title: E. coli W3110 igrown n LB plus glucose at OD 1.0 (WT_Glc2)

city: College Park

first: liang

last: wang

state: MD

address: 5115F Plant Science Building

institute: CBR, UMBI

description: Gene expression data for W3110 wild type grown in LB plus 0.8% glucose at OD 1.0

hyb_protocol: Hybridization solution mix was made with the labeled cDNA according to the manufacturer's instructions (Affymetrix, Inc., Santa Clara, CA), and the mixture was hybridized to the E. coli antisense genome arrays at 45°C for 16 h. A GeneChip fluidics station (Affymetrix, Inc., Santa Clara, CA) was used to automate the washing and staining of the arrays. Sequentially, the arrays were stained with ImmunoPure streptavidin (Pierce Biotechnology, Inc., Rockford, IL), antistreptavidin goat antibody (Vector Laboratories, Inc., Burlingame, CA), and R-phycoerythrin streptavidin (Molecular Probes, Inc., Eugene, OR).

scan_protocol: The probe arrays were scanned using the Affymetrix GeneArray scanner.

data_processing: Microarray data were analyzed with the Affymetrix Microarray Suite software version 5.1 (Affymetrix, Inc., Santa Clara, CA) and the four-comparison survival method. The fluorescence of each array was normalized by scaling total chip fluorescence intensities to a common value of 500. For each growth condition, two independent experimental cell cultures (wild type) were compared with two independent control groups ( luxS mutant), and four comparisons were made. The fold change for each gene was calculated as the ratio of signal intensity for the wild type to the signal intensity for the  luxS mutant. The reported value for the fold change is the average of the four comparisons. Genes with a consistent increase or decrease in all comparisons were determined and used for the analysis.

organism: Escherichia coli

source_name: E. coli W3110 grown in LB plus 0.8% glucose at OD 1.0

label_protocol: The Enzo BioArray terminal labeling kit with biotin-ddUTP (Affymetrix, Inc., Santa Clara, CA) was used to label the 3' termini of the fragmented cDNA.

growth_protocol: After inoculation from overnight cultures, E. coli cells were grown in LB medium plus 0.8% glucose at at 30C with shaking at 250 rpm in 50 ml flasks  until OD 1.0.

extract_protocol: Extraction of total RNA was performed using Qiagen Rneasy Mini Kit according to the manufacturer's instructions.

treatment_protocol: Cells were harested and stored at -80C

tag: E. coli W3110 wild type


title: E. coli W3110 grown in in LB plus glucose at OD 1.0 (WT_Glc1)

status: Public on Nov 14 2007

type: RNA

country: USA

label: biotin

molecule: total RNA

tag: W3110 wild type


title: SmutansUA159_Control_rep1

city: Gainesville

first: Robert

middle: A

last: Burne

state: FL

address: 1600 SW Archer Rd. BOX 100424

department: Oral Biology

institute: University of Florida

description: Streptococcus mutans wildtype, untreated control

hyb_protocol: Hybridizations were carried out in a Maui 4-chamber hybridization system (BioMicro Systems, Salt Lake City, Utah) for 17 h at 42ºC. The slides were then washed according to TIGR protocols (SOT# M008, version 2.1, 11/06).

scan_protocol: The slides were scanned using a GenePix Scanner (Axon Instruments Inc., Union City, CA).

data_processing: After the slides were scanned, single-channel images were loaded into TIGR Spotfinder software and overlayed. A spot grid was created according to TIGR specifications and manually adjusted to fit all spots within the grid, then the intensity values of each spot were measured. Data was normalized using Microarray data analysis software (MIDAS), by using LOWESS and Iterative Log Mean Centering with default settings.

organism: Streptococcus mutans

source_name: S. mutans UA159 control condition

label_protocol: All RNAs were purified as described above and used to generate cDNA according to the protocol provided by TIGR with the following minor modifications. The amount of RNA in each reaction was increased to 10 ug and the molar ratio of dTTP:aa-dUTP was increased to 1:2. SuperscriptIII Reverse transcriptase (Invitrogen, Gaithersburg, MD) was used to increase cDNA yields. Purified cDNAs were coupled with indocarbocyanine (Cy3)-dUTP

growth_protocol: Cells were grown in the chemically-defined medium FMC  to an OD600 of 0.3 in 5% CO2 at 37°C

extract_protocol: RNA from S. mutans was isolated from 50 ml cultures that were grown under the desired conditions and harvested by centrifugation at 4ºC. Pelleted cells were resuspended in 400 ml of DEPC-treated water, 800 ml of RNA protect reagent (QIAGEN, Inc., Chatsworth, CA) was added, and the samples were incubated at room temperature for 5 min with vortexing for 10 s at one minute intervals. Cells were then pelleted, resuspended in 500 ml Tris-EDTA (50:10) buffer and transferred to 1.5 ml screw cap tubes containing sterile glass beads (0.1 mm avg. diam.; Biospec, Bartlesville, OK), 100 ml of 1% SDS and 650 ml of acid phenol:chloroform (5:1). The mixture was homogenized in a Bead Beater twice for 40 s and chilled on ice for 2 min between cycles. Samples were centrifuged for 30 min at maximum speed at 4ºC. Two additional hot acid phenol:chloroform extractions were performed followed by one extraction with chloroform:isoamyl-alcohol (24:1). RNA was precipitated in the presence of isopropanol and sodium acetate, pH 5, at -20ºC for 1 h, followed by multiple washes with 70% ethanol, one wash with 99% ethanol, and drying in vacuo. RNA was resuspended in DEPC-treated water and digested with DNAseI (Ambion, Austin, TX). The RNA was re-purified and treated on-column with DNaseI using the RNeasy mini kit (QIAGEN, Inc., Chatsworth, CA), as recommended by the supplier.

treatment_protocol: Untreated (control)

tag: Strain UA159

source_name: S. mutans UA159 grown in BHI

label_protocol: All RNAs were purified as described above and used to generate cDNA according to the protocol provided by TIGR with the following minor modifications. The amount of RNA in each reaction was increased to 10 ug and the molar ratio of dTTP:aa-dUTP was increased to 1:2. SuperscriptIII Reverse transcriptase (Invitrogen, Gaithersburg, MD) was used to increase cDNA yields. Purified cDNAs were coupled with indocarbocyanine (Cy5)-dUTP

growth_protocol:
Growth Media: Brain Heart Infusion
OD600: 0.5
5% CO2, 37°C


title: SmutansJLrelA_Control_rep3

description: Streptococcus mutans relA mutant, no mupricin treatment

source_name: S. mutans JLrelA control condition

tag: Strain JLrelA

value: relA kockout


title: E. coli W3110 luxS mutant in LB plus glucose at OD 1.0 (luxS-_Glc2)

description: Gene expression data for W3110 luxS mutant grown in LB plus 0.8% glucose at OD 1.0

data_processing: Microarray data were analyzed with the Affymetrix Microarray Suite software version 5.1 (Affymetrix, Inc., Santa Clara, CA) and the four-comparison survival method (7). The fluorescence of each array was normalized by scaling total chip fluorescence intensities to a common value of 500. For each growth condition, two independent experimental cell cultures (wild type) were compared with two independent control groups ( luxS mutant), and four comparisons were made. The fold change for each gene was calculated as the ratio of signal intensity for the wild type to the signal intensity for the  luxS mutant. The reported value for the fold change is the average of the four comparisons. Genes with a consistent increase or decrease in all comparisons were determined and used for the analysis.

source_name: E. coli W3110 luxS mutant in LB plus 0.8% glucose at OD 1.0

tag: E. coli W3110 luxS::Kan


title: pRb-/-;p53-/-.1

city: Madrid

first: Ramon

last: Garcia-Escudero

state: Madrid

address: Avda Complutense 40, Ed 70A

department: Biomedical Innovation

country: Spain

institute: CIEMAT

description: RNA integrity tested with Bioanalyzer (Agilent)

hyb_protocol: Affymetrix Protocol

scan_protocol: Affymetrix Protocol

data_processing: RMA transformed, log2 signal values mean centered with mean=0 and stdv=1

organism: Mus musculus

source_name: Whole skin tissue

label_protocol: Affymetrix Protocol

extract_protocol: Mouse skin tissue, obtained after birth, was preserved in RNAlater (Ambion), and disrupted and homogenized using Mixer Mill MM301 (Retsch). Total RNA was extracted and purified from 30 mg of skin using RNeasy Fibrous Tissue Mini kit (Qiagen) following manufacturers' recommendations

tag: Age

value: newborn

tag: Strain

value: FVB

tag: Tissue

value: skin


title: E. coli W3110 luxS mutant grown in LB plus glucose at OD 1.0 (luxS-_Glc1)

source_name: E. coli W3110 luxS mutant grown in LB plus 0.8% glucose at OD 1.0


title: SmutansJLrelA_Control_rep2


title: pRb+/+;p53-/-.2

tag: Trp53 F1/F10; K14 cre


title: pRb-/-;p53-/-.2


title: SmutansJLrelA_Control_rep1


title: pRb-/-;p53+/+.2

tag: Rb F19/F19; K14 cre


title: E. coli luxS mutant grown in LB at OD 2.4 (luxS-_LB2)

description: Gene expression data for E. coli W3110 luxS mutant grown in LB medium at OD 2.4

growth_protocol: After inoculation from overnight cultures, E. coli cells were grown in LB medium at at 30C with shaking at 250 rpm in 50 ml flasks  until OD 2.4.


title: E. coli W3110 wild type grown in LB medium at OD 2.4 (WT_LB3)

description: Gene expression data for W3110 wild type grown in LB medium at OD 2.4

source_name: E. coli W3110 in LB medium at OD 2.4


title: pRb-/-;p53+/+.1


title: pRb+/+;p53-/-.1


title: E. coli W3110 luxS mutant grown in LB medium at OD 2.4 (luxS-_LB1)

description: Gene expression data for W3110 luxS mutant grown in LB medium at OD 2.4

source_name: E. coli W3110 luxS mutant in LB medium at OD 2.4


title: E. coli W3110 in LB medium at OD 2.4 (WT_LB2)

description: Gene expression data for W3110 wild type in LB medium at OD 2.4


title: Control2

label: Biotin

extract_protocol: Mouse skin tissue, obtained after birth, was preserved in RNAlater (Ambion), and disrupted and homogenized using Mixer Mill MM301 (Retsch). Total RNA was extracted and purified from 30 mg of skin using RNeasy Fibrous Tissue Mini kit (Qiagen) following manufacturers' recommendations.


title: pRb-/-;p107-/-2

status: Public on Nov 13 2007

hyb_protocol: Affymetrix protocol

scan_protocol: Affymetrix protocol

data_processing: RMA normalization, log2 transformed, probe value mean-centered, mean=0, stdv=1

label_protocol: Affymetrix protocol

tag: age

tag: strain

tag: tissue


title: pRb+/+;p107-/-.2


title: Control1


title: pRb-/-;p107-/-.1

source_name: pooled primary keratinocyte cultures

growth_protocol: Primary keratinocytes were obtained from newborn mice as described in Santos et al J Biol Chem 277, 19122 (2002). Pooled cells from five different mice of same genotype were cultured in low Ca2+ medium for no longer that 3 days without passage.

extract_protocol: Total RNA was extracted with Trizol and purified using RNeasy Mini kit (Qiagen) following manufacturers' recommendations.


title: pRb-/-;p107-/-.2


title: pRb-/-;p107+/+.1


title: pRb+/+;p107-/-.1


title: pRb-/-;p130-/-.2


title: pRb-/-;p130-/-.1


title: pRb-/-;p107-/-1


title: pRb-/-;p107+/+.2


title: wild type 3


title: pRb-/-.2


title: wild type 1


title: wild type 2


title: pRb+/+.1


title: pRb+/+.2


title: pRb-/-.1


title: vervet magnocellular layer 2

status: Public on Nov 12 2007

city: Montreal

first: Pascal

last: Lachance

state: QC

address: 1205 Ave. Dr. Penfield

department: Psychology

country: Canada

institute: McGill University

description: vervet monkey samples cross-species hybridized to the Rhesus Macaque Genome Array (Affymetrix)

hyb_protocol: Standard Affymetrix protocol, processed at the McGill University and Genome Quebec Innovation Center.

data_processing: CHP file obtained from MAS 5.0

organism: Chlorocebus aethiops vervet

source_name: lateral geniculate nucleus magnocellular heterogeneous layer

extract_protocol:
Laser-capture microdissected heterogeneous layer (mixture of neuronal cell bodies, neuropil, glia) from the magnocellular layer of the Lateral Geniculate Nucleus. Cells identified by cresyl violet staining.
Sample consists of roughly 3000 pulses from LCM, total RNA amplified through two rounds using the Affymetrix system.

treatment_protocol: normal animals

tag: Adult male vervet monkey (Cercopithecus aethipos), internal identifier number 05-1-2.


