title: UC2x, stage15, biological rep5


title: FRM, stage15, biological rep3

status: Public on Dec 14 2006

type: RNA

city: Coimbra

first: Tiago

middle: R

last: Magalhaes

address: Departamento Zoologia, Universidade Coimbra

department: Centro Neurociências

country: Portugal

institute: Universidade de Coimbra

description: This microarray belongs to an experiment that aims to understand the genetic pathways in axon guidance mechanisms. 17 mutants and transgenics of axon guidance genes were analyzed, with 4 to 5 technical replicates for each mutant or transgenic.

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

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

data_processing: The RMA algorithm was used as in Irizarry2003, NAR,31-e15, with the recommended quantile-quantile normalization procedure

label: biotin

molecule: polyA RNA

organism: Drosophila melanogaster

source_name: Drosophila embryos stage 15

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

extract_protocol: RNA was extracted from the embryos using QIAGEN columns according to manufacturer recommendations.

tag: FRM is a robo GOF, obtained by crossing the driver elavGal4 with a UAS chimeric construct, which combines the robo cytoplasmic domain with the frazzled extracellular domain (in Bashaw1999-917 Cell,97-917).


title: UC2x, stage15, biological rep2

tag: UC2x is a stronger comm GOF than UC (also in this experiment) and was obtained using two copies of UAS-comm crossed with the driver elavGal4 (unpublished, more information with the depository).


title: FRM, stage15, biological rep4


title: UC, stage15, biological rep1

tag: UC is the comm GOF, and corresponds to a single copy of the UAS-comm construct crossed with the driver elavGal4 (Kidd1998 Neuron,20-25}.


title: UC2x, stage15, biological rep3


title: UR, stage15, biological rep2

tag: UR is a robo Gain of Function that results from crossing UAS-robo flies with the postmitotic neural driver elavGal4 flies (Kidd1998, Kidd92-205).


title: FRM, stage15, biological rep2


title: FRM, stage15, biological rep1


title: UC2x, stage15, biological rep4


title: Expression data from ankle tissue of C3H mice at 1 week of infection

city: Salt Lake City

first: Janis

last: Weis

state: UT

address: 15 North Medical Drive East

department: Pathology

country: USA

institute: University of Utah

description: expression during the development of arthritis

hyb_protocol: standard Affymetrix procedure

scan_protocol: standard Affymetrix procedure

data_processing:
MAS 5
Algorithm: Statistical
HZ=4 VZ=4 BG=4 SmoothFactorBG=100 Epsilon=0.5 BF= Alpha1=0.05 Alpha2=0.065 Tau=0.015 Gamma1H=0.0045 Gamma1L=0.0045 Gamma2H=0.006 Gamma2L=0.006 Perturbation=1.1 TGT=500 NF=1.000000000000 SF=14.211210250854 SFGene=All
Background=Avg:29.22,Stdev:0.63,Max:30.8,Min:28.0  Noise=Avg:1.70,Stdev:0.08,Max:1.9,Min:1.6  RawQ=1.37  Corner+=Avg:63,Count:32 Corner-=Avg:12911,Count:32 Central-=Avg:12491,Count:9

molecule: total RNA

organism: Mus musculus

source_name: ankle tissue at 1 weeks of infection

label_protocol: standard Affymetrix procedure

extract_protocol: total RNA extracted from ankle tissues

tag: C3H/HeN mice


title: UC, stage15, biological rep2


title: UC, stage15, biological rep4


title: Expression data from ankle tissue of C3H mice at 2 week of infection

data_processing:
MAS 5
Algorithm: Statistical
HZ=4 VZ=4 BG=4 SmoothFactorBG=100 Epsilon=0.5 BF= Alpha1=0.05 Alpha2=0.065 Tau=0.015 Gamma1H=0.0045 Gamma1L=0.0045 Gamma2H=0.006 Gamma2L=0.006 Perturbation=1.1 TGT=500 NF=1.000000000000 SF=12.772108078003 SFGene=All
Background=Avg:28.48,Stdev:0.67,Max:29.7,Min:27.0  Noise=Avg:1.66,Stdev:0.05,Max:1.8,Min:1.6  RawQ=1.40  Corner+=Avg:103,Count:32 Corner-=Avg:12700,Count:32 Central-=Avg:12236,Count:9

source_name: ankle tissue at 2 weeks of infection


title: Expression data from ankle tissue of C3H mice at 0 week of infection (uninfected)

data_processing:
MAS 5
Algorithm: Statistical
HZ=4 VZ=4 BG=4 SmoothFactorBG=100 Epsilon=0.5 BF= Alpha1=0.05 Alpha2=0.065 Tau=0.015 Gamma1H=0.0045 Gamma1L=0.0045 Gamma2H=0.006 Gamma2L=0.006 Perturbation=1.1 TGT=500 NF=1.000000000000 SF=9.281411170959 SFGene=All
Background=Avg:36.85,Stdev:0.96,Max:39.1,Min:34.6  Noise=Avg:2.40,Stdev:0.41,Max:4.4,Min:2.0  RawQ=1.54  Corner+=Avg:72,Count:32 Corner-=Avg:13443,Count:32 Central-=Avg:12788,Count:9

source_name: ankle tissue at 0 weeks of infection (uninfected)


title: Expression data from ankle tissue of C3H mice at 4 week of infection

data_processing:
MAS 5
Algorithm: Statistical
HZ=4 VZ=4 BG=4 SmoothFactorBG=100 Epsilon=0.5 BF= Alpha1=0.05 Alpha2=0.065 Tau=0.015 Gamma1H=0.0045 Gamma1L=0.0045 Gamma2H=0.006 Gamma2L=0.006 Perturbation=1.1 TGT=500 NF=1.000000000000 SF=8.055230140686 SFGene=All
Background=Avg:38.71,Stdev:1.22,Max:41.2,Min:36.0  Noise=Avg:2.06,Stdev:0.10,Max:2.4,Min:1.8  RawQ=1.63  Corner+=Avg:60,Count:32 Corner-=Avg:14650,Count:32 Central-=Avg:13354,Count:9

source_name: ankle tissue at 4 weeks of infection


title: Untreated Control 12weeks AffyGC RMA rep7

status: Public on Dec 13 2006

city: London

first: Mie

middle: A

last: Rizig

address: Windeyer Institute of Medical Sciences,46 Cleveland Street

department: Mental Health Sciences

country: United Kingdom

institute: University Collage London

description: Gene expression data from the right forebrain of C57BL mouse received no treatment for twelve weeks.

hyb_protocol: Following fragmentation, 15 microg of cRNA were hybridized for 16 hr at 45C on Affymetrix GeneChip Mouse Expression Array 430A . GeneChips were washed and stained in the Affymetrix Fluidics Station 400.

scan_protocol: GeneChips were scanned using the Affymetrix GeneChip Scanner 3000.

data_processing: Normalized expression values were calculated from probe level data (CEL file) using the function of Robust Multi-Array expression measure using sequence information GC-RMA (Wu et al 2004, www.bepress.com/jhubiostat/paper4).Computational steps were implemented in the R-language. GCRMA and it's associated packages were provided by the open source bioconductor project.

source_name: Right brain hemisphere ,C57BL male mouse,Un-treated Control,12 weeks

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 8-10 microg total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).

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

treatment_protocol: Mouse was housed individually in a separate cage under a 12-h light–dark cycle and permitted food and water ad libitum. A period of acclimatization for two weeks was allowed prior to any experimental procedure. No treatment was added to the animal’s drinking water .The weight and water consumption was measured weekly. At the end of 12 weeks animal was sacrificed by cervical dislocation and the brain was removed, dissected into left and right hemispheres and cerebellum, flash frozen in liquid nitrogen and stored at -80 C for future use.

tag: Tissue

value: Forebrain (left hemisphere), Strain:C57BL, Gender:Male, Age:4weeks, Drug treatment:Untreated-Control, Period of treatment:12 weeks, Method of sacrificing:cervical dislocation.


title: UC, stage15, biological rep3


title: Untreated Control 12weeks AffyGC RMA rep10

hyb_protocol: Following fragmentation, 10 microg of cRNA were hybridized for 16 hr at 45C on GeneChip Drosophila Genome Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 400.


title: Untreated Control 12weeks AffyGC RMA rep9


title: Untreated Control 12weeks AffyGC RMA rep8


title: Untreated Control 12weeks AffyGC RMA rep6


title: Untreated Control 12weeks AffyGC RMA rep4


title: Untreated Control 12weeks AffyGC RMA rep3


title: Untreated Control 12weeks AffyGC RMA rep5


title: Untreated Control 12weeks AffyGC RMA rep2


title: Untreated Control 12weeks AffyGC RMA rep1


title: Haloperidol treated 12weeks AffyGC RMA rep4

description: Gene expression data from the right forebrain of C57BL mouse received Haloperidol oral treatment for twelve weeks.

source_name: Right brain hemisphere ,C57BL male mouse,Haloperidol treated,Orally,12 weeks

treatment_protocol: Mouse was housed individually in a separate cage under a 12-h light–dark cycle and permitted food and water ad libitum. A period of acclimatization for two weeks was allowed prior to any experimental procedure. Haloperidol was added to the animal’s drinking water in a concentration equivalent to the highest human clinical dosage (i.e. 1.6 mg/kg/day for Clozapine) and delivered through light protected bottles. The weight and water consumption was measured weekly and the drug concentration in water was adjusted accordingly .At the end of 12 weeks animal was sacrificed by cervical dislocation and the brain was removed, dissected into left and right hemispheres and cerebellum, flash frozen in liquid nitrogen and stored at -80 C for future use.

