymetrix MAS5 algorithm to control the quality of hybridization and synthesis steps. Raw data were analysed using R/Bioconductor and normalized by GC-RMA method.

label: Biotin

source_name: Eye

label_protocol: Biotinylated cRNA were prepared according to the Affymetrix two round amplification  protocol from 100ng total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).

extract_protocol: Trizol extraction of total RNA was performed according to the manufacturer's instructions. A cleanup with RNAII Nucleospin was conducted according Macherey Nagel recommendations

tag: tissue

value: Uveal Melanoma (eye)

tag: age

value: male

tag: eye

value: left

tag: tumor location

value: on equator

tag: tumor diameter (mm)

tag: tumor thickness (mm)

tag: tumor cell type

value: epithelioid

tag: retinal detachment

tag: extrascleral extension

tag: chromosome 3 status

tag: months to endpoint

tag: metastasis

value: yes


title: Uveal Melanoma MU9

value: female

value: no

value: disomy


title: Jurkat_T_Cells_stimulated_12H_Pol-II_rep1_slide1of2

type: genomic

city: Forest Hill

first: Russell

middle: Leigh

last: McInnes

state: Vic

address: 347 Burwood Hwy

department: LSCA

country: Australia

institute: Agilent Technologies

description: Biological replicate 1 of 2 on the first of the two slides that comprise a single 2-colour ChIP-chip experiment

hyb_protocol: Labelling, hybridization and scanning were performed as described in the Mammalian ChIP-on-chip protocol (version 9.1; Agilent Technologies)

scan_protocol: Scanned on Agilent G2565BA Scanner

data_processing: Scanned images were quantified with Agilent Feature Extraction software under the default chip-chip protocol (version 9.5.3).

molecule: genomic DNA

source_name: Total IP 12H

growth_protocol: Human Jurkat T cells were cultured in RPMI 1640 medium (Gibco) supplemented with 10% FCS, 2mM L-glutamine, 9 mM HEPES and antibiotics. Jurkat T cells were maintained in suspension at a density of 5 x 10^5 cells/ml. Cells were stimulated with 20 ng/ml of phorbol 12-myristate 13-acetate (P) (Boehringer Mannheim) and 1 µM Ca 2+ ionophore A23187 (I) (Sigma-Aldrich) for the times indicated.

extract_protocol: Pooled ChIP samples were subsequently amplified based on one round of the whole genome amplification method using the WGA2 kit (Sigma-Aldrich), as described previously (O'Geen, Hollenhorst, Dindot)

treatment_protocol: ChIP assays were performed following the protocol supplied by Upstate Biotechnology, as previously detailed (MCB 2009).

tag: cell line

value: Jurkat T Cells - stimulated

value: 12 hours

source_name: Pol-II 12H

tag: antibody

value: anti-Pol II (8WG16)

tag: sample type

value: DNA pooled from five independent ChIP assay experiments


title: egg_0min_mixed_rep1

description:
Agilent Feature Extraction file:
US84303580_251916110387_S01_GE1_105_Dec08_1_4.txt.
Gene expression in wild type zebrafish unfertilised egg, 0min, mixed sex, biological replicate 1.

source_name: unfertilised egg, 0min, mixed sex, replicate 1

tag: strain

value: wild type

tag: developmental stage

tag: developmental timing

tag: gender

value: mixed

tag: number of individuals per sample


title: Jurkat rep4

description: Jurkat 4

source_name: Jurkat lines expressing a Dox-inducible shRNA against TAL1 without Dox addition

tag: protocol

value: control (no Dox)


title: No_silver_ion__2a_1h

description: Gene expression of B. cereus without silver ion treatment 60 mins

tag: class

value: Gram-positive bacteria


title: No_silver_ion__1a_1/2h

description: Gene expression of B. cereus without silver ion treatment 30 mins


title: Creb-PomcCre-/-, biological rep2

city: NEW YORK

first: franck

last: oury

state: NY

address: HHSC 701W 168th street

country: USA

institute: Columbia University

description:
Creb-pomcCre-/-2_YW_(mouse430_2)
Creb-pomcCre-/- hypothalamus.

hyb_protocol: Samples were fragmented and hybridized to the array for 16h at 45oC in a rotating hybridization oven (60rpm) using the Affymetrix Eukaryotic Target Hybridization Controls and protocol. Slides were stained and washed as indicated in the Antibody Amplification Stain for Eukaryotic Targets protocol using the Affymetrix Fluidics Station FS450.  Affymetrix fluidics protocol, FS450_0001.

scan_protocol: Affymetrix GeneChIP Scanner 3000 7G.

data_processing: The resulting data were processed using R (version 2.10.0) and Bioconductor packages. The Bioconductor package affy was used for pre-processing and RMA normalization.  P-values and foldchanges were determined within R after non-specific filtering with the Bioconductor package genefilter.

organism: Mus musculus

source_name: Creb-PomcCre-/-

label_protocol: Starting with 100ng of total RNA, biotin-labeled cRNA was produced using the Affymetrix Express One-Cycle Target labeling kit according to the manufacturer's protocol.

growth_protocol: Mouse hypothalami were dissected from 3-month-old mice.

extract_protocol: Total RNA was extracted with Trizol followed by cleanup with miRNeasy(Qiagen) columns.

tag: genotype

value: Creb-PomcCre-/-

tag: age

value: 3 months

tag: tissue

value: hypothalamus


title: Creb-PomcCre-/-, biological rep3

description:
Creb-pomcCre-/-3_YW_(mouse430_2)
Creb-pomcCre-/- hypothalamus.


title: Jurkat +Dox rep1

description: Jurkat Dox+1

value: TAL1 knockdown


title: Wild type, biological rep2

description:
WT2_YW_(mouse430_2)
Wild type hypothalamus.

source_name: wild type


title: Creb-PomcCre-/-, biological rep1

description:
Creb-pomcCre-/-1_YW_(mouse430_2)
Creb-pomcCre-/- hypothalamus.


title: Jurkat rep1

description: Jurkat 1


title: Uveal Melanoma MU1

value: Uveal Melanoma (eye)

tag: eye

value: right

tag: tumor location

tag: tumor diameter (mm)

tag: tumor thickness (mm)

tag: tumor cell type

tag: retinal detachment

tag: extrascleral extension


title: Jurkat +Dox rep3

description: Jurkat Dox+3


title: Wild type, biological rep4

description:
WT4_YW_(mouse430_2)
Wild type hypothalamus.


title: Jurkat rep3

description: Jurkat 3


title: Wild type, biological rep3

description:
WT3_YW_(mouse430_2)
Wild type hypothalamus.


title: Jurkat +Dox rep2

description: Jurkat Dox+2


title: Jurkat rep2

description: Jurkat 2


title: Wild type, biological rep1

description:
WT1_YW_(mouse430_2)
Wild type hypothalamus.


title: Control Liver 29 (mRNA)

status: Public on Dec 13 2010

first: Andrew

last: Williams

address: 50 Columbine

institute: Health Canada

hyb_protocol: Agilent Mouse G4122F Microarrays (containing approximately 44,000 probes) were hybridized with 800 ng Cy5-labelled lung or liver RNA and 800 ng Cy3-labelled Universal Mouse Reference RNA (Stratagene, CA, USA), used as a common reference on all arrays.  Arrays were incubated overnight at 60 ºC in Agilent hybridization solution and washed according to manufacturer’s instructions.

scan_protocol: Arrays were scanned scanned on an Agilent G2505B scanner. Data were acquired using Agilent Feature Extraction software version 9.5.3.1.

data_processing: A reference design was used to analyse gene expression microarray data. The background fluorescence was measured using the ( - ) 3xSLv1 probes ; probes with median signal intensities less than the trimmed mean ( trim = 5% ) plus three trimmed standard deviations of the ( - ) 3xSLv1 probe were flagged as absent ( within the background signal ) . Data were normalized using lowess using the transform.madata function in the maanova library in R. ogRatio is the normalized log2 ratio ( sample/reference ) , RawCy3 is the raw median signal intensity for the reference, RawCy5 is the raw median signal intensity for the sample, Flag where 1 indicates if RawCy5 is below the mean plus 3 standard deviations of the negative control spots, NormCy3 is the lowess normalized data for the reference, and NormCy5 is the lowess normalized data for the reference.

source_name: Total RNA from mouse liver labeled with Cyanine-5 (red)

label_protocol: Individual 250 ng aliquots of RNA from each sample were amplified and labelled using the Low RNA Input Fluorescent Linear Amplification Kit (Agilent, (5184-3523)).

extract_protocol: RNA was isolated using the TRIzol reagent (Invitrogen Canada Inc., Burlington, ON, Canada), and further purified using RNeasy Mini Kits (Qiagen Inc., Mississauga, ON, Canada).  RNA was quantified using UV spectrophotometer, and quality was confirmed using the Agilent 2100 Bioanalyzer and RNA 6000 NanoLab Chip Kit (Agilent Technologies Canada Inc., Mississauga, ON, Canada).

value: part of liver

value: Female

value: equivalent to GD4

tag: strain

tag: date of exposure

tag: exposure

value: exposed to filtered air for 1 h/day for 11 consecutive days and were sacrificed 5 days following the last exposure.

tag: time post exposure

value: 5 days

tag: micro array id

tag: set

value: Hybridization Block 2

source_name: universal mouse reference total RNA (Catalog #740100; Stratagene, La Jolla, CA, USA)

extract_protocol:
purchased from Stratagene
Unknown: Material was purchased as total RNA (universal mouse reference total RNA (Stratagene, CA, USA))

tag: sample type

value: Total RNA derived from 11 mouse cell lines

tag: source

value: Stratagene, La Jolla, CA, USA


title: Control Liver 27 (mRNA)

tag: gender

tag: age


title: Caenorhabditis elegans L4

city: Toronto

first: Qun

last: Pan

address: 160 College St.

institute: University of Toronto

description: This Sample represents two hybridizations.

hyb_protocol: Mixed labeled cDNAs were added to hybridization buffer containing 1 M NaCl, 0.5% sodium sarcosine, 50 mM methyl ethane sulfonate (MES), pH 6.5, 33% formamide and 40 μg herring sperm DNA (Invitrogen, Carlsbad, USA). Hybridizations were carried out in a final volume of 0.5 ml injecting into an Agilent hybridization chamber at 42°C on a rotating platform in a hybridization oven (Robbins Scientific Corporation, Sunnyvale, USA) for 16–24 h. Slides were then washed (rocking for approximately 30 seconds in 6 × SSPE, 0.005% sarcosine, then rocking for approximately 30 seconds in 0.06 × SSPE) and scanned with a 4000A microarray scanner (Axon Instruments, Union City, USA).  Hybridizations were performed in duplicate or triplicate with fluor reversal: that is, each mRNA sample was examined 2-3 times, at least once in the Cy3 channel and at least once in the Cy5 channel, on separate arrays. Each array was hybridized with two samples simultaneously, each from a different developmental stage.

scan_protocol: Slides were scanned using an Agilent Microarray Scanner (Agilent, Palo Alto, USA).

data_processing: Images were processed by the Agilent Feature Extraction software (version 9.5). Variance stabilizing normalization (vsn) was performed using the vsn package within Bioconductor (version 2.1).