title: vervet parvocellular layer 1

scan_protocol: Standard Affymetrix protocol, processed at the McGill University and Genome Quebec Innovation Center.

source_name: lateral geniculate nucleus parvocellular heterogeneous layer

extract_protocol:
Laser-capture microdissected heterogeneous layer (mixture of neuronal cell bodies, neuropil, glia) from the parvocellular layer of the Lateral Geniculate Nucleus. Cells identified by cresyl violet staining.
Sample consists of roughly 3000 pulses from LCM, total RNA amplified through two rounds using the Affymetrix system.


title: pRb-/-.3


title: vervet magnocellular layer 1

label_protocol: Standard Affymetrix protocol, processed at the McGill University and Genome Quebec Innovation Center.

tag: Adult male vervet monkey (Cercopithecus aethipos), internal identifier number 03-1-2.


title: vervet parvocellular neuron 3

source_name: lateral geniculate nucleus parvocellular neuronal cell body

extract_protocol:
Laser-capture microdissected neuronal cell bodies from the parvocellular layer of the Lateral Geniculate Nucleus. Cells identified by cresyl violet staining.
Sample consists of roughly 3000 pulses from LCM, total RNA amplified through two rounds using the Affymetrix system.

tag: Adult male vervet monkey (Cercopithecus aethipos), internal identifier number 05-1-4.


title: vervet parvocellular neuron 2


title: vervet parvocellular neuron 1


title: vervet magnocellular neuron 3

source_name: lateral geniculate nucleus magnocellular neuronal cell body

extract_protocol:
Laser-capture microdissected neuronal cell bodies from the magnocellular layer of the Lateral Geniculate Nucleus. Cells identified by cresyl violet staining.
Sample consists of roughly 3000 pulses from LCM, total RNA amplified through two rounds using the Affymetrix system.


title: vervet magnocellular layer 3


title: vervet parvocellular layer 3


title: vervet parvocellular layer 2


title: vervet magnocellular neuron 2

tag: Adult male vervet monkey (Cercopithecus aethipos).


title: White turkey liver, replicate 2

city: Newark

first: Larry

middle: Albert

last: Cogburn

state: DE

address: 531 South College Ave.

department: Animal and Food Sciences

institute: University of Delaware

description: White turkey liver, replicate 2

hyb_protocol: The microarray was hybridized overnight at 42 degrees Celsius with Cy3-labeled cDNAs from the target sample and an aliquot of the Cy5-labeled reference RNA pool cDNAs using microarray hybridization buffer (Amersham Biosciences). On the following day, slides were sequentially washed with 1 x SSC, 0.2% SDS at 50 degrees Celsius for 10 min, then 0.1 x SSC and 0.2% SDS for 5 min at room temperature, and finally 0.1 x SSC for 1 min at room temperature. The slides were subsequently rinsed in dH2O and dried by centrifugation.

scan_protocol: The microarray slides were scanned using a 418 confocal laser scanner (Affymetrix) at 550 nM for Cy3 and 649 nM for Cy5 to generate two TIFF images for each slide. The photomultiplier tube (PMT) gain for the Cy3/Cy5 ratio was set to 1. For the high resolution scan for each dye, the PMT gain was set to achieve 1% saturation.

data_processing: For each microarray the high-resolution TIFF image files were analyzed using GenePix Pro V4.1 software (Axon Laboratories, Palo Alto, CA). The GenePix Pro report (gpr) files were automatically merged with Excel files containing the clone identification number/plate address and gene name/function (from the highest BLAST score). Data in the VALUE column is based on the loess-normalized ratio of the loess-normalized log2 Cy3 and Cy5 values. The values are based on non-background corrected median values.

organism: Meleagris gallopavo

source_name: White turkey liver

label_protocol: For indirect labeling, 25 micrograms of total RNA was converted to cDNA using random primers with the Amino Allyl cDNA Labeling Kit (Ambion), and Cy3-monoreactive NHS esters (Amersham Biosciences, Piscataway, NJ) were coupled to the cDNA. Labeled cDNA was purified from unincorporated dye using CyScribe GFX Purification Kit (Amersham Biosciences)

extract_protocol: Total RNA from frozen tissue sample was isolated using a RNeasy Midi kit (Qiagen, Valencia, CA) following the manufacturer's protocol with some modifications. The quantity of extracted total RNA was determined using a NanoDrop spectrophotometer (Wilmington, DE), and the quality of extracted total RNA was examined by microcapillary electrophoresis on a BioAnalyzer 2100 (Agilent, Wilmington, DE), where the ribosomal RNA ratio (28S/18S) was observed for integrity.

treatment_protocol: Tissue samples were excised immediately after death, snap frozen in liquid nitrogen, and stored at -80 degrees Celsius until RNA isolation.

tag: Medium white turkey liver

organism: Gallus gallus

source_name: Multi-tissues reference RNA pool

label_protocol: For each sample, a 25 microgram aliquot of reference RNA pool was labeled with Cy5 (red) as described above for channel 1. For each hybridization set, a final reference pool was made of all Cy5-labeled cDNA and equally aliquoted for all slides hybridized that day.

growth_protocol: Control Broiler chicks (Ross x Cobb)were raised under conditions described above for channel 1. Lean line chickens were derived from a population of broiler chickens divergently selected for high and low abdominal fat content (Leclercq et al., 1980).The lean line cockerels were raised in floor pens in a 4.4 x 3.9 square meter room. Pelleted feed (22% crude protein and 3100 kcal ME/kg) was provided throughout the experiment. After continuous light for the first two days, the birds were maintained on a 14L:10D light/dark cycle. Supplemental heat was provided by infrared gas and ambient temperature was progressively decreased from 32 degrees Celsius at hatching until 22 degrees Celsius was reached at 22 days. The birds were provided with water and feed ad libitum.

extract_protocol: Total RNA from frozen liver and hypothalamus samples was isolated using a RNeasy Midi kit (Qiagen, Valencia, CA) following the manufacturer's protocol with some modifications. Total RNA from breast muscle and abdominal fat was isolated using the guanidine thiocyanate lysis method and CsCl gradient purification. The quantity of extracted total RNA was determined using a NanoDrop spectrophotometer (Wilmington, DE) and the quality of extracted total RNA was examined by microcapillary electrophoresis on a BioAnalyzer 2100 (Agilent, Wilmington, DE), where the ribosomal RNA ratio (28S/18S) was examined for integrity.

treatment_protocol: Control broiler cockerels were used as a source of liver, breast muscle and abdominal fat. Lean line chickens were used as a source for hypothalamic samples. Tissues were excised immediately after death, snap frozen in liquid nitrogen, and stored at -80 degrees Celsius until RNA isolation.

tag: Multi-tissue RNA pool was made from an equal amount of total RNA from chicken liver, hypothalamus, breast muscle and abdominal fat.


title: White turkey liver, replicate 4

description: White turkey liver, replicate 4


title: White turkey hypothalamus, replicate 3

description: White turkey hypothalamus, replicate 3

source_name: White turkey hypothalamus

tag: Medium white turkey hypothalamus


title: White turkey hypothalamus, replicate 1

description: White turkey hypothalamus, replicate 1


title: White turkey liver, replicate 1

description: White turkey liver, replicate 1


title: White turkey liver, replicate 3

description: White turkey liver, replicate 3


title: Japanese quail liver, replicate 1

description: Japanese quail liver, replicate 1

organism: Coturnix japonica

source_name: Japanese quail liver

growth_protocol: Japanese quail maintained in Department at the University of Maryland, College Park were bred randomly and become sexually mature when 8 weeks old. Birds received feed (Purina Game Bird Startina or Layena) and water ad libitum. They were hatched and brooded under controlled conditions for 6 weeks, then transferred to colony cages in mixed sex groups. Starting at 9–12 months of age, birds were maintained singly and housed individually. The environment was temperature and light (15L: 9D; light on at 0600) controlled. Animals and facilities were maintained according to NIH Assurance Statement (A3270-01) and research was conducted under University of Maryland Institutional Animal Care and Use Committee approved protocols.

tag: Adult Japanese quail liver


title: White turkey hypothalamus, replicate 2

description: White turkey hypothalamus, replicate 2


title: Japanese quail liver, replicate 4

description: Japanese quail liver, replicate 4


title: Japanese quail liver, replicate 3

description: Japanese quail liver, replicate 3


title: Japanese quail hypothalamus, replicate 2

source_name: Japanese quail hypothalamus

growth_protocol: Japanese quail (maintained in Department at the University of Maryland, College Park) were bred randomly and become sexually mature when 8 weeks old. Birds received feed (Purina Game Bird Startina or Layena) and water ad libitum. They were hatched and brooded under controlled conditions for 6 weeks, then transferred to colony cages in mixed sex groups. Starting at 9–12 months of age, birds were maintained singly and housed individually. The environment was temperature and light (15L: 9D; light on at 0600) controlled. Animals and facilities were maintained according to NIH Assurance Statement (A3270-01) and research was conducted under University of Maryland Institutional Animal Care and Use Committee approved protocols.

tag: Adult Japanese quail hypothalamus


title: Tree sparrow liver, replicate 1

description: Tree sparrow liver, replicate 1

organism: Passer montanus

source_name: Tree sparrow liver

growth_protocol: The tree sparrows were from a captive colony maintained at the University of Maryland. Animals and facilities were maintained according to NIH Assurance Statement (A3270-01) and research was conducted under approved protocols from University of Maryland Institutional Animal Care and Use Committee.

tag: Adult tree sparrow liver


title: Tree sparrow liver, replicate 3

description: Tree sparrow liver, replicate 3


title: Duck liver, replicate 4

description: Duck liver, replicate 4

organism: Anas platyrhynchos

source_name: Duck liver

growth_protocol: Ducks were fed a standard ration ad libitum and raised in floor pens containing a wading pool.

tag: Duck liver (5 weeks of age)


title: Japanese quail hypothalamus, replicate 3

description: Japanese quail hypothalamus, replicate 19


title: American kestrel liver, replicate 1

description: American kestrel liver, replicate 1

organism: Falco sparverius

source_name: American kestrel liver

growth_protocol: American kestrels were offspring from a captive colony maintained at the U.S. Geological Survey Patuxent Wildlife Research Center (Laurel, MD, USA). Animals and facilities were maintained according to NIH Assurance Statement (A3270-01); research was conducted under approved protocols from the University of Maryland Institutional Animal Care and Use Committee.

tag: American kestrel liver


title: Japanese quail hypothalamus, replicate 1


title: vervet magnocellular neuron 1

tag: Adult male normal vervet monkey (Cercopithecus aethipos), internal identifier 03-1-2.


title: Tree sparrow liver, replicate 2

description: Tree sparrow liver, replicate 2


title: Japanese quail liver, replicate 2

description: Japanese quail liver, replicate 2


title: Duck liver, replicate 3

description: Duck liver, replicate 3


title: Duck hypothalamus, replicate 2

description: Duck hypothalamus, replicate 2

source_name: Duck hypothalamus

tag: Duck hypothalamus (5 weeks of age)


title: Duck liver, replicate 1

description: Duck liver, replicate 1


title: Chicken liver, 6 weeks of age, replicate 4

description: Chicken liver, replicate 4

source_name: Chick liver

growth_protocol: Chicks were fed a broiler starter ration and raised in a controlled-environment animal room under a 20-h light:4-h darkness photoperiod in a heated battery-brooder until 3 weeks of age. At 3 weeks of age, birds were transferred to wire cages within a controlled-temperature room with four birds per pen. Rooms were maintained under the same 20L:4D light/dark cycle from three to six weeks of age. The birds were provided with water and a commercial grower/finisher ration ad libitum.

treatment_protocol: Six-week old control broiler cockerels (Ross x Cobb) were used as a tissue source. Tissue samples were excised immediately after death, snap frozen in liquid nitrogen, and stored at -80 degrees Celsius until RNA isolation.

tag: Chicken liver, 6 weeks of age


title: Duck hypothalamus, replicate 1

description: Duck hypothalamus, replicate 1


title: American kestrel liver, replicate 3

description: American kestrel liver, replicate 3


title: Duck hypothalamus, replicate 3

description: Duck hypothalamus, replicate 3


title: Duck liver, replicate 2

description: Duck liver, replicate 2


title: American kestrel liver, replicate 2

description: American kestrel liver, replicate 2


title: Chicken hypothalamus, lean line, 9 weeks of age, replicate 2

description: Hypothalamus from lean line chickens, replicate 2

source_name: Chick hypothalamus

label_protocol: For indirect labeling, 25 micrograms of total RNA was converted to cDNA using random primers with the Amino Allyl cDNA Labeling Kit (Ambion), and Cy3-monoreactive NHS esters (Amersham Biosciences, Piscataway, NJ) were coupled to the cDNA. Labeled cDNA was purified from unincorporated dye using CyScribe GFX Purification Kit (Amersham Biosciences).