value: Forebrain (left hemisphere), Strain:C57BL, Gender:Male, Age:4weeks, Drug treatment:Typical antipsychotic Haloperidol, Dose:1.6mg/kg/day, Route of treatment:Oral treatment-Drug added to drinking Water, Period of treatment:12 weeks, Method of sacrificing:cervical dislocation.


title: Haloperidol treated 12weeks AffyGC RMA rep6


title: Haloperidol treated 12weeks AffyGC RMA rep5

value: Forebrain (left hemisphere), Strain:C57BL, Gender:Male, Age:4weeks, Drug treatment:Typical antipsychotic Haloperidol, Dose:1.6mg/kg/day, Route of treatment:Oral treatment-Drug added to drinking Water, Period of treatment:12 weeks, Method of sacrificing:cervical dislocation. At the end of the treatment, drug level in the plasma was assessed by Tandem mass Spectrometry LC- MS/MS.


title: Haloperidol treated 12weeks AffyGC RMA rep10


title: Haloperidol treated 12weeks AffyGC RMA rep1


title: Haloperidol treated 12weeks AffyGC RMA rep7


title: Clozapine treated 12weeks AffyGC RMA rep9

description: Gene expression data from the right forebrain of C57BL mouse received Clozapine oral treatment for twelve weeks..

source_name: Right brain hemisphere ,C57BL male mouse,Clozapine treated,Orally,12 weeks

treatment_protocol: Mouse was housed individually in a separate cage under a 12-h light–dark cycle and permitted food and water ad libitum. A period of acclimatization for two weeks was allowed prior to any experimental procedure. Clozapine was added to the animal’s drinking water in a concentration equivalent to the highest human clinical dosage (i.e. 12mg/kg/day for Clozapine) and delivered through light protected bottles. The weight and water consumption was measured weekly and the drug concentration in water was adjusted accordingly .At the end of 12 weeks animal was sacrificed by cervical dislocation and the brain was removed, dissected into left and right hemispheres and cerebellum, flash frozen in liquid nitrogen and stored at -80 C for future use.

value: Forebrain (left hemisphere), Strain:C57BL, Gender:Male, Age:4weeks, Drug treatment:Atypical antipsychotic Clozapine, Dose:12mg/kg/day, Route of treatment:Oral treatment-Drug added to drinking water, Period of treatment:12 weeks, Method of sacrificing:cervical dislocation.


title: Haloperidol treated 12weeks AffyGC RMA rep2


title: Haloperidol treated 12weeks AffyGC RMA rep8


title: Haloperidol treated 12weeks AffyGC RMA rep3


title: Clozapine treated 12weeks AffyGC RMA rep10

description: Gene expression data from the right forebrain of C57BL mouse received Clozapine oral treatment for twelve weeks.


title: Clozapine treated 12weeks AffyGC RMA rep8


title: Clozapine treated 12weeks AffyGC RMA rep7


title: Clozapine treated 12weeks AffyGC RMA rep6


title: Clozapine treated 12weeks AffyGC RMA rep5


title: 02.spretus.kidneyliver

city: Thuwal

first: Christian

middle: Robert

last: Voolstra

address: Red Sea Research Center

country: Saudi Arabia

institute: King Abdullah University of Science and Technology

description: gene expression in house mice, expression divergence in house mice in 3 tissues (brain, liver/kidney, testis)

data_processing: lowess normalization, sd reg (TIGR midas)

label: alexa fluor 555

source_name: C57Bl6 kidneyliver (pool of 9 individuals)

tag: reference channel

label: alexa fluor 647

organism: Mus spretus

tag: sample channel


title: Clozapine treated 12weeks AffyGC RMA rep3


title: Clozapine treated 12weeks AffyGC RMA rep4


title: Clozapine treated 12weeks AffyGC RMA rep1


title: 03.spretus.testis

source_name: C57BL6 testis (pool of 9 individuals)

source_name: Mus spretus testis


title: 03.cas.brain

source_name: C57Bl6 brain (pool of 9 individuals)


title: Haloperidol treated 12weeks AffyGC RMA rep9


title: 03.spretus.kidneyliver

source_name: C57BL6 kidneyliver (pool of 9 individuals)

source_name: Mus spretus kidneyliver


title: Clozapine treated 12weeks AffyGC RMA rep2


title: 03.spretus.brain

source_name: C57BL6 brain (pool of 9 individuals)

source_name: Mus spretus brain


title: 05.ssp.brain

source_name: Mus musculus ssp brain


title: 05.ssp.testis

source_name: Mus musculus ssp testis


title: 06.ssp.brain


title: 02.spretus.testis


title: 06.ssp.testis


title: 02.spretus.brain


title: 01.spretus.kidneyliver


title: 06.ssp.kidneyliver

source_name: Mus musculus ssp kidneyliver


title: 05.ssp.kidneyliver


title: 03.ssp.kidneyliver


title: 04.ssp.testis


title: 01.spretus.brain


title: 04.ssp.brain


title: 03.ssp.testis


title: 03.ssp.brain


title: 02.ssp.testis


title: 02.ssp.brain


title: 01.ssp.kidneyliver


title: 01.spretus.testis


title: 02.ssp.kidneyliver


title: 08.mus.kidneyliver

source_name: Mus musculus musculus kidneyliver


title: 07.mus.kidneyliver


title: 01.ssp.brain


title: Bsu168vDV1-E-2 replicate 1

type: genomic

city: Boston

first: Ashlee

last: Earl

state: MA

address: 200 Longwood Avenue D-1, Rm 219

institute: Harvard Medical School

description: Microarray-based comparative hybridization analysis of Bsu168, the sequenced representative of B. subtilis, with another B. subtilis isolate using Bsu168-specific microarrays.

hyb_protocol: Purified Cy5- and Cy3-labeled probes (Qiagen PCR Purification Kit, Valencia, CA) were mixed and concentrated to ~5mL using Microcon YM-30 columns (Millipore, Bedford, MA).  The following solutions were then added sequentially to each Cy5/Cy3 mixture: 1ml yeast tRNA (20?g/ml), 3.75ml 20X SSC, 0.75ml 10%SDS and 4.5ml 100% formamide.  The mixture was boiled for one minute and then a 13.5ml aliquot was added to the microarray and covered with a 22X22 mm coverslip.  The microarrays were incubated overnight inside a CMT-hybridization chamber submerged in a 42oC water bath.

scan_protocol: Slides were scanned for Cy5 and Cy3 fluorescence intensities using a ScanArray 5000 (Packard Instruments). Laser power and or PMT were adjusted such that the two channels were balanced.

data_processing:
Microarray images were loaded into Genepix 4.0 (Axon Instruments, Union City, CA) to calculate the Cy5 and Cy3 fluorescence intensity ratios for all gene spots. Within a given experiment, gene spots with fluorescence intensities less than the average of negative control spot intensities were excluded from analysis. Ratios above this cut-off were then transformed by log2. Normalization to compensate for slide-to-slide variation was accomplished by adjusting all ratios so that the median of all log2 ratios for a given experiment equaled zero.  Values reported here always reflect Bsu168/test strain ratio no matter which labeling regimen was used.
The raw datasets report information for all gene spots; however, data from 384 genes should be disregarded from all datasets due to a printing inconsistency.  These genes correspond to those in Row 16, Columns 1-18 and Row 15, Columns 13-18.  These datapoints can be easily eliminated from analysis of the raw data by sorting and deleting.  These data were already removed from the list below.

molecule: genomic DNA

organism: Bacillus spizizenii

label_protocol: Two micrograms of purified, Sau3AI digested gDNA were labeled with either Cy3- or Cy5-conjugated dCTP as described previously (Dziejman, M., E. et al. 2002. PNAS 99:1556-61).

extract_protocol: Mid- to late-log phase cultures grown in LB liquid medium were harvested by centrifugation.  Genomic DNA was extracted as described in Current Protocols in Molecular Biology (Wilson, K. 1997. John Wiley and Sons, New York.) except that resuspended cells were first incubated at 37oC for 10 minutes in the presence of lysozyme (2 mg/mL) and RNase (80 mg/mL).