label: Cy5, Cy3

organism: Caenorhabditis elegans

source_name: C. elegans larval stage 4

label_protocol: The mRNA (1–2 μg) was reverse-transcribed with random nonamer primers (1 μg per reaction) and T18VN (0.25 μg per reaction) to synthesize cDNA. The reaction contained a 1:1 mixture of 5-(3-aminoallyl) thymidine 5'-triphosphate (Sigma, St. Louis, USA) and thymidine triphosphate (TTP) in place of TTP alone. The cDNA products were bound to QIAquick PCR Purification columns (Qiagen, Hilden, Germany) following the manufacturer's instructions, washed three times with 80% ethanol, and eluted with water. Purified cDNA was reacted with N-hydroxy succinimide esters of Cy3 or Cy5 (Amersham Pharmacia Biotech, Piscataway, USA) following the manufacturer's instructions.

extract_protocol: RNA was extracted using homogenization and Trizol reagent (Invitrogen, Carlsbad, USA) following the instructions from the manufacturer. Prior to cDNA synthesis, polyA+ RNA was purified from total RNA using the Nucleotrap mRNA Midi kit (Clontech) as recommended by the manufacturer with two rounds of purification.

value: N2 Bristol

tag: genotype

value: wild-type

tag: tissue

value: whole body

tag: developmental stage


title: Caenorhabditis elegans L1

source_name: C. elegans larval stage 1


title: Caenorhabditis elegans L2

source_name: C. elegans larval stage 2


title: Nano-titanium Liver 9 (mRNA)

value: Exposed to 40 mg nanoTiO2/m3 for one hour/day for 11 consecutive days and were sacrificed 5 days following the last exposure.


title: Control Liver 34 (mRNA)

value: Hybridization Block 3


title: Caenorhabditis elegans adult

description: This Sample represents three hybridizations.

source_name: C. elegans adult

value: adult


title: Control Liver 37 (mRNA)

value: Hybridization Block 4


title: Control Liver 26 (mRNA)

value: Hybridization Block 1


title: Nano-titanium Liver 20 (mRNA)


title: Nano-titanium Liver 1 (mRNA)


title: Nano-titanium Liver 12 (mRNA)


title: Control Liver 45 (mRNA)


title: Caenorhabditis elegans L3

source_name: C. elegans larval stage 3


title: Control Liver 24 (mRNA)


title: Nano-titanium Liver 13 (mRNA)


title: Control Liver 32 (mRNA)


title: Control Lung 34 (mRNA)

source_name: Total RNA from mouse lungs labeled with Cyanine-5 (red)

value: part of lung


title: Nano-titanium Liver 19 (mRNA)

value: part of liver


title: Control Lung 23 (miRNA)

hyb_protocol: The labelled RNA samples were hybridised to an individual array on 8 x 15K format Agilent mouse miRNA array slides. Hybridizations were performed in SureHyb chambers (Agilent) at 55ºC for 20 hours.

scan_protocol: Arrays were washed, scanned at a resolution of 5 m using an Agilent G2505B scanner and data were acquired using Agilent Feature Extraction software version 9.5.3.1.

data_processing: Non-background subtracted raw data were quantile normalized (Bolstad et al. 2003). Present calls were determined as signals that were greater than 3 trimmed SDs above the trimmed mean of the the (-)3xSLv1 probes on the array. Probes with technical replicates for a miRNA were averaged using the median signal intensity.

source_name: Total RNA enriched with small RNA species from mouse lungs labeled with Cyanine-5 (red)

label_protocol: Freshly isolated individual total lung RNA samples from 8 controls and 8 treated groups were labelled using the Agilent miRNA Complete Labelling and Hybridisation Kit (Agilent Technologies, Inc.,). Briefly, 100 ng of total RNA was dephosphorylated by incubation with calf intestinal phosphatase at 37ºC for 30 min, denatured using 100 % DMSO at 100ºC for 5 min, then labelled with pCp-Cy3 using T4 ligase by incubation at 16ºC for 1 hour.

extract_protocol: The mirVana miRNA Isolation Kit (Ambion, Streetsville, ON, Canada) was used to prepare total RNA enriched with small RNA species from randomly selected left lung sections. All RNA samples showing A260/280 ratios between 2.0 and 2.1 were further analysed by Agilent 2100 Bioanalyzer (Agilent Technologies Inc.,) to determine RNA integrity. Only high quality RNA (28S/18S > 1.8) was used for further analysis.

treatment_protocol: Briefly, fortyfive time-mated, nulliparous, young adult female mice at GD 3 were housed in cages under controlled environmental conditions with 12 h light–dark cycles. Food and tap water were provided ad libitum. Exposure was performed by whole-body inhalation to 42.4 ± 2.9 (SEM) mg nanoTiO2/m3 or to the filtered air for 1 h/day from GD 8–18.   Seventeen of the time-mated mice (“NP3 females” in Hougaard et. al, submitted) that had not conceived, as evidenced by lack of pups and absence of interuterine implantation sites were used in the present study as a group of adult females. Animals were sacrificed by cardiac puncture five days after the last exposure. The lungs and livers were dissected, weighed, and snap frozen. Samples were stored at -80°C until analysis. All procedures complied with EC Directive 86/609/EEC and Danish law regulating experiments on animals (permission 2006/561-1123).

value: female

value: 11-13 weeks


title: Control Lung 32 (mRNA)

value: Female

value: equivalent to GD4

value: exposed to filtered air for 1 h/day for 11 consecutive days and were sacrificed 5 days following the last exposure.


title: Nano-titanium Lung 19 (mRNA)

value: 5 days

value: Total RNA derived from 11 mouse cell lines


title: Nano-titanium Lung 23 (mRNA)

value: Stratagene, La Jolla, CA, USA


title: Nano-titanium Lung 13 (mRNA)


title: Control Lung 20 (miRNA)

description: Total RNA enriched with small RNA species

value: Hybridization Block 2


title: Nano-titanium Lung 20 (mRNA)


title: Control Lung 29 (mRNA)


title: Nano-titanium Liver 23 (mRNA)


title: Nano-titanium Lung 13 (miRNA)


title: Rhodopseudomonas palustris CGA010_5

city: Seattle

first: Yasuhiro

last: Oda

state: WA

address: Box 357242, 1705 N.E. Pacific Street, HSB K-354

department: Department of Microbiology

institute: University of Washington

description: A high-density oligonucleotide microarray (Affymetrix GeneChip) was custom designed and manufactured by Affymetrix based on the sequences of ORFs and intergenic regions. cDNA synthesis, fragmentation, labeling, hybridization, and processing of custom designed Rhodopseudomonas palustris GeneChips were carried out as recommended by the manufacturer. Experiments were performed in duplicate with independently prepared samples.

hyb_protocol: As recommended by the manufacturer (Affymetrix)

data_processing: The Affymetrix GeneChip Operating Software Version 1.4 was used for initial data acquisition and processing.

label: Biotinylated GeneChip reagent

organism: Rhodopseudomonas palustris

source_name: Photosynthetic bacterium Rhodopseudomonas palustris

growth_protocol: Cells were grown in PM, anaerobically, with light. Succinate 10mM and 5.7 mM p-coumarate were used as carbon sources.

extract_protocol: Rhodopseudomonas palustris cells were disrupted by treatment with by bead beating and RNA was purified with the RNeasy Mini kit (Qiagen) including DNase treatment on the column.

value: wild type


title: Rhodopseudomonas palustris CGA010_6

scan_protocol: As recommended by the manufacturer (Affymetrix)

growth_protocol: Cells were grown in PM, anaerobically, with 10mM succinate and p-coumarate as carbon sources.


title: No_silver_ion__1b_1/2h

source_name: B. cereus without silver ion treatment 30 mins

tag: class

value: Gram-positive bacteria


title: CGH: Wm82-ISU-01_v_Williams_08_16_10

city: Saint Paul

first: Robert

middle: M

last: Stupar

state: MN

address: 1991 Upper Buford Circle

department: Agronomy and Plant Genetics

institute: University of Minnesota

hyb_protocol: Genomic DNA were hybridized to the microarray according to the manufacturer instructions (NimbleGen CGH User Guide Ver 5.1)

scan_protocol: Slides were scanned on the GenePix4000B Scanner (Molecular Devices, Sunnyvale, CA) according to the array manufacturer instructions (NimbleGen CGH User Guide Ver 5.1)

data_processing: The segMNT algorithm in the NimbleScan software (verison 2.5) was used to extract the raw data and normalize into log2 ratio value according to the default settings.

organism: Glycine max

source_name: Wm82-ISU-01, leaf tissue

label_protocol: 1ug of genomic DNA was labeled and amplified using the NimbleGen Dual-Color Labeling Kit according to the NimbleGen CGH User Guide Ver 5.1.

extract_protocol: Qiagen DNeasy

value: single plant leaf tissue

value: Williams 82; received from Randy Shoemaker lab (USDA-ARS at Iowa State University)

source_name: Williams, leaf tissue

value: Williams


title: CGH: Wm82-PU-01_v_Williams_08_16_10

source_name: Wm82-PU-01, leaf tissue

value: Williams 82; received from Scott Jackson lab (Purdue University)


title: Control Lung 27 (mRNA)

value: part of lung


title: Control Lung 12 (miRNA)

value: Hybridization Block 1


title: CGH: Wm82-SGC-01_v_Williams_08_16_10

source_name: Wm82-SGC-01, leaf tissue

value: Williams 82; received from Soybean Germplasm Center (Urbana, IL)


title: Control Lung 10 (miRNA)

value: female

value: 11-13 weeks


title: Control Lung 26 (mRNA)

value: Female

value: equivalent to GD4


title: Nano-titanium Lung 9 (miRNA)

value: Exposed to 40 mg nanoTiO2/m3 for one hour/day for 11 consecutive days and were sacrificed 5 days following the last exposure.

value: 5 days


title: CGH: Wm82-MN-01_v_Williams_08_16_10

source_name: Wm82-MN-01, leaf tissue

value: Williams 82; received from James Orf lab (University of Minnesota)


title: Nano-titanium Lung 12 (mRNA)

value: Hybridization Block 2

value: Total RNA derived from 11 mouse cell lines

value: Stratagene, La Jolla, CA, USA


title: Control Lung 8 (miRNA)

value: exposed to filtered air for 1 h/day for 11 consecutive days and were sacrificed 5 days following the last exposure.


title: Rhodopseudomonas palustris CGA010_4

label_protocol: As recommended by the manufacturer (Affymetrix)

value: wild type


title: Rhodopseudomonas palustris aadR mutant_3

value: aadR mutant


title: Rhodopseudomonas palustris aadR mutant_2


title: Nano-titanium Lung 7 (miRNA)


title: Control Lung 6 (miRNA)


title: Nano-titanium Lung 7 (mRNA)


title: Nano-titanium Lung 3 (miRNA)


title: Nano-titanium Lung 17 (miRNA)


title: Nano-titanium Lung 18 (miRNA)


title: Nano-titanium Lung 1 (miRNA)


title: Rhodopseudomonas palustris aadR mutant_1

growth_protocol: Cells were grown in PM medium to mid-exponential phase (anaerobically with light). Succinate 10 mM + p-coumarate 5.7 mM were used as carbon sources.


title: Nano-titanium Lung 1 (mRNA)


title: CGH: Kingwa_v_Williams_08_16_10

source_name: Kingwa, leaf tissue

value: single plant leaf tissue

value: Kingwa

value: Williams


title: CGH: Wm82-ISU-01_v_Wm82-SGC-01_05_07_10

value: Williams 82; received from Soybean Germplasm Center (Urbana, IL)

value: Williams 82; received from Randy Shoemaker lab (USDA-ARS at Iowa State University)


title: Control Lung 24 (mRNA)

value: part of lung

value: Hybridization Block 1


title: genomic DNA from patient #CCF052

status: Public on Dec 11 2010

city: Cleveland

first: Anna

last: Jankowska

state: Ohio

address: 9500 Euclid Ave.

