title: 1419_natural_infection_vs_1419_1x10^6_rep1

status: Public on May 20 2010

city: St Lucia

first: Ylva

last: Strandberg-Lutzow

state: Queensland

address: 306 Carmody Road

department: Livestock Industries

institute: CSIRO

description: Bovine mammary tissue

hyb_protocol: Hybridisation was performed at 44°C in a 50% formamide hybridisation buffer for 16 h. Following hybridisation, three washes were applied to the arrays: 2 x SSC, 0.1% SDS for 15 min, pre-warmed to 44°C; 0.2 x SSC for 15 min; and 0.06 x SSC for 5 min. Slides were washed once in high purity water to remove any remaining salt and dried in a centrifuge at 40xg for 5 min.

scan_protocol: Dried slides were scanned immediately using the GenePix. 4000 array scanner. GenePix.Pro software version 5.0 (Axon Instruments Inc.) was then used to process array images, align spots, integrate robot-spotting files with the microarray image, and export reports of spot intensity data. Slides were visually examined and spots with irregular morphology were excluded from data analysis

data_processing: After background correction and removal of flagged values, log base 2 expression ratios were mean centered and linear transformed to obtain the log and linear values given in the data table

organism: Bos taurus

source_name: Right Fore quarter Field mastitis mammary tissue

label_protocol: cDNA prepared from each sample using 20 ug of total RNA per dye channel per microarray, was reverse transcribed with Superscript III (Invitrogen) in the presence of 2-aminoallyl-dUTP (Sigma Chemical Co.) using both oligo-dT18 (2 .g) and pd(N)6 random hexamer (1 .g) (Amersham Bioscience, UK) to prime cDNA synthesis. First strand cDNAs were purified and subsequently labelled using n-hydroxysuccinate (NHS)-derivatized Cy3 and Cy5 dyes (Amersham Biosciences, UK)

extract_protocol: Approximately 3 g of frozen mammary tissue was placed in liquid nitrogen and pulverised with a hammer followed by the addition of 25 ml of Trizol reagent (Invitrogen). The tissue was homogenised using an Ultraturrex probe (IKA, 3 cycles of 2 seconds). RNA was then extracted from three equivalent samples from each udder quarter according to the Trizol manufacturer.s protocol (Invitrogen) and 1 mg aliquots were stored as pellets under ethanol at -80.C. RNA was further purified using an RNeasy Midi Kit (Qiagen) and each sample was treated twice with DNase I on column (Qiagen DNase I) and after elution (Ambion DNase I) to eliminate any contaminating genomic DNA.

tag: Natural Infection

source_name: Right Hind quarter infused with 1x10^6 Staphylococcus aureus mammary tissue

tag: Dose


title: 1419_natural_infection_vs_1419_1x10^5_rep2

type: RNA

country: Australia

description: Bovine mammary tissue

molecule: total RNA

source_name: Left Hind quarter infused with 1x10^5 Staphylococcus aureus mammary tissue


title: 1419_1x10^5_vs_1419_1x10^6_rep1

description: Bovine mammary tissue


title: shee217 | normal karyotype

status: Public on Jul 11 2010

city: Berlin

first: Lars

last: Bullinger

address: Augustenburger Platz 1

department: Hematology, Oncology and Tumorimmunology

country: Germany

institute: Charité

description:
Simple annotation: Myeloid Leukemias, Acute Myeloid Leukemia
Image: http://smd.stanford.edu/MicroArray/gifs/2003-08/44115.gif

hyb_protocol: not provided

scan_protocol: GenePix 4000B

data_processing: VALUE is Log (base 2) of the ratio of the median of Channel 2 (usually 635 nm) to Channel 1 (usually 532 nm)

organism: Homo sapiens

source_name: Stratagene common reference

extract_protocol: not provided

tag: reference

value: Stratagene common reference

label_protocol: RNA from AML specimens was labeled with Cy5-dUTP using an anchored poly(dT) primer during first strand reverse transcription (Superscript II).  A universal reference RNA (pooled from 11 cell lines) was labeled identically but with Cy3-dUTP

extract_protocol: Normal karyotype acute myeloid leukemia (NK-AML) specimens comprise Ficoll gradient purified mononuclear cells (>80% blasts) from diagnostic peripheral blood or bone marrow.  Total RNA was isolated from stored, frozen mononuclear AML-cell pellets using Trizol reagent, and RNA quality assessed by gel electrophoresis.; Protocol Type = Extract preparation

tag: source

tag: age

tag: overall survival time

value: 1282 days

tag: overall survival status

value: alive

tag: genotype

value: normal karyotype

tag: npm1 mutation status

value: WT

tag: gender

value: female

tag: runx1 mutation


title: 1419_control_vs_1419_1x10^5_rep1

description: Bovine mammary tissue

source_name: Control Left Fore quarter mammary tissue

value: control


title: 1419_control_vs_1419_natural_infection_rep1

description: Bovine mammary tissue


title: 1419_natural_infection_vs_1419_1x10^5_rep1

description: Bovine mammary tissue


title: 1419_control_vs_1419_1x10^5_rep2

description: Bovine mammary tissue


title: 1419_control_vs_1419_1x10^6_rep1

description: Bovine mammary tissue


title: 1592_1x10^4_vs_1592_1x10^6_rep1

description: Bovine mammary tissue

source_name: Right Fore quarter infused with 1x10^4 Staphylococcus aureus mammary tissue


title: 1419_control_vs_1419_natural_infection_rep2

description: Bovine mammary tissue


title: 1419_control_vs_1419_1x10^6_rep2

description: Bovine mammary tissue


title: 1419_1x10^5_vs_1419_1x10^6_rep2

description: Bovine mammary tissue


title: 1419_natural_infection_vs_1419_1x10^6_rep2

description: Bovine mammary tissue


title: 1592_1x10^4_vs_1592_1x10^5_rep2

description: Bovine mammary tissue


title: 1592_control_vs_1592_1x10^5_rep1

description: Bovine mammary tissue


title: 1592_1x10^4_vs_1592_1x10^5_rep1

description: Bovine mammary tissue


title: 1592_control_vs_1592_1x10^4_rep1

description: Bovine mammary tissue


title: 1592_control_vs_1592_1x10^6_rep1

description: Bovine mammary tissue


title: 1592_control_vs_1592_1x10^6_rep2

description: Bovine mammary tissue


title: 1592_1x10^5_vs_1592_1x10^6_rep2

description: Bovine mammary tissue


title: 1592_control_vs_1592_1x10^4_rep2

description: Bovine mammary tissue


title: 1592_1x10^4_vs_1592_1x10^6_rep2

description: Bovine mammary tissue


title: Sporadic - C017

status: Public on Jul 05 2010

type: genomic

city: Hamburg

first: Simon

middle: A

last: Joosse

address: Martinistrasse 52

department: Institute of Tumor Biology

institute: University Medical Center Hamburg-Eppendorf

hyb_protocol: As described in: Automated array-CGH optimized for archival formalin-fixed, paraffin-embedded tumor material. Joosse SA, et al. BMC Cancer. 2007 Mar 7;7:43.

molecule: genomic DNA

extract_protocol:
As decribed in:
Prediction of BRCA1-association in hereditary non-BRCA1/2 breast carcinomas with array-CGH.
Joosse SA, et al.
Breast Cancer Res Treat. 2008 Aug 14

tag: organ

value: mammary gland

tag: disease

value: infiltrating ductal carcinoma

tag: type

value: sporadic breast tumor

source_name: Women reference pool

tag: sample

value: A pool of DNA from peripheral blood lymphocytes, from 7 apparently healthy women.

tag: cell type

value: peripheral blood lymphocytes

value: healthy appearing


title: 1592_control_vs_1592_1x10^5_rep2

description: Bovine mammary tissue


title: 1490_control_vs_1490_5x10^2_rep2

description: Bovine mammary tissue

label_protocol: cDNA prepared from each sample using 20 ug of total RNA per dye channel per microarray, was reverse transcribed with Superscript III (Invitrogen) in the presence of 2-aminoallyl-dUTP (Sigma Chemical Co.) using both oligo-dT18 (2 .g) and pd(N)6 random hexamer (1 .g) (Amersham Bioscience, UK) to prime cDNA synthesis. First strand cDNAs were purified and subsequently labelled using n-hydroxysuccinate (NHS)-derivatized Cy3 and Cy5 dyes (Amersham Biosciences, UK).

extract_protocol: Approximately 3 g of frozen mammary tissue was placed in liquid nitrogen and pulverised with a hammer followed by the addition of 25 ml of Trizol reagent (Invitrogen). The tissue was homogenised using an Ultraturrex probe (IKA, 3 cycles of 2 seconds). RNA was then extracted from three equivalent samples from each udder quarter according to the Trizol manufacturer.s protocol (Invitrogen) and 1 mg aliquots were stored as pellets under ethanol at -80.C. RNA was further purified using an RNeasy Midi Kit (Qiagen) and each sample was treated twice with DNase I on column (Qiagen DNase I) and after elution (Ambion DNase I) to eliminate any contaminating genomic DNA..

source_name: Right Fore quarter infused with 5x10^2 Staphylococcus aureus mammary tissue


title: 1592_1x10^5_vs_1592_1x10^6_rep1

description: Bovine mammary tissue


title: 1490_control_vs_1490_1x10^4_rep2

description: Bovine mammary tissue

source_name: Left Hind quarter infused with 1x10^4 Staphylococcus aureus mammary tissue


title: 1490_5x10^2_vs_1490_1x10^4_rep2

description: Bovine mammary tissue


title: 1490_1x10^4_vs_1490_1x10^5_rep1

description: Bovine mammary tissue

source_name: Right Hind quarter infused with 1x10^5 Staphylococcus aureus mammary tissue


title: 1490_5x10^2_vs_1490_1x10^4_rep1

description: Bovine mammary tissue


title: 1490_5x10^2_vs_1490_1x10^5_rep1

description: Bovine mammary tissue


title: 1490_control_vs_1490_1x10^5_rep1

description: Bovine mammary tissue


title: 1490_5x10^2_vs_1490_1x10^5_rep2

description: Bovine mammary tissue


title: 1490_control_vs_1490_1x10^4_rep1

description: Bovine mammary tissue


title: 1490_1x10^4_vs_1490_1x10^5_rep2

description: Bovine mammary tissue


title: BC136

status: Public on May 05 2010

city: Marseille

first: Pascal

last: FINETTI

state: BdR

address: 232 Bd Ste Marguerite

department: Centre de cancérologie de Marseille

country: France

institute: Institut Paoli-Calmettes

hyb_protocol: Affymetrix standard protocol

scan_protocol: Affymetrix standard protocol

data_processing: RMA using a quantile normalization

label: biotin

source_name: human breast cancer tumors

label_protocol: Affymetrix standard protocol

extract_protocol: Guanidinium isothiocyanate and centrifugation over a cesium chloride gradient.

tag: age at diagnosis

tag: histology

value: IDC

tag: pt

tag: pn

tag: sbr grade

tag: er ihc

tag: pr ihc

tag: p53 ihc

tag: ki67 ihc

tag: dfs evt

tag: dfs time (months)

tag: molecular subtype

value: LuminalA

tag: tissue

value: breast cancer tumor


title: 1490_control_vs_1490_1x10^5_rep2

description: Bovine mammary tissue

tag: Dose


status: Public on May 11 2010

city: Sendai, Aoba-ku

first: Yukako

last: Oma

address: Tsutsumidori-Amamiyamachi 1-1

country: Japan

institute: Tohoku University

description:
ChIP-chip data used for the mapping of Swr1-FLAG distribution on chromosome III-V and VI right arm.
Used baseline: WCE fraction (Swr1-FLAG distribution on chromosome III-V and VI right arm)

hyb_protocol: Following denaturation at 100 C for 10 minutes, cooling, and spin-down, 150 micro liter of hybridization mixuter was hybridized for 16 hours at 42 C on GeneChip rikDACF using hybridization oven 641 (affymetrix). Each hybridization mixture contains the end-labbeled DNA, 6xSSPE, 0.005% Triton X-100, 0.1 mg/ml herring sperm DNA, oligo B2 (affymetrix) and hybridization control (affymetrix).

scan_protocol: Washing and scanning protocol (mini-WS1v2) provided by Affymetrix was performed automatically on a fluidics station (GeneChip fluidics station 400, Affymetrix). Arrays were scanned by Hewlett-Packard GeneArray Scanner at an emission wavelength of 560 nm at 7.5 microM resolution.

data_processing:
Data analyses were performed as previously described in Methods in Enzymology, Elsevier Life Sciences (CA), vol.409,chapter 23,389-410 in “DNA Repair” (edited by J.Campbell) (2007).
Primary data analyses were carried out with GeneChip Operating Software (GCOS 1.4) using default settings to obtain signal and detection p-value.
To discriminate significant signals for binding to DNA, signals from the ChIP fraction scan were compared to the WCE fraction scan with GCOS 1.4 using default settings.