growth_protocol: Male chickens used in this study were from lean line, which were derived from a population of broiler chickens divergently selected for high and low abdominal fat content (Leclercq et al., 1980). Tissues were excised immediately after death, snap frozen in liquid nitrogen, and stored at -80 degree Celsius until RNA isolation. The LL cockerels were raised in floor pens in a 4.4 x 3.9 m2 room. Pelleted feed (22% crude protein and 3100 kcal ME/kg) was provided throughout the experiment. After continuous light for the first two days, the birds were maintained on a 14L:10D light/dark cycle. Supplemental heat was provided by infrared gas and ambient temperature was progressively decreased from 32 degree Celsius at hatching until 22 degree Celsius was reached at 22 days. The birds were provided with water and feed ad libitum.

extract_protocol: Total RNA from frozen tissue samples was isolated using a RNeasy Midi kit (Qiagen, Valencia, CA) following the manufacturer's protocol with some modifications. The quantity of extracted total RNA was determined using a NanoDrop spectrophotometer (Wilmington, DE), and the quality of extracted total RNA was examined by microcapillary electrophoresis on a BioAnalyzer 2100 (Agilent, Wilmington, DE), where the ribosomal RNA ratio (28S/18S) was observed for integrity.

treatment_protocol: Divergently selected for leanness.

tag: Chicken hypothalamus lean line cockerel, 9 weeks of age


title: Chicken liver, 6 weeks of age, replicate 3

description: Chicken liver, replicate 3


title: Chicken abdominal fat, 6 weeks of age, replicate 4

description: Chicken abdominal fat, replicate 4

source_name: Chick abdominal fat

extract_protocol: Total RNA was isolated from chicken abdominal fat by the guanidine thiocyanate lysis method and CsCl gradient purification (Chirgwin et al., 1979 Biochem. 18: 5294–5299). The quantity of extracted total RNA was determined using a NanoDrop spectrophotometer (Wilmington, DE), and the quality of extracted total RNA was examined by microcapillary electrophoresis on a BioAnalyzer 2100 (Agilent, Wilmington, DE), where the ribosomal RNA ratio (28S/18S) was examined for integrity.

tag: Chicken abdominal fat, 6 weeks of age

source_name: Multi-tissue reference RNA pool


title: Chicken hypothalamus, lean line, 9 weeks of age, replicate 1

description: Hypothalamus from lean line chickens, replicate 1


title: Chicken abdominal fat, 6 weeks of age, replicate 3

description: Chicken abdominal fat, replicate 3


title: Chicken hypothalamus, lean line, 9 weeks of age, replicate 4

description: Hypothalamus from lean line chickens, replicate 4


title: Chicken liver, 6 weeks of age, replicate 1

description: Chicken liver, replicate 1


title: Chicken hypothalamus, lean line, 9 weeks of age, replicate 3

description: Hypothalamus from lean line chickens, replicate 3


title: Chicken liver, 6 weeks of age, replicate 2

description: Chicken liver, replicate 2


title: Chicken breast muscle, 6 weeks of age, replicate 3

description: Chicken breast muscle, replicate 3

source_name: Chick breast muscle

extract_protocol: Total RNA was isolated from chicken breast muscle by the guanidine thiocyanate lysis method and CsCl gradient purification (Chirgwin et al., 1979 Biochem. 18: 5294–5299). The quantity of extracted total RNA was determined using a NanoDrop spectrophotometer (Wilmington, DE), and the quality of extracted total RNA was examined by microcapillary electrophoresis on a BioAnalyzer 2100 (Agilent, Wilmington, DE), where the ribosomal RNA ratio (28S/18S) was examined for integrity.

tag: Chicken breast muscle, 6 weeks of age


title: Chicken abdominal fat, 6 weeks of age, replicate 1

description: Chicken abdominal fat, replicate 1


title: Chicken breast muscle, 6 weeks of age, replicate 1

description: Chicken breast muscle, replicate 1


title: Gene expression in Peyers patches after orogatral Yersinia enterocolitica infection, three days after infection

status: Public on Nov 10 2007

city: Tuebingen

first: Erwin

last: Bohn

address: Elfriede-Aulhorn-Strasse 6

department: Institutefor Medical Microbiology and Hygiene

country: Germany

institute: University of Tuebingen

description: Affymetrix

hyb_protocol: Affymetrix

scan_protocol: Affymetrix

data_processing: Analysis wit Affymetrix microarray suite 5.0 and Data mining tool 3.0

label: Affymetrix

source_name: Peyers patches

label_protocol: Affymetrix

extract_protocol: Rneasy minikit (Qiagen, Hilden)

treatment_protocol: Standard affymetrix procedure

tag: Five C57BL/6 mice were infected for 72 hours with 10 million Yersinia enterocolitica strain WA-314. Pyers patches were isolated, pooled and RNA was isolated


title: Gene expression in Peyers patches after orogatral Yersinia enterocolitica infection, one day after infection

description: Affymetrix

tag: Five C57BL/6 mice were infected for 24 with 10 million Yersinia enterocolitica strain WA-314. Pyers patches were isolated, pooled and RNA was isolated


title: Chicken abdominal fat, 6 weeks of age, replicate 2

description: Chicken abdominal fat, replicate 2


title: Chicken breast muscle, 6 weeks of age, replicate 4

description: Chicken breast muscle, replicate 4


title: Chicken breast muscle, 6 weeks of age, replicate 2

description: Chicken breast muscle, replicate 2


title: Gene expression in Peyers patches after orogatral Yersinia enterocolitica infection, Uninfected

description: Affymetrix

tag: Peyers patches from five uninfected C57BL/6 mice was pooled and total RNA was prepared with the use of RNeasy mini kit


title: wild type 11 mM Pi 6h vs delta phoB(PHO80) 11 mM Pi 6h

status: Public on Nov 09 2007

city: Ribeirao Preto

first: Gustavo

middle: Henrique

last: Goldman

state: Sao Paulo

address: Avenida do Café, s/n

department: Departament of Pharmaceutical Sciences

country: Brazil

institute: Faculty of Pharmaceutical Sciences of Ribeirao Preto

description: Both strains were grown in minimal medium (11 mM phosphate) for 6 hours.

hyb_protocol: Labelled cDNAs were mixed with 8 µl Liquid block RPN3601 Batch 24 (GE Health Care), 5.5 µl SDS 2% and 19 µl SSC (20X) and the volume adjusted to 110 µl. Slides were hybridized overnight (42°C) in a Gene-Tac Hybridization Station (Genomic Solutions, Inc.) and washed in 2 X SSC 0,5% SDS, 0,5 X SSC and 0.05X SSC. All washing steps consisted of 1 min of flow, followed by 5 min of incubation.

scan_protocol: Slides were dried by centrifugation and subjected to fluorescent detection with a GMS 418 Array Scanner (Affymetrix Inc., Santa Clara, CA).

data_processing: TIFF images generated were analyzed using TIGR Spotfinder (http://www.tigr.org/software/) to obtain relative transcript levels. Spotfinder uses a dynamic thresholding algorithm to identify spots and calculate local background and integrated fluorescence intensity for each of the Cy3 and Cy5 channels. This software was also useful in eliminating questionable spots (those for which shape deviates significantly from that of a true circle, or those with a low signal-to-noise ratio). Spots with very low intensity in one channel (and thus aberrantly high or low expression ratios) were removed and eliminated from future analyses. Data was normalized using a local regression technique LOWESS (LOcally WEighted Scatterplot Smoothing) for hybridizations with RNA-based samples and SD regularization of the Cy5/Cy3 ratio across all sectors (blocks) of the array using a software tool MIDAS (http://www.tigr.org/softlab). Following normalization, the values for each gene's in-slide replicates were averaged (median variance <0.01), and corresponding flip-dye replicates were averaged to compensate for dye-specific effects. Differentially expressed genes at the 95% confidence level were determined using intensity-dependent Z-scores (with Z=1.96) as implemented in MIDAS and the union of all genes identified at each time point were considered significant in this experiment. The resulting data were organized and visualized based on similar expression vectors in genes using Euclidean distance and hierarchical clustering with average linkage clustering method to view the whole data set and k-means to group the genes in 60 clusters with TIGR MEV (multi experiment viewer), also available at http://www.tigr.org/software.

organism: Aspergillus fumigatus

source_name: wild type conidia grown in Minimal Medium 11 mM Pi for 6 hours

label_protocol: The RNA samples were extracted with Trizol and further purified with the RNA easy kit (Qiagen, Germany) and directly labelled by incorporation of Cy3-dUTP (GE Health Care). Briefly, 30 µg of total RNA were mixed with 20 µg of Randon Primer Hexamers (GE Health Care), 100 pmol of oligo dTV (Invitrogen) and 40 U of RNAse Out (Invitrogen) in a volume of 13 µl and heated to 70°C for 5 minutes. The tubes were briefly chilled to 4°C for 5 minutes and mixed with 5 µl of 5X first-strand buffer (Invitrogen), 2 µl DTT (Invitrogen), 2 µl dNTP (5 mM dATP, 5 mM dCTP, 5 mM dGTP, 2 mM dTTP), 2 µl of Suprescript II (Invitrogen) and 1 µl of Cy3-dUTP (25 nmol). The reaction was then incubated at 25°C for 5 minutes and at 42°C for 3 hours. The RNA was degraded by adding 2.5 µl EDTA (0.5 M, pH 8) and 5 µl NaOH (1M) following an incubation at 37°C for 40 minutes. The resulting first strand cDNA was purified and concentrated using a Microcon YM-50 cartridge.

extract_protocol: At each time point the germlings were harvested by centrifugation and quickly frozen in liquid nitrogen. For total RNA isolation, the germlings were disrupted by grinding in liquid nitrogen and total RNA was extracted with Trizol reagent (Invitrogen). Ten micrograms of RNA from each treatment were then fractionated in 2.2 M formaldehyde, 1.2% agarose gel, stained with ethidium bromide, and then visualized with UV-light. The presence of intact 25S and 17S ribosomal RNA bands was used as a criterion to assess the integrity of the RNA.

tag: For the microarray experiments, 1.0 x 10(10) conidia of A. fumigatus wild type [delta akuB(ku80)] strain were used to inoculate 300 ml of liquid cultures (Minimal Medium with 11 and 0.1mMol/L of phosphate) in 500-ml erlenmeyer flasks and allow to grow for 6 and 10 hours in a reciprocal shaker (250 rpm) at 37°C. After this time, the cultures were centrifuged and their RNA were extracted. Hybridization experiments were competitive using probes derived from wild type and deltaphoB(PHO80) strain grown for 6 and 10 hours in MM with 11 and 0.1mmol/L of phosphate, using the wild type strain as the reference RNA for every hybridizantion. Normalized signal intensities were used to generate relative hybridization ratios (query/reference).

source_name: delta phoB(PHO80) conidia grown in Minimal Medium 11 mM Pi for 6 hours

label_protocol: The RNA samples were extracted with Trizol and further purified with the RNA easy kit (Qiagen, Germany) and directly labelled by incorporation of Cy5-dUTP (GE Health Care). Briefly, 30 µg of total RNA were mixed with 20 µg of Randon Primer Hexamers (GE Health Care), 100 pmol of oligo dTV (Invitrogen) and 40 U of RNAse Out (Invitrogen) in a volume of 13 µl and heated to 70°C for 5 minutes. The tubes were briefly chilled to 4°C for 5 minutes and mixed with 5 µl of 5X first-strand buffer (Invitrogen), 2 µl DTT (Invitrogen), 2 µl dNTP (5 mM dATP, 5 mM dCTP, 5 mM dGTP, 2 mM dTTP), 2 µl of Suprescript II (Invitrogen) and 1 µl of Cy5-dUTP (25 nmol). The reaction was then incubated at 25°C for 5 minutes and at 42°C for 3 hours. The RNA was degraded by adding 2.5 µl EDTA (0.5 M, pH 8) and 5 µl NaOH (1M) following an incubation at 37°C for 40 minutes. The resulting first strand cDNA was purified and concentrated using a Microcon YM-50 cartridge.

tag: For the microarray experiments, 1.0 x 10(10) conidia of A. fumigatus delta phoB(PHO80) strain were used to inoculate 300 ml of liquid cultures (Minimal Medium with 11 and 0.1mMol/L of phosphate) in 500-ml erlenmeyer flasks and allow to grow for 6 and 10 hours in a reciprocal shaker (250 rpm) at 37°C. After this time, the cultures were centrifuged and their RNA were extracted. Hybridization experiments were competitive using probes derived from wild type and delta phoB(PHO80) strain grown for 6 and 10 hours in MM with 11 and 0.1mmol/L of phosphate, using the wild type strain as the reference RNA for every hybridizantion. Normalized signal intensities were used to generate relative hybridization ratios (query/reference).