department: THOR

institute: Cleveland Clinic,Taussig Cancer Institute

description: 5hmC high (wt)

hyb_protocol: Bisulphite converted DNA was amplified, fragmented and hybridised to Illumina Infinium Human Methylation27 Beadchip using standard Illumina protocol

scan_protocol: Arrays were imaged using BeadArray Reader using standard recommended Illumina scanner setting

data_processing: BeadStudio software v3.2

label: Cy5 and Cy3

source_name: bone marrow

label_protocol: Standard Illumina Protocol

extract_protocol: Genomic DNA was extracted and purified from controls and patients samples using the ArchivePure DNA Blood kit (5Prime) as per the manufacturer's instructions

value: male

value: bone marrow

tag: disease state

value: MPN

tag: 5hmc levels

value: high

tag: tet2 genotype

value: wt


title: genomic DNA from patient #CCF070

description: 5hmC high (wt)

value: female

value: MDS/MPN


title: genomic DNA from patient #CCF053

description: 5hmC high (wt)


title: KM-H2 cells, CIITA-BX648577 knockdown, replicate 1

city: Vancouver

first: Christian

last: Steidl

state: BC

address: 675 West 10th Avenue

department: Pathology

institute: BC Cancer Agency

description:
Pleural effusion cells
Gene expression data of transduced KM-H2 cells

hyb_protocol: Following fragmentation, cRNA were hybridized O/N at 45C on Affymetrix GeneChip HG 2.0 Plus Arrays. GeneChips were washed and stained in the Affymetrix Fluidics Station 450.

scan_protocol: GeneChips were scanned using Affymetrix GeneChip Scanner

data_processing: The data were analyzed using DCHIP, http://biosun1.harvard.edu/complab/dchip/

source_name: Hodgkin lymphoma cell line KM-H2

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol (1-cycle)

growth_protocol: KMH2: Standard conditions (www.dsmz.de)

extract_protocol: Qiagen Allprep extraction columns

treatment_protocol: RNA interference (RNAi) with the CIITA-BX648577 fusion transcript was performed using lentiviral transduction of a vector (pGIPZ-sh.FLJ27352, clone V2LHS_212659, Thermo Scientific Open Biosystems, Huntsville, AL, USA) expressing a small hairpin RNA (shRNA: TGCTGTTGACAGTGAGCGCGACAGCCACCTCACTATCAAATAGTGAAGCCACAGATGTATTTGATAGTGAGGTGGCTGTCTTGCCTACTGCCTCGGA) that, after processing to mature siRNA, interferes with BX648577 exon 2 sequence as part of the CIITA-BX648577 fusion transcript.

tag: cell line

tag: treatment

value: CIITA-BX648577 knockdown


title: genomic DNA from patient #CCF033

description: 5hmC high (wt)

value: de novo AML


title: genomic DNA from patient #CCF029

description: 5hmC high (wt)


title: KM-H2 cells, Non-silencing control, replicate 1

value: Non-silencing control


title: KM-H2 cells, CIITA-BX648577 knockdown, replicate 2


title: genomic DNA from patient #CCF034

description: 5hmC high (wt)


title: genomic DNA from patient #CCF041

description: 5hmC high (wt)


title: genomic DNA from patient #CCF037

description: 5hmC high (wt)

value: MDS


title: MDA231-LM1-3461

status: Public on Dec 12 2010

city: Houston

first: Xiang

last: Zhang

state: TX

address: One Baylor Plaza, BCM600

department: Lester and Sue Smith Breast Center

institute: Baylor College of Medicine

description:
Agilent Feature Extraction file (original name):
XZ_3461_slide369_S01_CGH-v4_91.txt

hyb_protocol: Five micrograms of labelled tumor and reference samples were combined and hybridized onto the arrays according to Agilent's instructions.

scan_protocol: Arrays were scanned using G2565 scanners from Agilent following the manufacturer's instruction.

data_processing: Agilent feature extraction software (v 7.5.1) was used for spot quality filtering and normalization. The extracted intensity values were converted to log2 ratios with medians centered to 0.

source_name: Lung metastatic derivatives of MDA231

label_protocol: Five micrograms of digested and purified tumour and reference DNA was labelled with Cy5-dUTP and Cy3-dUTP, respectively, in a random priming reaction using BioPrime DNA Labelling System.

growth_protocol: DME 10% FCS

extract_protocol: Genomic DNA was extracted with Qiagen DNAeasy Blood and Tissue Kit following the manufacturer's protocol

value: MDA231 breast cancer cell line

tag: cell type

value: Lung metastatic derivatives

source_name: reference DNA

extract_protocol: Purchased reference.

tag: reference

value: Human female genomic DNA


title: genomic DNA from patient #CCF038

description: 5hmC high (wt)


title: KM-H2 cells, Non-silencing control, replicate 2


title: genomic DNA from patient #CCF026

description: 5hmC high (wt)


title: genomic DNA from patient #CCF036

description: 5hmC high (wt)


title: MDA231-LM2-4181

description:
Agilent Feature Extraction file (original name):
XZ_4180_slide366_S01_CGH-v4_91.txt


title: CN34-BrM-2A

description:
Agilent Feature Extraction file (original name):
XZ_Br2A_slide378_S01_CGH-v4_91.txt

source_name: Brain metastastatic derivatives of CN34

value: CN34 breast cancer cell line

value: Brain metastastatic derivatives


title: CN34-Parental

description:
Agilent Feature Extraction file (original name):
XZ_CN34_slide370_S01_CGH-v4_91.txt

source_name: Parental CN34 cells

value: Parental cells


title: genomic DNA from patient #CCF074

description: 5hmC high (wt)

value: male


title: CN34-BrM-2C

description:
Agilent Feature Extraction file (original name):
XZ_Br2C_slide379_S01_CGH-v4_91.txt


title: genomic DNA from patient #CCF028

description: 5hmC high (wt)

value: bone marrow


title: MDA231-LM2-4175

description:
Agilent Feature Extraction file (original name):
XZ_4175_slide359_S01_CGH-v4_91.txt


title: MDA231-Parental-rep-2

description:
Agilent Feature Extraction file (original name):
XZ_ATCCU_slide356_S01_CGH-v4_91.txt

source_name: Parental MDA231 cells


title: MDA231-LM2-4142

description:
Agilent Feature Extraction file (original name):
XZ_4142_slide357_S01_CGH-v4_91.txt


title: MDA231-LM2-4173

description:
Agilent Feature Extraction file (original name):
XZ_4173_slide358_S01_CGH-v4_91.txt


title: CN34-BrM-2E

description:
Agilent Feature Extraction file (original name):
XZ_Br2E_slide380_S01_CGH-v4_91.txt


title: MDA231-BrM-831

description:
Agilent Feature Extraction file (original name):
XZ_831_slide367_S01_CGH-v4_91.txt

source_name: Brain metastastatic derivatives of MDA231


title: MDA231-BrM-923

description:
Agilent Feature Extraction file (original name):
XZ_923_slide368_S01_CGH-v4_91.txt


title: Post-selection DP thymocytes Replicate 1

city: La Jolla

first: Steven

last: Head

state: CA

address: 10550 N Torrey Pines Rd

institute: The Scripps Research Institute

hyb_protocol: Following fragmentation, 10 ug of cRNA were hybridized for 16 hr at 45C on the scrGLYCO-v1GeneChip Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 400.

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

data_processing: The data were analyzed with Microarray Suite version 5.0 (MAS 5.0) using Affymetrix default analysis settings and global scaling as normalization method. The trimmed mean target intensity of each array was arbitrarily set to 100.

source_name: Post-selection (post-positive selection) DP thymocytes

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 6 ug total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).

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

tag: Sex

value: OT-I TCR transgenic

value: 8 weeks

value: healthy


title: Activated CD8 T cells Replicate 2

source_name: naïve OT-I T cells activated for 48 hours in vitro with cognate antigen (SIINFEKL peptide/Kb) (displayed on cell sized latex beads) in the presence of IL-2 and IL-12

value: female


title: Activated CD8 T cells Replicate 1


title: Pancreatic tumor cell line Suit-2, replicate C

source_name: Human pancreatic tumor cell line Suit-2

value: unknown

value: Human pancreatic tumor cell line Suit-2

value: Tumor


title: Pre-selection DP thymocytes Replicate 2

source_name: Highly purified pre-selection CD4+8+ thymocytes

value: TCRa-/-


title: Pancreatic tumor cell line Suit-2, replicate B


title: MDA231-Parental-rep-1

description:
Agilent Feature Extraction file (original name):
XZ_ATCC_slide355_S01_CGH-v4_91.txt


title: Naive CD8 T cells Replicate 1

source_name: Naïve (CD44lo) CD8 T cells from  lymph node


title: Pre-selection DP thymocytes Replicate 1


title: genomic DNA from patient #CCF031

description: 5hmC high (wt)

value: MDS/MPN

value: high

value: wt


title: genomic DNA from patient #CCF065

description: 5hmC high (wt)


title: genomic DNA from patient #CCF072

description: 5hmC high (wt)

value: sAML


title: Pancreatic tumor cell line ASPC-1, replicate C

source_name: Human pancreatic tumor cell line ASPC-1

value: Human pancreatic tumor cell line ASPC-1


title: genomic DNA from patient #CCF058

description: 5hmC high (mutant)

value: MDS

value: mutant


title: genomic DNA from patient #CCF030

description: 5hmC high (wt)


title: genomic DNA from patient #CCF027

description: 5hmC high (wt)

value: de novo AML


title: genomic DNA from patient #CCF051

description: 5hmC high (mutant)

value: bone marrow


title: genomic DNA from patient #CCF004

description: 5hmC low (mutant)

value: male

value: low


title: genomic DNA from patient #CCF032

description: 5hmC high (wt)


title: genomic DNA from patient #CCF040

description: 5hmC high (wt)


title: genomic DNA from patient #CCF059

description: 5hmC high (wt)


title: Pancreatic tumor cell line Suit-2, replicate A


title: genomic DNA from patient #CCF085

description: 5hmC low (mutant)

source_name: peripheral blood

value: peripheral blood


title: genomic DNA from patient #CCF019

description: 5hmC low (wt)

value: female


title: genomic DNA from patient #CCF073

description: 5hmC high (wt)


title: HS2ST (+/+) 3

description: P1 mouse brain wild type for Heparan sulfate 2-O sulfotransferase (HS2ST)

data_processing: The data were analyzed with RMA Express using the default settings (Background Adjust - Yes, Normalization - Quantile, Summarization Method - Median Polish, Store Residuals - unchecked).