tag: NRRL B-23050 Death Valley National Monument, California/USDA

organism: Bacillus subtilis subsp. subtilis

tag: trpC2, sequenced strain


title: Bsu168vBsu168 replicate 3


title: Bsu168vDV1-E-2 replicate 3


title: 08.mus.testis

source_name: Mus musculus musculus testis


title: 07.mus.testis


title: Bsu168vBsu168 replicate 2


title: Bsu168vDV1-E-2 replicate 2


title: 05.mus.brain

source_name: Mus musculus musculus brain


title: 04.mus.kidneyliver


title: 04.ssp.kidneyliver


title: Bsu168vBsu168 replicate 1


title: 03.mus.testis


title: 04.mus.brain


title: Bsu168vNCIB3610 replicate 3

tag: Marburg, Germany “wild” strain/BGSC


title: 04.mus.testis


title: 03.mus.kidneyliver


title: 08.mus.brain

source_name: Mus musculus domesticus brain


title: 05.mus.kidneyliver


title: 07.mus.brain


title: 05.mus.testis


title: Bsu168vNCIB3610 replicate 2


title: Bsu168vBsu168eps::tet replicate 2

tag: Bsu168 derivative containing a 15 gene deletion of the eps operon


title: Bsu168vBsu168eps::tet replicate 3


title: Bsu168vBsu168eps::tet replicate 1


title: Bsu168vNCIB3610 replicate 1


title: 01.ssp.testis


title: Bsu168vTU-B-10 replicate 2

tag: NRRL B-23049 Saharan Desert Nefta, Egypt/USDA


title: Bsu168vTU-B-10 replicate 3


title: Bsu168vTU-B-10 replicate 1


title: Bsu168vT-89-48 replicate 2

tag: NRRL B-23076 Sonoran Desert, Tumamoc Hill, Arizona/USDA


title: 03.mus.brain


title: 02.mus.testis


title: 02.mus.brain


title: Bsu168vT-89-48 replicate 3


title: 13.dom.kidneyliver

source_name: Mus musculus domesticus kidneyliver


title: 10.dom.brain


title: 06.dom.kidneyliver


title: Bsu168vW23replicate 3

tag: NRRL B-14472 USDA


title: 06.dom.testis

source_name: Mus musculus domesticus testis


title: 13.dom.brain


title: 10.dom.kidneyliver


title: 06.dom.brain


title: Bsu168vW23replicate 2


title: 10.dom.testis


title: 02.mus.kidneyliver


title: 13.dom.testis


title: Bsu168vW23replicate 1


title: Bsu168vDV1-F-3 replicate 3

description: Microarray-based comparative hybridization analysis of Bsu168, the sequenced representative of B. subtilis, with a B. vallismortis isolate using Bsu168-specific microarrays.

organism: Bacillus vallismortis

tag: NRRL B-14890 Death Valley National Monument, California/USDA


title: Bsu168vT-89-48 replicate 1


title: Bsu168vDV1-F-3 replicate 2


title: Bsu168vDV1-F-3 replicate 1


title: Bsu168vMGla47 replicate 3

tag: Mendenhall Glacier, Alaska


title: Bsu168vMGla47 replicate 2


title: Bsu168vMGla47 replicate 1


title: Bsu168vRO-FF-1 replicate 3

tag: NRRL B-23074 Mojave Desert Rosamond, California/USDA


title: 05.dom.brain


title: 05.dom.testis


title: 03.dom.brain


title: 01.dom.testis


title: Bsu168vDV1-B-1 replicate 3

tag: NRRL B-23054 Death Valley National Monument, California/USDA


title: 03.dom.kidneyliver


title: 03.dom.testis


title: 03.cas.testis

source_name: Mus musculus castaneus testis


title: 05.dom.kidneyliver


title: 01.dom.brain


title: 02.cas.testis

source_name: C57BL6 testis (pool of 9 individuals)


title: Bsu168vRO-FF-1 replicate 2


title: Bsu168vN10 replicate 4

tag: Mediterranean Sea, Coast of Egypt/BGSC


title: 03.cas.kidneyliver

source_name: Mus musculus castaneus kidneyliver


title: 02.cas.kidneyliver


title: 01.dom.kidneyliver


title: Bsu168vDV1-B-1 replicate 2


title: 02.cas.brain

source_name: Mus musculus castaneus brain


title: Bsu168vRO-FF-1 replicate 1


title: Bsu168vRO-E-2 replicate 1

tag: NRRL B-23055 Mojave Desert Rosamond, California/USDA


title: Bsu168vN10 replicate 3


title: Bsu168vN10 replicate 1


title: Bsu168vHLR11 replicate 3

tag: Hyallite Reservoir, MT


title: Bsu168vN10 replicate 2


title: 01.cas.kidneyliver

source_name: C57BL6 kidneyliver (pool of 9 individuals)


title: Bsu168vHLR11 replicate 2


title: Bsu168vHLR11 replicate 1


title: Bsu168vRO-E-2 replicate 3


title: Bsu168vHIKEM-7 replicate 3

tag: Kee, Hawaii


title: Bsu168vDV1-B-1 replicate 1


title: Bsu168vRO-E-2 replicate 2


title: Bsu168vHIKEM-7 replicate 2


title: LEA1_13myc_SUB2_85_ChIP

first: Michael

middle: J

last: Moore

address: 200 Longwood Ave

department: Systems Biology

description: This data represents combined raw values from three independent biological replicates.

hyb_protocol: Hybridized at 37 degrees overnight in DIG Easy Hyb Mix (Roche) supplemented with ssDNA and yeast tRNA.

scan_protocol: GenePIX 4000B

data_processing: Biological replicates were combined and analyzed with Rosetta Resolver.

organism: Saccharomyces cerevisiae

source_name: IP

label_protocol: Cy5- or Cy3-UTP was incorporated by Klenow treatment in the presence of a Random hexamer mix (Bioprime Kit from Invitrogen). IP fraction was labeled with Cy5; WCE with Cy3. In at least one biological replicate fluorophores were reversed.

growth_protocol:
1.) Vegetative growth in SC-HIS media to O.D.=0.6.
2.) Warmed to 37C for 5 minutes.
3.) Grown at 37C for 30 minutes prior to fixation.

extract_protocol:
1.) Chromatin IP
2.) Linker mediated PCR amplification

treatment_protocol: Cells were heat shocked to deplete them of functional SUB2.

tag: anti-myc chromatin IP from LEA1-13myc strain with plasmid-borne sub2-85 temperature sensitive allele.

source_name: WCE

extract_protocol: Linker mediated PCR of WCE fraction.

tag: whole chromatin extract from LEA1-13myc strain with plasmid-borne sub2-85 temperature sensitive allele.


title: BRR2_13myc_SUB2_WT_ChIP

treatment_protocol: Cells were heat shocked as for sub2-85 ts strain, to control for effects of temperature change.

tag: anti-myc chromatin IP from BRR2-13myc strain with plasmid-borne SUB2 (WT).

tag: whole chromatin extract BRR2-13myc strain with plasmid-borne SUB2 (WT).


title: MUD2_13myc_RNAse_ChIP

growth_protocol: Vegetative growth in rich media to O.D.=0.6.

treatment_protocol: Fixation shortened to sensitive cross-linked complexes to RNAse digestion.  WCE was treated with RNAse prior to ChIP.

tag: anti-myc chromatin IP of MUD2-13myc strain.  WCE was treated with RNAse prior to ChIP.

treatment_protocol: Fixation shortened to sensitive cross-linked complexes to RNAse digestion.  WCE was treated with RNAse prior to amplification.

tag: whole chromatin extract of MUD2-13myc strain.  WCE was treated with RNAse prior amplification.


title: PRP40_9myc_RNAse_ChIP

treatment_protocol: Fixation shortened to sensitive cross-linked complexes to RNAse digestion.  Samples were treated with RNase.

tag: anti-myc chromatin IP from PRP40-9myc strain.  WCE was treated with RNAse prior to ChIP.

tag: Whole chromatin extract from PRP40-9myc strain.  WCE was treated with RNAse prior to ChIP.


title: BRR2_13myc_SUB2_85_ChIP

tag: anti-myc chromatin IP from BRR2-13myc strain with plasmid-borne sub2-85 temperature sensitive allele.

tag: whole chromatin extract from BRR2-13myc strain with plasmid-borne sub2-85 temperature sensitive allele.


title: LEA1_13myc_SUB2_WT_ChIP

tag: anti-myc chromatin IP from LEA1-13myc strain with plasmid-borne SUB2 (WT)

tag: whole chromatin extract from LEA1-13myc strain with plasmid-borne SUB2 (WT)


title: PRP40_9myc_del_cbc_ChIP

tag: anti-myc chromatin IP from PRP40-9myc strain lacking both subunits (CBP80 and CBP20) of cap-binding-complex (CBC)

tag: whole chromatin extract from PRP40-9myc strain lacking both subunits (CBP80 and CBP20) of cap-binding-complex (CBC)


title: PRP40_9myc_no_RNAse_ChIP

treatment_protocol: Fixation shortened as for RNAse treatment protocol, such that these samples could properly control for RNAse treatment.

tag: anti-myc chromatin IP of PRP40-9myc strain.  Served as control for RNAse sensitivity assay for Prp40.

tag: whole chromatin extract of PRP40-9myc strain.  Served as control for RNAse sensitivity assay for Prp40.


title: MUD2_13myc_no_RNAse_ChIP

treatment_protocol: Fixation shortened as for RNAse treatment protocol, such that these samples properly controlled for RNAse treatment.

tag: anti-myc chromatin IP from MUD2-13myc strain.  These samples served as a control for RNAse treatment.

tag: Whole chromatin extract from MUD2-13myc strain.  These samples served as a control for RNAse treatment.


title: 01.cas.brain

source_name: C57BL6 brain (pool of 9 individuals)


title: PRP40_3myc_6AU_ChIP

growth_protocol: Vegetative growth in SC-URA media to O.D.=0.6.

treatment_protocol: Treated with 80ug/ml 6-azauracil, a transcription elongation inhibitor, for 30 minutes prior to fixation.