title: Mp313E_Rep0

city: Mississippi State

first: Rowena

middle: Yvette

last: Kelley

state: MS

address: 810 Highway 12 East

department: Corn Host Plant Resistance Research Unit (CHPRRU)

institute: USDA-ARS

description: MP313E dye swap

hyb_protocol: 1. 40 μl of the cDNA mixture (20µl of each label), 3 μl Liquid Block™ (Amersham Pharmacia Biotech, Piscataway, NJ), 5 μl 20X SSC and 2 μl 2% (wt/vol) sodium dodecyl sulfate (SDS) were combined, denatured at 95˚C for 2 min and immediately transferred to ice. 2. Array slides were denatured at 65˚C for 30 seconds before application of hybridization mixture, slides were then covered with hybrislips (Sigma-Aldrich, St. Louis, MO), transferred to a pre-warmed hybridization oven (Fisher Scientific, Pittsburg, PA), and incubated overnight at 60˚C. 3. The hybridized slides were washed on an orbital shaker using 2X SSC, 0.5% (wt/vol) SDS for 5 min; 0.5X SSC for 5 min; and 0.05X SSC for 5 min, spun dry at low speed in a centrifuge (100 x g) for 5 min and scanned using a GenePix Personal 4100A (Molecular Devices Corporation, Union City, CA).

scan_protocol: Scanned on an Gene Pix Pro scanner. Images were quantified using Gene Pix Pro Feature Extraction Software (version 4000B).

data_processing:
All data were analyzed with SAS Version 9.1.3 (SAS, Cary, NC). The null hypothesis was defined as follows:
Null Hypothesis:
(resistant inoculated expression)/(resistant uninoculated expression)=(susceptible inoculated expression)/(susceptible uninoculated expression)
The median expression level for each spot intensity was log transformed and analyzed by analysis of variance for a split plot design. The main unit was genotype and the subunit treatment was inoculation. Dye was treated as a fixed effect in the model to account for differences in dyes.  Therefore, Genotype, Inoculation Treatment, Genotype x Inoculation, Treatment, and Dye were fixed effects in the analysis. Rep (replication), Genotype x Rep, Spots (Rep Genotype), Rep x Inoculation, Treatment (Genotype), and residual subsampling error were the components of error in the analysis. F-test for Genotype x Inoculation Treatment interaction was used to address the null hypothesis shown in the above equation. The mathematical notation for Genotype x Inoculation treatment was H0: Log (YR,I) - Log (YR,UI) =  Log (YS,I) - Log (YS,UI), where Y= “Yes, expressed gene”, and R, S, I, and UI refer to resistant, susceptible, inoculated, and non-inoculated, respectively.

organism: Zea mays

source_name: Total RNA from inoculated Mp313E maize ear labeled with Cyanine-3 (green).

label_protocol:
Isolated mRNAs were used for the preparation of Cy3-labeled and Cy5-labeled cDNA probes.
Reverse transcription (RT) PCR (Invitrogen, Carlsbad, CA) was used with random hexamer primers to generate cDNAs from the mRNA templates as per manufacturer’s instructions.
The labeled cDNAs were collected and used immediately for hybridization.

growth_protocol: The maize inbred lines Va35 and Mp313E were selected for this study. Maize line Va35 has yellow kernels and is susceptible to infection by A. flavus (Henderson 1976). Mp313E is a white dent inbred line and was released primarily as a source of resistance to kernel infection by A. flavus (Scott and Zummo 1990).  Seeds of Va35 and Mp313E are maintained by the United States Department of Agriculture, Agricultural Research Service, Corn Host Plant Resistance Research Unit (USDA-ARS-CHPRRU) at the R. R. Foil Plant Science Farm, Mississippi State, MS. The field experimental design was a randomized complete block arranged as a split plot with three replicates.  The treatment design was a 2 x 2 factorial with two genotypes (resistant Mp313E and susceptible Va35) and two inoculation treatments (inoculated and non-inoculated).  Genotypes were planted in main plots, and inoculation treatments were applied to subplots. Rows were 4 meters long and spaced 0.97 meters apart with a fallow alley of 1 meter.  Primary ears on all plants in all plots were self-pollinated. Plots received supplemental furrow irrigation throughout the growing season to mitigate drought stress. Herbicides and fertilizer (application based on soil tests) were applied according to standard cultural practices in corn for a continuous production system in northern Mississippi.

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

treatment_protocol:
A. flavus isolate NRRL 3357 (ATCC # 200026; SRRC 167), a wild-type strain widely used in laboratory and field studies, known to produce high levels of aflatoxin in corn grain (Windham and Williams 2002), was chosen for this study. Cultures were grown on sterile corn cob grits in 500-ml flasks, each containing 50 g of grits and 100 ml of H2O, and incubated at 28°C for 3 weeks. Conidia were washed from the grits using 500 ml sterile distilled water containing 0.02% (vol/vol) Tween 20 (Sigma-Aldrich Co., USA) (polyoxyethylene (20) sorbitol monolaurate) and filtered through four layers of sterile cheesecloth.  Conidial concentrations were determined by hemacytometer and adjusted with sterile distilled water to 9 x 107 conidia/ml. Inoculum not used immediately were refrigerated at 4°C. All maize lines were inoculated 14 days after pollination (DAP) using the side-needle technique (Zummo and Scott 1989).
Plants in inoculated plots were injected between the husks and the kernels with 3.4 ml of a suspension containing approximately 3 x 108 spores/ml of A. flavus conidia. Typically, two to three kernels are damaged by this inoculation technique.

tag: Mp313E resistant line

tag: Inoculum

tag: Age

value: day 2 after inoculation

tag: Tissue

value: Maize ear

source_name: Total RNA from uninoculated Mp313E maize ear labeled with Cyanine-5 (red).

treatment_protocol: All maize lines were in the non-inoculated plots were collected 16 days after pollination (DAP).

value: day 16 after pollination


title: Mp313E_Rep2

description: MP313E Rep 2

source_name: Total RNA from inoculated Mp313E maize ear labeled with Cyanine-5 (red).

source_name: Total RNA from uninoculated Mp313E maize ear labeled with Cyanine-3 (green).


title: wild type 0.1 mM Pi 10h vs delta phoB(PHO80) 0.1 mM Pi 10h dye swap

description: Both strains were grown in modified minimal medium (0.1 mM phosphate) for 10 hours.

source_name: wild type conidia grown in Minimal Medium 0.1 mM Pi for 10 hours

label_protocol: The RNA samples were extracted with Trizol and further purified with the RNA easy kit (Qiagen, Germany) and directly labelled by incorporation of Cy5-dUTP (GE Health Care). Briefly, 30 µg of total RNA were mixed with 20 µg of Randon Primer Hexamers (GE Health Care), 100 pmol of oligo dTV (Invitrogen) and 40 U of RNAse Out (Invitrogen) in a volume of 13 µl and heated to 70°C for 5 minutes. The tubes were briefly chilled to 4°C for 5 minutes and mixed with 5 µl of 5X first-strand buffer (Invitrogen), 2 µl DTT (Invitrogen), 2 µl dNTP (5 mM dATP, 5 mM dCTP, 5 mM dGTP, 2 mM dTTP), 2 µl of Suprescript II (Invitrogen) and 1 µl of Cy3-dUTP (25 nmol). The reaction was then incubated at 25°C for 5 minutes and at 42°C for 3 hours. The RNA was degraded by adding 2.5 µl EDTA (0.5 M, pH 8) and 5 µl NaOH (1M) following an incubation at 37°C for 40 minutes. The resulting first strand cDNA was purified and concentrated using a Microcon YM-50 cartridge.

source_name: deltaphoB(PHO80) conidia grown in Minimal Medium 0.1 mM Pi for 10 hours

label_protocol: The RNA samples were extracted with Trizol and further purified with the RNA easy kit (Qiagen, Germany) and directly labelled by incorporation of Cy3-dUTP (GE Health Care). Briefly, 30 µg of total RNA were mixed with 20 µg of Randon Primer Hexamers (GE Health Care), 100 pmol of oligo dTV (Invitrogen) and 40 U of RNAse Out (Invitrogen) in a volume of 13 µl and heated to 70°C for 5 minutes. The tubes were briefly chilled to 4°C for 5 minutes and mixed with 5 µl of 5X first-strand buffer (Invitrogen), 2 µl DTT (Invitrogen), 2 µl dNTP (5 mM dATP, 5 mM dCTP, 5 mM dGTP, 2 mM dTTP), 2 µl of Suprescript II (Invitrogen) and 1 µl of Cy5-dUTP (25 nmol). The reaction was then incubated at 25°C for 5 minutes and at 42°C for 3 hours. The RNA was degraded by adding 2.5 µl EDTA (0.5 M, pH 8) and 5 µl NaOH (1M) following an incubation at 37°C for 40 minutes. The resulting first strand cDNA was purified and concentrated using a Microcon YM-50 cartridge.


title: Mp313E_Rep1

description: MP313E Rep 1


title: wild type 11 mM Pi 10h vs delta phoB(PHO80) 11 mM Pi 10h dye swap

description: Both strains were grown in minimal medium (11 mM phosphate) for 10 hours.

source_name: wild type conidia grown in Minimal Medium 11 mM Pi for 10 hours

source_name: delta phoB(PHO80) conidia grown in Minimal Medium 11 mM Pi for 10 hours


title: wild type 0.1 mM Pi 10h vs delta phoB(PHO80) 0.1 mM Pi 10h

source_name: delta phoB(PHO80) conidia grown in Minimal Medium 0.1 mM Pi for 10 hours


title: wild type 11 mM Pi 10h vs delta phoB(PHO80) 11 mM Pi 10h


title: wild type 11 mM Pi 6h vs delta phoB(PHO80) 11 mM Pi 6h dye swap


title: Va35_Rep3

description: VA35 Rep 3

source_name: Total RNA from inoculated Va35 maize ear labeled with Cyanine-5 (red).

tag: Va35 seceptible line

source_name: Total RNA from uninoculated Va35 maize ear labeled with Cyanine-3 (green).


title: wild type 0.1 mM Pi 6h vs delta phoB(PHO80) 0.1 mM Pi 6h

description: Both strains were grown in modified minimal medium (0.1 mM phosphate) for 6 hours.

source_name: wild type conidia grown in Minimal Medium 0.1 mM Pi for 6 hours

source_name: delta phoB(PHO80) conidia grown in Minimal Medium 0.1 mM Pi for 6 hours


title: Mp313E_Rep3

description: MP313E Rep 3


title: wild type time 0 min vs wild type time 10 min upon 200 mM CaCl2 dye swap

description: This hybridization used the wild type RNA from time point zero (16 hours grown) as reference. The test RNA was obtained from a 16 hours culture treated for 10 minutes with 200 mM calcium chloride.

hyb_protocol: Labelled cDNAs were mixed with 8 µl Liquid block RPN3601 Batch 24 (GE Health Care), 5.5 µl SDS 2% and 19 µl SSC (20X) and the volume adjusted to 110 µl. Slides were hybridized overnight (42°C) in a Gene-Tac Hybridization Station (Genomic Solutions, Inc.) and washed in 2x SSC 0,5% SDS, 0,5x SSC and 0.05x SSC. All washing steps consisted of 1 min of flow, followed by 5 min of incubation

data_processing: TIFF images generated were analyzed using TIGR Spotfinder (http://www.tigr.org/software/) to obtain relative transcript levels. Spotfinder uses a dynamic thresholding algorithm to identify spots and calculate local background and integrated fluorescence intensity for each of the Cy3 and Cy5 channels. This software was also useful in eliminating questionable spots (those for which shape deviates significantly from that of a true circle, or those with a low signal-to-noise ratio). Spots with very low intensity in one channel (and thus aberrantly high or low expression ratios) were removed and eliminated from future analyses. Data was normalized using a local regression technique LOWESS (LOcally WEighted Scatterplot Smoothing) for hybridizations with RNA-based samples and SD regularization of the Cy5/Cy3 ratio across all sectors (blocks) of the array using a software tool MIDAS (http://www.tigr.org/softlab). Following normalization, the values for each gene's in-slide replicates were averaged (median variance <0.01), and corresponding flip-dye replicates were averaged to compensate for dye-specific effects. Differentially expressed genes at the 95% confidence level were determined using intensity-dependent Z-scores (with Z=1.96) as implemented in MIDAS and the union of all genes identified at each time point were considered significant in this experiment. The resulting data were organized and visualized based on similar expression vectors in genes using Euclidean distance and hierarchical clustering with average linkage clustering method to view the whole data set and k-means to group the genes in clusters with TIGR MEV (multi experiment viewer), also available at http://www.tigr.org/software.