organism: Saccharomyces cerevisiae

source_name: Chromatin-immunoprecipitated DNA

label_protocol: End labeling by terminal transferase

growth_protocol: The cells were grown to a density of 1.0x10e7 cells/ml at 30C in 200 ml YPD media.

extract_protocol:
The cells were disrupted using Multi-Beads Shocker (YASUI KIKAI, Osaka) with 0.5-mm glass beads. Whole cell extract was sonicated to obtain average 500 bp genomic DNA fragments.
The WCE was immunoprecipitated with anti-FLAG monoclonal antibody M2 (Sigma-Aldrich Co., St Louis, MO) coupled to Dynabeads (Invitrogen, protein A Dynabeads).
The chromatin immunoprecipitated sample was incubated overnight at 37C with proteinase K, incubated for 6 hrs at 65C, extracted with phenol/chloroform/isoamylalcohol, precipitated, and resuspended in TE and incubated with RNaseA.
The DNA was amplified by PCR after random priming as previously described (Katou et al., 2003, nature). The amplified DNA was digested with Dnase I to a mean size of 100 bp. After DNase I inactivation at 95ºC, fragments were labelled with indicated label protocol.

treatment_protocol: The culture was fixed in 1% formaldehyde for 10 min at room temperature and then treated with 125 mM glycine for 5 min at room temperature. The cells were washed with 40 ml of ice-cold TBS twice, freezed with liquid nitrogen, and stored at -80C.

tag: W303 background strain expressing FLAG tagged Swr1 protein.

tag: Strain

tag: Culture

value: grown to log phase.


title: WCE fraction (Swr1-FLAG cells, chromosome III-V and VI right arm.)

description:
WCE fraction used for the nomalization of ChIP fraction
Baseline of Swr1-FLAG distribution on chromosome III-V and VI right arm.

hyb_protocol: Following denaturation at 100 C for 10 minutes, cooling, and spin-down, 150 micro liter of hybridization mixuter was hybridized for 16 hours at 42 C on GeneChip rikDACF using hybridization oven 644 (affymetrix). Each hybridization mixture contains the end-labbeled DNA, 6xSSPE, 0.005% Triton X-100, 0.1 mg/ml herring sperm DNA, oligo B2 (affymetrix) and hybridization control (affymetrix).

data_processing:
Data analyses were performed as previously described in Methods in Enzymology, Elsevier Life Sciences (CA), vol.409,chapter 23,389-410 in “DNA Repair” (edited by J.Campbell) (2010).
Primary data analyses were carried out with GeneChip Operating Software (GCOS 1.4) using default settings to obtain signal and detection p-value.

source_name: Fragmented whole genomic DNA

extract_protocol:
The cells were disrupted using Multi-Beads Shocker (YASUI KIKAI, Osaka) with 0.5-mm glass beads. Whole cell extract was sonicated to obtain average 500 bp genomic DNA fragments.
0.7% of the WCE was used for a preparation of WCE fraction (input).
The WCE fraction were incubated overnight at 37C with proteinase K, incubated for 6 hours at 65C, extracted with phenol/chloroform/isoamylalcohol, precipitated, and resuspended in TE and incubated with RNaseA.
The DNA was amplified by PCR after random priming as previously described (Katou et al., 2003, nature). The amplified DNA was digested with Dnase I to a mean size of 100 bp. After DNase I inactivation at 95ºC, fragments were labelled with indicated label protocol.


title: BC140

tag: age at diagnosis

value: ILC


title: BC145

tag: histology

value: Normal


title: BC137

tag: pt


title: 1490_control_vs_1490_5x10^2_rep1

description: Bovine mammary tissue


description:
ChIP-chip data used for the mapping of Arp6-FLAG distribution on chromosome III-V and VI right arm.
Used baseline: WCE fraction (Arp6-FLAG distribution on chromosome III-V and VI right arm)

hyb_protocol: Following denaturation at 100 C for 10 minutes, cooling, and spin-down, 150 micro liter of hybridization mixuter was hybridized for 16 hours at 42 C on GeneChip rikDACF using hybridization oven 640 (affymetrix). Each hybridization mixture contains the end-labbeled DNA, 6xSSPE, 0.005% Triton X-100, 0.1 mg/ml herring sperm DNA, oligo B2 (affymetrix) and hybridization control (affymetrix).

data_processing:
Data analyses were performed as previously described in Methods in Enzymology, Elsevier Life Sciences (CA), vol.409,chapter 23,389-410 in “DNA Repair” (edited by J.Campbell) (2006).
Primary data analyses were carried out with GeneChip Operating Software (GCOS 1.4) using default settings to obtain signal and detection p-value.
To discriminate significant signals for binding to DNA, signals from the ChIP fraction scan were compared to the WCE fraction scan with GCOS 1.4 using default settings.

tag: W303 background strain expressing FLAG tagged Arp6 protein.


title: BC126

tag: pn

tag: sbr grade


title: WCE fraction (Arp6-FLAG cells, chromosome III-V and VI right arm.)

description:
WCE fraction used for the nomalization of ChIP fraction
Baseline of Arp6-FLAG distribution on chromosome III-V and VI right arm.

hyb_protocol: Following denaturation at 100 C for 10 minutes, cooling, and spin-down, 150 micro liter of hybridization mixuter was hybridized for 16 hours at 42 C on GeneChip rikDACF using hybridization oven 643 (affymetrix). Each hybridization mixture contains the end-labbeled DNA, 6xSSPE, 0.005% Triton X-100, 0.1 mg/ml herring sperm DNA, oligo B2 (affymetrix) and hybridization control (affymetrix).

data_processing:
Data analyses were performed as previously described in Methods in Enzymology, Elsevier Life Sciences (CA), vol.409,chapter 23,389-410 in “DNA Repair” (edited by J.Campbell) (2009).
Primary data analyses were carried out with GeneChip Operating Software (GCOS 1.4) using default settings to obtain signal and detection p-value.


title: BC134

tag: er ihc

tag: pr ihc


title: WCE fraction (Arp6-FLAG swr1 deletion cells, chromosome III-V and VI right arm.)

description:
WCE fraction used for the nomalization of ChIP fraction
Baseline of Arp6-FLAG distribution on chromosome III-V and VI right arm in swr1 delta cells.

hyb_protocol: Following denaturation at 100 C for 10 minutes, cooling, and spin-down, 150 micro liter of hybridization mixuter was hybridized for 16 hours at 42 C on GeneChip rikDACF using hybridization oven 645 (affymetrix). Each hybridization mixture contains the end-labbeled DNA, 6xSSPE, 0.005% Triton X-100, 0.1 mg/ml herring sperm DNA, oligo B2 (affymetrix) and hybridization control (affymetrix).

data_processing:
Data analyses were performed as previously described in Methods in Enzymology, Elsevier Life Sciences (CA), vol.409,chapter 23,389-410 in “DNA Repair” (edited by J.Campbell) (2011).
Primary data analyses were carried out with GeneChip Operating Software (GCOS 1.4) using default settings to obtain signal and detection p-value.

tag: W303 background strain (swr1 delta) expressing FLAG tagged Arp6 protein.


title: Arp6-FLAG distribution on chromosome III-V and VI right arm in swr1 mutant.

description:
ChIP-chip data used for the mapping of Arp6-FLAG distribution on chromosome III-V and VI right arm in swr1 delta cells.
Used baseline: WCE fraction (Arp6-FLAG distribution on chromosome III-V and VI right arm in swr1 delta cells)

hyb_protocol: Following denaturation at 100 C for 10 minutes, cooling, and spin-down, 150 micro liter of hybridization mixuter was hybridized for 16 hours at 42 C on GeneChip rikDACF using hybridization oven 642 (affymetrix). Each hybridization mixture contains the end-labbeled DNA, 6xSSPE, 0.005% Triton X-100, 0.1 mg/ml herring sperm DNA, oligo B2 (affymetrix) and hybridization control (affymetrix).

data_processing:
Primary data analyses were carried out with GeneChip Operating Software (GCOS 1.4) using default settings to obtain signal and detection p-value.
To discriminate significant signals for binding to DNA, signals from the ChIP fraction scan were compared to the WCE fraction scan with GCOS 1.4 using default settings.


title: Sporadic - C032

hyb_protocol: As described in: Joosse SA, van Beers EH, Nederlof PM. Automated array-CGH optimized for archival formalin-fixed, paraffin-embedded tumor material. BMC Cancer. 2007 Mar 7;7:43.

label_protocol: According to the manufacturer's instructions (Kreatech Biotechnology, Amsterdam).

extract_protocol: DNA is extracted as described in: Joosse et al. (submitted)

tag: organ

value: mammary gland; breast

tag: disease

tag: sample

tag: cell type


title: BC123

tag: p53 ihc

tag: ki67 ihc

tag: dfs evt

tag: dfs time (months)

tag: molecular subtype

tag: tissue


title: BC120

tag: age at diagnosis

tag: histology

tag: pt


title: BC118

value: TUB

tag: pn

tag: sbr grade


title: BC115

tag: er ihc

tag: pr ihc


title: BC110


title: BC111


title: BC100


title: BC105


title: BC106


title: BC91


title: BC90


title: BC97


title: BC96


title: BC103


title: BC84


title: BC104


title: BC83

value: MIX


title: BC81


title: BC77


title: BC82


title: BC80


title: BC73


title: BC78


title: BC61

value: LuminalB


title: BC58


title: BC265

value: Basal


title: BC68


title: BC56


title: BC266


title: BC64


title: BC67


title: BC28


title: BC264


title: BC263


title: BC260

value: other


title: BC261


title: BC262


title: BC32


title: BC40


title: BC254


title: BC256


title: BC259


title: BC257


title: BC255


title: BC258


title: BC250


title: BC251


title: BC245


title: BC247


title: BC249


title: BC248


title: BC243


title: BC237


title: BC236


title: BC238


title: BC222


title: BC241


title: BC227


title: BC253


title: BC229


title: BC230


title: BC223


title: BC224


title: BC239


title: BC225


title: BC244


title: BC228


title: BC246


title: BC234


title: BC231


title: BC242


title: BC252


title: BC235


title: BC232


title: BC226


title: BC233


title: BC240


title: BC219


title: BC216


title: BC217


title: BC215

value: MUC


title: BC218


title: BC201


title: BC211


title: BC213


title: BC220


title: BC209


title: BC202


title: BC204


title: BC212


title: BC205


title: BC197


title: BC208


title: BC206


title: BC210


title: BC198


title: BC214


title: BC207


title: BC200


title: BC193


title: BC192


title: BC196


title: BC195


title: BC203


title: BC221


title: BC199


title: BC194


title: BC188


title: BC183


title: BC190


title: BC189


title: BC185


title: BC182


title: BC186


title: BC179


title: TAX577002 [32K]

status: Public on Apr 14 2010

city: Lund

first: Johan

last: Staaf

address: Medicon Village

country: Sweden

institute: SCIBLU - Swegene Centre for Integrative Biology at Lund University

hyb_protocol: BAC aCGH microarrays were hybridized as described (PMID: 17334996)

scan_protocol: Arrays were washed as described (PMID: 17334996) and fluorescence was recorded using an Agilent G2565AA microarray scanner (Agilent Technologies).

data_processing: TIFF images were analyzed using the Gene Pix Pro software (Axon Instruments, Foster City, CA), and the quantified data matrix was loaded into a local installation of BioArray Software Environment (BASE) (http://base.thep.lu.se). Quantified intensities for tumor sample were stored as channel 1 (ch1) whereas quantified intensities for reference sample were stored as channel 2 (ch2). Median spot pixel values were used to calculate background corrected intensities (Int1 and Int2). Spots were removed and regarded as missing values if either: (1) flagged during image-analysis, (2) background corrected intensity value >64999 signal units, (3) background corrected intensity value <0 signal units, or (4) spot diameter less than 40 um. For spots, log ratio (M) was calculated as log2(Int1/Int2) and average intensity (A) was calculated as log10(Int1*Int2)/2. Normalization was performed by applying popLowess (PMID: 17953745). Spots from dye-swap hybridization were merged using geometric mean of ratios after normalization.

source_name: Breast tumor

label_protocol: Fluorescently labeled DNA targets for hybridization were prepared according to manufacturers' instructions using the Invitrogen Bioprime aCGH Genomic Labeling Module as described (PMID:17334996).

extract_protocol: Genomic DNA from fresh frozen breast tumor tissue was extracted using overnight digestion with proteinase K at 55C, followed by repeated phenol-chloroform purification steps. Genomic DNA from FFPE tissue was extracted according to protocol from http://cancer.ucsf.edu/array/protocols/index.php. DNA concentrations were measured using a Nanodrop.