tag: anti-myc chromatin IP of PRP40-3myc strain treated with 6AU

tag: Whole chromatin extract of PRP40-3myc strain treated with 6AU


title: 01.cas.testis


title: Bsu168vHIKEM-7 replicate 1

tag: trpC2, sequenced strain

tag: Kee, Hawaii


title: Bsu168v1431 replicate 1

tag: Moscow, Russia/BGSC


title: Bsu168v1431 replicate 3


title: Bsu168v1440 replicate 3


title: PRP40_3myc_spt5_194_ChIP

tag: anti-myc chromatin IP of PRP40-3myc; spt5-194 strain.

tag: whole chromatin extract of PRP40-3myc; spt5-194 strain.


title: Bsu168v1431 replicate 2


title: Bsu168vBS5 replicate 2

tag: tetracyclin sensitive strain derived from CU2189


title: Bsu168v1440 replicate 1


title: Bsu168vBS5 replicate 1


title: Bsu168vBS5 replicate 3


title: PRP40_3myc_control_ChIP

description: This data represents combined raw values from four independent biological replicates.

tag: anti-myc chromatin IP of PRP40-3myc strain.

tag: whole chromatin extract of PRP40-3myc strain.


title: RNAPII_ChIP

description: This data represents combined raw values from two independent biological replicates.

tag: Chromatin IP of RNAPII-specific subunit RPB3 with anti-RPB3 antibody (Neoclone) from PSY2156 strain (W303).

tag: Whole chromatin extract from PSY2156 (W303) strain.


title: SPT5_9myc_ChIP

tag: anti-myc chromatin IP of SPT5-9myc strain.

tag: whole chromatin extract of SPT5-9myc strain.


title: Untreated Control 4weeks AffyGC RMA rep3

description: Gene expression data from the right forebrain of C57BL mouse received no treatment for four weeks.

hyb_protocol: Following fragmentation, 15 microg of cRNA were hybridized for 16 hr at 45C on Affymetrix GeneChip Murine Genome U74 Version 2 Set MG-U74A. GeneChips were washed and stained in the Affymetrix Fluidics Station 400.

scan_protocol: GeneChips were scanned using GeneArray Scanner (Agilent Technologies, Foster City, CA)..

source_name: Right brain hemisphere ,C57BL male mouse,Un-treated Control,4 weeks

treatment_protocol: Mouse was housed individually in a separate cage under a 12-h light–dark cycle and permitted food and water ad libitum. A period of acclimatization for two weeks was allowed prior to any experimental procedure. No treatment was added to the animal’s drinking water .The weight and water consumption was measured weekly. At the end of 4 weeks animal was sacrificed by cervical dislocation and the brain was removed, dissected into left and right hemispheres and cerebellum, flash frozen in liquid nitrogen and stored at -80 C for future use.

tag: Tissue

value: Forebrain (left hemisphere), Strain:C57BL, Gender:Male, Age:4weeks, Drug treatment:Untreated-Control, Period of treatment:4 weeks, Method of sacrificing:cervical dislocation.


title: Bsu168vnatto replicate 2

source_name: natto

tag: IFO3335 Japan


title: Haloperidol treated 4weeks AffyGC RMA rep3

description: Gene expression data from the right forebrain of C57BL mouse received Haloperidol oral treatment for four weeks.

source_name: Right brain hemisphere ,C57BL male mouse,Haloperidol treated,Orally,4 weeks

treatment_protocol: Mouse was housed individually in a separate cage under a 12-h light–dark cycle and permitted food and water ad libitum. A period of acclimatization for two weeks was allowed prior to any experimental procedure. Haloperidol was added to the animal’s drinking water in a concentration equivalent to the highest human clinical dosage (i.e. 1.6 mg/kg/day) and delivered through light protected bottles. The weight and water consumption was measured weekly and the drug concentration in water was adjusted accordingly .At the end of 4 weeks animal was sacrificed by cervical dislocation and the brain was removed, dissected into left and right hemispheres and cerebellum, flash frozen in liquid nitrogen and stored at -80 C for future use.

value: Forebrain (left hemisphere), Strain:C57BL, Gender:Male, Age:4weeks, Drug treatment:Typical antipsychotic Haloperidol, Dose:1.6mg/kg/day, Route of treatment:Oral treatment-Drug added to drinking Water, Period of treatment:4 weeks, Method of sacrificing:cervical dislocation.


title: SUB2_3HA_ChIP

description: This data represents combined raw values from six independent biological replicates.

tag: anti-HA chromatin IP of SUB2-3HA strain.

tag: Whole chromatin extract of SUB2-3HA strain.


title: PRP2_13myc_ChIP

tag: anti-myc chromatin IP of PRP2-13myc strain.

tag: Whole chromatin extract of PRP2-13myc strain.


title: PRP19_9myc_ChIP

tag: anti-myc chromatin IP of PRP19-9myc strain.

tag: whole chromatin extract of PRP19-9myc strain.


title: Bsu168v1440 replicate 2

tag: Moscow, Russia/BGSC

tag: trpC2, sequenced strain


title: BRR2_13myc_ChIP

tag: anti-myc chromatin IP of BRR2-13myc strain.

tag: whole chromatin extract of BRR2-13myc strain.


title: Haloperidol treated 4weeks AffyGC RMA rep2

hyb_protocol: Following fragmentation, 10 microg of cRNA were hybridized for 16 hr at 45C on Affymetrix GeneChip Murine Genome U74 Version 2 Set MG-U74A. GeneChips were washed and stained in the Affymetrix Fluidics Station 400.


title: Untreated Control 4weeks AffyGC RMA rep1


title: PRP5_9myc_ChIP

tag: anti-myc chromatin IP of PRP5-9myc strain.

tag: Whole chromatin extract of PRP5-9myc strain.


title: Bsu168vnatto replicate 3


title: Clozapine treated 4weeks AffyGC RMA rep3

description: Gene expression data from the right forebrain of C57BL mouse received Clozapine oral treatment for four weeks..

source_name: Right brain hemisphere ,C57BL male mouse,Clozapine treated,Orally,4 weeks

treatment_protocol: Mouse was housed individually in a separate cage under a 12-h light–dark cycle and permitted food and water ad libitum. A period of acclimatization for two weeks was allowed prior to any experimental procedure. Clozapine was added to the animal’s drinking water in a concentration equivalent to the highest human clinical dosage (i.e. 12mg/kg/day for Clozapine) and delivered through light protected bottles. The weight and water consumption was measured weekly and the drug concentration in water was adjusted accordingly .At the end of 4 weeks animal was sacrificed by cervical dislocation and the brain was removed, dissected into left and right hemispheres and cerebellum, flash frozen in liquid nitrogen and stored at -80 C for future use.

value: Forebrain (left hemisphere), Strain:C57BL, Gender:Male, Age:4weeks, Drug treatment:Atypical antipsychotic Clozapine, Dose:12mg/kg/day, Route of treatment:Oral treatment-Drug added to drinking water, Period of treatment:4 weeks, Method of sacrificing:cervical dislocation.


title: Untreated Control 4weeks AffyGC RMA rep2


title: Clozapine treated 4weeks AffyGC RMA rep1

scan_protocol: GeneChips were scanned using GeneArray Scanner (Agilent Technologies, Foster City, CA).


title: Clozapine treated 4weeks AffyGC RMA rep2

description: Gene expression data from the right forebrain of C57BL mouse received Clozapine oral treatment for four weeks.


title: Bsu168vRO-NN-1 replicate 3

tag: NRRL B-14823 isolated Mojave Desert Rosamond, California/USDA


title: lea1_13myc_ChIP

tag: anti-myc chromatin IP of lea1-13myc strain.

tag: Whole chromatin extract of Lea1-13myc strain.


title: Bsu168vRO-NN-1 replicate 2


title: Bsu168vnatto replicate 1


title: Bsu168vRO-NN-1 replicate 1


title: Haloperidol treated 4weeks AffyGC RMA rep1

description: Gene expression data from the right forebrain of C57BL mouse received Haloperidol oral treatment four weeks.

tag: Tissue


title: Mud2_13myc_ChIP

tag: anti-myc ChIP of Mud2-13myc strain

tag: Whole chromatin extract from Mud2-13myc strain.


title: BBP_3HA_ChIP

hyb_protocol: Hybridized at 37 degrees in DIG Easy Hyb Mix (Roche) overnight.

label_protocol: Cy5- or Cy3-UTP was incorporated by Klenow treatment in the presence of a Random hexamer mix (Bioprime Kit from Invitrogen).  IP fraction was labeled with Cy5; WCE with Cy3.  In at least one biological replicate fluorophores were reversed.

extract_protocol:
1.)  Chromatin IP
2.)  Linker mediated PCR amplification

tag: anti-HA ChIP of BBP-3HA strain

tag: Whole chromatin extract from BBP-3HA strain.


title: PRP40_9myc_ChIP

description: This data represents combined raw values from five independent biological replicates.

tag: anti-myc ChIP of Prp40-9myc strain

tag: Whole chromatin extract from Prp40-9myc strain.


title: Pt_3 non-acne Normal Skin

status: Public on Dec 12 2006

city: Hershey

first: Wei

last: Zhao

state: PA

address: 600 Centerview Drive,  A210

institute: Penn State University

description: Check paper for details. "Gene Array Expression Profiling in Acne Lesions Reveals Marked Upregulation of Genes Involved in Inflammation and Matrix Remodeling". Journal of Investigative Dermatology (JID 126:1071-1079, 2006).