source_name: wild type conidia grown for 16 hours plus 10 min upon 200 mM CaCl2

label_protocol: The RNA samples were extracted with Trizol and further purified with the RNeasy kit (Qiagen, Germany) and directly labelled by incorporation of Cy3-dUTP (GE Health Care). Briefly, 30 µg of total RNA were mixed with 20 µg of Randon Primer Hexamers (GE Health Care), 100 pmol of oligo dTV (Invitrogen) and 40 U of RNAse Out (Invitrogen) in a volume of 13 µl and heated to 70°C for 5 minutes. The tubes were briefly chilled to 4°C for 5 minutes and mixed with 5 µl of 5X first-strand buffer (Invitrogen), 2 µl DTT (Invitrogen), 2 µl dNTP (5 mM dATP, 5 mM dCTP, 5 mM dGTP, 2 mM dTTP), 2 µl of Suprescript III (Invitrogen) and 1 µl of Cy3-dUTP (25 nmol). The reaction was then incubated at 25°C for 5 minutes and at 42°C for 3 hours. The RNA was degraded by adding 2.5 µl EDTA (0.5 M, pH 8) and 5 µl NaOH (1M) following an incubation at 37°C for 40 minutes. The resulting first strand cDNA was purified and concentrated using a Microcon YM-50 cartridge.

extract_protocol: At each time point the micelia was harvested by centrifugation and quickly frozen in liquid nitrogen. For total RNA isolation, the micelia weas disrupted by grinding in liquid nitrogen and total RNA was extracted with Trizol reagent (Invitrogen). Ten micrograms of RNA from each treatment were then fractionated in 2.2 M formaldehyde, 1.2% agarose gel, stained with ethidium bromide, and then visualized with UV-light. The presence of intact 25S and 17S ribosomal RNA bands was used as a criterion to assess the integrity of the RNA.

tag: For the time course microarray experiments, 5.0 x 108 conidia of A. fumigatus wild type (CEA17 DakuBKU80) strain were inoculated in 300 ml of pre-warmed liquid cultures (YG) in 500-mL erlenmeyer flasks and allowed to grow for 16 hours in a reciprocal shaker (250 rpm) at 37°C. After this time, the control culture was centrifuged for mycelia recovery and RNA extraction. The other cultures were added by calcium chloride (200 mM) and incubated for 10 and 60 more minutes before centrifugation and RNA extraction. Hybridization experiments were competitive using probes derived from 10 and 60 minutes calcium chloride shock cultures using timepoint zero (control) as the reference RNA for every hybridizantion. Normalized signal intensities were used to generate relative hybridization ratios (query/reference).

source_name: wild type conidia grown for 16 hours

label_protocol: The RNA samples were extracted with Trizol and further purified with the RNeasy kit (Qiagen, Germany) and directly labelled by incorporation of Cy5-dUTP (GE Health Care). Briefly, 30 µg of total RNA were mixed with 20 µg of Randon Primer Hexamers (GE Health Care), 100 pmol of oligo dTV (Invitrogen) and 40 U of RNAse Out (Invitrogen) in a volume of 13 µl and heated to 70°C for 5 minutes. The tubes were briefly chilled to 4°C for 5 minutes and mixed with 5 µl of 5X first-strand buffer (Invitrogen), 2 µl DTT (Invitrogen), 2 µl dNTP (5 mM dATP, 5 mM dCTP, 5 mM dGTP, 2 mM dTTP), 2 µl of Suprescript III (Invitrogen) and 1 µl of Cy5-dUTP (25 nmol). The reaction was then incubated at 25°C for 5 minutes and at 42°C for 3 hours. The RNA was degraded by adding 2.5 µl EDTA (0.5 M, pH 8) and 5 µl NaOH (1M) following an incubation at 37°C for 40 minutes. The resulting first strand cDNA was purified and concentrated using a Microcon YM-50 cartridge.


title: Va35_Rep2

description: VA35 Rep 2


title: wild type 0.1 mM Pi 6h vs delta phoB(PHO80) 0.1 mM Pi 6h dye swap

tag: For the time course microarray experiments, 1.0 x 1010 conidia of A. fumigatus wild type [delta akuB(ku80)] strain were used to inoculate 300 ml of liquid cultures (Minimal Medium with 11 and 0.1mMol/L of phosphate) in 500-ml erlenmeyer flasks and allow to grow for 6 and 10 hours in a reciprocal shaker (250 rpm) at 37°C. After this time, the cultures were centrifuged and their RNA were extracted. Hybridization experiments were competitive using probes derived from wild type and deltaphoB(PHO80) strain grown for 6 and 10 hours in MM with 11 and 0.1mmol/L of phosphate, using the wild type strain as the reference RNA for every hybridizantion. Normalized signal intensities were used to generate relative hybridization ratios (query/reference).

tag: For the time course microarray experiments, 1.0 x 1010 conidia of A. fumigatus delta phoB(PHO80) strain were used to inoculate 300 ml of liquid cultures (Minimal Medium with 11 and 0.1mMol/L of phosphate) in 500-ml erlenmeyer flasks and allow to grow for 6 and 10 hours in a reciprocal shaker (250 rpm) at 37°C. After this time, the cultures were centrifuged and their RNA were extracted. Hybridization experiments were competitive using probes derived from wild type and delta phoB(PHO80) strain grown for 6 and 10 hours in MM with 11 and 0.1mmol/L of phosphate, using the wild type strain as the reference RNA for every hybridizantion. Normalized signal intensities were used to generate relative hybridization ratios (query/reference).


title: Va35_Rep1

description: VA35 Rep 1


title: Ewing Sarcoma Tumour Sample # 26

type: genomic

first: Juan

middle: C

last: Cigudosa

address: C/ Melchor Fernández Almagro, 3

department: Human Cancer Genetics

institute: Centro Nacional de Investigaciones Oncologicas - CNIO

description: Primary Ewing Sarcoma Tumour Sample # 26

hyb_protocol: Human cot-1 DNA, Agilent 10x blocking agent and Agilent 2x hybridization buffer were added, and samples were applied to microarrays enclosed in Agilent SureHyb-enabled hybridization chambers. After hybridization, slides were washed sequential

scan_protocol: Scanned on an Agilent G2565AA scanner. Images were quantified using Agilent Feature Extraction Software (version v:8.1).

data_processing: LINEAR normalized, background subtracted data obtained from log2 of processed Red signal/processed Green signal. Agilent software was used.

molecule: genomic DNA

organism: Homo sapiens

source_name: Genomic DNA from ES sample # 26 labeled with Cyanine-5 (red).

label_protocol: 2 µg of genomic DNA were primed with 20 µl of 2.5x Random Primers at 95°C for 5 min and then labeled with 5µl of 10x dUTP mix, 3 µl of Cy3 or Cy5 dUTP (1.0mM) and 1 µl of Exp-Klenow at 37ºC for 3 hours.

extract_protocol: Total DNA extracted using Quiagen's Tissue Kit following manufacturer's instructions

tag: Sex

value: Female

tag: Site

value: Vertebra

tag: Stage

value: M (metastatic)

tag: Genetic

value: ST (stable)

value: Yes

tag: Status

value: A (alive)

tag: Follow-up

source_name: Genomic DNA from pooled pheripheral blood of ten healthy female donors

tag: control; pooled peripheral blood of ten healthy female donors


title: Ewing Sarcoma Tumour Sample # 24

description: Primary Ewing Sarcoma Tumour Sample # 24

source_name: Genomic DNA from ES sample # 24 labeled with Cyanine-5 (red).

value: Rib

value: UNST (unstable)

value: DOD (dead of disease)


title: Va35_Rep0

description: VA35 dye swap

source_name: Total RNA from inoculated Va35 maize ear labeled with Cyanine-3 (green).

source_name: Total RNA from uninoculated Va35 maize ear labeled with Cyanine-5 (red).


title: Ewing Sarcoma Tumour Sample # 25

description: Primary Ewing Sarcoma Tumour Sample # 25

source_name: Genomic DNA from ES sample # 25 labeled with Cyanine-5 (red).

value: Shoulder

value: No


title: Ewing Sarcoma Tumour Sample # 23

description: Ewing Sarcoma Tumour Sample # 23

source_name: Genomic DNA from ES sample # 23 labeled with Cyanine-5 (red).


title: Ewing Sarcoma Tumour Sample # 22

description: Primary Ewing Sarcoma Tumour Sample # 22

source_name: Genomic DNA from ES sample # 22 labeled with Cyanine-5 (red).


title: wild type time 0 min  vs wild type time 10 min upon 200 mM CaCl2

extract_protocol: At each time point the micelia was harvested by centrifugation and quickly frozen in liquid nitrogen. For total RNA isolation, the micelia weas disrupted by grinding in liquid nitrogen and total RNA was extracted with Trizol reagent (Invitrogen). Ten micrograms of RNA from each treatment were then fractionated in 2.2 M formaldehyde, 1.2% agarose gel, stained with ethidium bromide, and then visualized with UV-light. The presence of intact 25S and 17S ribosomal RNA bands was used as a criterion to assess the integrity of the RNA


title: Ewing Sarcoma Tumour Sample # 18

description: GSM205917 and GSM207908 represent Primary Ewing Sarcoma Tumor # 18

source_name: Genomic DNA from ES sample # 18 labeled with Cyanine-5 (red).

value: Male

value: Pelvis

value: L (localized)


title: Ewing Sarcoma Tumour Sample # 20

description: Primary Ewing Sarcoma Tumour Sample # 20

source_name: Genomic DNA from ES sample # 20 labeled with Cyanine-5 (red).


title: Ewing Sarcoma Tumour Sample # 19

description: Primary Ewing Sarcoma Tumour Sample # 19

source_name: Genomic DNA from ES sample # 19 labeled with Cyanine-5 (red).


title: Ewing Sarcoma Tumour Sample # 13

description: GSM205913 and GSM207903 represent Primary Ewing Sarcoma Tumor # 13

source_name: Genomic DNA from ES sample # 13 labeled with Cyanine-5 (red).

value: Radius


title: Ewing Sarcoma Tumour Sample # 16

description: GSM205915 and GSM207906 represent Primary Ewing Sarcoma Tumor # 16

source_name: Genomic DNA from ES sample # 16 labeled with Cyanine-5 (red).

tag: Ewing Sarcoma


title: Ewing Sarcoma Tumour Sample # 10

description: GSM205911 and GSM207901 represent Primary Ewing Sarcoma Tumor # 10

source_name: Genomic DNA from ES sample # 10 labeled with Cyanine-5 (red).

value: Femur


title: Ewing Sarcoma Tumour Sample # 17

description: GSM205916 and GSM207907 represent Primary Ewing Sarcoma Tumor # 17

source_name: Genomic DNA from ES sample # 17 labeled with Cyanine-5 (red).


title: Ewing Sarcoma Tumour Sample # 9

description: GSM205910 and GSM207900 represent Recurrent Ewing tumour Tumor # 9

source_name: Genomic DNA from ES sample # 9 labeled with Cyanine-5 (red).


title: Ewing Sarcoma Tumour Sample # 21

description: Primary Ewing Sarcoma Tumour Sample # 21

source_name: Genomic DNA from ES sample # 21 labeled with Cyanine-5 (red).


title: Ewing Sarcoma Tumour Sample # 12

description: GSM205912 and GSM207902 represent Primary Ewing Sarcoma Tumor # 12

source_name: Genomic DNA from ES sample # 12 labeled with Cyanine-5 (red).

value: Soft tissue


title: Ewing Sarcoma Tumour Sample # 14

description: GSM205914 and GSM207904 represent Recurrent Ewing tumour Tumor # 14

source_name: Genomic DNA from ES sample # 14 labeled with Cyanine-5 (red).

source_name: Genomic DNA from pooled pheripheral blood of ten healthy female donors


title: Ewing Sarcoma Tumour Sample # 15

description: Ewing Sarcoma Tumour Sample # 15

source_name: Genomic DNA from ES sample # 15 labeled with Cyanine-5 (red).

value: Female


title: Ewing Sarcoma Tumour Sample # 6

description: GSM205907 and GSM207897 represent Primary Ewing Sarcoma Tumor # 6

source_name: Genomic DNA from ES sample # 6 labeled with Cyanine-5 (red).


title: Ewing Sarcoma Tumour Sample # 8

description: GSM205909 and GSM207899 represent Primary Ewing Sarcoma Tumor # 8

source_name: Genomic DNA from ES sample # 8 labeled with Cyanine-5 (red).

value: Clavicle


title: HLH-1 induction 6 hour replicate A (11-16-04_Krause_Celegans_H6A)

status: Public on Nov 08 2007

city: Bethesda

first: WeiPing

last: Chen

address: Bldg 8, Room 1A11, NIDDK/NIH

department: GCL

institute: NIDDK/NIH

description: 6 hour replicate A

hyb_protocol: standard Affymetrix procedure

scan_protocol: standard Affymetrix procedure

data_processing: Affymetrix Microarray Suite version 5.0

organism: Caenorhabditis elegans

label_protocol: standard Affymetrix procedure

extract_protocol: standard Affymetrix procedures

tag: embryonic heat shock HLH-1


title: HLH-1 induction 6 hour replicate B (11-16-04_Krause_Celegans_H6B)

description: 6 hour replicate B


title: Ewing Sarcoma Tumour Sample # 7

description: GSM205908 and GSM207898 represent Primary Ewing Sarcoma Tumor # 7

source_name: Genomic DNA from ES sample # 7 labeled with Cyanine-5 (red).