tag: disease state

value: Breast cancer (HER2+)

tag: er

tag: osbin

tag: os

tag: age

tag: ln

tag: primary

tag: grade

source_name: promega male reference

tag: reference

value: Promega male reference DNA


title: TAX577419 [32K]


title: TAX577354 [32K]


title: TAX577422 [32K]


title: TAX577712 [32K]


title: TAX577715 [38K]


title: BC180

tag: p53 ihc

tag: ki67 ihc

tag: dfs evt

tag: dfs time (months)

tag: molecular subtype

tag: tissue


title: TAX577421 [32K]


title: TAX577700 [32K]


title: TAX577719 [38K]


title: TAX577691 [38K]


title: TAX577065 [32K]


title: TAX577703 [38K]


title: TAX577654 [32K]


title: TAX577423 [32K]


title: TAX577605 [38K]


title: TAX577544 [32K]

description: This case was hybridized as replicates (non-dye-swap).


title: TAX577652 [32K]


title: TAX577705 [38K]


title: TAX577717 [32K]


title: TAX577721 [32K]


title: TAX577277 [33K]


title: TAX577280 [33K]

description: This case was hybridized as dye-swap replicates.

label: Cy3, Cy5

label: Cy5, Cy3


title: TAX577646 [33K]


title: Brucella melitensis 16M vjbR mutant strain

status: Public on Apr 12 2010

city: Namur

first: Jean-Jacques

last: Letesson

address: rue de Bruxelles, 61

department: URBM

country: Belgium

institute: University of Namur

description: Bacterial cultures were made in triplicate. Triplicate RNA samples of the strain were mixed and one chip was made.

hyb_protocol: After labeling, hybridize the sample. Then wash the array before scanning and extracting data. Step 1. Hybridization of Cy-labeled DNA 1.  Set MAUI Hybridization System to 42°C. With the cover closed, allow at least 3 hours for the heat block temperature to stabilize at 42°C. 2.  If the sample was dried to a pellet at the end of labeling (Chapter 3, step 24 on page 13), resuspend in 11.3µl VWR water. Vortex well and spin down to collect contents in bottom of tube. 3.  Using the NimbleGen Hybridization Kit, add the following reagents to the sample:  Hybridization Solution  Volume  Cy3-labeled sample  11.3µl  2X Hybridization Buffer 19.5µl  Hybridization Component A 7.8µl  Cy3 CPK6 48mer Oligo, 50nM 0.4µl  Total 39.0µl 4.  Vortex well (~15 seconds) and then spin to collect the contents in the bottom of the tube. Incubate at 95C for 5 minutes, protected from light. 5.  Vortex, spin, and place the tube in the MAUI 42C sample block and incubate at this temperature until you are ready for sample loading. 6.  Place the MAUI Mixer SL Hybridization Chamber on the array using the provided assembly/disassembly tool and carefully follow MAUI setup instructions. Use the braying tool to remove all air bubbles from the adhesive gasket around the outside of the hybridization chamber. 7.  Place the array and Hybridization Chamber assembly in one of the MAUI bays and allow 30 seconds for the assembly to come to temperature. 8.  Load the sample using the pipette supplied with the MAUI Hybridization Station; follow the manufacturer’s instructions. During loading, a small amount of the sample should flow out of the outlet port. 9.  Confirm there are no bubbles in the chamber.  If there are bubbles, gently massage any bubbles to either end, away from the center of the array. Avoid applying too much pressure because this will force liquid out of the ports. Return the loaded array into one of the MAUI bays. 10.  Let equilibrate for 30 seconds. Wipe off any sample leakage at the ports with a clean tissue. Adhere MAUI stickers to both ports, making sure to press on both stickers simultaneously to seal. 11.  Close the bay clamp and select mix mode B. Hold down the mix button to start mixing. Confirm that the mixing is in progress before closing the cover. 12.  Hybridize the sample overnight (16 to 20 hours).

scan_protocol: This chapter describes the protocol for scanning NimbleGen one-color arrays with a GenePix 4000B Scanner and associated software. 1.  Turn on the scanner using the power switch on the back right side. 2.  Launch the GenePix software 10 minutes before scanning to allow lasers to warm. 3.  Open the scanner door and the slide carriage. 4.  Place the array in the slide carriage so that the array is face down and the barcode end is closest to you. 5.  Move the black lever on the left side of the carriage to the left until the array is lying flat in the carriage. 6.  Release the lever so that the array is gently pushed to the right side of the carriage and held firmly. 7.  Close the carriage (you should hear a click) and slide the scanner door shut. 8.  Open the Hardware Settings dialog box (right side, second icon from the bottom or Alt + Q). Select the following settings for scanning:   Wavelength 532 (Cy3)   PMT Gain = 650   Power (%) = 100   Pixel size (µm) = 5   Lines to average = 1   Focus position (µm) = 0 9.  Under Image on the left side of the screen, select the 532 wavelength. 10.  Perform a preview scan by clicking on the fast-forward icon (or Alt + P). 11.  Stop the scan once the full array is in view by pressing the stop icon or the Esc key. 12.  Switch to zoom mode by clicking on the array image and pressing Z on the keyboard. Click and drag the cursor over the region in which you want to zoom. To restore the prior view, press Alt + Z. Center the entire array image in the viewing window. 13.  Under Tools on the left side of the screen, click the Scan Area icon (or Ctrl + V). Click and drag to define a box that surrounds the array image. The box dimensions should be stretched to create a box just slightly larger than the array image. It is critical to include all of the corner probes within the scan image. 14.  Scan the array by clicking the play icon (or Alt + D). While scanning, set the zoom to view the whole image and adjust the brightness and contrast of the displayed image to eliminate visible saturation. 15.  Zoom the view to a region scanned under the most recent PMT settings and click on the Histogram tab (top of the left side of the screen) to check global intensity of the features.    On the top left side of the screen, make sure the 532 wavelength boxes are checked.   On the left side of the screen, under Y-Axis, make sure that the Log Axis box is checked.   You want the curve to have 1e-5 normalized counts at the 65,000-intensity level (saturation). This means that you have ~1% to 2% of the features saturated.   If the normalized counts at the 65,000-intensity level are less than 1e-5, increase the PMT Gain value (Hardware Settings dialog box).   If the normalized counts at the 65,000-intensity level are more than  1e-5, decrease the PMT Gain value (Hardware Settings dialog box).  The histogram graphs the region of the image viewable on-screen in the image tab. If the histogram is no longer changing, either the scan is complete or area on the image has been scanned. 16.  After the PMT settings are properly adjusted, stop the preview scan and do not save this image. Restart the scan under the new settings. 17.  Save the images using the following naming convention:      NNNNN_XXXXX_532.tif  where NNNNN is the barcode number on the slide and XXXXX is user-defined text. 18.  After scanning, remove the array from the scanner. Store arrays in a dark desiccator in the event that rescanning is necessary.

data_processing:
All of the analysis was performed using the statistical program R (normalisation etc ?), using the package stats (R_Development_Core_Team 2006).
Chip data used were preprocessed using the RMA algorithm (Irizarry, Hobbs et al. 2003), as provided by NimbleGen™ Systems, Inc.
Two pairwise comparisons were performed (∆vjbr-wt) and (∆babr-wt). For each comparison, the foldchange was computed as the ratio of intensity averages (mutant/wt). A Student T test was used as statistical analysis, for over-expression and under-expression.
Genes presenting both a foldchange greater than 1.3 (or below 0.7) and a statistical test significant at the alpha level 0.005 were selected as under/over-expressed between the two conditions.

organism: Brucella melitensis bv. 1 str. 16M

label_protocol:
1.  Prepare the following three solutions:
10X TE (10mM Tris-HCl, 1mM EDTA)
50X dNTP Mix (VWR deionized water  250µl, 10X TE  50µl, 100 mM dGTP  50µl, 100 mM dTTP  50µl, 100 mM dCTP  50µl)
Random 9mer Buffer (VWR deionized water  8.6ml, 1M Tris-HCl  1.25ml, 1M MgCl   125µl, 2-Mercaptoethanol  17.5µl)

2.  Dilute Cy3 dye-labeled 9mers to 1 O.D./42µl Random 9mer Buffer. Aliquot 40µl in 0.2ml thin-walled PCR tubes and store at -20°C.
3.  Assemble the following components in separate 0.2ml thin-walled PCR tubes:
 cDNA 1µg
 Cy3-9mer Primers 40µl
 VWR water To volume
 Total 80µl
4.  Heat-denature samples in a thermocycler at 98C for 10 minutes. Quick-chill in an ice-water bath for 2 minutes.
5.  Prepare the following dNTP/Klenow Master Mix:

50X dNTP mix  10µl
 VWR deionized water   8µl
 Klenow (50U/µl)  2µl
 Total  20µl
6.  Add the dNTP/Klenow Master Mix to the denatured samples from step 4. Keep on ice.

 Reaction volume from step 4    80µl
 dNTP/Klenow Master Mix  20µl
 Total  100µl
7.  Mix well by pipetting 10 times.
8.  Spin down at low RPM to force contents to bottom of tube.
9.  Incubate at 37C for 2 hours (3 hours maximum) in a thermocycler protected from light.
10.  Stop the reaction by addition of 0.5M EDTA.

 Reaction volume from step 6   100µl
 Stop Solution (0.5M EDTA)  10µl
 Total  110µl
11.  Add 5M NaCl to each tube.

Reaction volume from step 10   110µl
 5M NaCl  11.5µl
 Total  121.5µl
12.  Vortex briefly, spin down, and transfer to a 1.5ml tube containing isopropanol.

 Reaction volume from step 11   121.5µl
 Isopropanol  110µl
 Total  231.5µl
13.  Vortex well. Incubate 10 minutes at room temperature, protected from light.
14.  Centrifuge at maximum speed of 12,000 x g for 10 minutes. Remove supernatant with pipette.
15.  Rinse pellet with 500µl 80% ice-cold ethanol. Dislodge pellet from tube wall.
16.  Centrifuge at maximum speed of 12,000 x g for 2 minutes. Remove supernatant with pipette.
17.  SpeedVac on low heat for 5 minutes until dry, protected from light.
18.  Store labeled samples at -20°C, protected from light.
19.  Rehydrate pellets in 25µl VWR deionized water.
20.  Vortex for 30 seconds and quick-spin to collect contents at bottom of tube. Continue to vortex or let sit at room temperature, protected from light, until the pellet is completely rehydrated, then vortex again, and quickly spin.
21.  Determine the concentration of each sample. The concentration can be determined using the following formula:
Concentration (µg/ml) = A260nm x 50 x Dilution Factor
22.  Based on the concentration, calculate the amount of sample required for hybridization per the following table and place in 1.5ml tube:
Sample Requirements   Amount : Prokaryotic sample  1.5µg
23.  If you’re ready to perform the hybridization, correct the sample volume to 11.3µl and proceed to Chapter 4, Microarray Hybridization & Washing.
a.  If the volume is smaller than 11.3µl, bring the volume up to 11.3µl with VWR water. Mix and spin down to collect contents in bottom of tube.
b.  If the volume is greater than 11.3µl, dry the contents in a SpeedVac on low heat, protected from light. Resuspend the sample in 11.3µl VWR water and vortex to dissolve. Mix and spin down to collect contents in bottom of tube.
24.  If you’re not ready to perform the hybridization, dry the contents in a SpeedVac on low heat, protected from light. Store samples at -20C until ready for hybridization.

growth_protocol: Brucella melitensis vjbR mutant strain was grown with shaking at 37°C in 2YT medium (10% yeast extract, 10 g l-1 tryptone, 5 g l-1 NaCl) from an initial optical density at 600 nm (OD600) of 0.05. Cultures was incubated at 37°C with shaking to an OD600 of 0.75.

extract_protocol: Total RNA were extracted from wt, ∆vjbR and ∆babR (all cultured in triplicate) as following: 45 ml of culture (OD600 of 0.75) were centrifuged at 3500 rpm for 15 min. Bacterial pellets were resuspended in 100 µl SDS 10% and 20 µl proteinase K (20 mg ml-1) and incubated at 37°C with shaking for 1h. Five milliliters of TRIzol® Reagent (Invitrogen) were added and suspensions were vigorously shacked. After 10 min of incubation at 65°C, 1 ml chloroform was added to the suspensions and the mixes were shaked and incubated at room temperature for 5-10 min. Samples were centrifuged at 14.000 rpm for 15 min at 4°C. Then, 2.5 ml propan-2-ol were added to the aqueous phases and samples were stored overnight at -20°C. After centrifugation for 30 min at 14.000 rpm at 4°C, pellets were washed with 75% ethanol RNase free. Supernatants were discarded and pellets were dried 15 min at room temperature. Total RNA samples were resuspended in 100 µl RNase free water, incubated 10 min at 55°C and stored at -80°C. The integrity of the RNA and the absence of DNA were checked by gel electrophoresis.