hyb_protocol: Biotinylated cRNA, produced through in vitro transcription, was fragmented and hybridized to an Affymetrix human U133A 2.0 microarray.

scan_protocol: The arrays were processed on a GeneChip Fluidics Station 450 and scanned on an Affymetrix GeneChip Scanner (Santa Clara, CA).

data_processing: The expression signals were normalized by using the R-Affy package from Bioconductor version 1.1 to remove background noise and non-biological variations among arrays. The background noise was removed from the PM probe intensities using the "RMA" method. Normalization was performed using the quantile normalization method (Bolstad et al., 2003).  To remove outlier probes and summarize probe intensities within one probe set into a single expression value, the "Tukey Biweight" method was applied to the background adjusted, normalized PM intensities. Expression values were obtained based on PM intensities and not PM-MM intensities, because PM and MM intensities were found to be highly correlated, which suggested that MM intensities were composed of background noise as well as probe-specific signals. Significant gene expression alterations were identified using the computer software Significance Analysis of Microarrays (Tusher et al., 2001).

organism: Homo sapiens

source_name: patient skin sample-non-acne Normal Skin

label_protocol: Biotinylated cRNA, produced through in vitro transcription.

extract_protocol: Total RNA was isolated from skin samples and DNase treated using the RNeasy Fibrous Tissue Kit (Qiagen Inc., Valencia, CA). RNA was ethanol precipitated and quantified using a spectrophotometer. Approximately 2 mug of total RNA from each sample was used to generate double-stranded cDNA using a T7-oligo (dT) primer.

tag: non-acne patient normal skin sample


title: Pt_5 non-acne Normal Skin


title: Pt_2 non-acne Normal Skin


title: Genome-wide distribution of yeast RNA polymerase II and its control by Sen1 helicase - Sen1 mutant strain

city: Madison

first: Christopher

middle: Lawrence

last: Warren

state: WI

address: 1400 University Ave

institute: University of Wisconsin - Madison

description:
Microarrays were synthesized by NimbleGen Systems, Inc. using a Maskless Array Synthesizer (Singh-Gasson et al., 1999). Homopolymer (T10) linkers were covalently attached to monohydroxysilane glass slides, and 24-mer oligonucleotides were then synthesized on the
homopolymers. The microarrays contain ~194,000 features that “tile” the Saccharomyces cerevisiae S288C genome, with each feature consisting of many copies of an oligonucleotide complementary to a specific sequence in the genome. Sequences were spaced every 60-120
base pairs along the genome, alternating between the Watson and Crick strands. Probe sequences were chosen within this 60 bp range by NimbleGen to maximize each probe’s genomic uniqueness. The microarray was based on Saccharomyces Genome Database (SGD;
www.yeastgenome.org) genomic sequence as of 6/1/03. Analysis of the ChIP-chip data converted those probe sequences to SGD version 11/1/05 of the genomic sequence and annotation.
Information pertaining to each probe is included with the raw data.

hyb_protocol: Equal masses (2.5 μg each) of purified, Cy-dye labeled input and IP samples from a single strain were mixed and hybridized to microarrays. A duplicate hybridization was performed with the dye labels swapped; i.e. with Cy3 coupled to IP DNA and Cy5 coupled to input DNA in experiment 1 and Cy5 coupled to IP DNA and Cy3 coupled to input DNA in experiment 2. Hybridization was carried out for 16 hr. at 45˚C.

scan_protocol: Scanner is Axon 4000B and extraction software is NimbleScan  2.1.

data_processing: Raw data from each channel on each array was scaled so that the mean of the cy3 and cy5 channels are equal.  An average of the two replicates (dye swap) was taken for each probe.  Natural log ratios of these averaged scaled intensities for each feature on the array was then taken.  Nine positionally adjacent ln ratios were averaged together (four 5’ and four 3’ of a given position) to get a smoothed value for each chromosomal probe.  This final smoothed natural log ratio averaged from two replicates (through dye swap) are the VALUE column of the data.

source_name: ChIP-chip of Polymerase II genomic localization in a Sen1 mutant strain of S. cerevisiae

label_protocol: Amplified samples (500 ng) were labeled with the Bioprime Array-CGH kit (Invitrogen) using 48 μM amino-allyl dUTP (Sigma), 72 μM dTTP, and 120 μM each of dATP, dCTP, and dGTP, yielding 4 to 5 μg of modified DNA, 80% of which was then coupled to NHS esters (Amersham RPN 5661) of Cy3 (input) and Cy5 (IP).

extract_protocol:
Experiments were performed with the sen1-E1597K mutant strain, it’s wild type parent 46alpha, or a Sen1-myc13 derivative of 46alpha. A one-step integration strategy was used to introduce a myc13-kanMX6 cassette (Longtine et al., 1998) at the 3’ end of the chromosomal SEN1 locus in yeast strains 46alpha and EJS101-9c. Proper integration was confirmed by PCR and sequencing. Preparation and immunoprecipitation of cross-linked chromatin were based on a procedure described previously (Komarnitsky et al., 2000). Thirty ml of 11.1% formaldehyde in 100 mM NaCl, 1 mM EDTA, 50 mM HEPES-KOH was added to 300 ml of yeast culture grown in YEPD at 30˚C to mid-log phase. The mixture was incubated at room temperature for 20 min, followed by addition of 45 ml of 3 M glycine, 20 mM Tris base for 5 min. Cells were harvested by centrifugation, washed twice with 100 ml TBS (20 mM Tris-HCl, pH 7.5, 150 mM NaCl) and once in 10 ml FA lysis buffer (50 mM HEPES KOH, pH 7.5, 150 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate) + 0.1% SDS. Cells were resuspended in 1 ml FA lysis buffer + 0.5% SDS, and lysed by vortexing with 1.5 ml glass beads (212-300 μm diameter) 20 to 30 times for 30 seconds each, with 30 seconds on ice between vortexing. The lysate was added to 6.5 ml FA lysis buffer + 0.1% SDS, then ultracentrifuged for 20 minutes at 45,000 rpm, 4˚C in a 70.1 Ti rotor. The pellet was resuspended in 8 ml FA lysis buffer + 0.1% SDS, and ultracentrifugation was repeated. The pellet was then resuspended in 1.5 ml FA lysis buffer + 0.1% SDS, and the chromatin was sheared by sonication to an average fragment size of 200-500 base pairs using a Model 100 sonic dismembrator (Fisher Scientific) or a Sonifier 250 (Branson). The sonicated sample was then added to 6.5 ml FA lysis buffer + 0.1% SDS, and chromatin fragments were recovered in the supernatant after ultracentrifugation as above. Samples were stored in 800 μl aliquots at –70˚C.
Immunoprecipitation was performed on 750 μl crosslinked chromatin with salt concentration adjusted to 275 mM NaCl. Samples were incubated 6-14 hrs. with rotation at 4˚C with ~ 10 μl bed volume of protein A-Sepharose (protein G-Sepharose for 1Y26 antibody) and 5-10 μl of monoclonal antibody 9E10 against the myc epitope (Upstate Biochemicals #05-419) or 7 μl of 1Y26 antibody against the Rpb3 subunit of Pol II (1Y26 was a gift from Nancy Thompson and Richard Burgess, University of Wisconsin). Immunoprecipitates were washed once each with 1.4 ml of the following: 1) FA lysis buffer/275 mM NaCl/0.1% SDS, 2) FA lysis buffer/500 mM NaCl/0.1% SDS, 3) 10 mM Tris-HCl pH 8.0/0.25 M LiCl/1 mM EDTA/0.5% NP-40/0.5% Na-deoxycholate, and 4) 10 mM Tris-HCl, pH 8.0/1 mM EDTA (TE). Chromatin was eluted from protein A-Sepharose by incubation in 200 μl of 50 mM Tris pH 7.5, 1 mM EDTA, 1 % SDS at 65˚ C for 10 minutes. Twenty μl of 10 mg/ml pronase (Roche) was added to the supernatant and incubated for 1 hour at 42˚C, followed by incubation at 65˚C for 5 hours to reverse crosslinks. Samples were extracted with an equal volume of phenol/chloroform/isoamyl alcohol (25:24:1), pH 7.9 (Ambion), followed by chloroform. DNA was precipitated with 2.5 volumes of EtOH in the presence of 20 μg glycogen, and resuspended in 200 μl TE buffer (50 μl for Sen1-myc13 immunoprecipitates). Samples of input chromatin were treated similarly.
Cross-linked chromatin prepared from 46alpha and sen1-E1597K mutant cultures grown in YEPD at 30˚C (semi-permissive for sen1 mutant) was immunoprecipitated with monoclonal anti-Rpb3 1Y26 and protein G-Sepharose as described above. Input (2 μl) and immunoprecipitated (IP, approx. 2/3 of material recovered from IP of a 750 μl aliquot) samples were amplified by ligation-mediated PCR and labeled as described (Oberley and Farnham, 2003). Briefly, DNA samples were treated with T4 DNA polymerase to create blunt ends and ligated to annealed primers LMPCR1 (5’- dGCGGTGACCCGGGAGATCTGAATTC-3’) and LMPCR2 (5’-dGAATTCAGATC-3’). Amplification was carried out with primer LMPCR1 in two 15-cycle stages, with purification of products between stages using the Qiaquick PCR purification kit (Qiagen), yielding 3-5 μg of DNA from input and IP samples.