value: Vertebra

value: M (metastatic)


title: Ewing Sarcoma Tumour Sample # 3

description: GSM205906 and GSM207894 represent Primary Ewing Sarcoma Tumor # 3

source_name: Genomic DNA from ES sample # 3 labeled with Cyanine-5 (red).

value: ST (stable)

value: Yes


title: Ewing Sarcoma Tumour Sample # 2

description: GSM205905 and GSM207893 represent Primary Ewing Sarcoma Tumor # 2

source_name: Genomic DNA from ES sample # 2 labeled with Cyanine-5 (red).


title: Ewing Sarcoma Tumour Sample # 1

description: GSM205904 and GSM207892 represent Primary Ewing Sarcoma Tumor # 1

source_name: Genomic DNA from ES sample # 1 labeled with Cyanine-5 (red).


title: HLH-1 induction 4 hour replicate B (11-16-04_Krause_Celegans_H4B)

description: 4 hour replicate B


title: HLH-1 induction 6 hour replicate C (11-16-04_Krause_Celegans_H6C)

description: 6 hour replicate C


title: HLH-1 induction 4 hour replicate A (11-16-04_Krause_Celegans_H4A)

description: 4 hour replicate A


title: Ewing Sarcoma Tumour Sample # 5

description: Ewing Sarcoma Tumour Sample # 5

source_name: Genomic DNA from ES sample # 5 labeled with Cyanine-5 (red).


title: HLH-1 induction 4 hour replicate C (11-16-04_Krause_Celegans_H4C)

description: 4 hour replicate C


title: HLH-1 induction 2 hour replicate C (11-16-04_Krause_Celegans_H2C)

description: 2 hour replicate C


title: Ewing Sarcoma Tumour Sample # 4

description: Ewing Sarcoma Tumour Sample # 4

source_name: Genomic DNA from ES sample # 4 labeled with Cyanine-5 (red).


title: HLH-1 induction 2 hour replicate B (11-16-04_Krause_Celegans_H2B)

description: 2 hour replicate B


title: HLH-1 induction 2 hour replicate A (11-16-04_Krause_Celegans_H2A)

description: 2 hour replicate A


title: HLH-1 induction 0 hour replicate A (11-16-04_Krause_Celegans_H0A)

description: 0 hour replicate A


title: HLH-1 induction 0 hour replicate C (11-16-04_Krause_Celegans_H0C)

description: 0 hour replicate C


title: Untreated Testes, biological rep 2

status: Public on Nov 07 2007

city: NEW YORK

first: C YAN

last: CHENG

state: NY

address: 1230 YORK AVENUE

institute: POPULATION COUNCIL

description: gene expression data from normal rat testis

hyb_protocol: Standard Affymetrix protocol

scan_protocol: Standard Affymetrix protocol

data_processing: The data were analyzed with Microarray Suite version 5.0 (MAS 5.0) using Affymetrix default analysis settings and global scaling as normalization method. The trimmed mean target intensity of each array was arbitrarily set to 250.

organism: Rattus norvegicus

source_name: normal rat testis

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol

growth_protocol: Adult male SD rats

extract_protocol: Trizol extraction of total RNA was performed according to the manufacturer's instructions followed by Qiagen's Rneasy kit purification

treatment_protocol: 3 mg/kg b.w. cadmium chloride injected via i.p.

tag: normal adult rat testis


title: WT CS CNS 0-24hr adult rep2

status: Public on Nov 06 2007

city: Los Angeles

first: Thomas

middle: Daniel

state: CA

address: 1050 Childs Way Room 201B

department: Molecular and Computational Biology

institute: University of Southern California

hyb_protocol: Slides were post-processed using the Nunc SuperChip Aldehyde protocol (Thermo Fisher Scientific). Targets were dried and resuspended in formamide (25mM), 3M NaCl and 0.3M sodium citrate buffer (SSC, 3.3X), SDS (1.1%), Denhardts (5.56X), and polyA solution (8.88mM), and boiled for 2 minutes. The resuspended labeled cDNA was applied to the microarrays and allowed to hybridize 16-18 hours at 42°C. Slides were washed in 300mls of a 1.5% SDS, 1XSSC solution for 5 minutes, followed by a 5 minute wash in 0.2XSSC, and twice for 10 minutes in dH2O and spun dry.

scan_protocol: Slides were scanned with 635nm and 532nm lasers using a Genepix 4100A scanner (Axon Instruments). Data was analyzed using Genepix 5.0 software and the R Bioconductor software package v.2.3.1.

data_processing: Poor quality features, identified by visualization of the scanned microarray, were removed from the raw data before analysis using Genepix software. Filtering using R software identified and kept features containing >75% of total pixels above 1 standard deviation above background for further study. Data was normalized using the Bioconductor lowess method and data was log2 transformed using R software. Recipricols of dye-flip hybridizations were calculated to compare all replicates in the same direction.

organism: Drosophila melanogaster

source_name: male w; Berlin wild type 0-24 hour adult CNS tissue

label_protocol: Total RNA (20ug) was used as template to make cDNA in the presence of Cy labeled nucleotides (direct labeling).  Direct labeling was achieved through a two-hour reverse transcription reaction at 42°C. Final concentrations are indicated in parentheses: oligo dT primer (Operon, 3.75 mM), dithiothreitol (Invitrogen, 10mM), First Strand Buffer (Invitrogen, 1X), dNTPs minus dTTP (Invitrogen, 0.5mM), dTTP (Invitrogen, 50nM), Cy-labeled dUTP (Perkin-Elmer, 0.625nM), and Superscript II reverse transcriptase (Invitrogen, 10U/mL). The reaction was quenched using NaOH (167mM) and EDTA (83mM), and purified using the Qiaquick PCR purification kit (Qiagen). For dissected CNS tissues, cRNA probes were made using the Amino Allyl MessageAmp II aRNA Amplification Kit (Ambion).

extract_protocol: For experiments in which RNA was extracted from adult head tissues, flies were collected under CO2 anesthesia (ZT 2) and allowed 8 hours (ZT 10) to recover to minimize identifying genes induced by exposure to CO2, before being snap frozen in liquid nitrogen. All collections were timed such that all adult flies were between 8- 24 hours post-eclosion when snap frozen. For experiments performed using RNA from dissected CNS tissues, animals were lightly anesthetized at the time of dissection and dissections were performed rapidly. For experiments in which RNA was extracted from adult CNS tissues, brains and ventral nerve cords were dissected in 1XPBS and snap frozen in liquid nitrogen. Total RNA was extracted from fly heads or dissected CNS tissues by homogenization and extraction using TRIzol reagent (Invitrogen). All experiments included dye-flip hybridizations, to minimize identifying genes because of differences in dye incorporation, with one half of the comparisons labeled in one dye orientation and the other half the other dye orientation. All experiments were performed using at least four independent biological samples of ~100 fly heads each for head experiments and ~20 dissected brains and VNCs for CNS experiments; for the experiments comparing wild type male versus female, and fru P1 male versus wild type male eight and six independent biological samples were used respectively.

tag: Strain

value: w; Berlin

tag: Gender

value: male

value: 0-24 hour

value: CNS

source_name: fru P1 male 0-24 hour adult CNS tissue

value: fru w12/Cham5


title: Testes, 8 hours after cadmium treatment, biological rep 1

description: gene expression data from rat testis 8 h after cadmium treatment

source_name: 8 hour after cadmium treatment

tag: early to medium response after cadmium treatment


title: WT wBerlin CNS 0-24hr adult rep4

source_name: male CS wild type 0-24 hour adult CNS tissue

value: Canton-S

tag: Age

tag: Tissue


title: Testes, 20 hours after cadmium treatment, biological rep 2

description: gene expression data from rat testis 20 h after cadmium treatment

source_name: 20 hour after cadmium treatment

tag: medium to late response after cadmium treatment


title: Untreated Testes, biological rep 1


title: WT wBerlin CNS 0-24hr adult rep6


title: HLH-1 induction 0 hour replicate B (11-16-04_Krause_Celegans_H0B)

description: 0 hour replicate B


title: WT wBerlin CNS 0-24hr adult rep5


title: Untreated Testes, biological rep 3


title: WT wBerlin head 0-24hr adult rep8

source_name: male w; Berlin wild type 0-24 hour adult head tissue

value: head

source_name: fru P1 male 0-24 hour adult head tissue


title: dsx null head XY 0-24hr adult rep3

source_name: dsx null XY 0-24 hour adult head tissue

value: dsx d+r3/M+r15 XY

source_name: male CS wild type 0-24 hour adult head tissue


title: dsx null head XY 0-24hr adult rep2


title: WT wBerlin head 0-24hr adult rep1


title: WT CS CNS 0-24hr adult rep1


title: Testes, 8 hours after cadmium treatment, biological rep 2


title: Testes, 20 hours after cadmium treatment, biological rep 1


title: WT CS CNS 0-24hr adult rep3


title: WT wBerlin head 0-24hr adult rep4


title: WT wBerlin head 0-24hr adult rep6


title: dsx null head XY 0-24hr adult rep4


title: WT wBerlin head 0-24hr adult rep7


title: WT wBerlin head 0-24hr adult rep3


title: WT wBerlin head 0-24hr adult rep2


title: dsx null head XY 0-24hr adult rep1


title: WT CS CNS adult rep3

source_name: female CS wild type 0-24 hour adult CNS tissue

value: female


title: dsxD head 0-24hr adult rep3

source_name: dsxD 0-24 hour adult head tissue

value: dsxD XX

source_name: female CS wild type 0-24 hour adult head tissue


title: WT CS head 0-24hr adult rep4


title: dsx null head XX 0-24hr adult rep2

source_name: dsx null XX 0-24 hour adult head tissue

value: dsx d+r3/M+r15 XX


title: dsx null head XX 0-24hr adult rep3


title: dsxD head 0-24hr adult rep2


title: WT wBerlin head 0-24hr adult rep5


title: WT Berlin head 0-24hr adult rep6

value: Berlin

source_name: female Berlin wild type 0-24 hour adult head tissue


title: dsx null head XX 0-24hr adult rep4


title: WT Berlin head 0-24hr adult rep5

source_name: male Berlin wild type 0-24 hour adult head tissue


title: tra head 0-24hr adult rep2

source_name: tra XX 0-24 hour adult head tissue

value: tra XX


title: dsxD head 0-24hr adult rep4


title: dsx null head XX 0-24hr adult rep1


title: WT CS CNS adult rep1


title: dsxD head 0-24hr adult rep1


title: tra head 0-24hr adult rep4


title: WT CS CNS adult rep2


title: tra head 0-24hr adult rep1


title: WT Berlin head 0-24hr adult rep7


title: tra head 0-24hr adult rep3


title: WT Berlin head 0-24hr adult rep8


title: WT CS CNS adult rep4


title: WT CS head 0-24hr adult rep1


title: WT CS head 0-24hr adult rep3


title: WT CS head 0-24hr adult rep2


title: AKR/J rev

city: South San Francisco

first: Gene

last: Cutler

address: 1120 Veterans Blvd

department: Lead Discovery

institute: Amgen Inc

hyb_protocol: The Maui hybridization system was used (BioMicro Systems, Salt Lake City UT), and the samples were prepared with Agilent’s 2X hybridization. Post-collection, the sample was diluted with 1x TE buffer to a volume of 22.8 μl. To this material, 28.5 μl of Agilent 2x hybridization buffer was added. The genomic DNA mixture was then denatured at 990C for 2 minutes prior to incubation for 30 minutes at 370C to allow Cot-1 pre-annealing. Deionized formamide (Sigma, St Louis MO, # F9037) was added to a final formamide concentration of 10% (v/v) and a final hybridization volume of 57 μl. To remove any precipitates, samples were centrifuged at 16,000 x g for 5 minutes. Hybridization mixtures and the BioMicro assembly jig were pre-warmed to 60°C as recommended in the BioMicro protocol. Barcode adhesives were removed from the Agilent arrays (active side only) to ensure proper sealing of the Maui hybridization lids (BioMicro, catalog # 02-A008-09). 50 μl of each sample was loaded onto an array and hybridized at 60°C for 17 hours in the 4-bay hybridization station (catalog # 02-A002-02) . The arrays were washed and dried according to the standard protocol supplied by Agilent. The arrays were then disassembled in a slide-staining dish containing wash 1 (0.5x SSC, 0.005% Triton X-102) and transferred to a slide rack in a second staining dish containing wash 1. The wash was stirred with a magnetic stir bar and stir plate for 5 minutes at ambient temperature. The arrays were transferred to a 37oC incubator in a third staining dish of pre-warmed wash 2 (0.1x SSC, 0.005% Triton X-102) and stirred for 5 minutes. The arrays were then removed slowly to minimize droplet formation. Residual solution was blown from the surface of the arrays with a stream of nitrogen gas as required.

scan_protocol: The arrays were scanned with an Agilent scanner and images were analyzed with the Feature Extraction software (Agilent, Palo Alto CA).