tag: brucella melitensis 16m vjbr mutant strain; od600nm


title: TAX577718 [38K]


title: TAX577146 [33K]


title: TAX577498 [33K]


title: TAX577688 [33K]


title: TAX577716 [38K]


title: TAX577678 [33K]


title: TAX577683 [33K]


title: CD34+ ET 12

status: Public on Apr 06 2010

city: Modena

first: Rossella

last: Manfredini

address: Via Gottardi 100

department: Life Sciences

country: Italy

institute: Centre for Regenerative Medicine

description: Two-cycle target labeling assays, as well as the Affymetrix HG-U133A GeneChip arrays hybridization, staining, and scanning, were performed, using Affymetrix standard protocols (Affymetrix, Santa Clara, CA).

hyb_protocol: The fragmented cRNA was then hybridized to an identical lot of Affymetrix HG-U133A GeneChip arrays for 16 hours. GeneChips were washed and stained using the instrument’s standard Eukaryotic GE WS2v4 protocol, using antibody-mediated signal amplification.

scan_protocol: GeneChip were finally scanned using the Affymetrix GeneChip scanner 3000, enabled for High-Resolution Scanning.

data_processing: Robust multi-array average (RMA) procedure was applied to the entire set of raw signals (i.e. .CEL files) in order to background adjust and normalize microarray intensities and to generate gene expression values.

source_name: Bone Marrow CD34+ progenitor cells

label_protocol: Two-cycle target labeling assays were performed, using Affymetrix standard protocols (Affymetrix, Santa Clara,CA). Briefly, biotin-labeled target synthesis was performed, starting from 100 ng of total cellular RNA, according to the protocol supplied by the manufacturer (Affymetrix, Santa Clara, CA). Labeled cRNA was purified using Affymetrix GeneChip Sample Cleanup Module and fragmented (20 µg) as described in the Affymetrix GeneChip protocol. Disposable RNA chips (Agilent RNA 6000 Nano LabChip kit, Agilent Technologies, Waldbrunn, Germany) and Agilent 2100 Bioanalyzer were used to determine the cRNA concentration/quality as well as to optimize the cRNA fragmentation.

extract_protocol: Total RNA from CD34+ cells was extracted using RNeasy Micro Kit (Qiagen, Valencia, CA) following the protocol supplied by the manufacturer. Disposable RNA chips (Agilent RNA 6000 Nano LabChip kit, Agilent Technologies, Waldbrunn, Germany) were used to determine the concentration and purity/integrity of RNA samples using Agilent 2100 Bioanalyzer.

tag: CD34+ hematopoietic progenitors purified from bone marrow of Essential Thrombocythemia (ET) patient with the JAK2V617F mutation


title: CD34+ ET 9

tag: CD34+ hematopoietic progenitors purified from bone marrow of Essential Thrombocythemia (ET) patient without the JAK2V617F mutation


title: CD34+ ET 6


title: CD34+ ET 5


title: CD34+ ET 8


title: CD34+ ET 13


title: BC184

tag: age at diagnosis

tag: histology

tag: pt

tag: pn

tag: sbr grade

tag: er ihc

tag: pr ihc


title: CD34+ ET 1


title: Brucella melitensis 16M wild type

source_name: Brucella melitensis 16M wild type strain ; OD600nm : 0.75

growth_protocol: Brucella melitensis wild type strain was grown with shaking at 37°C in 2YT medium (10% yeast extract, 10 g l-1 tryptone, 5 g l-1 NaCl) from an initial optical density at 600 nm (OD600) of 0.05. Cultures was incubated at 37°C with shaking to an OD600 of 0.75.


title: Brucella melitensis 16M babR mutant strain

growth_protocol: Brucella melitensis babR mutant strain was grown with shaking at 37°C in 2YT medium (10% yeast extract, 10 g l-1 tryptone, 5 g l-1 NaCl) from an initial optical density at 600 nm (OD600) of 0.05. Cultures was incubated at 37°C with shaking to an OD600 of 0.75.

tag: brucella melitensis 16m babr mutant strain; od600nm


title: CD34+ ET 7


title: CD34+ ET 10


title: CD34+ ET 16


title: CD34+ ET 14


title: CD34+ ET 4


title: CD34+ ET 2


title: CD34+ ET 11


title: CD34+ ET 3


title: CD34+ ET 15


title: CD34+ CTR 4

tag: CD34+ hematopoietic progenitors purified from bone marrow of healthy control


title: CD34+ CTR 2


title: CD34+ CTR 1


title: CD45+/Sca1+_Muscle_rep2

description: Synthesis of biotinylated cRNA targets, arrays hybridization (Mouse430A GeneChip, Affymetrix, Santa Clara,CA), staining and scanning were performed using Affymetrix standard protocols starting from 1μg of total RNA.

hyb_protocol: The fragmented cRNA was then hybridized to an identical lot of Affymetrix Mouse430A GeneChip arrays for 16 hours. GeneChips were washed and stained using the instrument’s standard Eukaryotic GE WS2v4 protocol, using antibody-mediated signal amplification.

data_processing: Scaling to TGT value=250. The GeneChip-Operating-Software (GCOS) absolute analysis algorithm was used to determine the mRNA expression levels (Signal), while the GCOS comparison analysis algorithm was used in order to compare gene expression levels between two samples.

organism: Mus musculus

source_name: CD45+/Sca1+ cells isolated from the muscle

label_protocol: One-cycle target labeling assays were performed, using Affymetrix standard protocols (Affymetrix, Santa Clara,CA). Briefly, biotin-labeled target synthesis was performed, starting from 1 µg of total cellular RNA, according to the protocol supplied by the manufacturer (Affymetrix, Santa Clara, CA). Labeled cRNA was purified using Affymetrix GeneChip Sample Cleanup Module and fragmented (20 µg) as described in the Affymetrix GeneChip protocol. Disposable RNA chips (Agilent RNA 6000 Nano LabChip kit, Agilent Technologies, Waldbrunn, Germany) and Agilent 2100 Bioanalyzer were used to determine the cRNA concentration/quality as well as to optimize the cRNA fragmentation.

extract_protocol: Total RNA was extracted using RNeasy Micro Kit (Qiagen, Valencia, CA) following the protocol supplied by the manufacturer. Disposable RNA chips (Agilent RNA 6000 Nano LabChip kit, Agilent Technologies, Waldbrunn, Germany) were used to determine the concentration and purity/integrity of RNA samples using Agilent 2100 Bioanalyzer.

tag: Strain

value: C57BL/6, Age:6-8 weeks old, Tissue:muscle


title: CD45+/Sca1+_Muscle_+Ctx/BMT_rep2

treatment_protocol: Injury with Cardiotoxin (Ctx) and BM transplantation (BMT)

tag: strain


title: CD45+/Sca1+_Muscle_rep1


title: CD45+/Sca1+_BM_rep2

source_name: CD45+/Sca1+ cells isolated from the bone marrow

value: C57BL/6, Age:6-8 weeks old, Tissue:bone marrow


title: CD45+/Sca1+_Muscle_+Ctx_rep1

treatment_protocol: Injury with Cardiotoxin (Ctx)


title: CD45+/Sca1+_Muscle_+Ctx_rep2


title: RAW 264.7 – infected with Brucella abortus for 4hrs – replicate date 07252001

status: Public on Jan 01 2010

city: Madison

first: Gary

last: Splitter

state: WI

address: 1656 Linden Dr

department: Pathobiological Sciences

country: USA

institute: University of Wisconsin - Madison

description: See other fields.

hyb_protocol: Labeled cRNA was hybridized to GeneChip® Murine Genome U74A microarrays (Affymetrix). GeneChip® hybridization, washing and scanning was performed by the University of WI –Biotechnology Center, Gene Expression Center (University of WI, Madison) utilizing all Affymetrix protocols and procedures.  Washing and streptavidin-phycoerythrin staining was completed on a GeneChip Fluidics station 400 (Affymetrix).

scan_protocol: GeneChips were scanned with HP 2500 GeneArray Scanner and signal output analysis completed with MAS5.0 (Affymetrix).

data_processing: Signal output analysis completed with U74A Mask Files and MAS5.0 (Affymetrix).  For original analysis Eskra, et al. Infection and Immunity 71(3), 2003 the difference calls were assigned values and summed across the six comparisons.

source_name: RAW 264.7, infected with Brucella abortus, 4 hours

label_protocol: Target RNA was prepared according to protocols in the Affymetrix GeneChip® Expression Analysis technical manual (Affymetrix, Santa Clara, CA).  In vitro synthesis of biotin-labeled cRNA was completed with Enzo BioArray HighYield RNA Transcript Labeling Kit (Affymetrix).  In vitro transcription clean up was completed over an RNeasy spin column (Qiagen).  Labeled cRNA mixed with fragmentation buffer was incubated at 94°C for 35 minutes.  Final RNA concentration in fragmentation buffer ranged from 0.5-1.1 μg/μL.  Visual confirmation of isolation and fragmentation was completed with agarose gel electrophoresis.

growth_protocol:
RAW 264.7 (TIB-71, ATCC) mouse macrophage (Mφ) cell line was maintained at 37°C with 5% CO2 in RPMI 1640 (Invitrogen) supplemented with 10% Fetal Bovine Serum, 0.2mM L-glutamine, 100U penicillin/mL, 100ug streptomycin/mL (Sigma).  Macrophages were subcultured as necessary by removing old media, washing with PBS, adding fresh media, scraping cells and diluting cells 1:3 to 1:20.
B. abortus was grown in 12- by 75-mm tubes on a shaker platform in BBL Brucella broth (BD Bioscience).  Cultures for infection were grown at 37°C for 3 days prior to infection.

extract_protocol: Total RNA was isolated from macrophage cultures utilizing RNeasy Mini Kit (Qiagen) according to manufacturer’s protocol with the following modifications.  Cells were lysed in RLT and then centrifuged to remove intact bacterial cells.  DNase treated RNA was converted to double stranded cDNA using a synthesis kit (Gibco) except that T-7-(dT)24 oligomer (Genset Corp.) was used.  The double stranded cDNA was isolated with the GeneChip® Sample cleanup module.

treatment_protocol: RAW 264.7 cells for microarray were plated in 75cm2 flasks 12-24hr prior to infection, in supplemented RPMI 1640 without antibiotics at an approximate concentration of 6x 10^6 cells per flask.  Macrophage cells were washed once with PBS prior to infection and 5 mL fresh media added.  One mL of actively growing Brucella culture was added to the macrophages (MOI ~100:1).  Infections were allowed to incubate for 4 hours at 37°C with 5% CO2.  Cells were washed three times with PBS to remove extracellular bacteria and then cells were lysed for RNA collection (RNeasy Mini Kit (Qiagen, Valencia, CA)).

tag: designation

value: RAW 264.7 macrophages

tag: atcc

tag: growth properties

value: adherent

value: Abelson murine leukemia virus-induced tumor

tag: antigen expression

value: BALB/c mice

tag: bacteria

value: Brucella abortus, Gram negative, facultative intracellular pathogen


title: RAW 264.7 – infected with Brucella abortus for 4hrs – replicate date 04192001b

description: See other fields.

tag: tissue


title: CD45+/Sca1+_Muscle_+Ctx/BMT_rep1


title: CD45+/Sca1+_BM_rep1


title: RAW 264.7 – infected with Brucella ovis for 4hrs – replicate date 08122003

description: See other fields.

hyb_protocol: Labeled cRNA was hybridized to GeneChip® Test3 arrays and Murine Genome U74Av2 microarrays (Affymetrix). GeneChip® hybridization, washing and scanning was performed by the University of WI –Biotechnology Center, Gene Expression Center (University of WI, Madison) utilizing all Affymetrix protocols and procedures.  Washing and streptavidin-phycoerythrin staining was completed on a GeneChip Fluidics station 400 (Affymetrix).

data_processing: Signal output analysis completed with MAS5.0 (Affymetrix).  For statistical analysis arrays were compiled and processed with EBarrays, a statistical analysis package in the comprehensive R archive network.

source_name: RAW 264.7, infected with Brucella ovis, 4 hours

growth_protocol:
RAW 264.7 (TIB-71, ATCC) mouse macrophage (Mφ) cell line was maintained at 37°C with 5% CO2 in RPMI 1640 (Invitrogen) supplemented with 10% Fetal Bovine Serum, 0.2mM L-glutamine, antibiotic-antimycotic (100 U/mL penicillin G, 100 μg/mL streptomycin, 0.25 μg/mL amphotericin B, Gibco), 1mM sodium pyruvate (SAFC Biosciences), and MEM amino acids (Hyclone).  Macrophages were subcultured as necessary by removing old media, washing with PBS, adding fresh media, scraping cells and diluting cells 1:3 to 1:20.
B. melitensis, B. neotomae, B. ovis were grown in 12- by 75-mm tubes on a shaker platform in BBL Brucella broth (BD Bioscience).  Cultures for infection were grown at 37°C for 2 days prior to infection.