tag: The cup1Δ sen1-E1597K (nrd2-1) strain and its cup1Δ SEN1 parent 46alpha MATalpha cup1 Δ ura3 his3 trp1 lys2 ade2 leu2; (Lesser and Guthrie, 1993) were described previously (Steinmetz and Brow, 1996). The nrd2-1 strain was obtained from a derivative of 46alpha with switched mating type, so the nrd2-1 strain is MATa. EJS101-9c is a NRD1 haploid progeny from the heterozygous nrd1Δ::HIS3/NRD1 diploid strain EJS101 (Steinmetz and Brow, 1996). Strain CD4KO (rrp6::KanMX, imd1::HIS3, imd3::KanMX) was provided by M. Ares and was described previously (Davis and Ares, 2006). CD4KO was transformed with pRS316 to provide a functional copy of the URA3 gene, which enhanced the response to 6-azauracil. The strain was grown on SC Ura medium to maintain the plasmid.

source_name: S. cerevisiae ChIP-chip of Polymerase II genomic localization in a Sen1 mutant strain


title: Genome-wide distribution of yeast RNA polymerase II and its control by Sen1 helicase - WT strain

source_name: Saccharomyces cerevisiae ChIP-chip Polymerase II Immunoprecipitated DNA in Wild-Type S. cerevisiae


title: Pt_5 Acne Lesion

description: The inclusion criteria for the acne lesion group included (a) men and women aged 18-45 years with inflammatory acne on their back, (b) subjects without other skin disease in the biopsy area, (c) subjects who were willing to have skin biopsies performed from their back, and (d) subjects who have not been treated with isotretinoin for acne within the previous 6 months.

source_name: patient skin sample-Acne Lesion

tag: acne skin sample


title: Pt_5 Normal Skin

source_name: patient skin sample-Normal Skin

tag: normal skin sample


title: Pt_4 Normal Skin


title: Pt_4 Acne Lesion


title: Pt_3 Normal Skin


title: Pt_6 Normal Skin


title: PRP11_13myc_ChIP

tag: anti-myc chromatin IP of PRP11-13myc strain.

tag: Whole chromatin extract of PRP11-13myc strain.


title: Pt_2 Acne Lesion

source_name: Patient skin sample-Acne Lesion


title: Pt_2 Normal Skin


title: Pt_4 non-acne Normal Skin


title: Pt_3 Acne Lesion


title: Pt_1 Acne Lesion


title: Pt_1 non-acne Normal Skin


title: Pt_1 Normal Skin


title: pf09_44

status: Public on Dec 11 2006

city: San Diego

first: Hongfan

last: Jin

state: CA

address: 5501 Oberlin Drive

department: Biotechnology

institute: The Dow Chemical Company

description: This is the replicate 2 of the comparison of I2m vs I0

data_processing: log2 ratio of lowess normalized data

organism: Pseudomonas fluorescens

source_name: I0: the 0 hr control sample

tag: Pseudomonas fluorescens DC454 control without MMC treatment 0 hr sample

source_name: I2m: the mmc-treated 2 hr sample

tag: Pseudomonas fluorescens DC454 MMC treated 2 hr sample


title: pf09_40

description: This is the replicate 2 of the comparison of I1mvs I0

source_name: I1m: 1hr MMC-treated sample

tag: Pseudomonas fluorescens DC454 MMC treated 1 hr sample


title: pf09_41

description: This is the replicate 1 of the comparison of I2c vs I0

source_name: I2c:the 2 hr control sample

tag: Pseudomonas fluorescens DC454 control without MMC treatment 2 hr sample


title: pf09_42

description: This is the replicate 2 of the comparison of I2c vs I0

source_name: I2c: the 2 hr control sample


title: pf09_43

description: This is the replicate 1 of the comparison of I2m vs I0

source_name: I2m: the MMC-treated 2hr sample


title: Pt_6 Acne Lesion


title: Rice Bala 1ppm Arsenate Rep3

city: Aberdeen

first: Gareth

last: Norton

address: St Machar drive

department: Plant and Soil Science

institute: University of Aberdeen

description: Gene expression data from rice roots (variety Bala) grown in hydroponics for one week with 1ppm arsenate, rep3

hyb_protocol: Hybridisation, washing and staining procedures of the Affymetrix GeneChip Rice Genome arrays were carried out by the Affymetrix core facility of the University of Aberdeen, as described in the Affymetrix technical manual.

scan_protocol: Scanning procedures of the Affymetrix GeneChip Rice Genome arrays were carried out by the Affymetrix core facility of the University of Aberdeen, as described in the Affymetrix technical manual.

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

organism: Oryza sativa

source_name: Rice Hydroponic Roots 1 Week Old

label_protocol: RNA was processed for use with the Affymetrix GeneChip Rice Genome array according to the manufactures protocol. Briefly, 5 ug of total RNA was used in the one-cycle cDNA synthesis reaction. After the generation of double stranded cDNA an in vitro transcription reaction was performed to generate biotinylated cRNA. (Expression Analysis Technical Manual, 2004, Affymetrix).

extract_protocol: 100 mg of root material was pooled from four separate plants and RNA was extracted using the TRIzol method (Invitrogen) followed by further purification and DNase treatment using the RNeasy clean up kit (Qiagen).

treatment_protocol: Germinated seedlings were grown for 1 week in hydroponics with 1ppm arsenate under 12 hr of light as described in Dasgupta T, Hossain SA, Meharg AA and Price AH. (2004) An arsenate tolerance gene on chromosome 6 of rice. New Phytologist 163: 45-49

tag: Rice variety Bala, 1 week in hydroponics with 1ppm arsenate, whole root system


title: pf09_37

description: This is the replicate 1 of the comparsion of 1 hr vs 0 hr samples for the control without MMC treatment

data_processing: log 2 ratio of lowess normalized data

source_name: I1c: the 1 hr control sample

tag: Pseudomonas fluorescens DC454 control without MMC treatment 1 hr sample

source_name: I0: control 0 hr sample


title: Pt_6 non-acne Normal Skin


title: MCF7_dose_HRG_10.0nM_90min

status: Public on Dec 09 2006

city: Yokohama

first: Mariko

last: Okada

address: W518, 1-7-22, Suehiro-cho, Tsurumi-ku

country: Japan

institute: RIKEN RCAI

description: Early gene expression (up to 90min) induced by growth hormone stimulation.

hyb_protocol: Standard Affymetrix Protocols (Affymetrix GeneChip Expression Analysis Technical Manual) was used. Complete protocal can be found manufacturer's web site (http://www.affymetrix.com/support/technical/manual/expression_manual.affx) (section2, 2.2.p3-8 & 2.3.p3-14)

scan_protocol: GeneChips were scanned using the GeneChip Scanner 3000 with default parameter.

data_processing: The data were analyzed with GeneChip Operating System (GCOS) (ver.1.2) using Affymetrix default analysis settings and global scaling as normalization method. The Single-Array Expression Analysis function was used and the trimmed mean target intensity of each array was arbitrarily set to 500.

source_name: breast epithelial cell, adherent, HRG, 10.0nM, 90min

label_protocol: Standard Affymetrix Protocols (Affymetrix GeneChip Expression Analysis Technical Manual) was used. Complete protocal can be found manufacturer's web site (http://www.affymetrix.com/support/technical/manual/expression_manual.affx) (section2, chapter1-p13-18, & 2.1.p32-33,p36-40)

growth_protocol: Prior to the growth factor (GF) treatment, cells were serum starved for 16-18 hours with serum-free DMEM medium and incubated at 37C/5% CO2. The medium was again changed 2 hr prior to the GF treatment.

extract_protocol: Total RNA was isolated using TRIzol reagent (Invitrogen) and then purified using QIAGEN Rneasy kit.

treatment_protocol: Before treatments, medium were changed with serum-free medium again and incubated at 37C for 2 hours. Followed by incubation, added each ligand and incubated for each time.

tag: Cell line

value: MCF-7 (Obtained from ATCC (American Type Culture Collection)), Organ:mammary gland; breast, Cell type:epithelial, Disease:adenocarcinoma, Derived from metastatic site:pleural effusion, Age:69 years adult, Gender:female, Ethnicity:Caucasian


title: Rice Bala 0ppm Arsenate (Control) Rep3

description: Gene expression data from rice roots (variety Bala) grown in hydroponics for one week with 0ppm arsenate, rep3

treatment_protocol: Germinated seedlings were grown for 1 week in hydroponics with 0ppm arsenate under 12 hr of light as described in Dasgupta T, Hossain SA, Meharg AA and Price AH. (2004) An arsenate tolerance gene on chromosome 6 of rice. New Phytologist 163: 45-49