data_processing: A loess normalization (R function loess, span=.4) is performed on log transformed non-flagged data.

source_name: AKR/J

label_protocol: 3ug of test and reference genomic DNA samples were digested with 30 units AluI (New England Biolabs, Ipswich MA) and 30 units RsaI (New England Biolabs). All digests were performed in a volume of 100 μl and incubated for at least 16 hours at 37°C. Digested genomic DNA was purified using PCR purification columns (Qiagen, Valencia CA, Catalog #28104). A 1μl aliquot of each digest was analyzed on a DNA 7500 cassette using a Bioanalyzer 2100 (Agilent Technologies). Complete digestion resulted in a smear of fragments between 200-800 bp. DNA was quantified on a ND-1000 spectrophotometer (NanoDrop Technologies, Wilmington DE) prior to vacuum concentration to a reduced volume of 21 μl. Digested genomic DNA was labeled using the BioPrime DNA labeling System (Invitrogen, Carlsbad CA) according to the protocol supplied with the Agilent microarrays. To the digested DNA, 20μl of 2.5X random octomer (750 µg/ml) was added and incubated at 99°C for 7 minutes and cooled on ice for 5 minutes. To this reaction, 5μl of 10X low “C” dNTP Mix (120 μM each of dATP, dGTP and dTTP, and 60 μM of CTP), 1 μl exo-Klenow (40 U/μl) and 3 μl of either 60 μM Cy3-dCTP or 60 μM Cy5-dCTP (Amersham, Piscataway NJ) was added and incubated for 2hrs at 37°C. Following incubation, each labeled test (male, Cy5) and reference (male, Cy3) samples were combined appropriately and mixed with 400 μl of 1X TE prior to purification. Replicates were performed for each analysis as dye-flips, where the reference sample was Cy5-labeled and the test sample was Cy3-labeled. Samples were loaded onto YM-30 column (Millipore Catalog # 42410) and spun at 10,000 x g for 10 minutes. An additional 500 μl of 1X TE was added to the column and spun at 10,000 x g for 10 minutes. The column was inverted into a fresh tube and a spun for one-minute at 10,000 x g to collect the samples. Labeled genomic DNA was mixed with 50 μg of Cot-1 DNA (Invitrogen), 100 μg of Yeast tRNA (Invitrogen), 50 μl of 10X Control Targets (Agilent Technologies) and 1X TE for a final volume of 500 μl. Samples were placed in new YM-30 columns and spun at 14,000 x g for 20 minutes. The column was inverted into a fresh tube and a spun for one-minute at 14,000 x g to collect the sample. Dye incorporation (as measured by the ND-1000) typically exceeded 30 pmol/μl for Cy3 and 20 pmol/μl for Cy5.

extract_protocol: DNA samples were obtained from Jackson Laboratories

tag: Male, AKR/J

tag: Male, C57BL/6J


title: MSM/Ms rev

data_processing: The log2 ratio data for values which were not flagged were saved.

source_name: MSM/Ms

tag: Male, MSM/Ms


title: C57L/J 2

tag: Male, C57L/J


title: NON/LtJ rev

source_name: NON/LtJ

tag: Male, NON/LtJ


title: NOD/LtJ rev

source_name: NOD/LtJ

tag: Male, NOD/LtJ


title: SJL/J rev

source_name: SJL/J

tag: Male, SJL/J


title: MSM/Ms


title: A/J rev

source_name: A/J

tag: Male, A/J


title: 129S1/SvlmJ rev

tag: Male, 129S1/SvlmJ


title: PL/J

source_name: PL/J

tag: Male, PL/J


title: SEA/GnJ rev

source_name: SEA/GnJ

tag: Male, SEA/GnJ


title: DBA/2J rev

tag: Male, DBA/2J


title: NON/LtJ


title: DBA/1J rev

tag: Male, DBA/1J


title: A/J


title: Balb/cJ rev

source_name: Balb/cJ

tag: Male, Balb/cJ


title: DBA/2J


title: 129S1/SvlmJ


title: C3H/HeJ rev

tag: Male, C3H/HeJ


title: PERA/EiJ rev

source_name: PERA/EiJ

tag: Male, PERA/EiJ


title: AKR/J


title: C57BL/10J

tag: Male, C57BL/10J


title: RIIIS/J

source_name: RIIIS/J

tag: Male, RIIIS/J


title: C3H/HeJ


title: Pool of 384 individuals (Illumina)

city: Brisbane

first: Stuart

last: Macgregor

state: Queensland

address: 300 Herston Road

department: Genetic Epidemiology

country: Australia

description:
Two main file types are included with this submission - 1) the 30 raw data files, 2) a file which summarizes the allele frequency estimate from all 3 arrays for the control pool
Raw data:
The raw data are the 30 files .txt files each beginning 15... These are data from 3 arrays, where in each case the SNPs on the array are split across 10 'strands'. The 3 replicate arrays on this pool are technical replicates. The uploaded files have raw red/green scores which is used to compute the allele frequency estimates for the pool. Unlike Affymetrix where each array has up to 10 PAF estimates per array, the number of PAF estimates (i.e. pairs of R and G scores) varies widely on Illumina arrays, from 1 up to ~55, with an average of ~18. Unlike the Affy arrays, where all of the replicate PAF scores are put on one line for a given SNP, each R and G pair are put on a separate line. The total data for a given snp on one array hence consists of several lines (up to ~55 per snp). As with the affy arrays, multiple PAF estimates are required to get the technical error down to a reasonable level so we use 3 arrays per pool - this means there are potentially up to ~150 PAF estimates per SNP, with an average of 3*18=54. As with the affy arrays, 3 arrays were used so there were 3 hybs (generating up to 150 data points per SNP, with an average of ~54).
For each snp, the raw data files contain multiple rows for each of the red/green beadscores for that snp - for any given snp the number of red/green scores varies considerably (from 1 up to a maximum of ~55, average is ~18). This file would typically contain roughly 500k lines (since there is an average of ~18 red/green scores per snp and ~31k snps, there are approx 18* 31000 ~ 550k lines). The SNP names are given the illumina name (e.g. 10008); information on rs names is included in the summary file.
Processed data:
The summary file, forgeoconpoolfreq.txt, contains the rs and Illumina names (Code), along with the physical position, chromosome and estimate of allele frequency for the pool based on the raw data from all 3 arrays used.  The estimate of pooling allele frequency (PAF) comes from PAF=R/(R+G) where R and G are the red and green intensities respectively. The PAFs were normalized to ensure that the mean allele frequency was 0.5 over each strand of the array (i.e. over each of the 10 sets of ~30k SNPs on the array).

hyb_protocol: Array data. SNPs were genotyped on DNA pool samples using Illumina HumanHap300 arrays. Arrays were treated according to standard protocols.

scan_protocol: Arrays were scanned at Queensland Institute of Medical Research by Anjali Henders and Zhen Zhen Zhao.

data_processing:
Raw probe intensities were dumped from Illumina array software and analysed using methods described in Macgregor et al Nucleic Acids Research, 2006, Vol. 34, No. 7 e55
Pooling array frequencey = red/(red+green)

source_name: pool of 384 individuals

label_protocol: standard illumina protocol

extract_protocol: DNA was extracted from each of 384 individuals. DNA concentrations were measured using PicoGreen (Molecular Probes) for the quantitation of double-stranded DNA in solution on a Fluoroskan Ascent CF plate reader (Labsystems, Chicago). Concentrations of DNA samples were carefully adjusted by serial dilutions to a ﬁnal concen- tration of 25 ng/ml (M SD ¼ 25.19 ± 0.55). Individual DNA samples were tested in at least two PCR to ensure samples containing high quality DNA.

tag: control group

label_protocol: standard Illumina protocol


title: NOD/LtJ

tag: Male, C57BL/6J


title: SPRET/EiJ rev

source_name: SPRET/EiJ

tag: Male, SPRET/EiJ


title: RIIIS/J rev


title: SEA/GnJ


title: PERA/EiJ


title: Balb/cJ


title: NZW/LacJ

source_name: NZW/LacJ

tag: Male, NZW/LacJ


title: 129Sv/J

tag: Male, 129Sv/J


title: SPRET/EiJ


title: C57L/J

tag: Male, C57L/J


title: 129Sv/J rev


title: MA/MyJ rev

source_name: MA/MyJ

tag: Male, MA/MyJ


title: FVB/Ntac

source_name: FVB/Ntac

tag: Male, FVB/Ntac


title: LP/J rev

source_name: LP/J

tag: Male, LP/J


title: DBA/1J


title: LP/J


title: PWK/PhJ rev

source_name: PWK/PhJ

tag: Male, PWK/PhJ


title: FVB/Ntac rev


title: PWK/PhJ


title: JF1/Ms rev

tag: Male, JF1/Ms


title: CAST/EiJ

source_name: CAST/EiJ

tag: Male, CAST/EiJ


title: I/LnJ

source_name: I/LnJ

tag: Male, I/LnJ


title: CAST/EiJ rev


title: I/LnJ rev


title: JF1/Ms


title: MA/MyJ


title: C57BL/6J


title: WSB/EiJ

source_name: WSB/EiJ

tag: Male, WSB/EiJ


title: C57BL/6J rev


title: CZECHII/EiJ rev

source_name: CZECHII/EiJ

tag: Male, CZECHII/EiJ


title: CZECHII/EiJ


title: CE/J rev

source_name: CE/J

tag: Male, CE/J


title: KK/HIJ rev

source_name: KK/HIJ

tag: Male, KK/HIJ


title: C57BR/cdJ rev

tag: Male, C57BR/cdJ


title: C57BR/cdJ

tag: Male, C57BL/6J


title: KK/HIJ


title: BUB/BnJ rev

source_name: BUB/BnJ

tag: Male, BUB/BnJ


title: WSB/EiJ rev


title: BUB/BnJ


title: CE/J


title: MOLF/EiJ rev

source_name: MOLF/EiJ

tag: Male, MOLF/EiJ


title: MOLF/EiJ


title: SWR/J rev

source_name: SWR/J

tag: Male, SWR/J


title: SJL/J

source_name: SJL/J

tag: Male, SJL/J


title: NZW/LacJ rev

tag: Male, NZW/LacJ


title: SWR/J


title: CBA/J rev

source_name: CBA/J

tag: Male, CBA/J


title: SM/J rev

source_name: SM/J

tag: Male, SM/J


title: C57BL/10J rev

tag: Male, C57BL/10J


title: BTBR T+tf/J rev

source_name: BTBR T+tf/J

tag: Male, BTBR T+tf/J


title: BTBR T+tf/J


title: C57BLKS/J rev

tag: Male, C57BLKS/J


title: C57BLKS/J

tag: Male, C57BL//6J


title: CBA/J


title: WT strain grown in YPD under Normoxia (08-15-06_Philpott_YG_S98_9)

first: Olga

last: Protchenko

address: 10 Center Dr, Bldg.10-Rm.9B-05

department: NIDDK

institute: National Institutes of health

description: Gene expression data from wild type strain under normoxic conditions

organism: Saccharomyces cerevisiae

source_name: 9-WT strain grown in YPD under Normoxia

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

tag: BY4742 strain


title: hem1 strain grown in YPD w/o  ALA (08-15-06_Philpott_YG_S98_4)

description: Gene expression data from hem1 strain under heme deplete conditions (-ALA)

source_name: 4- hem1 strain grown in YPD w/o  ALA

tag: BY4742 hem1 delta strain


title: WT strain grown in YPD under Normoxia (08-15-06_Philpott_YG_S98_8)

source_name: 8-WT strain grown in YPD under Normoxia


title: NZB/BINJ

source_name: NZB/BINJ

tag: Male


title: PL/J rev

source_name: PL/J

tag: Male, PL/J

tag: Male, C56BL/6J


title: SM/J

tag: Male, C57BL/6J


title: C58/J rev

hyb_protocol: The Maui hybridization system was used (BioMicro Systems, Salt Lake City UT), and the samples were prepared with Agilent’s 2X hybridization. Post-collection, the sample was diluted with 1x TE buffer to a volume of 22.8 μl. To this material, 28.5 μl of Agilent 2x hybridization buffer was added. The genomic DNA mixture was then denatured at 990C for 2 minutes prior to incubation for 30 minutes at 370C to allow Cot-1 pre-annealing.  Deionized formamide (Sigma, St Louis MO, # F9037) was added to a final formamide concentration of 10% (v/v) and a final hybridization volume of 57 μl.  To remove any precipitates, samples were centrifuged at 16,000 x g for 5 minutes. Hybridization mixtures and the BioMicro assembly jig were pre-warmed to 60°C as recommended in the BioMicro protocol. Barcode adhesives were removed from the Agilent arrays (active side only) to ensure proper sealing of the Maui hybridization lids (BioMicro, catalog # 02-A008-09).  50 μl of each sample was loaded onto an array and hybridized at 60°C for 17 hours in the 4-bay hybridization station (catalog # 02-A002-02) . The arrays were washed and dried according to the standard protocol supplied by Agilent. The arrays were then disassembled in a slide-staining dish containing wash 1 (0.5x SSC, 0.005% Triton X-102) and transferred to a slide rack in a second staining dish containing wash 1. The wash was stirred with a magnetic stir bar and stir plate for 5 minutes at ambient temperature. The arrays were transferred to a 37oC incubator in a third staining dish of pre-warmed wash 2 (0.1x SSC, 0.005% Triton X-102) and stirred for 5 minutes. The arrays were then removed slowly to minimize droplet formation. Residual solution was blown from the surface of the arrays with a stream of nitrogen gas as required.