extract_protocol: Total RNA was isolated from macrophage cultures utilizing RNeasy Mini Kit (Qiagen) according to manufacturer’s protocol with the following modifications.  Cells were lysed in RLT and then centrifuged to remove intact bacterial cells.  RNA from two independent infections was pooled for each target preparation. DNase treated RNA was converted to double stranded cDNA using a synthesis kit (Gibco) except that T-7-(dT)24 oligomer (Genset Corp.) was used.  The double stranded cDNA was isolated with the GeneChip® Sample cleanup module.

treatment_protocol: RAW 264.7 cells for microarray were plated in 75cm2 flasks 12-24hr prior to infection, in supplemented RPMI 1640 without antibiotics at an approximate concentration of 5x 10^6 cells per flask.  Macrophage cells were washed once with PBS prior to infection and 5 mL fresh media added.  One mL of actively growing Brucella culture was added to the macrophages (MOI ~1000:1).  Infections were allowed to incubate for 4 hours at 37°C with 5% CO2.  Cells were washed once with PBS to remove extracellular bacteria and then cells were lysed for RNA collection (RNeasy Mini Kit (Qiagen, Valencia, CA)).

value: Brucella ovis, Gram negative, facultative intracellular pathogen


title: RAW 264.7 – uninfected control – replicate date 04192001

description: See other fields.

source_name: RAW 264.7, uninfected, 4 hours

growth_protocol: RAW 264.7 (TIB-71, ATCC) mouse macrophage (Mφ) cell line was maintained at 37°C with 5% CO2 in RPMI 1640 (Invitrogen) supplemented with 10% Fetal Bovine Serum, 0.2mM L-glutamine, 100U penicillin/mL, 100ug streptomycin/mL (Sigma).  Macrophages were subcultured as necessary by removing old media, washing with PBS, adding fresh media, scraping cells and diluting cells 1:3 to 1:20.

extract_protocol: Total RNA was isolated from macrophage cultures utilizing RNeasy Mini Kit (Qiagen) according to manufacturer’s protocol.  DNase treated RNA was converted to double stranded cDNA using a synthesis kit (Gibco) except that T-7-(dT)24 oligomer (Genset Corp.) was used.  The double stranded cDNA was isolated with the GeneChip® Sample cleanup module.

treatment_protocol: RAW 264.7 cells for microarray were plated in 75cm2 flasks 12-24hr prior to infection, in supplemented RPMI 1640 without antibiotics at an approximate concentration of 6x 10^6 cells per flask.  Macrophage cells were washed once with PBS and 5 mL fresh media added.  Cells were incubated for 4 hours at 37°C with 5% CO2.  Cells were washed once with PBS and then lysed for RNA collection (RNeasy Mini Kit (Qiagen, Valencia, CA)).


title: RAW 264.7 – uninfected control – replicate date 07252001

description: See other fields.


title: RAW 264.7 – infected with Brucella abortus for 4hrs – replicate date 04192001a

description: See other fields.


title: RAW 264.7 – infected with Brucella ovis for 4hrs – replicate date 10142003

description: See other fields.

hyb_protocol: Labeled cRNA was hybridized to GeneChip® Test3 arrays and Murine Genome U74Av2 microarrays (Affymetrix). GeneChip® hybridization, washing and scanning was performed by the University of WI –Biotechnology Center, Gene Expression Center (University of WI, Madison) utilizing all Affymetrix protocols and procedures.  Washing and streptavidin-phycoerythrin staining was completed on a GeneChip Fluidics station 450 (Affymetrix).

scan_protocol: GeneChips were scanned with HP 2500 GeneArray Scanner and signal output analysis completed with MAS5.0 as a part of GCOS v1 (Affymetrix).

data_processing: Signal output analysis completed with MAS5.0 as a part of GCOS v1 (Affymetrix).  For statistical analysis arrays were compiled and processed with EBarrays, a statistical analysis package in the comprehensive R archive network.


title: SJL/J vs C57BL/6J Kidney replicate2

status: Public on Dec 31 2009

city: University of New South Wales

first: Mark

last: Cowley

state: NSW

address: School of BABS, Building D26

department: Biotechnology and Biomolecular Sciences

institute: University of New South Wales

hyb_protocol: Labeled cDNA was directly compared on a single glass slide two-color microarray containing the Compugen Mouse OligoLibrary representing 21,997 genes and Lucidea Universal ScoreCard (Clive and Vera Ramaciotti Center for Gene Function Analysis, UNSW, Sydney, Australia), in 100 ul of DIGEasy buffer (Roche, Basel, Switzerland) with 5 uL each yeast tRNA and calf thymus DNA as blockers (Invitrogen, Mt. Waverley, Vic, Australia). Utility controls from the Lucidea Scorecard were not used, and therefore served as additional negative controls. Hybridized microarrays were washed in 1xSSC, three times in 1xSSC, 0.1%SDS at 50 deg. C, and three times in 1xSSC, dried by centrifugation.

scan_protocol: GenePix 4000B microarray scanner (Axon Instruments, Union City, CA, USA).

data_processing: Image analysis was performed with the Spot image analysis software version 2 (CSIRO, Australia). All further data processing and statistical analyses were performed using R version 2.0.0 (17). Gene expression data were morph background corrected and log2 transformed. Data for controls and the 232 replicated spots of the housekeeping gene Gapd (NM_008084) were removed prior to normalization to avoid bias. All 31 slides were normalized using print-tip loess (R/limma package) and then applied quantile normalization across arrays.

label: Alexa Fluor 647

source_name: SJL/J whole Kidney RNA (pool of 5)

label_protocol: Equal amounts of total RNA from five replicates of each strain were mixed to give a tissue pool representative of the genetic background. 20 ug of pooled RNA was reverse transcribed (SuperScript Indirect cDNA Labeling System, Invitrogen, Mt. Waverley, Vic, Australia) and fluorescently labeled with Alexa Fluor 647.

extract_protocol: Total RNA was extracted according to the manufacturer's instructions with Qiagen Midi Extraction Kits (Qiagen, Australia); purity and integrity was assessed by OD_260/OD_280 readings greater than 2 and intact rRNA bands analysis (Agilent Bioanalyzer, Agilent, Forest Hills, Vic, Australia), respectively.

value: SJL/J

tag: Gender

value: male

tag: Age

value: Eight weeks

tag: Tissue

value: Whole Brain

label: Alexa Fluor 555

source_name: C57BL/6J whole Kidney RNA (pool of 5)

label_protocol: Equal amounts of total RNA from five replicates of each strain were mixed to give a tissue pool representative of the genetic background. 20 ug of pooled RNA was reverse transcribed (SuperScript Indirect cDNA Labeling System, Invitrogen, Mt. Waverley, Vic, Australia) and fluorescently labeled with Alexa Fluor 555.

value: Whole Kidney


title: SJL/J vs C57BL/6J Liver replicate3

source_name: SJL/J whole Liver RNA (pool of 5)

source_name: C57BL/6J whole Liver RNA (pool of 5)

value: Whole Liver


title: RAW 264.7 – infected with Brucella neotomae for 4hrs – replicate date 06012004

description: See other fields.

scan_protocol: GeneChips were scanned with Affymetrix GeneChip Scanner 3000 and signal output analysis completed with MAS5.0 as a part of GCOS v1.1.1 (Affymetrix).

data_processing: Signal output analysis completed with MAS5.0 as a part of GCOS v1.1.1 (Affymetrix).  For statistical analysis arrays were compiled and processed with EBarrays, a statistical analysis package in the comprehensive R archive network.

source_name: RAW 264.7, infected with Brucella neotomae, 4 hours

value: Brucella neotomae, Gram negative, facultative intracellular pathogen


title: RAW 264.7 – infected with Brucella ovis for 4hrs – replicate date 06012004

description: See other fields.


title: SJL/J vs C57BL/6J Liver replicate2


title: RAW 264.7 – infected with Brucella neotomae for 4hrs – replicate date 08262003

description: See other fields.


title: SJL/J vs C57BL/6J Kidney replicate3


title: RAW 264.7 – infected with Brucella neotomae for 4hrs – replicate date 08122003

description: See other fields.


title: SJL/J vs C57BL/6J Liver replicate1


title: SJL/J vs C57BL/6J Kidney replicate1


title: SJL/J vs C57BL/6J Brain replicate3

source_name: SJL/J whole Brain RNA (pool of 5)

source_name: C57BL/6J whole Brain RNA (pool of 5)


title: CD34+ CTR 3


title: atmyc2 cop1-6_Seedling_BL+ABA_1

city: New Delhi

first: Sudip

last: Chattopadhyay

address: Aruna Asaf Ali Marg

country: India

institute: National Institute For Plant Genome Research

description:
atmyc2 cop1-6 double mutant seedlings grown under constant blue-light along with abscisic acid, biological rep 1
10-day-old atmyc2 cop1-6 double mutant Arabidopsis seedlings grown under constant blue-light and presence of abscisic acid

hyb_protocol: Following fragmentation, 10 ug of cRNA were hybridized for 16 h at 45 degree Celsius on Affymetrix ATH1 Arabidopsis Genome Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 450 using script EukGE-WS2v4_450

scan_protocol: GeneChips were scanned using GeneChip Scanner 3000

data_processing: The data were analyzed with GeneChip Operating Software (GCOS 1.4) using Affymetrix default analysis settings and global scaling as normalization method. The mean target intensity of each array was arbitrarily set to 100

label: Biotin

organism: Arabidopsis thaliana

source_name: 10-day-old atmyc2 cop1-6 double mutant seedlings

label_protocol: Biotinylated cRNA was prepared according to the standard Affymetrix protocol from 5 ug total RNA (Expression Analysis Technical Manual, Affymetrix).

growth_protocol: Seeds were surface sterilized with 2% sodium hypochlorite, thoroughly washed with RO (reverse-osmosis) water and spread on MS plates supplemented with abscisic acid (0.5 µm). The plates were kept at 4 degree Celsius for 3-5 days for cold treatment and transferred to constant blue-light at 22 ± 1 degree Celsius

extract_protocol: Extraction of total RNA was performed using Trizol, according to the manufacturer's instructions

treatment_protocol: Seedlings were grown under constant blue-light (35 µmol/sec/m2) and presence of abscisic acid (0.5 µm) for 10 days at 22 ± 1 degree Celsius

tag: Ecotype

value: Columbia, Developmental stage:10-day-old seedlings


title: WT_Seedling_BL+ABA_2

description:
Wild-type seedlings grown under constant blue-light along with abscisic acid, biological rep 2
10-day-old wild-type Arabidopsis seedlings grown under constant blue-light and presence of abscisic acid

source_name: 10-day-old wild-type seedlings


title: cop1-6_Seedling_BL_1

description:
cop1-6 mutant seedlings grown under constant blue-light, biological rep 1
10-day-old cop1-6 mutant Arabidopsis seedlings grown under constant blue-light

source_name: 10-day-old cop1-6 mutant seedlings

growth_protocol: Seeds were surface sterilized with 2% sodium hypochlorite, thoroughly washed with RO (reverse-osmosis) water and spread on MS plates. The plates were kept at 4 degree Celsius for 3-5 days for cold treatment and transferred to constant blue-light at 22 ± 1 degree Celsius

treatment_protocol: Seedlings were grown under constant blue-light (35 µmol/sec/m2) for 10 days at 22 ± 1 degree Celsius


title: SJL/J vs C57BL/6J Brain replicate1


title: atmyc2 cop1-6_Seedling_BL+ABA_2

description:
atmyc2 cop1-6 double mutant seedlings grown under constant blue-light along with abscisic acid, biological rep 2
10-day-old atmyc2 cop1-6 double mutant Arabidopsis seedlings grown under constant blue-light and presence of abscisic acid


title: atmyc2_Seedling_BL+ABA_2

description:
atmyc2 mutant seedlings grown under constant blue-light along with abscisic acid, biological rep 2
10-day-old atmyc2 mutant Arabidopsis seedlings grown under constant blue-light and presence of abscisic acid

source_name: 10-day-old atmyc2 mutant seedlings


title: cop1-6_Seedling_BL_2

description:
cop1-6 mutant seedlings grown under constant blue-light, biological rep 2
10-day-old cop1-6 mutant Arabidopsis seedlings grown under constant blue-light