value: unknown

value: WT

value: healthy

value: whole brain


title: Pancreatic tumor cell line ASPC-1, replicate A

value: Tumor


title: Pancreatic tumor cell line ASPC-1, replicate B


title: Naive CD8 T cells Replicate 2

value: OT-I TCR transgenic

value: 8 weeks


title: HS2ST (+/-) 2

description: P1 mouse brain heterozygous for Heparan sulfate 2-O sulfotransferase (HS2ST) loss of function mutation: Replicate 2

value: Heterozygous HS2ST loss of function mutation


title: genomic DNA from patient #CCF086

description: 5hmC low (mutant)

value: MDS/MPN


title: Laser ablation + 16-18 hours healing (right eye) Replicate 3

source_name: Wounded

tag: mouse strain

value: Corneal epithelial cells

value: 105 days

value: Wounded


title: HS2ST (+/-) 1

description: P1 mouse brain heterozygous for Heparan sulfate 2-O sulfotransferase (HS2ST) loss of function mutation: Replicate 1


title: genomic DNA from patient #CCF112

description: 5hmC low (mutant)

value: bone marrow

value: mutant


title: Laser ablation + 16-18 hours healing (right eye) Replicate 1


title: Laser ablation + 16-18 hours healing (right eye) Replicate 2


title: genomic DNA from patient #CCF016

description: 5hmC low (wt)

value: male

value: low

value: wt


title: genomic DNA from patient #CCF011

description: 5hmC low (mutant)

value: female

value: de novo AML


title: genomic DNA from patient #CCF080

description: 5hmC low (mutant)


title: Normal Cornea (Left eye) Replicate 3

source_name: Normal

value: unknown

value: WT

value: Healthy


title: genomic DNA from patient #CCF018

description: 5hmC low (wt)

source_name: bone marrow

value: MDS


title: genomic DNA from patient #CCF082

description: 5hmC low (mutant)


title: Normal Cornea (Left eye) Replicate 2


title: genomic DNA from patient #CCF014

description: 5hmC low (wt)


title: Pool #4, B Labeling, Hyb day #3

source_name: Pool #4, B Labeling, Hyb day #3

value: WT C57/Bl

value: healthy

value: 8 weeks

value: brain


title: Pool #4, C Labeling, Hyb day #3

source_name: Pool #4, C Labeling, Hyb day #3


title: Pool #4, A Labeling, Hyb day #3

source_name: Pool #4, A Labeling, Hyb day #3


title: Pool #3, C Labeling, Hyb day #3

source_name: Pool #3, C Labeling, Hyb day #3


title: Pool #3, B Labeling, Hyb day #3

source_name: Pool #3, B Labeling, Hyb day #3


title: Pool #3, A Labeling, Hyb day #3

source_name: Pool #3, A Labeling, Hyb day #3


title: Pool #2, C Labeling, Hyb day #3

source_name: Pool #2, C Labeling, Hyb day #3


title: Pool #2, B Labeling, Hyb day #3

source_name: Pool #2, B Labeling, Hyb day #3


title: Pool #2, A Labeling, Hyb day #3

source_name: Pool #2, A Labeling, Hyb day #3


title: genomic DNA from patient #CCF083

description: 5hmC low (mutant)

value: MDS/MPN


title: genomic DNA from patient #CCF002

description: 5hmC low (mutant)


title: genomic DNA from patient #CCF106

description: 5hmC low (mutant)


title: genomic DNA from patient #CCF003

description: 5hmC low (mutant)


title: genomic DNA from patient #CCF087

description: 5hmC low (mutant)


title: genomic DNA from patient #CCF105

description: 5hmC low (mutant)


title: genomic DNA from patient #CCF057

description: 5hmC low (mutant)


title: genomic DNA from patient #CCF054

description: 5hmC low (wt)


title: genomic DNA from patient #CCF009

description: 5hmC low (mutant)


title: Normal Cornea (Left eye) Replicate 1

value: Corneal epithelial cells

value: 105 days


title: Pool #1, C Labeling, Hyb day #3

source_name: Pool #1, C Labeling, Hyb day #3


title: Pool #4, B Labeling, Hyb day #2

source_name: Pool #4, B Labeling, Hyb day #2


title: Pool #4, C Labeling, Hyb day #2

source_name: Pool #4, C Labeling, Hyb day #2


title: genomic DNA from patient #CCF084

description: 5hmC low (mutant)

value: bone marrow

value: mutant


title: genomic DNA from patient #CCF006

description: 5hmC low (mutant)

value: low


title: genomic DNA from patient #CCF012

description: 5hmC low (mutant)

value: male


title: Pool #4, A Labeling, Hyb day #2

source_name: Pool #4, A Labeling, Hyb day #2


title: genomic DNA from patient #CCF005

description: 5hmC low (mutant)

value: sAML


title: genomic DNA from patient #CCF021

description: 5hmC low (wt)

value: wt


title: Pool #3, B Labeling, Hyb day #2

source_name: Pool #3, B Labeling, Hyb day #2


title: genomic DNA from patient #CCF023

description: 5hmC low (wt)


title: genomic DNA from patient #CCF001

description: 5hmC low (mutant)


title: Pool #3, C Labeling, Hyb day #2

source_name: Pool #3, C Labeling, Hyb day #2


title: Pool #3, A Labeling, Hyb day #2

source_name: Pool #3, A Labeling, Hyb day #2


title: Pool #2, C Labeling, Hyb day #2

source_name: Pool #2, C Labeling, Hyb day #2


title: Pool #2, B Labeling, Hyb day #2

source_name: Pool #2, B Labeling, Hyb day #2


title: Pool #1, C Labeling, Hyb day #2

source_name: Pool #1, C Labeling, Hyb day #2


title: Pool #2, A Labeling, Hyb day #2

source_name: Pool #2, A Labeling, Hyb day #2


title: Pool #1, A Labeling, Hyb day #2

source_name: Pool #1, A Labeling, Hyb day #2


title: Pool #4, C Labeling, Hyb day #1

source_name: Pool #4, C Labeling, Hyb day #1


title: Pool #1, B Labeling, Hyb day #2

source_name: Pool #1, B Labeling, Hyb day #2


title: genomic DNA from patient #CCF088

description: 5hmC low (mutant)

source_name: bone marrow

value: female


title: Pool #3, C Labeling, Hyb day #1

source_name: Pool #3, C Labeling, Hyb day #1


title: genomic DNA from healthy bone marrow #CCF049

description: control

value: healthy control


title: Pool #3, A Labeling, Hyb day #1

source_name: Pool #3, A Labeling, Hyb day #1


title: Pool #2, C Labeling, Hyb day #1

source_name: Pool #2, C Labeling, Hyb day #1


title: genomic DNA from healthy bone marrow #CCF137

description: control


title: genomic DNA from healthy bone marrow #CCF136

description: control


title: Pool #2, B Labeling, Hyb day #1

source_name: Pool #2, B Labeling, Hyb day #1


title: genomic DNA from healthy bone marrow #CCF139

description: control


title: genomic DNA from healthy bone marrow #CCF117

description: control


title: Pool #4, A Labeling, Hyb day #1

source_name: Pool #4, A Labeling, Hyb day #1


title: genomic DNA from healthy bone marrow #CCF135

description: control


title: genomic DNA from healthy bone marrow #CCF138

description: control


title: genomic DNA from healthy bone marrow #CCF140

description: control


title: Pool #4, B Labeling, Hyb day #1

source_name: Pool #4, B Labeling, Hyb day #1


title: genomic DNA from healthy bone marrow #CCF132

description: control


title: Pool #1, B Labeling, Hyb day #1

source_name: Pool #1, B Labeling, Hyb day #1


title: genomic DNA from healthy bone marrow #CCF119

description: control


title: Pool #1, A Labeling, Hyb day #1

source_name: Pool #1, A Labeling, Hyb day #1


title: genomic DNA from patient #CCF081

description: 5hmC low (mutant)


title: Pool #1, C Labeling, Hyb day #1

source_name: Pool #1, C Labeling, Hyb day #1


title: genomic DNA from healthy bone marrow #CCF050

description: control


title: genomic DNA from healthy bone marrow #CCF131

description: control


title: genomic DNA from healthy bone marrow #CCF046

description: control


title: genomic DNA from healthy bone marrow #CCF134

description: control


title: Pavement cells; 5 week old plants; Rep 1

city: Golm

first: Berit

last: Ebert

address: Am Mühlenberg 1

department: Willmitzer

country: Germany

institute: Max-Planck Institute of Molecular Plant Physiology

description: Gene expression data from epidermis cells

hyb_protocol: standard Affymetrix protocol

scan_protocol: standard Affymetrix protocol

data_processing: The data were analyzed with Bioconductor using the RMA normalization procedure

organism: Arabidopsis thaliana

source_name: Pavement cells, Arabidopsis leave

label_protocol:
Amplification (one round) was performed using the ArcturusRiboAmp Kit, labelling and hybridisation were performed with the GeneChip Expression 3' Amplification reagents for IVT labeling Kit according to the manufacturer’s (Affymetrix, CA)
The GeneChip Expression 3' Amplification reagents for IVT labeling Kit according to the manufacturer’s was then used to synthesize biotin-labeled cRNA from template cDNA by in vitro transcription.

growth_protocol: Arabidopsis plants were cultivated in growth chambers at 20oC/18°C under a 16 h light/8 h dark cycle until seed set.

extract_protocol: Extraction of total RNA was performed using the Quiagen Micro RNA Kit according to the manufacturer's instructions

treatment_protocol: Pavement cells of Arabidopsis leaves were sampled using microcapilaries as described in Brandt et al., 1999

tag: Genotype

tag: Age

value: 5 weeks old plants


title: genomic DNA from healthy bone marrow #CCF130

description: control

tag: gender


title: genomic DNA from healthy bone marrow #CCF128

description: control


title: genomic DNA from healthy bone marrow #CCF129

description: control


title: Trichome cells; 5 week old plants; Rep 1

description: Gene expression data trichomes

source_name: Trichome cells, Arabidopsis leave

treatment_protocol: Trichome cells of Arabidopsis leaves were sampled using microcapilaries as described in Brandt et al., 1999


title: genomic DNA from healthy bone marrow #CCF127

description: control


title: genomic DNA from healthy bone marrow #CCF133

description: control


title: genomic DNA from healthy bone marrow #CCF126

description: control


title: genomic DNA from healthy bone marrow #CCF048

description: control


title: genomic DNA from healthy bone marrow #CCF141

description: control


title: genomic DNA from healthy bone marrow #CCF125

description: control


title: CASMC_Control_Pair3

city: Boston

first: James

middle: R.

last: Stone

state: MA

address: Simches 8236, 185 Cambridge Street

institute: Massachusetts General Hospital

description: The resulting CEL files were summarized using Affymetrix Expression Console (version 1.1).

hyb_protocol: The labeled fragmented DNA was hybridized to the Exon Arrays for 16 hours in GeneChip Hybridization oven 640 at 45oC with rotation (60 rpm).  The hybridized samples were washed and stained using Affymetrix fluidics station 450. The first stain with streptavidin-R-phycoerythrin was followed by signal amplification using a biotinilated goat anti-streptavidin antibody followed by another SAPE staining (Hybridization, Washing and Staining Kit, Affymetrix, Santa Clara, CA).

scan_protocol: Microarrays were immediately scanned using Affymetrix GeneArray Scanner 3000 7G Plus (Affymetrix, Santa Clara, CA).

data_processing: RMA (Robust Multi-Array Analysis - Irizarry RA, Hobbs B, Collin F, Beazer-Barclay YD, Antonellis KJ, Scherf U, Speed TP. Biostatistics. 2003 Apr;4(2):249-64) algorithm was used to generate gene-level data and iterPLIER algorithm (http://www.affymetrix.com/support/technical/whitepapers/ exon_gene_signal_estimate_whitepaper.pdf) was used to generate the exon-level data.