tag: Rice variety Bala, 1 week in hydroponics, whole root system


title: MCF7_dose_HRG_10.0nM_60min

source_name: breast epithelial cell, adherent, HRG, 10.0nM, 60min


title: MCF7_dose_HRG_1.0nM_15min

source_name: breast epithelial cell, adherent, HRG, 1.0nM, 15min


title: MCF7_dose_HRG_10.0nM_5min

source_name: breast epithelial cell, adherent, HRG, 10.0nM, 5min


title: MCF7_dose_HRG_1.0nM_60min

source_name: breast epithelial cell, adherent, HRG, 1.0nM, 60min


title: pf09_39

description: This is the replicate 1 of the comparison of I1mvs I0

data_processing: log2 ratio lowess normalized data

source_name: I1m:the 1 hr MMC-treated sample

tag: Pseudomonas fluorescens D454 MMC-treated 1 hr sample

source_name: I0: 0 hr control sample

tag: Pseudomonas fluorescens D454 control 0 hr sample


title: MCF7_dose_HRG_1.0nM_90min

source_name: breast epithelial cell, adherent, HRG, 1.0nM, 90min


title: MCF7_dose_HRG_10.0nM_45min

source_name: breast epithelial cell, adherent, HRG, 10.0nM, 45min


title: MCF7_dose_HRG_10.0nM_10min

source_name: breast epithelial cell, adherent, HRG, 10.0nM, 10min


title: MCF7_dose_HRG_1.0nM_30min

source_name: breast epithelial cell, adherent, HRG, 1.0nM, 30min


title: Rice Azucena 1ppm Arsenate Rep3

description: Gene expression data from rice roots (variety Azucena) grown in hydroponics for one week with 1ppm arsenate, rep3

growth_protocol: Dasgupta T, Hossain SA, Meharg AA and Price AH. (2004) An arsenate tolerance gene on chromosome 6 of rice. New Phytologist 163: 45-49

tag: Rice variety Azucena, 1 week in hydroponics with 1ppm arsenate, whole root system


title: MCF7_dose_HRG_10.0nM_30min

source_name: breast epithelial cell, adherent, HRG, 10.0nM, 30min


title: MCF7_dose_HRG_10.0nM_15min

source_name: breast epithelial cell, adherent, HRG, 10.0nM, 15min


title: MCF7_dose_HRG_1.0nM_45min

source_name: breast epithelial cell, adherent, HRG, 1.0nM, 45min


title: Rice Azucena 0ppm Arsenate (Control) Rep3

description: Gene expression data from rice roots (variety Azucena) grown in hydroponics for one week with 0ppm arsenate, rep3

source_name: Rice Hydroponic Roots 1 Week Old

tag: Rice variety Azucena, 1 week in hydroponics, whole root system


title: MCF7_dose_HRG_1.0nM_5min

source_name: breast epithelial cell, adherent, HRG, 1.0nM, 5min


title: MCF7_dose_HRG_1.0nM_10min

source_name: breast epithelial cell, adherent, HRG, 1.0nM, 10min


title: MCF7_dose_HRG_0.5nM_60min

source_name: breast epithelial cell, adherent, HRG, 0.5nM, 60min


title: MCF7_dose_HRG_0.5nM_90min

source_name: breast epithelial cell, adherent, HRG, 0.5nM, 90min


title: MCF7_dose_HRG_0.5nM_5min

source_name: breast epithelial cell, adherent, HRG, 0.5nM, 5min


title: MCF7_dose_HRG_0.1nM_90min

source_name: breast epithelial cell, adherent, HRG, 0.1nM, 90min


title: MCF7_dose_HRG_0.5nM_10min

source_name: breast epithelial cell, adherent, HRG, 0.5nM, 10min


title: MCF7_dose_HRG_0.1nM_10min

source_name: breast epithelial cell, adherent, HRG, 0.1nM, 10min


title: MCF7_dose_EGF_10.0nM_90min

source_name: breast epithelial cell, adherent, EGF, 10.0nM, 90min


title: MCF7_dose_HRG_0.5nM_15min

source_name: breast epithelial cell, adherent, HRG, 0.5nM, 15min


title: MCF7_dose_HRG_0.1nM_5min

source_name: breast epithelial cell, adherent, HRG, 0.1nM, 5min


title: MCF7_dose_HRG_0.1nM_60min

source_name: breast epithelial cell, adherent, HRG, 0.1nM, 60min


title: MCF7_dose_EGF_1.0nM_60min

source_name: breast epithelial cell, adherent, EGF, 1.0nM, 60min


title: MCF7_dose_EGF_1.0nM_90min

source_name: breast epithelial cell, adherent, EGF, 1.0nM, 90min


title: MCF7_dose_HRG_0.1nM_45min

source_name: breast epithelial cell, adherent, HRG, 0.1nM, 45min


title: MCF7_dose_EGF_10.0nM_15min

source_name: breast epithelial cell, adherent, EGF, 10.0nM, 15min


title: MCF7_dose_EGF_10.0nM_30min

source_name: breast epithelial cell, adherent, EGF, 10.0nM, 30min


title: MCF7_dose_EGF_1.0nM_45min

source_name: breast epithelial cell, adherent, EGF, 1.0nM, 45min


title: MCF7_dose_EGF_10.0nM_5min

source_name: breast epithelial cell, adherent, EGF, 10.0nM, 5min


title: MCF7_dose_HRG_0.1nM_15min

source_name: breast epithelial cell, adherent, HRG, 0.1nM, 15min


title: MCF7_dose_HRG_0.1nM_30min

source_name: breast epithelial cell, adherent, HRG, 0.1nM, 30min


title: MCF7_dose_EGF_1.0nM_30min

source_name: breast epithelial cell, adherent, EGF, 1.0nM, 30min


title: MCF7_dose_EGF_10.0nM_45min

source_name: breast epithelial cell, adherent, EGF, 10.0nM, 45min


title: MCF7_dose_EGF_1.0nM_15min

source_name: breast epithelial cell, adherent, EGF, 1.0nM, 15min


title: MCF7_dose_EGF_10.0nM_10min

source_name: breast epithelial cell, adherent, EGF, 10.0nM, 10min


title: pf09_38

description: this is the replicate 2 of the comparison of 1 hr vs 0 hr sample for the control without MMC treatment

source_name: I0: control 0 hr sample

source_name: I1c: the 1 hr control sample


title: MCF7_dose_EGF_1.0nM_5min

source_name: breast epithelial cell, adherent, EGF, 1.0nM, 5min


title: MCF7_dose_EGF_1.0nM_10min

source_name: breast epithelial cell, adherent, EGF, 1.0nM, 10min


title: MCF7_dose_HRG_0.5nM_45min

source_name: breast epithelial cell, adherent, HRG, 0.5nM, 45min


title: MCF7_dose_EGF_0.5nM_45min

source_name: breast epithelial cell, adherent, EGF, 0.5nM, 45min


title: MCF7_dose_EGF_0.5nM_90min

source_name: breast epithelial cell, adherent, EGF, 0.5nM, 90min


title: MCF7_dose_EGF_0.5nM_30min

source_name: breast epithelial cell, adherent, EGF, 0.5nM, 30min


title: MCF7_dose_HRG_0.5nM_30min

source_name: breast epithelial cell, adherent, HRG, 0.5nM, 30min


title: MCF7_dose_EGF_0.5nM_5min

source_name: breast epithelial cell, adherent, EGF, 0.5nM, 5min


title: MCF7_dose_control_for_HRG

source_name: breast epithelial cell, adherent, without stimulus, control for HRG


title: MCF7_dose_EGF_0.5nM_60min

source_name: breast epithelial cell, adherent, EGF, 0.5nM, 60min


title: MCF7_dose_EGF_0.1nM_5min

source_name: breast epithelial cell, adherent, EGF, 0.1nM, 5min


title: MCF7_dose_EGF_0.1nM_45min

source_name: breast epithelial cell, adherent, EGF, 0.1nM, 45min


title: MCF7_dose_EGF_0.5nM_15min

source_name: breast epithelial cell, adherent, EGF, 0.5nM, 15min


title: MCF7_dose_EGF_0.1nM_10min

source_name: breast epithelial cell, adherent, EGF, 0.1nM, 10min


title: MCF7_dose_EGF_0.1nM_60min

source_name: breast epithelial cell, adherent, EGF, 0.1nM, 60min


title: MCF7_dose_EGF_0.1nM_90min

source_name: breast epithelial cell, adherent, EGF, 0.1nM, 90min


title: MCF7_dose_EGF_0.5nM_10min

source_name: breast epithelial cell, adherent, EGF, 0.5nM, 10min


title: MCF7_dose_control_for_EGF

source_name: breast epithelial cell, adherent, without stimulus, control for EGF


title: Diffuse large B cell lymphoma VUMC_L082_b33_s12

status: Public on Dec 08 2006

city: Amsterdam

first: Daoud

last: Sie

address: De Boelelaan 1117

country: Netherlands

institute: Vrije Universiteit Medical Center

description: see also Muris et al. submitted

hyb_protocol: Hybridization protocol available via www.vumc.nl/microarrays Also described Muris et al. submitted

scan_protocol: Microarray Scanner G2505B (Agilent Technologies), default settings

data_processing: Image analysis and feature extraction was done with BlueFuse for Microarrays (version 2.0 (3276), BlueGnome Limited, Cambridge, United Kingdom). Fused values of in duplicate spotted oligos were taken and intensities were divided by the mean intensity of each channel. Next, data were inter array centered by dividing the ratios for each gene across the sample set by the mean observed value. The resulting ratios were log-transformed (base 2) and processed into Stanford file format (rows, mean values for the in duplicate measured oligos; columns, mRNA samples). The resulting dataset contained expression ratios of 14487 genes.