title: atmyc2 cop1-6_Seedling_BL_2

description:
atmyc2 cop1-6 double mutant seedlings grown under constant blue-light, biological rep 2
10-day-old atmyc2 cop1-6 double mutant Arabidopsis seedlings grown under constant blue-light


title: atmyc2_Seedling_BL+ABA_1

description:
atmyc2 mutant seedlings grown under constant blue-light along with abscisic acid, biological rep 1
10-day-old atmyc2 mutant Arabidopsis seedlings grown under constant blue-light and presence of abscisic acid


title: atmyc2 cop1-6_Seedling_BL_1

description:
atmyc2 cop1-6 double mutant seedlings grown under constant blue-light, biological rep 1
10-day-old atmyc2 cop1-6 double mutant Arabidopsis seedlings grown under constant blue-light


title: cop1-6_Seedling_BL+ABA_2

description:
cop1-6 mutant seedlings grown under constant blue-light along with abscisic acid, biological rep 2
10-day-old cop1-6 mutant Arabidopsis seedlings grown under constant blue-light and presence of abscisic acid


title: SJL/J vs C57BL/6J Brain replicate2


title: WT_Seedling_BL_1

description:
Wild-type seedlings grown under constant blue-light, biological rep 1
10-day-old wild-type Arabidopsis seedlings grown under constant blue-light


title: Sustained L-type Calcium Channel Blockade Alters Gene Regulation in the Adult Mouse Ventricle-male vehicle1

city: Lexington

first: Elizabeth

middle: Anne

last: Schroder

state: KY

address: 800 Rose Street, MS508

department: Physiology

institute: University of Kentucky

description: vehicle treated heart tissue

hyb_protocol: standard Affymetrix procedures

scan_protocol: standard Affymetrix procedures

data_processing: GeneChip Operating Software

source_name: heart, left ventricle, free wall, 24 hours vehicle (0.02% ascorbic acid) treatment

label_protocol: standard Affymetrix procedures

extract_protocol: Left ventricular free wall from male mice was rapidly excised and either snap frozen at -80oC or used immediately for RNA isolation. Total RNA was isolated using the RNAqueous -4PCR kit (Ambion) and quantitated spectrophotometrically at 260nm. Contaminating genomic DNA was eliminated by DNase treatment (Ambion). RNA quality was assessed using the Agilent 2100 Bioanalyzer.

treatment_protocol: Male ICR mice (12-14 weeks age) weighing between 25 and 30 grams were anesthetized with a ketamine/xylazine mixture ( i.p.) allowing the subcutaneous implantation of miniosmotic pumps (Alzet, model 2001). The pumps were filled with vehicle (0.02% ascorbic acid). After treatment animals were anesthetized and weighed. Hearts were excised, rinsed, blotted dry, weighed, and then frozen on dry ice and the stored at -80oC until studied. Animals were anesthetized and euthanized according to animal protocols approved by the University of Kentucky Institutional Animal Care and Use Committee. This investigation conforms with the Guide for the Care and Use of Laboratory Animals published by the US National Institutes of Health (NIH Publication NO. 85-23, revised 1996).

value: ICR

value: Male

value: 12-14 weeks

value: Heart/ left ventricle free wall

tag: Treatment

value: 24 hours vehicle (0.02% ascorbic acid)


title: WT_Seedling_BL+ABA_1

description:
Wild-type seedlings grown under constant blue-light along with abscisic acid, biological rep 1
10-day-old wild-type Arabidopsis seedlings grown under constant blue-light and presence of abscisic acid


title: atmyc2 cop1-6_Seedling_Dark_2

description:
atmyc2 cop1-6 double mutant seedlings grown under constant dark, biological rep 2
10-day-old atmyc2 cop1-6 double mutant Arabidopsis seedlings grown under constant dark

growth_protocol: Seeds were surface sterilized with 2% sodium hypochlorite, thoroughly washed with RO (reverse-osmosis) water and spread on MS plates. The plates were kept at 4 degree Celsius for 3-5 days for cold treatment and transferred to constant dark at 22 ± 1 degree Celsius

treatment_protocol: Seedlings were grown under constant dark for 10 days at 22 ± 1 degree Celsius


title: atmyc2_Seedling_BL_1

description:
atmyc2 mutant seedlings grown under constant blue-light, biological rep 1
10-day-old atmyc2 mutant Arabidopsis seedlings grown under constant blue-light


title: Sustained L-type Calcium Channel Blockade Alters Gene Regulation in the Adult Mouse Ventricle- Male verapamil 3

description: verapamil treated heart tissue

source_name: heart, left ventricle, free wall, 24 hours verapamil (3.6 mg/kg/day) treatment

treatment_protocol: Male ICR mice (12-14 weeks age) weighing between 25 and 30 grams were anesthetized with a ketamine/xylazine mixture ( i.p.) allowing the subcutaneous implantation of miniosmotic pumps (Alzet, model 2001). The pumps were filled with verapamil (3.6 mg/kg/day). After treatment animals were anesthetized and weighed. Hearts were excised, rinsed, blotted dry, weighed, and then frozen on dry ice and the stored at -80oC until studied. Animals were anesthetized and euthanized according to animal protocols approved by the University of Kentucky Institutional Animal Care and Use Committee. This investigation conforms with the Guide for the Care and Use of Laboratory Animals published by the US National Institutes of Health (NIH Publication NO. 85-23, revised 1996).

value: 24 hours verapamil (3.6mg/kg/day)


title: atmyc2_Seedling_BL_2

description:
atmyc2 mutant seedlings grown under constant blue-light, biological rep 2
10-day-old atmyc2 mutant Arabidopsis seedlings grown under constant blue-light


title: Sustained L-type Calcium Channel Blockade Alters Gene Regulation in the Adult Mouse Ventricle- male verapamil 1

description: verapamil treated heart tissue

value: 24 hours verapamil (3.6 mg/kg/day)


title: Sustained L-type Calcium Channel Blockade Alters Gene Regulation in the Adult Mouse Ventricle- male vehicle 2

description: vehicle treated heart tissue


title: atmyc2 cop1-6_Seedling_Dark_1

description:
atmyc2 cop1-6 double mutant seedlings grown under constant dark, biological rep 1
10-day-old atmyc2 cop1-6 double mutant Arabidopsis seedlings grown under constant dark


title: Sustained L-type Calcium Channel Blockade Alters Gene Regulation in the Adult Mouse Ventricle- Male verapamil 2

description: verapamil treated heart tissue


title: WT_Seedling_BL_2

description:
Wild-type seedlings grown under constant blue-light, biological rep 2
10-day-old wild-type Arabidopsis seedlings grown under constant blue-light


title: Sustained L-type Calcium Channel Blockade Alters Gene Regulation in the Adult Mouse Ventricle- Vehicle 3

description: vehicle treated heart tissue

extract_protocol: Left ventricular free wall from female mice was rapidly excised and either snap frozen at -80oC or used immediately for RNA isolation. Total RNA was isolated using the RNAqueous -4PCR kit (Ambion) and quantitated spectrophotometrically at 260nm. Contaminating genomic DNA was eliminated by DNase treatment (Ambion). RNA quality was assessed using the Agilent 2100 Bioanalyzer.

treatment_protocol: Female ICR mice (12-14 weeks age) weighing between 25 and 30 grams were anesthetized with a ketamine/xylazine mixture ( i.p.) allowing the subcutaneous implantation of miniosmotic pumps (Alzet, model 2001). The pumps were filled with vehicle (0.02% ascorbic acid). After treatment animals were anesthetized and weighed. Hearts were excised, rinsed, blotted dry, weighed, and then frozen on dry ice and the stored at -80oC until studied. Animals were anesthetized and euthanized according to animal protocols approved by the University of Kentucky Institutional Animal Care and Use Committee. This investigation conforms with the Guide for the Care and Use of Laboratory Animals published by the US National Institutes of Health (NIH Publication NO. 85-23, revised 1996).

value: Female


title: Sustained L-type Calcium Channel Blockade Alters Gene Regulation in the Adult Mouse Ventricle- Vehicle 2

description: vehicle treated heart tissue


title: Sustained L-type Calcium Channel Blockade Alters Gene Regulation in the Adult Mouse Ventricle Ver-3

description: verapamil treated heart tissue

treatment_protocol: Female ICR mice (12-14 weeks age) weighing between 25 and 30 grams were anesthetized with a ketamine/xylazine mixture ( i.p.) allowing the subcutaneous implantation of miniosmotic pumps (Alzet, model 2001). The pumps were filled with verapamil (3.6 mg/kg/day). After treatment animals were anesthetized and weighed. Hearts were excised, rinsed, blotted dry, weighed, and then frozen on dry ice and the stored at -80oC until studied. Animals were anesthetized and euthanized according to animal protocols approved by the University of Kentucky Institutional Animal Care and Use Committee. This investigation conforms with the Guide for the Care and Use of Laboratory Animals published by the US National Institutes of Health (NIH Publication NO. 85-23, revised 1996).


title: atmyc2_Seedling_Dark_2

description:
atmyc2 mutant seedlings grown under constant dark, biological rep 2
10-day-old atmyc2 mutant Arabidopsis seedlings grown under constant dark


title: cop1-6_Seedling_BL+ABA_1

description:
cop1-6 mutant seedlings grown under constant blue-light along with abscisic acid, biological rep 1
10-day-old cop1-6 mutant Arabidopsis seedlings grown under constant blue-light and presence of abscisic acid


title: cop1-6_Seedling_Dark_1

description:
cop1-6 mutant seedlings grown under constant dark, biological rep 1
10-day-old cop1-6 mutant Arabidopsis seedlings grown under constant dark


title: Sustained L-type Calcium Channel Blockade Alters Gene Regulation in the Adult Mouse Ventricle- male vehicle 3

description: vehicle treated heart tissue


title: cop1-6_Seedling_Dark_2

description:
cop1-6 mutant seedlings grown under constant dark, biological rep 2
10-day-old cop1-6 mutant Arabidopsis seedlings grown under constant dark


title: Sustained L-type Calcium Channel Blockade Alters Gene Regulation in the Adult Mouse Ventricle-VER1

description: verapamil treated heart tissue

source_name: heart, left ventricle, free wall, 24 hours verapmil (3.6 mg/Kg/day) treatment

value: 24 hours verapamil (3.6 mg/kg)


title: WT_Seedling_Dark_1

description:
Wild-type seedlings grown under constant dark, biological rep 1
10-day-old wild-type Arabidopsis seedlings grown under constant dark


title: Sustained L-type Calcium Channel Blockade Alters Gene Regulation in the Adult Mouse Ventricle- VER2

description: verapamil treated heart tissue


title: WT_Seedling_Dark_2

description:
Wild-type seedlings grown under constant dark, biological rep 2
10-day-old wild-type Arabidopsis seedlings grown under constant dark


title: hyperglycemia, donor II

status: Public on Dec 24 2009

city: Oslo

first: Hege

last: Thoresen

address: Sem Sælandsvei 3

department: School of Pharmacy

country: Norway

institute: University of Oslo

hyb_protocol: Standard Illumina hybridization protocol

scan_protocol: Standard Illumina scanning protocol

data_processing: BeadStudio version 3.1.2.0 from Illumnia and the Gene Expression module 3.2.7.  Program R and Bioconductor packages.

source_name: myotubes from satellite cells from m. obliquus internus abdominis

label_protocol: Biotinylated cRNA were prepared with the Ambion MessageAmp kit for Illumina arrays

growth_protocol: The cells were cultured in DMEM-Glutamax (5.5 mmol/l glucose) with 2 % FCS, 2 % Ultroser G, penicillin/streptomycin (P/S) and amphotericin B until 70-80 % confluence. Myoblast differentiation to myotubes was then induced by changing medium to DMEM-Glutamax (5.5 mmol/l glucose) with 2 % FCS, 34 pmol/l insulin, P/S and amphotericin B. Experiments were performed after 8 days of differentiation, and pre-incubation with hyperglycemia (20 mmol/l glucose) was started after 4 days.

extract_protocol: Total RNA was prepared from primary myotubes from three donors using Agilent Total RNA isolation kit according to the supplier’s protocol (Agilent Technologies). RNA was used individually and RNA integrity was checked on chip analysis (Agilent 2100 bioanalyzer, Agilent Technologies, Santa Clara, CA, USA) according to the manufacturer's instructions. RNA was judged as suitable for array hybridization only if samples exhibited intact bands corresponding to the 18S and 28S ribosomal RNA subunits, and displayed no chromosomal peaks or RNA degradation products (RNA Integrity Number > 9.0).

treatment_protocol: From day 4-8 during the differentiation period the cells were treated with 5.5 mM glucose (control, normoglycemia) or 20 mM glucose (hyperglycemia). Isolation of RNA was performed on day 8.