source_name: Human Coronary Artery Smooth Muscle Cells, vector control, pair group number 3

label_protocol: Affymetrix Whole Transcript Terminal Labeling Kit

extract_protocol: QIANEN's RNeasy kit

value: coronary artery smooth muscle cell


title: CASMC_Knockdown_Pair3

source_name: Human Coronary Artery Smooth Muscle Cells, Knockdown, pair group number 3


title: genomic DNA from healthy bone marrow #CCF045

description: control


title: CASMC_Knockdown_Pair1

source_name: Human Coronary Artery Smooth Muscle Cells, Knockdown, pair group number 1


title: genomic DNA from healthy bone marrow #CCF124

description: control


title: CASMC_Knockdown_Pair2

source_name: Human Coronary Artery Smooth Muscle Cells, Knockdown, pair group number 2


title: CASMC_Control_Pair2

source_name: Human Coronary Artery Smooth Muscle Cells, vector control, pair group number 2


title: Maltose_vs_Glucose_dyeswap_rep2

city: Oak Ridge

first: Charlotte

last: Wilson

state: Tennessee

address: 1 Bethel Valley Road

department: Biosciences Bldg 1520

country: New Zealand

institute: ORNL

hyb_protocol: The labelled samples were mixed and competitively hybridised using an Agilent Gene Expression Hybridisation Kit (part number: 5188-5242) in an Agilent hybridization oven (G2545A) at 65°C for 24 hours.

scan_protocol: Microarrays were scanned using the Agilent Microarray Scanner System (G2505B) and Agilent scan control software version 7.0 at a resolution of 5 micrometres

data_processing: Generated scans were converted to data files with Agilent's Feature Extraction software (Version 9.1) which performs a lowess normalization. Microarray data was processed as described by García de la Nava et al 2003 and van Hijum et al 2005. Differential gene expression was determined by Cyber-T test (Long 2001). Genes with a P value <0.001 and greater than two fold difference under the two conditions were considered differentially expressed.

organism: Limosilactobacillus reuteri subsp. rodentium

source_name: Lactobacillus reuteri 100-23 grown in the presence of glucose

label_protocol: Two micrograms of total RNA was labelled with either Cy5- or Cy3-dCTP during first-strand synthesis with SuperScript II (Invitrogen) reverse transcriptase kit following the manufacturer’s protocol. Labelled cDNA was purified using a MinElute PCR purification kit (Qiagen) and the dye incorporation rate was determined by NanoDrop ND-1000 UV-Vis spectrophotometer.

growth_protocol: Lactobacillus reuteri 100-23 was cultured in Lactobacilli MRS medium (Difco, Becton Dickinson Co., Sparks, MD, USA) incubated anaerobically at 37oC. Lactobacilli MRS medium was made from scratch and contained either glucose or maltose at 2% (w/v) according to experimental requirements. Liquid media were filter-sterilised, agar media were autoclaved.

extract_protocol: Bacterial cells were harvested by brief centrifugation at 12,000 x g and washed with 750 µl of RNAprotect reagent (Qiagen). The bacterial cells were disrupted by bead beating (5000 rpm, 2 x 40 seconds) in TRIzol™ reagent (Invitrogen) and extracted with chloroform. RNA was precipitated with iso-propanol and dried after removal of iso-propanol. The dried RNA was then dissolved in nuclease-free water. The RNA was further purified using the RNeasy Mini Kit (Qiagen) and DNase treated using a DNA-free kit (Ambion). RNA quality was assessed using a Bioanalyzer 2100 instrument (Agilent Technologies) and quantified by NanoDrop ND-1000 (Nanodrop).

treatment_protocol: Total RNA was extracted from cultures of strain 100-23 grown in Lactobacilli MRS medium containing 2% (w/v) glucose or maltose for four hours.

tag: growth stage

value: exponential

tag: carbon source

value: glucose

source_name: Lactobacillus reuteri 100-23 grown in mMRS with maltose

value: maltose


title: Maltose_vs_Glucose_dyeswap_rep1


title: CASMC_Control_Pair1

source_name: Human Coronary Artery Smooth Muscle Cells, vector control, pair group number 1


title: genomic DNA from healthy bone marrow #CCF044

description: control

source_name: bone marrow

tag: tissue


title: Maltose_vs_Glucose_ rep2


title: 18 hours antibody C7 exposure 4

city: San Sebastian

first: Iñigo

last: Fernandez de Larrinoa

state: Guipuzcoa

address: Pº Manuel de Lardizabal #3

department: Química Aplicada

country: Spain

institute: Universidad del País Vasco (UPV/EHU)

description: Biological replicate 4 of 4

hyb_protocol: A mixture containing 5µl each Cy3 and Cy5-labeled cDNA samples plus 5µl herring sperm DNA (10 mg/ml) was heat denatured at 95°C for 2 min and quickly cooled on ice. Forty microliters of hybridization buffer were added to the DNA probe, and the solution was placed onto DNA microarrays slides, which were covered with a rimmed cover glass (Erie Scientific, Portsmouth, NH). Slides were placed in a hybridization chamber (Corning, Palo Alto, CA) and hybridized for 24 h at 42°C by immersion in a water bath. Following hybridization, the slides were immersed in 2x SSC, 1% SDS for 15 min. and washed twice for 5 min each in 1x SSC at room temperature. The slides were dried by centrifugation at 1100 rpm for 5 min. Four dye-swap comparative analysis were performed.

scan_protocol: The microarrays were scanned using a GenePix4000B microarray scanner (Axon Instruments) at a 10-µm resolution. GenePix Pro 4.0 analysis software (Axon Instruments) was used to locate spots in the microarray with the appropriate grid and to obtain the two Cy3/Cy5 image TIFF files. All images were further processed using GenePix 4.0 software according to manufacturer instructions. Low-quality spots were automatically flagged by Genepix Pro 4.0.

data_processing: Data were evaluated with the GeneSpring version 5.0 software (Silicon Genetics, Redwood City, CA) and normalized in a per-chip (to the 50 percentile) and per-gene (to the median) way. Median F635, F532, B635, and B532 of each spot were loaded into GeneSpring for calculation of signal and background intensities. Signals were log transformed and per-chip normalized by an intensity-dependent method (Lowess) applied to the print-tip region.

organism: Candida albicans

source_name: candida not exposed

label_protocol: Fifteen micrograms of total RNA was labeled in a total volume of 40 µl including 1x first-strand buffer (Invitrogen), 0.2 pmol C. albicans-specific primer mix (Eurogentec), 1.5 mM (each) dATP, dTTP, and dGTP, 25 µM dCTP, 10 mM dithiothreitol, 1 µl RNasin, and 37.5 µM Cy5-dCTP or Cy3-dCTP (Amersham). The mixture was incubated at 65°C for 5 min and then at 42°C for 5 min. One microliter of RNasin (Promega) and 1 µl Superscript II reverse transcriptase (Invitrogen) were added and incubated at 42°C for 1.5 h. Following the addition of another 1 µl of Superscript II, the incubation was continued for 1.5 h at 42°C. The reaction was stopped by the addition of 5 mM EDTA, pH 8.0, and 0.5 M NaOH and incubation at 65°C for 20 min and was neutralized by adding 0.5 M acetic acid. The labeled cDNAs were purified using a QIAquick PCR purification kit (QIAGEN) and concentrated by filtration through microcon columns (Millipore) to a volume of 10 µl. The amount of cDNA as well as the incorporation of Cy3 and Cy5 dyes into cDNA targets was quantified on a spectrophotometer using Ultra-micro cuvettes.

growth_protocol: A saturated culture of C. albicans SC5314 grown overnight in modified Lee’s medium was diluted to an optical density at 600 nm of approximately 0.1 and split in two aliquots. Monoclonal antibody C7 was added to one of the cultures at 12.5 µg/ml final concentration whereas the second one remained untreated. Both cultures were incubated for 18 h at 37ºC before harvesting cells.

extract_protocol: Cells were harvested by centrifugation, transferred to a pre-cooled teflon vessel and quickly chilled in liquid nitrogen. Cells were disrupted by shaking with a tungsten carbide bead at 2000 rpm for 2 minutes in a micro-dismembrator and RNA was extracted with Trizol Reagent.

treatment_protocol: Monoclonal antibody C7 was added at 12’5 ug/ml final concentration in order to inhibit fungal growth and avoiding fungicidal effect

tag: strain

tag: growth conditions

value: maintained on solid YPD medium and incubated at 30ºC

source_name: candida exposed to antibody C7


title: 18 hours antibody C7 exposure 2

description: Biological replicate 2 of 4


title: genomic DNA from healthy bone marrow #CCF122

description: control

tag: disease state


title: 18 hours antibody C7 exposure 1

description: Biological replicate 1 of 4


title: Maltose_vs_Glucose_ rep1


title: genomic DNA from healthy bone marrow #CCF042

description: control


title: 18 hours antibody C7 exposure 3

description: Biological replicate 3 of 4


title: S. salar_5_dpf_rep2

status: Public on Dec 10 2010

city: Victoria

first: Ben

middle: F

last: Koop

address: PO Box 3020 STN CSC

department: Biology

institute: The University of Victoria

description: Biological replicate 2 of 3

hyb_protocol: cRNA fragmentation mixtures were created following the LIQA kit instructions, using 825 ng of experimental sample and 825 ng of reference pool. These mixtures were incubated at 60 ºC for 30 minutes. After cooling on ice for one minute, hybridization mixtures were prepared by adding 2x GEx Hybridization Buffer HI-RPM and mixing well by pipetting. These reactions were loaded in random arrangements with respect to time point onto 44K oligo salmonid microarrays (Agilent-025055) using Agilent SureHyb Hybridization Chambers. Each of the 4x44K arrays on the microarray slides had 100 uL of hybridization reaction added. The hybridization reactions were allowed to occur for 17 hours at 65 ºC. Slide washes were performed as per the manufacturer’s instructions, including an ozone-protection step using the Agilent Stabilization and Drying Solution.

scan_protocol: Slides were scanned as soon as possible on a ScanArray Express (Perkin Elmer) scanner at 5 µm resolution using a PMT setting of 80 in both channels, a black threshold of 1800, and a full color threshold of 26.8. Slides were stored in a low ozone chamber (typically <5 ppb) until scanned. Images were quantified using Imagene 8.0 (Biodiscovery).

data_processing: Background corrected signal medians converted to threshold of 1.0, log2 transformed, Lowess normalized, baseline transformation to median.

organism: Salmo salar

source_name: Whole tissue, 5 days post fertilization

label_protocol: Cy5 labeled experimental cRNA samples was generated using an Agilent Low Input Quick Amp (LIQA) kit, following the manufacturer’s instructions. For each time point, 40 ng of total RNA from three individuals was used to generate first-strand cDNA. Agilent Spike-In B control RNA was included in each reaction. After the denaturation step (10 min at 65 ºC) and cRNA synthesis step (2 hr at 40 ºC), the reactions were incubated at 70 ºC for 15 minutes to inactivate the AffinityScript enzyme, and subsequently stored at -80 ºC until further use. For the labeling reactions, thawed cRNA samples were each mixed with 16 µL of Transcription Master Mix cocktail containing Cy5 dye, and incubated at 40 ºC for two hours. Purification was performed using Qiagen RNeasy mini spin columns, eluting in 30 µL of RNase-free water. For the generation of the reference pool, equimolar amounts from the three individuals in each time point were pooled to give 120 ng of total RNA used in each first-strand reaction. Spike-In A control RNA was included in each reaction. After labeling with Cy3 and column purification as above, a common reference pool was created by including 2.8 µg of cRNA from each time point, except for D2, for which only 1.3 µg of material was available due to the small size of the animal at this stage.