label_protocol: 3ug of test and reference genomic DNA samples were digested with 30 units AluI (New England Biolabs, Ipswich MA) and 30 units RsaI (New England Biolabs). All digests were performed in a volume of 100 μl and incubated for at least 16 hours at 37°C. Digested genomic DNA was purified using PCR purification columns (Qiagen, Valencia CA, Catalog #28104). A 1μl aliquot of each digest was analyzed on a DNA 7500 cassette using a Bioanalyzer 2100 (Agilent Technologies). Complete digestion resulted in a smear of fragments between 200-800 bp. DNA was quantified on a ND-1000 spectrophotometer (NanoDrop Technologies, Wilmington DE) prior to vacuum concentration to a reduced volume of 21 μl. Digested genomic DNA was labeled using the BioPrime DNA labeling System (Invitrogen, Carlsbad CA) according to the protocol supplied with the Agilent microarrays.  To the digested DNA, 20μl of 2.5X random octomer (750 µg/ml) was added and incubated at 99°C for 7 minutes and cooled on ice for 5 minutes.  To this reaction, 5μl of 10X low “C” dNTP Mix (120 μM each of dATP, dGTP and dTTP, and 60 μM of CTP), 1 μl exo-Klenow (40 U/μl) and 3 μl of either 60 μM Cy3-dCTP or 60 μM Cy5-dCTP (Amersham, Piscataway NJ) was added and incubated for 2hrs at 37°C.  Following incubation, each labeled test (male, Cy5) and reference (male, Cy3) samples were combined appropriately and mixed with 400 μl of 1X TE prior to purification.  Replicates were performed for each analysis as dye-flips, where the reference sample was Cy5-labeled and the test sample was Cy3-labeled. Samples were loaded onto YM-30 column (Millipore Catalog # 42410) and spun at 10,000 x g for 10 minutes.  An additional 500 μl of 1X TE was added to the column and spun at 10,000 x g for 10 minutes.  The column was inverted into a fresh tube and a spun for one-minute at 10,000 x g to collect the samples.  Labeled genomic DNA was mixed with 50 μg of Cot-1 DNA (Invitrogen), 100 μg of Yeast tRNA (Invitrogen), 50 μl of 10X Control Targets (Agilent Technologies) and 1X TE for a final volume of 500 μl.  Samples were placed in new YM-30 columns and spun at 14,000 x g for 20 minutes. The column was inverted into a fresh tube and a spun for one-minute at 14,000 x g to collect the sample. Dye incorporation (as measured by the ND-1000) typically exceeded 30 pmol/μl for Cy3 and 20 pmol/μl for Cy5.

tag: Male, C58/J


title: NZB/BINJ rev


title: hem1 strain grown in YPD w/o ALA (08-15-06_Philpott_YG_S98_3)

source_name: 3-hem1 strain grown in YPD w/o ALA

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


title: hem1 strain grown in YPD with 250 uM ALA (08-15-06_Philpott_YG_S98_2)

description: Gene expression data from hem1 strain under heme replete conditions (+ ALA)

source_name: 2  hem1 strain grown in YPD with 250 uM ALA


title: Pool of 384 individuals (Affymetrix)

institute: Queensland Institute of Medical Research

description: 384 individual pool

hyb_protocol: Array data. SNPs were genotyped on DNA pool samples using Affymetrix Genechip HindIII arrays. Arrays were treated according to standard protocols (Affymetrix, San Diego).

scan_protocol: Arrays were scanned at the Australian Genotyping Research Facility using Affymetrix GTYPE software.

data_processing: Raw probe intensities were dumped from Affymetrix GTYPE software and analysed using methods described in Macgregor et al Nucleic Acids Research, 2006, Vol. 34, No. 7 e55

label: Control Group

source_name: pool of 384 individuals

label_protocol: standard Affymtrix protocol

treatment_protocol: Control group

tag: Control Group


title: WT strain grown in YPD under Hypoxia (08-15-06_Philpott_YG_S98_10)

description: Gene expression data from wild type strain under hypoxic conditions

source_name: 10-WT strain grown in YPD under Hypoxia


title: C58/J


title: WT strain grown in YPD under Hypoxia (08-15-06_Philpott_YG_S98_11)

source_name: 11-WT strain grown in YPD under Hypoxia


title: hem1 strain grown in YPD with 250 uM ALA (08-15-06_Philpott_YG_S98_1)

source_name: 1-hem1 strain grown in YPD with 250 uM ALA


title: Rpl14a_knockout_rep1

city: Boston

first: Suzanne

last: Komili

state: MA

address: 44 Binney St.

institute: Harvard University

description: Replicate #1 of Rpl14a deletion mRNA sample used for analysis of ribosomal protein knockouts.

hyb_protocol: The purified cRNA is fragmented in order to facilitate the subsequent hybridization step. The fragmented cRNA is added to a hybridization solution containing several biotinylated control oligonucleotides (for quality control), and hybridized to a microarray chip overnight at 45°C. The chips are then transferred to a fluidics instrument that performs washes to remove cRNA that has not hybridized to its complementary oligonucleotide probe. The bound cRNA is then fluorescently labeled using phycoerythrin-conjugated streptavidin (SAPE); additional fluors are then added using biotinylated anti-streptavidin antibody and additional SAPE.

scan_protocol: Each cRNA bound at its complementary oligonucleotide is excited using a confocal laser scanner, and the positions and intensities of the fluorescent emissions are captured. These measures provide the basis of subsequent biostatistical analysis.

data_processing: Gene Chip Operating System's Statistical Expression algorithm is used to analyze the cell intensities and generate both expression values and presence/absence calls

source_name: Rpl14a deletion cells grown in rich media at 30C

label_protocol: RNA is converted into cDNA using a T7 promoter-tailed oligo-dT primer in the synthesis of the first cDNA strand; second strand cDNA synthesis is then carried out. The double-stranded cDNA is used as the template in an in vitro transcription (IVT) reaction catalyzed by T7 polymerase and containing biotinylated CTP and UTP in addition to the four unmodified ribonucleoside triphosphates. The biotinylated complementary RNA (cRNA) is purified from the IVT reaction mixture using the RNeasy system (Qiagen). The cRNA is quantified spectrophotometrically and purity of the cRNA is also assessed by spectrophometric measurements.

growth_protocol: 50mL cultures were grown in YPD to early log phase (OD260 0.2-0.4) at 30oC., centrifuged at room temperature and flash-frozen using liquid nitrogen.

extract_protocol: mRNA was extracted from cell pellets using the hot phenol method and precipitated overnight in ethanol

tag: Rpl14a::KAN in BY4741 background


title: Rps18a_knockout_rep2

description: Replicate #2 of Rps18a deletion mRNA sample used for analysis of ribosomal protein knockouts.

source_name: Rps18a deletion cells grown in rich media at 30C

tag: Rps18a::KAN in BY4741 background


title: Rps18b_knockout_rep1

description: Replicate #1 of Rps18b deletion mRNA sample used for analysis of ribosomal protein knockouts.

source_name: Rps18b deletion cells grown in rich media at 30C

tag: Rps18b::KAN in BY4741 background


title: Rpl22b_knockout_rep1

description: Replicate #1 of Rpl22b deletion mRNA sample used for analysis of ribosomal protein knockouts.

source_name: Rpl22b deletion cells grown in rich media at 30C

tag: Rpl22b::KAN in BY4741 background


title: Rpl12a_knockout_rep2

description: Replicate #2 of Rpl12a deletion mRNA sample used for analysis of ribosomal protein knockouts.

source_name: Rpl12a deletion cells grown in rich media at 30C

tag: Rpl12a::KAN in BY4741 background


title: Rpl12a_knockout_rep1

description: Replicate #1 of Rpl12a deletion mRNA sample used for analysis of ribosomal protein knockouts.


title: Rpl14a_knockout_rep2

description: Replicate #2 of Rpl14a deletion mRNA sample used for analysis of ribosomal protein knockouts.


title: Rpl22a_knockout_rep1

description: Replicate #1 of Rpl22a deletion mRNA sample used for analysis of ribosomal protein knockouts.

source_name: Rpl22a deletion cells grown in rich media at 30C

tag: Rpl22a::KAN in BY4741 background


title: Rps18b_knockout_rep2

description: Replicate #2 of Rps18b deletion mRNA sample used for analysis of ribosomal protein knockouts.


title: Rpl22a_knockout_rep2

description: Replicate #2 of Rpl22a deletion mRNA sample used for analysis of ribosomal protein knockouts.


title: Rps0a_knockout_rep1

description: Replicate #1 of Rps0a deletion mRNA sample used for analysis of ribosomal protein knockouts.

source_name: Rps0a deletion cells grown in rich media at 30C

tag: Rps0a::KAN in BY4741 background


title: Rpl12b_knockout_rep2

description: Replicate #2 of Rpl12b deletion mRNA sample used for analysis of ribosomal protein knockouts.

source_name: Rpl12b deletion cells grown in rich media at 30C

tag: Rpl12b::KAN in BY4741 background


title: Rps0a_knockout_rep2

description: Replicate #2 of Rps0a deletion mRNA sample used for analysis of ribosomal protein knockouts.


title: Rps0b_knockout_rep1

description: Replicate #1 of Rps0b deletion mRNA sample used for analysis of ribosomal protein knockouts.

source_name: Rps0b deletion cells grown in rich media at 30C

tag: Rps0b::KAN in BY4741 background


title: Rps0b_knockout_rep2

description: Replicate #2 of Rps0b deletion mRNA sample used for analysis of ribosomal protein knockouts.


title: Rpl22b_knockout_rep2

description: Replicate #2 of Rpl22b deletion mRNA sample used for analysis of ribosomal protein knockouts.


title: WT_rep2

description: Replicate #2 of wild-type mRNA sample used for analysis of ribosomal protein knockouts.

hyb_protocol: The purified cRNA is fragmented in order to facilitate the subsequent hybridization step.  The fragmented cRNA is added to a hybridization solution containing several biotinylated control oligonucleotides (for quality control), and hybridized to a microarray chip overnight at 45°C.  The chips are then transferred to a fluidics instrument that performs washes to remove cRNA that has not hybridized to its complementary oligonucleotide probe. The bound cRNA is then fluorescently labeled using phycoerythrin-conjugated streptavidin (SAPE); additional fluors are then added using biotinylated anti-streptavidin antibody and additional SAPE.

source_name: Wild-type cells grown in rich media at 30C

label_protocol: RNA is converted into cDNA using a T7 promoter-tailed oligo-dT primer in the synthesis of the first cDNA strand; second strand cDNA synthesis is then carried out.  The double-stranded cDNA is used as the template in an in vitro transcription (IVT) reaction catalyzed by T7 polymerase and containing biotinylated CTP and UTP in addition to the four unmodified ribonucleoside triphosphates.  The biotinylated complementary RNA (cRNA) is purified from the IVT reaction mixture using the RNeasy system (Qiagen).  The cRNA is quantified spectrophotometrically and purity of the cRNA is also assessed by spectrophometric measurements.

tag: Strain


title: Rpl7b_knockout_rep1

description: Replicate #1 of Rpl7b deletion mRNA sample used for analysis of ribosomal protein knockouts.

source_name: Rpl7b deletion cells grown in rich media at 30C

tag: Rpl7b::KAN in BY4741 background


title: Rpl7a_knockout_rep2

description: Replicate #2 of rpl7a- mRNA sample used for analysis of ribosomal protein knockout expression levels.

source_name: Rpl7a deletion cells grown in rich media at 30C

tag: RPL7A::KAN in BY4741 background


title: Rpl12b_knockout_rep1

description: Replicate #1 of Rpl12b deletion mRNA sample used for analysis of ribosomal protein knockouts.


title: Rpl7a_knockout_rep1

description: Replicate #1 of rpl7a- mRNA sample used for analysis of ribosomal protein knockout expression levels.

tag: Rpl7a::KAN in BY4741 background


title: WT_rep1

description: Replicate #1 of wild-type mRNA sample used for analysis of ribosomal protein knockouts.


title: Rps18a_knockout_rep1

description: Replicate #1 of Rps18a deletion mRNA sample used for analysis of ribosomal protein knockouts.


title: Rpl7b_knockout_rep2

description: Replicate #2 of Rpl7b deletion mRNA sample used for analysis of ribosomal protein knockouts.