tag: cell type


title: normoglycemia, donor III

value: myotubes from satellite cells from m. obliquus internus abdominis


title: Sustained L-type Calcium Channel Blockade Alters Gene Regulation in the Adult Mouse Ventricle

description: vehicle treated heart tissue

extract_protocol: Left ventricular free wall from female mice was rapidly excised and either snap frozen at -80oC or used immediately for RNA isolation.    Total RNA was isolated using the RNAqueous -4PCR kit (Ambion) and quantitated spectrophotometrically at 260nm.  Contaminating genomic DNA was eliminated by DNase treatment (Ambion).  RNA quality was assessed using the Agilent 2100 Bioanalyzer.

treatment_protocol: Female ICR mice (12-14 weeks age) weighing between 25 and 30 grams were anesthetized with a ketamine/xylazine mixture ( i.p.) allowing the subcutaneous implantation of miniosmotic pumps (Alzet, model 2001).  The pumps were filled with vehicle (0.02% ascorbic acid).  After treatment animals were anesthetized and weighed.  Hearts were excised, rinsed, blotted dry, weighed, and then frozen on dry ice and the stored at -80oC until studied.    Animals were anesthetized and euthanized according to animal protocols approved by the University of Kentucky Institutional Animal Care and Use Committee.  This investigation conforms with the Guide for the Care and Use of Laboratory Animals published by the US National Institutes of Health (NIH Publication NO. 85-23, revised 1996).


title: hyperglycemia, donor I


title: hyperglycemia, donor III


title: atmyc2_Seedling_Dark_1

description:
atmyc2 mutant seedlings grown under constant dark, biological rep 1
10-day-old atmyc2 mutant Arabidopsis seedlings grown under constant dark


title: normoglycemia, donor II


title: normoglycemia, donor I


title: spleen_Yaa_rep3

status: Public on Dec 15 2009

city: Oklahoma City

first: Bart

last: Frank

state: OK

institute: OMRF

description: The Yaa gene is on the Y chromosome and is responsible for accelerated autoimmune disease in mice with susceptible genetic backgrounds

hyb_protocol: Standard Affymetrix hybridization plus EukGE-WS2v5 fluidics protocol

scan_protocol: Affymetrix 7G scanner

data_processing: Affymetrix MAS5.0 normalization & transforming algorithm

source_name: spleen cells from C57Bl/6 male with Yaa mutation

label_protocol: Affymetrix One cycle target labeling kit

extract_protocol: Combination of Trizol protocol followed by Qiagen RNeasy column

tag: Sex

value: male

value: whole spleen tissue

tag: note

value: RNA in this one sample was prepared from spleenocytes from two different spleens


title: MC 6h 1

city: Bergisch Gladbach

first: Silvia

last: Rueberg

address: Friedrich-Ebert-Str. 68

department: Genomic Services

institute: Miltenyi Biotec GmbH

description: Cy3 and Cy5 labeled (sample_ch1 and sample_ch2) cDNA were combined  and hybridzed on the PIQOR ™ custom_CTID88 Microarray, mouse,

hyb_protocol: PIQOR ™ Microarray pretreatment: microarrays were heated in distilled H20 at 95 °C for 2 min, microarrays were transfered into ethanol for at least 30 seconds and then dried by using pressured air. Hybridization: Hybridization was performed using a GeneTAC hybridization station (PerkinElmer, Rodgau-Juegesheim, Germany). Microarrays were prehybridized with 100 µl Prehybridization Solution at 65 °C

scan_protocol: Image capture and signal quantification of hybridized PIQOR (TM) microarrays were done with the ScanArray 3000 Microarray Scanner (GSI Lumonics, Watertown,MA, USA) and the ImaGene software Version 4.1 (BioDiscovery, Los Angeles, CA, USA).

data_processing: Mean signal and mean local background intensities were obtained for each spot of the microarray images using the ImaGene software Version 4.1 (Biodiscovery, Los Angeles, CA, USA). Low-quality spots were flagged and excluded from data analysis. Unflagged spots were analysed with the PIQORTM Analyzer software (Miltenyi Biotec, Bergisch Gladbach, Germany). The PIQOR Analyzer allows automated data processing of the raw data text files derived from the ImaGene software. This includes background subtraction to obtain the net signal intensity, data normalization, and calculation of the Cy5/Cy3 ratios. Subsequently, the mean of the ratios of the four corresponding spots representing the same cDNA was computed. The ratios were normalized using the LOWESS method. As an additional quality filtering step, only genes were taken into account for the calculation of the Cy5/Cy3 ratio that have at least in one channel a signal intensity that was at least 2-fold higher than the mean background.

source_name: control:liver, pool of 9 untreated mice

label_protocol: mRNA was isolated from 100 µg total RNA (Oligotex mRNA Mini Kit, Qiagen, Hilden, Germany), mRNA was reverse transcribed in the presence of Cy3-dCTPs according to the PIQOR ™ User manual (Miltenyi Biotec, Bergisch Gladbach, Germany)

extract_protocol: NucleoSpin RNA L Kit (Macherey-Nagel GmbH and Co KG, Dueren, Germany)

tag: wildtype C57BL/6 (8-10 weeks), liver, untreated

source_name: tissue: liver, treatment: 50 mg/kg MC (methylene chloride), p.o., timepoint: 6 h

tag: wildtype C57BL/6 (8-10 weeks), liver, biological replicate 1 of 3


title: MC 24h 3

source_name: tissue: liver, treatment: 50 mg/kg MC (methylene chloride), p.o., timepoint: 24 h

tag: wildtype C57BL/6 (8-10 weeks), liver, biological replicate 3 of 3


title: MC 6h 3


title: MC 24h 1


title: MC 24h 2

tag: wildtype C57BL/6 (8-10 weeks), liver, biological replicate 2 of 3


title: spleen_Yaa_rep1

source_name: spleen cells from B6.SB-Yaa/J male

tag: Strain


title: CoPPIX 24h 1

source_name: tissue: liver, treatment: 5 mg/kg CoPPIX (metalloporphyrin cobalt-protoporphyrin IX, i.p., timepoint: 24 h


title: CoPPIX 24h 2


title: CoPPIX 6h 3

source_name: tissue: liver, treatment: 5 mg/kg CoPPIX (metalloporphyrin cobalt-protoporphyrin IX, i.p., timepoint: 6 h


title: spleen_Yaa_rep2

source_name: spleen cells from C57Bl/6 males with Yaa mutation


title: Buffer treated leaves, biological replicate1

city: Hyderabad

first: Ramesh

middle: V.

last: Sonti

state: Andhra Pradesh

address: Uppal Road

department: Plant Microbe Interactions

institute: Centre for cellular and molecular biology (CCMB)

description: Expression profile of buffer treated rice leaves, 12 h after infiltration

hyb_protocol: The biotinylated cRNAs were fragmented and 15µg of this sample was hybridized to Gene Chip Rice Genome Array for 16h at 450C and 60rpm. The hybridized gene chips were subjected to washing and staining protocol using an Affymetrix fluidics station 450

scan_protocol: The gene chips were scanned on an Affymetrix Scanner 3000

data_processing: The CEL files generated by the Gene Chip Operating System (Affymetrix) were subjected to AvadisTM software version 4.3 (Strand Life Sciences, Bangalore, India) for further analysis. The PLIER and RMA algorithms were used for data normalization

organism: Oryza sativa

source_name: 20 days old Taichung Native-1 rice seedlings

label_protocol: 8µg of total RNA were converted into biotinylated cRNA using Affymetrix one cycle labeling kit as per instructions of the manufacturer (Expression analysis technical manual, 2001, Affymetrix)

growth_protocol: TN-1 rice seeds were germinated in petri plates containing moistened filter paper. Three days old seedlings were transferred to plastic pots containing soil and were grown for 15 days in a green house. Two days before the actual treatment, the pots containing the rice seedlings were transferred to a growth chamber (28oC; 80% relative humidity with 12/12h light/dark cycle)

extract_protocol: 20-30 leaf pieces covering the infiltrated zone from each of the treatments were harvested 12h after infiltration and subjected to total RNA isolation using Trizol (Invitrogen)

treatment_protocol: 30-40μl of either cellulase (5μg/ml) purified from the culture supernatant of wild type Xoo or 10mM potassium phosphate buffer (pH 6.0) were infiltrated to the adaxial surfaces of rice leaves with a 1ml hypodermal syringe without the needle (as described in Jha et al. 2007; MPMI vol 20, pp 31-40)

tag: Genotype

value: bacterial blight susceptible rice cultivar

value: 20 days old rice seedlings


title: spleen_WT_rep3

description: This study was designed to identify gene expression between spleen cells of male mice with and without the Yaa mutation.

source_name: spleen cells from C57Bl/6J wild-type male


title: spleen_Yaa_rep4


title: spleen_WT_rep4


title: spleen_WT_rep2


title: spleen_WT_rep1


title: Cellulase treated leaves, biological replicate3

description: Expression profile of cellulase treated rice leaves, 12 h after infiltration


title: Buffer treated leaves, biological replicate3


title: CoPPIX 6h 2


title: CoPPIX 6h 1


title: CO 24h 3

source_name: tissue: liver, treatment: 500 ppm CO (carbon monoxide), timepoint: 24 h


title: Cellulase treated leaves, biological replicate1


title: Buffer treated leaves, biological replicate2


title: Cellulase treated leaves, biological replicate2


title: CO 24h 1

tag: wildtype C57BL/6 (8-10 weeks),liver, biological replicate 1 of 3


title: CO 24h 2


title: CO 6h 3

source_name: tissue: liver, treatment: 500 ppm CO (carbon monoxide), timepoint: 6 h


title: CO 6h 2


title: CO 6h 1


title: BALB MCA 25 week_Biorep1

status: Public on Dec 12 2009

city: RTP

first: NIEHS

last: Microarray Core

state: NC

address: 111 T.W. Alexander Drive

department: DIR

institute: NIEHS

description: Gene expression analysis was conducted using Mouse 430A Genechip® arrays (Affymetrix, Santa Clara, CA).  Starting with 1ug of total RNA, biotin-labeled cRNA was produced using the Affymetrix 3’ Amplification One-Cycle Target labeling kit according to manufacturer’s protocol.  For each array, 15ug of amplified cRNAs were fragmented and hybridized to the array for 16 hours in a rotating hybridization oven 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.  Arrays were then scanned with an Affymetrix Scanner 3000.  Data was obtained using the Genechip® Operating Software.

scan_protocol: Slides were stained and washed as indicated in the Antibody Amplification Stain for Eukaryotic Targets protocol using the Affymetrix Fluidics Station FS450.  Arrays were then scanned with an Affymetrix Scanner 3000.  Data was obtained using the Genechip® Operating Software (Version 1.2.0.037).

data_processing: Since samples were hybridized to two different microarray platforms (MOE430A or MOE430Av2), data were merged based on common probeset identifiers. Principal component analysis (PCA) was performed to examine the effect of the two platforms. The primary component of variation defined the platform on which a sample was hybridized. This effect appeared independent of the genotype or treatment condition, therefore, singular value decomposition was used to adjust the data and minimize the effect of platform biases (Alter et al. 2000; Nielsen et al. 2002).  PCA plots after adjustment gave no evidence of residual platform effect. This adjusted data was further normalized by gc-RMA and used further as described below (Wu et al, 2004).

source_name: mouse RNA from lungs after MCA or MCA + BHT treatment

growth_protocol: Mice were allowed to acclimate for 1 wk prior to treatment. All animal use was conducted in facilities accredited by the Association for the Assessment and Accreditation of Laboratory Animal Care and approved by the NIEHS Animal Care and Use Committee and the MSU Institutional Animal Care and Use Committee. Mice were housed in shoebox cages in a humidity and temperature-controlled room and provided water and pelleted open-formula rodent diet NIH-07 (Zeigler Brothers, Gardners, PA.) ad libitum.

extract_protocol: Total RNA was isolated from the left lung lobe using the RNAeasy Mini Kits (Qiagen, Valencia, CA) and following the kit specifications, including Dnase 1 treatment.

treatment_protocol: For BHT-induced tumor promotion studies (Protocol 2), 10 ug MCA/gm (Sigma) body wt. was injected ip. on week one, followed by 150 mg BHT/kg or corn oil vehicle control on week two.  Five additional weekly 200 mg/kg BHT doses or vehicle control doses were administered to maximize tumor promotion (Brown et al, 1999).  The mice were sacrificed 26-28 wks after the MCA injection and thus during the tumor progression stage of carcinogenesis.

tag: Male 5-7 wk. old BALB/c (BALB; Tlr4 normal) and C.C3-Tlr4Lps-d/J (BALBLps-d ; Tlr4 dominant negative) mice were purchased from Jackson Laboratories (Bar Harbor, ME).