growth_protocol: Eggs from Atlantic salmon (McConnell (Mowi)) were obtained in November, 2009 from Marine Harvest United Hatchery (Fanny Bay, B.C., Canada). The eggs were fertilized by gently mixing the eggs and milt by hand and then washed with partial exchanges of water. Approximately 2000 fertilized eggs were then transferred and placed in Heath trays (Marisource) at the University of Victoria. The embryos and larvae were raised in fresh water at a temperature of 12°C and a flow rate of 200 liters/h.

extract_protocol: Total RNAs were extracted in TRIzol reagent (Invitrogen) by mixer-mill homogenization (Retsch) and spin-column purified using RNeasy Mini kits (Qiagen).

value: whole organism

tag: developmental stage

value: 5 days post fertilization

source_name: Pooled cRNA from whole tissues in all time points

tag: sample type


title: S. salar_30_dpf_rep3

description: Biological replicate 3 of 3

source_name: Whole tissue, 30 days post fertilization

value: 30 days post fertilization

value: Pooled cRNA from whole tissues in all time points


title: S. salar_15_dpf_rep3

description: Biological replicate 3 of 3

source_name: Whole tissue, 15 days post fertilization

value: 15 days post fertilization


title: S. salar_5_dpf_rep3

description: Biological replicate 3 of 3


title: S. salar_83_dpf_rep3

description: Biological replicate 3 of 3

source_name: Whole tissue, 83 days post fertilization

value: 83 days post fertilization


title: S. salar_53_dpf_rep3

description: Biological replicate 3 of 3

source_name: Whole tissue, 53 days post fertilization

value: 53 days post fertilization


title: S. salar_47_dpf_rep2

description: Biological replicate 2 of 3

source_name: Whole tissue, 47 days post fertilization

value: 47 days post fertilization


title: S. salar_89_dpf_rep3

description: Biological replicate 3 of 3

source_name: Whole tissue, 89 days post fertilization

value: 89 days post fertilization


title: S. salar_47_dpf_rep3

description: Biological replicate 3 of 3


title: S. salar_2_dpf_rep3

description: Biological replicate 3 of 3

source_name: Whole tissue, 2 days post fertilization

value: 2 days post fertilization


title: S. salar_89_dpf_rep2

description: Biological replicate 2 of 3


title: S. salar_5_dpf_rep1

description: Biological replicate 1 of 3


title: S. salar_10_dpf_rep3

description: Biological replicate 3 of 3

source_name: Whole tissue, 10 days post fertilization

value: 10 days post fertilization


title: S. salar_59_dpf_rep3

description: Biological replicate 3 of 3

source_name: Whole tissue, 59 days post fertilization

value: 59 days post fertilization


title: S. salar_10_dpf_rep2

description: Biological replicate 2 of 3


title: S. salar_30_dpf_rep2

description: Biological replicate 2 of 3


title: S. salar_20_dpf_rep3

description: Biological replicate 3 of 3

source_name: Whole tissue, 20 days post fertilization

value: 20 days post fertilization


title: S. salar_83_dpf_rep2

description: Biological replicate 2 of 3


title: S. salar_77_dpf_rep3

description: Biological replicate 3 of 3

source_name: Whole tissue, 77 days post fertilization

value: 77 days post fertilization


title: S. salar_10_dpf_rep1

description: Biological replicate 1 of 3


title: S. salar_71_dpf_rep2

description: Biological replicate 2 of 3

source_name: Whole tissue, 71 days post fertilization

value: 71 days post fertilization


title: S. salar_77_dpf_rep2

description: Biological replicate 2 of 3


title: S. salar_35_dpf_rep3

description: Biological replicate 3 of 3

source_name: Whole tissue, 35 days post fertilization

value: 35 days post fertilization


title: S. salar_71_dpf_rep3

description: Biological replicate 3 of 3


title: S. salar_25_dpf_rep3

description: Biological replicate 3 of 3

source_name: Whole tissue, 25 days post fertilization

value: 25 days post fertilization


title: S. salar_2_dpf_rep2

description: Biological replicate 2 of 3

value: whole organism


title: S. salar_41_dpf_rep3

description: Biological replicate 3 of 3

source_name: Whole tissue, 41 days post fertilization

value: 41 days post fertilization


title: S. salar_65_dpf_rep3

description: Biological replicate 3 of 3

source_name: Whole tissue, 65 days post fertilization

value: 65 days post fertilization


title: S. salar_20_dpf_rep2

description: Biological replicate 2 of 3

source_name: Pooled cRNA from whole tissues in all time points


title: S. salar_59_dpf_rep1

description: Biological replicate 1 of 3

value: Pooled cRNA from whole tissues in all time points


title: S. salar_65_dpf_rep2

description: Biological replicate 2 of 3


title: S. salar_53_dpf_rep2

description: Biological replicate 2 of 3


title: KM-H2

description: Genome-wide SNP/CN data of Hodgkin lymphoma cell line KM-H2

hyb_protocol: DNA was restriction digested, PCR amplified, fragmented, labeled and hybridized to each array according to the manufacturer's instructions.

data_processing: The Affymetrix SNP 6.0 data were processed using the R-based aroma.affymetrix package (Bengsston et al, Bioinformatics 2009). Total copy number was estimated according to the CRMA procedure, calibrating for allelic cross talk, nucleotide-position probe sequence effects, robust probe summarization and correction for fragment length effects.

source_name: Hodgkin lymphoma cell line KM-H2

label_protocol: standard Affymetrix protocol

value: Pleural effusion cells

tag: cell line


title: L-428

description: Genome-wide SNP/CN data of Hodgkin lymphoma cell line L-428

source_name: Hodgkin lymphoma cell line L-428


title: S. salar_35_dpf_rep2

description: Biological replicate 2 of 3


title: S. salar_59_dpf_rep2

description: Biological replicate 2 of 3


title: S. salar_15_dpf_rep2

description: Biological replicate 2 of 3

source_name: Whole tissue, 15 days post fertilization

value: 15 days post fertilization


title: S. salar_71_dpf_rep1

description: Biological replicate 1 of 3


title: S. salar_2_dpf_rep1

description: Biological replicate 1 of 3


title: S. salar_53_dpf_rep1

description: Biological replicate 1 of 3

source_name: Whole tissue, 53 days post fertilization

value: 53 days post fertilization


title: S. salar_25_dpf_rep2

description: Biological replicate 2 of 3


title: S. salar_89_dpf_rep1

description: Biological replicate 1 of 3


title: S. salar_41_dpf_rep2

description: Biological replicate 2 of 3


title: S. salar_20_dpf_rep1

description: Biological replicate 1 of 3


title: S. salar_77_dpf_rep1

description: Biological replicate 1 of 3


title: S. salar_41_dpf_rep1

description: Biological replicate 1 of 3


title: S. salar_83_dpf_rep1

description: Biological replicate 1 of 3

source_name: Whole tissue, 83 days post fertilization

value: 83 days post fertilization


title: S. salar_15_dpf_rep1

description: Biological replicate 1 of 3


title: S. salar_35_dpf_rep1

description: Biological replicate 1 of 3


title: S. salar_30_dpf_rep1

description: Biological replicate 1 of 3

source_name: Whole tissue, 30 days post fertilization

value: 30 days post fertilization


title: S. salar_25_dpf_rep1

description: Biological replicate 1 of 3


title: MA7

city: San Antonio

first: August

middle: Nolan

last: Blackburn

state: Texas

address: 7702 Floyd Curl

department: Medicine - Clinical Epidemiology

institute: University of Texas Health Science Center at San Antonio

hyb_protocol: Standard Illumina protocol

scan_protocol: Standard Illumina protocol

data_processing:
To normalize the data I used Illumina Genome Studio's Genotyping Module version 1.5.16. The data was normalized using a cluster file of 782 individuals of Mexican American descent who were genotyped as part of the San Antonio Family Heart and San Antonio Family Diabetes/Gallbladder studies.
To identify regions that differed in copy number between pair samples I used circular binary segmentation (CBS) applied to the absolute value of the difference between b-allele ratios. Applying CBS to b-allele ratio differences is a novel approach. To implement CBS I used DNAcopy 1.16.0. The statistical approach I used is described here: (VENKATRAMAN, E. S., and A. B. OLSHEN, 2007 A faster circular binary segmentation algorithm for the analysis of array CGH data. Bioinformatics 23: 657-663). I used PennCNV (Sat, 01 May 2010 version) to identify germline copy number variants in all of the samples. I used the available trio information and a posteriori trio calling to identify de novo CNVs.

label: C-Bio and A-DNP

label_protocol: Standard Illumina protocol

extract_protocol: To extract DNA we used a standard phenol-chloroform method. The detailed method is attached in a pdf document.

value: female

tag: parent twin pair


title: MA8

value: Male


title: MA9

tag: age of examination, not current age

tag: twin pair


title: E0-4_H3K9Me3_2 extraction2_array1

first: DCC

last: modENCODE

state: Ontario

address: MaRS Centre, South Tower, 101 College Street, Suite 800

institute: Ontario Institute for Cancer Research

description: channel ch1 is ChIP DNA; Antibody information listed below: official name: H3K9me3;target name: H3K9me3;host: Rabbit;antigen: Synthetic peptide conjugated to KLH derived from within residues 1 - 100 of Human Histone H3 tri methylated at K9.;clonal: Polyclonal;purified: Affinity;company: Abcam;catalog: ab8898;short description: Histone H3 (tri methyl K9) antibody - ChIP Grade (ab8898);

hyb_protocol: This protocol is used for hybridization of ChIP samples on 244K Agilent arrays.  The hybridization solution is composed of 5 ug each of Cy3- and Cy5-labeled DNA, 50 ug of salmon sperm DNA, Agilent Blocking Agent and Agilent Hyb Buffer. The mixture is heated to 95*C for 3 min and then 37*C for 30 minutes immediately prior to hybridization. Samples are hybridized for 40 hours at 65*C. After hybridization the slides are washed in Agilent Wash Buffer 1 for 5 minutes at room temperature and then in Agilent Wash Buffer 2 for 5 minutes at 31*C.

scan_protocol: Arrays are scanned on Agilent's DNA Microarray Scanner G2505B at a resolution of 5 microns. Data is extracted from the images using Agilent's Feature Extraction Software 9.5.3.1.

data_processing: Agilent ChIP data processing:KW:1 protocol. We utilized Agilent 244 K array sets for carrying out ChIP-chip experiments. The Drosophila melanogaster genome is tiled using 3 array sets with mean probe resolution of ~160 base pairs. We preprocessed microarray data to normalize it and reduce experimental noise as described in Qi et al. (Qi et al., Nature biotechnology 2006). In short, the raw intensities from each channel was divided by the median intensity from that channel before computing a ratio to arrive at a median adjusted ratio. We further subtracted average value of negative control probes on the array and then added one as a regularization parameter. For factors that mark regions (e.g. H3K27me3), the data was quantile normalized, replicates were merged and HMM based segmentation was carried out.

organism: Drosophila melanogaster

label_protocol: Attached as supplementary file

extract_protocol:
Collection of Drosophila embryos at time-point 0-4h of development for further ChIP experiments or RNA extraction.
Formaldehyde Cross-linking of Chromatin from any Drosophila tissue (Embryos, Larvae, Pupae, Adult flies).1. Homogenize the material (about 150-200 mg of embryos, pupae, adµlts, larvae, sufficient for four-five independent immuno-precipitations) in 5 ml of buffer A1 + 1.8% formaldehyde (290 µl of 37% solution) at room temperature using first a Potter homogenizer and then a Douncer with type A pestle (3 strokes). Wait 15? (total time starting from beginning of homogenization). Add glycin solution to 225 mM (540 µl of 2.5 M solution for 6 ml of cross-linked mixture), mix and incubate 5?. Put on ice. Keep on ice unless specified.2. Transfer the homogenate to a 15 ml tube. Centrifuge 5 min, at 4000 g at 4°C. Discard supernatant. Add 3 ml of buffer A1, resuspend pellet and spin down the same way. Repeat washing step 3 times.3. Wash once in 3 ml of lysis buffer without SDS. Spin down, 5 min at 4000g.4. Resuspend cross-linked material in 0.5 ml of lysis buffer and add SDS to 0.1% (2.5 µl) and N-lauroylsarcosine to 0.5% (12.5 µl). Incubate 10 min at 4°C in a rotating wheel.5. Sonicate in Polypropylene 15ml Falcon tubes according to .Diagenode instructions. The settings are: High power, 15 minutes, 30 seconds off, 30 seconds on. The water bath shoµld be kept cold either in a cold room or by a cold water pump.6. Rotate 10'. Transfer to eppendorf tubes and centrifuge 5', room temperature at maximum speed. Transfer supernatant to the new tube. Add another 0.5 ml of lysis buffer to the pellet, rotate for 10 min. Repeat centrifugation and combine supernatants. Centrifuge the combined supernatant 2x10 min at max speed. The supernatant from this stage will be referred as chromatin extract. Chromatin can be stored at ?80°C for several months at this stage. Add sodium azide to 0.02% for storage.7. [Optional : Purification of chromatin. Put chromatin extract in Centricon YM-100 column (4212), centrifuge 3x40' (or more) at 1000 g while adding lysis buffer. At least 3 vol. of lysis buffer should pass through column. Bring the final volume of chromatin extract to 1 ml by lysis buffer + protease inhibitors.][edit]
Solutions and MaterialsLysis buffer: (see above)5mg/ml RNaseA (DNase free)PAS suspension: 100 mg of CL-4B (Amersham, 17-0780-01) PAS shoµld be resuspended in 1 ml of lysis buffer + protease inhibitors + 0.1 mg/ml BSA for 50 %v suspension. Wash in lysis buffer 2-3 times, overall equilibration time 1h. Store up to one week at 4 C.TE: 10 mM Tris-HCl pH 8.0, 1 mM EDTAElution buffer1: 10 mM EDTA, 1% SDS, 50 mM Tris-Cl pH 8.Elution buffer2: TE+0.67% SDS.LiCL 4M? Protocol 1. To an amount of chromatin corresponding to 150 mg of biological material, suspended in a final volume of chromatin extract of 1 ml (in lysis buffer + protease inhibitors), add 100 µl of PAS suspension for preincubation. Incubate several hours or overnight at 4°C, then remove PAS. Crosslinked chromatin at this stage can be stored several days at 4°C or frozen at ?70°C.2. [Optional : Check for amount of DNA. From the 1 ml solution above, take a 100 µl aliquot, Add proteinase K up to 100 ug/ml and SDS to 1%, incubate 6h at 60°C, then 20 min at 70°C, add RNAse to 50 ug/ml and incubate additional 2h at 37°C. Phenol-chlorophorm extract, ethanol precipitate. Run on an agarose gel to check amount and size of DNA.]3. Separate chromatin sample into 4x250 µl aliquots (one aliquot is enough for one IP but the amount may be decreased further). Immunoprecipitate chromatin by adding the antibody (Ab) of interest (amount of Ab shoµld be determined empirically, usually the same concentration than for a successfµl IF experiement). Keep a control sample without Ab IP, named "Mock".4. Incubate 4 h at 4°C on a head to head rotating wheel, then add 50 µl of PAS suspension and incubate 4 h or overnight. Spin down PAS and proceed to washes.5. Wash PAS 4x with lysis buffer, followed by 2x with TE (without protease inhibitors). Each wash is for 5 min at 4°C, using 1 ml of solution.6. Elution of precipitated material. Spin down PAS. Add 100 µl of elution buffer, mix and incubate 10 min at 65°C. Spin down PAS and transfer supernatant to new tube. Add 150 µl of elution buffer 2 to PAS, mix, centrifuge at fµll speed and transfer eluate to a tube together with the eluate from the first centrifugation. The combined material is the "chromatin precipitate" (approx. 250 µl).7. Incubate precipitate 6 h (or overnight) at 65 C to reverse cross-links. Add 250 µl of Proteinase K solution, incubate at 50°C 2-3 h.8. Add 55 µl of 4M LiCl and 500 µl of phenol-chlorophorm. Mix and microfuge at fµll speed at RT. Transfer aqueous phase to a new tube and precipitate with 1 ml of 100% ethanol. Wash with 750 µl of 70% ethanol. Spin down and dry precipitate.9. Dissolve in 25 µl of water. This is the chromatin immunoprecipitate or ?Native ChIP? sample.
LM-PCR amplification of the native ChIP sample to obtain a sufficient quantity for further hybridization on microarrays.

tag: genotype

tag: Sex

value: Unknown


title: E0-4_H3K9Me3_3 extraction3_array1

description: channel ch1 is negative control for ChIP; Antibody information listed below: official name: No Antibody Control;target name: Not Applicable;host: None-Control;antigen: NONE;clonal: None-Control;purified: None-Control;company: None-Control;catalog: Not Applicable;reference: Not Applicable;short description: Specific Antibodies were omitted from this immunoprecipitation reaction (as a negative control).;


title: MA12


title: E0-4_H3K9Me3_1 extraction1_array1


title: MA5


title: MA3


title: MA11

value: Female


title: E0-4_H3K27Ac_2 extraction2_array2


title: FXRSHPko.5wk.rep2

first: Scott

middle: Andrew

last: Ochsner

address: One Baylor Plaza

department: Molecular and Cellular Biology

description: Gene expression in 5wk old FXR-/-/SHP-/- mouse liver

hyb_protocol: Labeled cRNA was hybridized to the Illumina mouse REF8 v1.1 Expression BeadChip using standard protocols (Illumina).

scan_protocol: Standard Illumina scanning protocol

data_processing: The probe profile output from the Illumina BeadStudio toolkit was read into R using the lumi BioC package. Then, the data was VST transformed (Lin, S.M., Du, P., Huber, W., and Kibbe, W.A. 2008. Model-based variance-stabilizing transformation for Illumina microarray data. Nucleic Acids Res 36:e11) followed by quantile normalization. The versions of software and BioC packages were as follows: R_2.8.0, lumi_1.8.2.

source_name: mouse liver

label_protocol: 500 ng of total RNA was reverse transcribed into cRNA and biotin-UTP labeled using the Illumina TotalPrep RNA Amplification Kit (Ambion). cRNA was quantified using an Agilent Bioanalyzer 2100

growth_protocol: Individual FXR-/- or SHP-/- mice were backcrossed with C57BL/6 mice to generate congenic C57BL/6 FXR-/- or SHP-/- mice. Congenic single knockout animals were bred with each other to generate FXR/SHP double knockout mice. Age-matched, male, DKO congenic and C57BL/6 wild-type (WT) mice were used throughout this study. WT control mice (C57BL/6) were purchased from Harlan Sprague Dawley, Inc. (Indianapolis, IN). Mice were sacrificed at 3 or 5 weeks after birth. Mice were housed on a standard 12-hour light/dark cycle and were fed normal chow and water ad libitum.

extract_protocol: Total RNA was prepared according to manufacturer’s protocol. (Trizol, Invitrogen). The RNA obtained was further purified using qiagen columns.

tag: age

value: 5 weeks

value: male

value: liver

tag: genome/variation

value: FXR-/-,SHP-/- double knock out


title: E0-4_H3K27Ac_1 extraction1_array2

description: channel ch1 is ChIP DNA; Antibody information listed below: official name: H3K27Ac;target name: H3K27Ac;host: Rabbit;antigen: Synthetic peptide conjugated to KLH derived from within residues 1 - 100 of Human Histone H3 acetylated at K27.;clonal: Polyclonal;purified: Affinity;company: Abcam;catalog: ab4729;short description: Histone H3 (acetyl K27) antibody - ChIP Grade;


title: E0-4_H3K27Ac_3 extraction3_array2


title: breast_cancer_T4

first: Kornelia

last: Polyak

address: 450 Brookline Ave

department: Medical Oncology

institute: Dana-Farber Cancer Institute

hyb_protocol: For each individual array, 200 ng of Cy3-labeled normal or tumor sample was mixed with 200 ng of Cy5-labeled reference sample and twocolor hybridization was performed according to the manufacturer's protocol (Agilent Technologies).

data_processing: Image analysis was conducted using Agilent Feature extraction software (version 10.5.1.1; Agilent Technologies), and processed signals and LogRatio were used for further analysis. To confirm the presence of antisense transcripts, a rigorous threshold was applied using the flag IsWellAboveBG.

molecule: polyA RNA

source_name: biopsy and surgery

label_protocol: Two hundred nanograms of mRNA was directly labeled with Cy3 using a ULS micro-RNA labeling kit (Kreatech) and fragmented with an RNA fragmentation kit (Ambion). For the reference sample, equal amounts of mRNA from the four normal samples were mixed together, labeled with Cy5.

extract_protocol: polyA RNA were purified from four (N1 to N4) normal organoid and four breast tumor samples as described previously (PMID: 17349583).