title: BALB MCA+BHT 25 week uninvolved_5315

hyb_protocol: 15ug of amplified cRNAs were fragmented and hybridized to the array for 16 hours in a rotating hybridization oven using the Affymetrix Eukaryotic Target Hybridization Controls and protocol.

label_protocol: Starting with 1ug of total RNA, biotin-labeled cRNA was produced using the Affymetrix 3’ Amplification One-Cycle Target labeling kit according to manufacturer’s protocol.


title: BALBlpsd BHT 3 day_Biorep3

source_name: mouse RNA treated with Oil or BHT for 1 or 3 days

treatment_protocol: To induce chronic inflammation, (Protocol 1), mice were injected intraperitoneally (ip.) with BHT (Sigma, St. Louis, MO) weekly for 4 weeks (150 mg/kg for the first dose followed by 200 mg/kg for the next 3 doses) and then sacrificed 1 or 3 days following the last BHT dose.


title: BALBlpsd BHT 1 day_Biorep2


title: BALB MCA+BHT 25 week tumor_5314


title: BALB_BHT_1_day_Biorep2


title: BALBlpsd Oil 3 day_Biorep3


title: BALB_Oil_3_day_Biorep3


title: BALBlpsd MCA+BHT 25 week uninvolved_5321


title: BALB_BHT_1_day_Biorep3


title: BALBlpsd MCA 25 week_Biorep1


title: BALBlpsd MCA+BHT 25 week tumor_5320


title: BALB MCA+BHT 25 week uninvolved_5313


title: BALBlpsd BHT 3 day_Biorep1


title: BALBlpsd BHT 1 day_Biorep3


title: BALBlpsd MCA+BHT 25 week tumor_5318


title: BALB_BHT_1_day_Biorep1


title: BALBlpsd Oil 3 day_Biorep2


title: BALBlpsd MCA 25 week_Biorep2


title: BALBlpsd Oil 3 day_Biorep1


title: BALBlpsd BHT 3 day_Biorep2


title: BALBlpsd MCA+BHT 25 week uninvolved_5319


title: BALBlpsd BHT 1 day_Biorep1


title: BALB BHT 3 day_Biorep2


title: BALB MCA 25 week_Biorep2


title: BALB_Oil_3_day_Biorep2


title: BALB BHT 3 day_Biorep3


title: BALB_Oil_3_day_Biorep1


title: Bdellovibrio bacteriovorus 109J sample 10

status: Public on Dec 09 2009

city: Nottingham

first: Carey

last: Lambert

state: Notts

address: Queen's Medical Centre

department: Genetics

country: United Kingdom

institute: Nottingham University

description: Biological replicate 5 of 5, technical replicate 2 of 2

hyb_protocol: Rinse the arrays in 50ml 2X SSPE for 5 minutes. Drain. Pre-Hybridise the array in 5ml hybridisation solution with 100ug/ml denatured salmon sperm DNA for at least 1 hour at 65°C. Add all labelled cDNA to 3ml hydridisation solution and denature for 10minutes at 95°C, add to array and hybridise overnight at 65°C Decant off hybridisation solution and wash 3 times in 40-50ml wash solution at room temperature Wash 3 times in 80-100ml wash solution at 65°C for 20 minutes Air dry array for 5 minutes Wrap in saran wrap

scan_protocol: Arrays were exposed to Molecular Dynamics phosphor screens for 2 days and scanned at 88um with Molecular Dynamics Phosphorimager 425 running Image Quant software

data_processing: The ArayVision software was used to analyse the phosphorimages with the template provided by Sigma Genosys for the arrays. Background correction was by using the area surrounding the template, 2 pixels distance and thickness, and normalisation was against all spots in the array

label: 33P dCTP

organism: Bdellovibrio bacteriovorus

source_name: Total RNA from  Bdellovibrio only control

label_protocol:
Add to a 0.5ml tube 4ul E. coli cDNA labeling primers (Sigma), reconstituted in 40ul sterile distilled water and RNA sample made up to 11ul
Anneal the primers to the RNA template by heating to 90°C for 2 minutes, then ramp to 42°C over a period of 20 minutes.
When the sample reaches 42°C, add 3ul 10X Reverse transcriptase buffer, 1ul 333uM dATP, 1ul 333uM dGTP, 1ul 333uM dTTP, 2ul 20uCi [alpha-33P] dCTP, 2ul AMV reverse transcriptase, final volume 30ul
Incubate 42°C for 2-3 hours
apply the sample to Bio-Rad Micro Bio-Spin Chromatography Columns for removal of unincorporated radiolabel

extract_protocol: IFR protocol, eluting in 50µl and DNase treating with Ambion DNA-free kit. http://www.ifr.ac.uk/Safety/Microarrays/protocols.html

treatment_protocol:
1) Grow the Bdellovibrio in a “prey lysate” of 1L Ca/HEPES, 75ml overnight E. coli YT culture and 50ml of a previous lysate in a 2L conical flask with 200rpm shaking in a 29°C incubator. Grow E. coli in 100ml YT in a 250ml flask overnight at 37°C in the shaking incubator.
2) Check that the lysate is fully lysed by microscopy (i.e. that virtually all the E. coli and bdelloplasts are gone and there are lots of motile attack phase bdellovibrios).
3) Pre-warm a benchtop centrifuge at 29°C.
4) When lysate is fully lysed, spin 15,000g for 30 mins.
5) Spin down the E. coli in the benchtop centrifuge in 50ml falcon tubes, 5,000g, 10 mins at 29°C
6) Resuspend 100ml of E. coli in 100ml Ca/HEPES then incubate at 29ºC in a 500ml flask with 200rpm shaking for 3 hours to stabilise expression.
7) Resuspend all of the bdellovibrios in 100ml Ca/HEPES buffer and incubate at 29°C in 500ml conical flask with 200rpm shaking for 3 hours to stabilise expression.
8) After 2.5 hours incubation, remove 35ml from each, spin 5,000g for 5 minutes, filter 3 times through a 0.22µm filter to give supernatant which is returned to incubate at 29°C
9) Pre-label 15ml falcon tubes for the samples and add 1ml Phenol/ethanol solution (5% phenol 95% ethanol, v/v).
10) To start the infection, after 3 hours of pre-incubation, quickly add to the pre-incubated flasks:
* the TEST with 25ml E. coli and 25ml bdellovibrios
* the S17-1 ONLY with 25ml E. coli and 25ml bdellovibrio supernatant
* the BDELLOVIBRIO ONLY with 25ml bdellovibrio and 25ml E. coli supernatant
11) At 15 minutes quickly take 4ml samples from each flask and add them to the phenol/ethanol tubes. Mix tubes well by inversion and put on ice for at least 45mins.
12) After the samples have been on ice for at least 45 minutes, spin 10 mins, 3320g
15) Pour off the supernatant, leaving about 1ml to resuspend pellet carefully and transfer to microcentrifuge tube on ice.
16) Spin 13K 2 mins at 4ºC and then pipette off remaining supernatant carefully.
17) Store the tube containing the pellet at -80°C.

tag: Total RNA from Bdellovibrio bacteriovorus 109J cells


title: Bdellovibrio bacteriovorus 109J sample 9

description: Biological replicate 5 of 5, technical replicate 1 of 2


title: Bdellovibrio bacteriovorus 109J sample 8

description: Biological replicate 4 of 5, technical replicate 2 of 2


title: Bdellovibrio bacteriovorus 109J sample 3

description: Biological replicate 2 of 5, technical replicate 1 of 2


title: Bdellovibrio bacteriovorus 109J sample 2

description: Biological replicate 1 of 5, technical replicate 2 of 2


title: Test sample 9

organism:
Escherichia coli
Bdellovibrio bacteriovorus

source_name: Total RNA from  Bdellovibrio infection on E. coli sample

tag: Total RNA from E. coli and Bdellovibrio cells


title: Bdellovibrio bacteriovorus 109J sample 4

description: Biological replicate 2 of 5, technical replicate 2 of 2


title: Bdellovibrio bacteriovorus 109J sample 1

description: Biological replicate 1 of 5, technical replicate 1 of 2


title: Test sample 10


title: Bdellovibrio bacteriovorus 109J sample 7

description: Biological replicate 4 of 5, technical replicate 1 of 2


title: Bdellovibrio bacteriovorus 109J sample 5

description: Biological replicate 3 of 5, technical replicate 1 of 2


title: Bdellovibrio bacteriovorus 109J sample 6

description: Biological replicate 3 of 5, technical replicate 2 of 2


title: Test sample 8


title: E. coli sample 10

organism: Escherichia coli

source_name: Total RNA from E. coli only control

tag: Total RNA from pre-incubated E. coli cells


title: Test sample 6


title: Test sample 4


title: Test sample 3


title: E. coli sample 9


title: E. coli sample 8


title: E. coli sample 6


title: E. coli sample 7


title: Test sample 5


title: Test sample 7


title: E. coli sample 4


title: E. coli sample 5


title: Test sample 2


title: Test sample 1


title: E. coli sample 3


title: E. coli sample 2


title: E. coli sample 1


title: MC 6h 2


title: Bdellovibrio bacteriovorus attack phase replicate 3

description: Biological replicate 3 of 3

hyb_protocol: see the Nimblegen website for details

scan_protocol: see the Nimblegen website for details

data_processing: Normalized gene calls using the RMA package, see the Nimblegen website for details

organism: Bdellovibrio bacteriovorus HD100

source_name: Total RNA from attack phase

label_protocol: see the Nimblegen website for details

extract_protocol: IFR protocol, eluting in 50µl and DNase treating with Ambion DNA-free kitSee the Institute of Food Research (Norwich, UK) for more details.

treatment_protocol:
1) Grow the Bdellovibrio in a “prey lysate” of 1L Ca/HEPES, 60ml overnight E. coli YT culture and 50ml of a previous lysate in a 2L conical flask with 200rpm shaking in a 29°C incubator. Grow E. coli in 100ml YT in a 250ml flask overnight at 37°C in the shaking incubator.
2) Check that the lysate is fully lysed by microscopy (i.e. that virtually all the E. coli and bdelloplasts are gone and there are lots of motile attack phase bdellovibrios).
3) Pre-warm a benchtop centrifuge at 29°C.When lysate is fully lysed, spin 15,000g for 30 mins.
4) When lysate is fully lysed, spin 15,000g for 30 mins.
5) Spin down the E. coli in the benchtop centrifuge in 50ml falcon tubes, 5,000g, 10 mins at 29°C
6) Resuspend 50ml of E. coli in 50ml Ca/HEPES and dilute to OD600 of 1.0 then incubate at 29ºC in a 500ml flask with 200rpm shaking for 3 hours to stabilise expression.
7) Resuspend all of the bdellovibrios in 100ml Ca/HEPES buffer and incubate at 29°C in 500ml conical flask with 200rpm shaking for 3 hours to stabilise expression.
8) Pre-label 15ml falcon tubes for the samples and timepoints and add 1ml Phenol/ethanol solution (5% phenol 95% ethanol, v/v).
9) To start the infection, after 3 hours of pre-incubation, quickly add to the pre-incubated flasks:
* the TEST with 40ml E. coli and 50ml bdellovibrios and 30ml Ca/HEPES (pre-incubated)
* the S17-1 ONLY with 40ml E. coli and 80ml Ca/HEPES
* the BDELLOVIBRIO ONLY with 50ml bdellovibrio and 70ml Ca/HEPES
10) At each timepoint quickly take 4ml from each flask and add it to the phenol/ethanol tubes (for each sample), and quickly return the flasks to the incubator. Mix tubes well by inversion and put on ice for at least 45mins.
11) At each timepoint, also make a microscope slide of the test to check lysate synchrony and make a note of estimated percentage of E. coli, bdelloplasts and attack phase bdellovibrio
12) After the samples for RNA have been on ice for at least 1 hour, spin 10 mins, 3320g
15) Pour off the supernatant, leaving about 1ml to resuspend pellet carefully and transfer to microcentrifuge tube on ice.
16) Spin 13K 2 mins at 4ºC and then pipette off remaining supernatant carefully.
17) Store the tube containing the pellet at -80°C.

tag: Total RNA from concentrated pre-incubated attack phase cells


title: Bdellovibrio bacteriovorus bdelloplast replicate 1

description: Biological replicate 1 of 3

source_name: Total RNA from bdelloplast

extract_protocol: IFR protocol, DNase treating with Ambion DNA-free kitSee the Institute of Food Research (Norwich, UK) for more details.

tag: Total RNA from 30 minute sample of a semi-synchronous infection of B. bacteriovorus on E. coli S17-1


title: BALB BHT 3 day_Biorep1


title: Bdellovibrio bacteriovorus attack phase replicate 1

description: Biological replicate 1 of 3


title: Bdellovibrio bacteriovorus bdelloplast replicate 2

description: Biological replicate 2 of 3