value: breast

value: cancer

source_name: pooled polyA RNA from four normal organids from reduction mammoplasty

value: normal


title: breast_cancer_T5

scan_protocol: Image analysis was conducted using Agilent Feature extraction software (version 10.5.1.1; Agilent Technologies)


title: SHPko.5wk.rep1

description: Gene expression in 5wk old SHP-/- mouse liver

tag: gender

value: SHP-/-


title: FXRko.5wk.rep2

description: Gene expression in 5wk old FXR-/- mouse liver

value: FXR-/-


title: FXRSHPko.5wk.rep1


title: S. salar_47_dpf_rep1

description: Biological replicate 1 of 3

source_name: Whole tissue, 47 days post fertilization

value: whole organism

value: 47 days post fertilization

value: Pooled cRNA from whole tissues in all time points


title: SHPko.5wk.rep2


title: breast_cancer_T2


title: MA6


title: MA2


title: breast_cancer_T3


title: FXRko.5wk.rep1


title: E12-16_PolII_1 extraction1_array3

extract_protocol:
Collection of Drosophila embryos at time-point 12-16h of development for further ChIP experiments  or RNA extraction.
Formaldehyde Cross-linking of Chromatin from any Drosophila tissue (Embryos, Larvae, Pupae, Adult flies).1. Homogenize the material (about 150-200 mg of embryos, pupae, adµlts, larvae, sufficient for four-five independent immuno-precipitations) in 5 ml of buffer A1 + 1.8% formaldehyde (290 µl of 37% solution) at room temperature using first a Potter homogenizer and then a Douncer with type A pestle (3 strokes). Wait 15? (total time starting from beginning of homogenization). Add glycin solution to 225 mM (540 µl of 2.5 M solution for 6 ml of cross-linked mixture), mix and incubate 5?. Put on ice. Keep on ice unless specified.2. Transfer the homogenate to a 15 ml tube. Centrifuge 5 min, at 4000 g at 4°C. Discard supernatant. Add 3 ml of buffer A1, resuspend pellet and spin down the same way. Repeat washing step 3 times.3. Wash once in 3 ml of lysis buffer without SDS. Spin down, 5 min at 4000g.4. Resuspend cross-linked material in 0.5 ml of lysis buffer and add SDS to 0.1% (2.5 µl) and N-lauroylsarcosine to 0.5% (12.5 µl). Incubate 10 min at 4°C in a rotating wheel.5. Sonicate in Polypropylene 15ml Falcon tubes according to .Diagenode instructions. The settings are: High power, 15 minutes, 30 seconds off, 30 seconds on. The water bath shoµld be kept cold either in a cold room or by a cold water pump.6. Rotate 10'. Transfer to eppendorf tubes and centrifuge 5', room temperature at maximum speed. Transfer supernatant to the new tube. Add another 0.5 ml of lysis buffer to the pellet, rotate for 10 min. Repeat centrifugation and combine supernatants. Centrifuge the combined supernatant 2x10 min at max speed. The supernatant from this stage will be referred as chromatin extract. Chromatin can be stored at ?80°C for several months at this stage. Add sodium azide to 0.02% for storage.7. [Optional : Purification of chromatin. Put chromatin extract in Centricon YM-100 column (4212), centrifuge 3x40' (or more) at 1000 g while adding lysis buffer. At least 3 vol. of lysis buffer should pass through column. Bring the final volume of chromatin extract to 1 ml by lysis buffer + protease inhibitors.][edit]
Solutions and MaterialsLysis buffer: (see above)5mg/ml RNaseA (DNase free)PAS suspension: 100 mg of CL-4B (Amersham, 17-0780-01) PAS shoµld be resuspended in 1 ml of lysis buffer + protease inhibitors + 0.1 mg/ml BSA for 50 %v suspension. Wash in lysis buffer 2-3 times, overall equilibration time 1h. Store up to one week at 4 C.TE: 10 mM Tris-HCl pH 8.0, 1 mM EDTAElution buffer1: 10 mM EDTA, 1% SDS, 50 mM Tris-Cl pH 8.Elution buffer2: TE+0.67% SDS.LiCL 4M? Protocol 1. To an amount of chromatin corresponding to 150 mg of biological material, suspended in a final volume of chromatin extract of 1 ml (in lysis buffer + protease inhibitors), add 100 µl of PAS suspension for preincubation. Incubate several hours or overnight at 4°C, then remove PAS. Crosslinked chromatin at this stage can be stored several days at 4°C or frozen at ?70°C.2. [Optional : Check for amount of DNA. From the 1 ml solution above, take a 100 µl aliquot, Add proteinase K up to 100 ug/ml and SDS to 1%, incubate 6h at 60°C, then 20 min at 70°C, add RNAse to 50 ug/ml and incubate additional 2h at 37°C. Phenol-chlorophorm extract, ethanol precipitate. Run on an agarose gel to check amount and size of DNA.]3. Separate chromatin sample into 4x250 µl aliquots (one aliquot is enough for one IP but the amount may be decreased further). Immunoprecipitate chromatin by adding the antibody (Ab) of interest (amount of Ab shoµld be determined empirically, usually the same concentration than for a successfµl IF experiement). Keep a control sample without Ab IP, named "Mock".4. Incubate 4 h at 4°C on a head to head rotating wheel, then add 50 µl of PAS suspension and incubate 4 h or overnight. Spin down PAS and proceed to washes.5. Wash PAS 4x with lysis buffer, followed by 2x with TE (without protease inhibitors). Each wash is for 5 min at 4°C, using 1 ml of solution.6. Elution of precipitated material. Spin down PAS. Add 100 µl of elution buffer, mix and incubate 10 min at 65°C. Spin down PAS and transfer supernatant to new tube. Add 150 µl of elution buffer 2 to PAS, mix, centrifuge at fµll speed and transfer eluate to a tube together with the eluate from the first centrifugation. The combined material is the "chromatin precipitate" (approx. 250 µl).7. Incubate precipitate 6 h (or overnight) at 65 C to reverse cross-links. Add 250 µl of Proteinase K solution, incubate at 50°C 2-3 h.8. Add 55 µl of 4M LiCl and 500 µl of phenol-chlorophorm. Mix and microfuge at fµll speed at RT. Transfer aqueous phase to a new tube and precipitate with 1 ml of 100% ethanol. Wash with 750 µl of 70% ethanol. Spin down and dry precipitate.9. Dissolve in 25 µl of water. This is the chromatin immunoprecipitate or ?Native ChIP? sample.
LM-PCR amplification of the native ChIP sample to obtain a sufficient quantity for further hybridization on microarrays.


title: E12-16_PolII_3 extraction3_array3

description: channel ch1 is ChIP DNA; Antibody information listed below: official name: PolII;target name: RNA Polymerase II;host: Mouse;antigen: The antibody 8WG16 was developed using RNA polymerase (RNA pol II) purified from wheat germ extract. It recognizes the C-terminal heptapeptide repeat present on the largest subunit of pol II. Clone 8WG16 has been used to investigate the role of the C-terminal domain of the 220kD RNA pol II subunit in the formation of the transcription complex.;clonal: Polyclonal;purified: Affinity;company: Covance;catalog: 8wg16;short description: Monoclonal Antibody against RNA Polymerase II;


title: MA10


title: MA4


title: E12-16_PolII_2 extraction2_array3


title: E8-12_CBP_3 extraction3_array2

description: channel ch1 is ChIP DNA; Antibody information listed below: official name: CBP antibody;target name: nejire;host: Mouse;

extract_protocol:
Collection of Drosophila embryos at time-point 12-16h of development for further ChIP experiments or RNA extraction.
Formaldehyde Cross-linking of Chromatin from any Drosophila tissue (Embryos, Larvae, Pupae, Adult flies).1. Homogenize the material (about 150-200 mg of embryos, pupae, adµlts, larvae, sufficient for four-five independent immuno-precipitations) in 5 ml of buffer A1 + 1.8% formaldehyde (290 µl of 37% solution) at room temperature using first a Potter homogenizer and then a Douncer with type A pestle (3 strokes). Wait 15? (total time starting from beginning of homogenization). Add glycin solution to 225 mM (540 µl of 2.5 M solution for 6 ml of cross-linked mixture), mix and incubate 5?. Put on ice. Keep on ice unless specified.2. Transfer the homogenate to a 15 ml tube. Centrifuge 5 min, at 4000 g at 4°C. Discard supernatant. Add 3 ml of buffer A1, resuspend pellet and spin down the same way. Repeat washing step 3 times.3. Wash once in 3 ml of lysis buffer without SDS. Spin down, 5 min at 4000g.4. Resuspend cross-linked material in 0.5 ml of lysis buffer and add SDS to 0.1% (2.5 µl) and N-lauroylsarcosine to 0.5% (12.5 µl). Incubate 10 min at 4°C in a rotating wheel.5. Sonicate in Polypropylene 15ml Falcon tubes according to .Diagenode instructions. The settings are: High power, 15 minutes, 30 seconds off, 30 seconds on. The water bath shoµld be kept cold either in a cold room or by a cold water pump.6. Rotate 10'. Transfer to eppendorf tubes and centrifuge 5', room temperature at maximum speed. Transfer supernatant to the new tube. Add another 0.5 ml of lysis buffer to the pellet, rotate for 10 min. Repeat centrifugation and combine supernatants. Centrifuge the combined supernatant 2x10 min at max speed. The supernatant from this stage will be referred as chromatin extract. Chromatin can be stored at ?80°C for several months at this stage. Add sodium azide to 0.02% for storage.7. [Optional : Purification of chromatin. Put chromatin extract in Centricon YM-100 column (4212), centrifuge 3x40' (or more) at 1000 g while adding lysis buffer. At least 3 vol. of lysis buffer should pass through column. Bring the final volume of chromatin extract to 1 ml by lysis buffer + protease inhibitors.][edit]
Solutions and MaterialsLysis buffer: (see above)5mg/ml RNaseA (DNase free)PAS suspension: 100 mg of CL-4B (Amersham, 17-0780-01) PAS shoµld be resuspended in 1 ml of lysis buffer + protease inhibitors + 0.1 mg/ml BSA for 50 %v suspension. Wash in lysis buffer 2-3 times, overall equilibration time 1h. Store up to one week at 4 C.TE: 10 mM Tris-HCl pH 8.0, 1 mM EDTAElution buffer1: 10 mM EDTA, 1% SDS, 50 mM Tris-Cl pH 8.Elution buffer2: TE+0.67% SDS.LiCL 4M? Protocol 1. To an amount of chromatin corresponding to 150 mg of biological material, suspended in a final volume of chromatin extract of 1 ml (in lysis buffer + protease inhibitors), add 100 µl of PAS suspension for preincubation. Incubate several hours or overnight at 4°C, then remove PAS. Crosslinked chromatin at this stage can be stored several days at 4°C or frozen at ?70°C.2. [Optional : Check for amount of DNA. From the 1 ml solution above, take a 100 µl aliquot, Add proteinase K up to 100 ug/ml and SDS to 1%, incubate 6h at 60°C, then 20 min at 70°C, add RNAse to 50 ug/ml and incubate additional 2h at 37°C. Phenol-chlorophorm extract, ethanol precipitate. Run on an agarose gel to check amount and size of DNA.]3. Separate chromatin sample into 4x250 µl aliquots (one aliquot is enough for one IP but the amount may be decreased further). Immunoprecipitate chromatin by adding the antibody (Ab) of interest (amount of Ab shoµld be determined empirically, usually the same concentration than for a successfµl IF experiement). Keep a control sample without Ab IP, named "Mock".4. Incubate 4 h at 4°C on a head to head rotating wheel, then add 50 µl of PAS suspension and incubate 4 h or overnight. Spin down PAS and proceed to washes.5. Wash PAS 4x with lysis buffer, followed by 2x with TE (without protease inhibitors). Each wash is for 5 min at 4°C, using 1 ml of solution.6. Elution of precipitated material. Spin down PAS. Add 100 µl of elution buffer, mix and incubate 10 min at 65°C. Spin down PAS and transfer supernatant to new tube. Add 150 µl of elution buffer 2 to PAS, mix, centrifuge at fµll speed and transfer eluate to a tube together with the eluate from the first centrifugation. The combined material is the "chromatin precipitate" (approx. 250 µl).7. Incubate precipitate 6 h (or overnight) at 65 C to reverse cross-links. Add 250 µl of Proteinase K solution, incubate at 50°C 2-3 h.8. Add 55 µl of 4M LiCl and 500 µl of phenol-chlorophorm. Mix and microfuge at fµll speed at RT. Transfer aqueous phase to a new tube and precipitate with 1 ml of 100% ethanol. Wash with 750 µl of 70% ethanol. Spin down and dry precipitate.9. Dissolve in 25 µl of water. This is the chromatin immunoprecipitate or ?Native ChIP? sample.
LM-PCR amplification of the native ChIP sample to obtain a sufficient quantity for further hybridization on microarrays.


title: MA1


title: WT.5wk.rep2

description: Gene expression in 5wk old wild type mouse liver

value: wild type


title: E8-12_CBP_2 extraction2_array2