source_name: universal human reference RNA

label_protocol:
labeling protocol available via www.vumc.nl/microarrays
Also described in Muris et al. submitted

extract_protocol: see www.stratagene.com

tag: comprised of 10 cell lines for human giving broad gene coverage

source_name: lymph node

label_protocol:
labeling protocol available via www.vumc.nl/microarrays
also described in Muris et al. submitted

extract_protocol:
RNA isolation protocol available via www.vumc.nl/microarrays
also described in Muris et al. submitted

tag: diffuse large B cell lymphoma

tag: described in Muris et al. submitted


title: MCF7_dose_EGF_0.1nM_30min

source_name: breast epithelial cell, adherent, EGF, 0.1nM, 30min


title: MCF7_dose_EGF_0.1nM_15min

source_name: breast epithelial cell, adherent, EGF, 0.1nM, 15min


title: Diffuse large B cell lymphoma VUMC_L074_b33_s55

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L079_b33_s57

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L073_b33_s54

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L071_b33_s53

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L070_b33_s52

description: see also Muris et al. submitted


title: Hyperplastic lymphoid tissue VUMC_L090_b33_s14

description: see also Muris et al. submitted

source_name: tonsil

tag: hyperplastic lymphoid tissue


title: Diffuse large B cell lymphoma VUMC_L060_b33_s24

description: see also Muris et al. submitted


title: Hyperplastic lymphoid tissue VUMC_L091_b33_s50

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L077_b33_s56

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L093_b33_s59

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L088a_b33_s58

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L088b_b33_s60

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L086_b33_s13

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L058_b33_s49

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L057_b33_s37

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L050_b33_s47

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L059_b33_s11

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L051_b33_s35

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L053_b33_s10

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L056_b33_s48

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L054_b33_s23

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L052_b33_s36

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L043_b33_s34

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L041_b33_s8

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L044_b33_s46

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma  VUMC_L046_b33_s9

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L042_b33_s45

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L030_b33_s44

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L029_b33_s33

description: see also Muris et al. submitted

tag: Diffuse large B cell lymphoma


title: Diffuse large B cell lymphoma VUMC_L023a_b33_s43

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L020_b33_s20

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L010b_b33_s6

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L018_b33_s5

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L017_b33_s42

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L023b_b33_s7

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L048_b33_s22

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L013b_b33_s19

description: see also Muris et al. submitted


title: Klf9WT_jejunum_PND30_3

city: Little Rock

first: Rijin

last: Xiao

state: AR

address: 2020 Marshall Street

institute: Nutrition Center, Arkansas Chidren's Hospital

description: Bacterial sequence-derived probes on the arrays served as external controls for hybridization, whereas the housekeeping genes β-actin and GAPDH served as endogenous controls and for monitoring the quality of the RNA target.

hyb_protocol: 10 ug of cRNA was hybridized for 16 hours to an Affymetrix (Santa Clara, CA) mouse 430A GeneChip (5 chips per genotype group), followed by incubations with streptavidin-conjugated phycoerythrin, and then with polyclonal anti-streptavidin antibody coupled to phycoerythrin.

scan_protocol: GeneChips were scanned using an Agilent GeneArray laser scanner. Images were analyzed using Affymetrix MAS 5.0 software.

data_processing: To compare array data between GeneChips, we scaled the average of the fluorescent intensities of all probes on each array to a constant target intensity of 500

label: Biotin

source_name: Mouse jejunum, PND30

label_protocol: 8 ug total RNA was used for cDNA synthesis using a T7-(deoxythymidine)24 primer and Superscript II (Life Technologies, Inc., Gaithersburg, MD). The resulting cDNA was used with the ENZO BioArray High Yield RNA Transcript labeling kit (ENZO, Farmingdale, NY) to synthesize biotin-labeled cRNA. The cRNA was purified on RNeasy spin columns (QIAGEN) and subjected to chemical fragmentation (size range of 35 to 200 bp). Three replicate cRNA targets were made in parallel starting from each RNA pool.

extract_protocol: TriTRIzol extraction procedure, followed by a cleanup using QIAGEN RNeasy Mini kit.

tag: Klf9 wild type (Klf9WT) mice jejunum; Age

value: PND 30


title: Klf9WT_jejunum_PND30_2


title: Diffuse large B cell lymphoma VUMC_L021_b33_s31

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L024_b33_s51

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L010a_b33_s40

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L026_b33_s32

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L014a_b33_s18

description: see also Muris et al. submitted


title: Klf9WT_jejunum_PND30_4


title: Klf9WT_jejunum_PND30_5


title: Diffuse large B cell lymphoma VUMC_L014b_b33_s30

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L025_b33_s21

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L013a_b33_s41

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L09_b33_s29

description: see also Muris et al. submitted


title: Klf9KO_jejunum_PND30_4

tag: Klf9 mutant (Klf9KO) mice jejunum; Age


title: Klf9WT_jejunum_PND30_1


title: Diffuse large B cell lymphoma VUMC_L08_b33_s17

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L05_b33_s16

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L07_b33_s39

description: see also Muris et al. submitted


title: Klf9KO_jejunum_PND30_2

source_name: Mouse jejunum PND30

tag: Klf9 mutant (Klf9KO) mouse jejunum; Age


title: Diffuse large B cell lymphoma VUMC_L04_b33_s28

description: see also Muris et al. submitted


title: Klf9KO_jejunum_PND30_5


title: Klf9KO_jejunum_PND30_3


title: Diffuse large B cell lymphoma  VUMC_L02_b33_s15

description: see also Muris et al. submitted


title: Diffuse large B cell lymphoma VUMC_L03_b33_s27

description: see also Muris et al. submitted


title: Klf9KO_jejunum_PND30_1

tag: Klf9 wild type (Klf9WT) and mutant (Klf9KO) mice jejunum; Age


title: Diffuse large B cell lymphoma VUMC_L01_b33_s3

description: see also Muris et al. submitted


title: MF9

status: Public on Dec 06 2006

city: Vancouver

first: Kendy

last: Wong

state: British Columbia

address: 675 West 10th Avenue

department: Cancer Genetics

country: Canada

institute: BC Cancer Research Center

description: single male reference genomic DNA competitively hybridized against single female genomic DNA (control experiment)

data_processing: three-step normalization procedure; (Khojasteh et al BMC Bioinformatics 2005;6:274)

label: Cyanine 3

source_name: Mref

extract_protocol: QIAamp DNA Blood Maxi Kit (QIAGEN Inc.)

tag: single male reference genomic DNA

label: cyanine 5

source_name: Fgenomic

tag: single female genomic DNA


title: M_Mpool

description: single male genomic DNA hybridized against male pooled genomic DNA

source_name: male pooled genomic

tag: male pooled genomic


title: M_Fpool

description: single male genomic DNA hybridized against female pooled genomic DNA

source_name: female pooled genomic

tag: female pooled genomic DNA


title: F_Fpool

description: single female genomic DNA hybridized against female pooled genomic DNA


title: self_hyb_1

description: self versus self hybridization


title: MF10


title: self_hyb_2

description: self versus self hybridization


title: self_hyb_6

description: self versus self hybridization


title: self_hyb_5

description: self versus self hybridization


title: MF7


title: F_Mpool

description: single female genomic DNA hybridized against male pooled genomic DNA

tag: male pooled genomic DNA


title: self_hyb_4

description: self versus self hybridization


title: MF8


title: MF6


title: MF2


title: S41_dyeswap

description:
cyanine 3 = reference
cyanine 5 = sample

tag: B-lymphocyte cell line DNA


title: S14_dyeswap


title: MF3


title: MF4


title: S38_dyeswap


title: self_hyb_3

description: self versus self hybridization


title: MF5


title: MF1


title: S44_dyeswap


title: S11_dyeswap


title: S33_dyeswap

tag: single male genomic DNA


title: S39_dyeswap


title: S7_dyeswap


title: S16_dyeswap


title: S8_dyeswap


title: S17_dyeswap


title: F1_dyeswap

tag: B-lymphocyte cell line DNA; paternal grandfather of pedigree XC01423


title: S12_dyeswap


title: F3_dyeswap

tag: B-lymphocyte cell line DNA; maternal grandfather of pedigree XC01423


title: F2_dyeswap

tag: B-lymphocyte cell line DNA; paternal grandmother of pedigree XC01423


title: S10_dyeswap


title: S3_dyeswap


title: S4_dyeswap

source_name: MRef


title: S1_dyeswap


title: S91

description:
cyanine 3 = sample
cyanine 5 = reference

tag: anonymous single female genomic DNA from blood

label: Cyanine 5


title: S84


title: S83


title: S81


title: S67


title: S40_dyeswap


title: S90


title: S87


title: S86


title: S89


title: S60


title: S65


title: S88


title: S82


title: S66


title: S85


title: S61


title: S63


title: S62


title: S76


title: S58


title: S59


title: S57


title: S54


title: S75


title: S55


title: S64


title: S53


title: S56


title: S50


title: S52


title: S51


title: S47


title: S46


title: S27

tag: anonymous single male genomic DNA from blood


title: S32


title: S25


title: S28


title: S30


title: S26


title: S48


title: S49


title: S31


title: S22


title: S24


title: S45


title: S29


title: S21


title: S20


title: S19


title: S80


title: S74


title: S18


title: S79


title: S73


title: S23


title: S72


title: S78


title: S71


title: S33


title: S36


title: S42


title: S70


title: Finch 3b; green: whole brain, red: HVC

city: New York

first: XiaoChing

last: Li

state: NY

address: 1230 York Avenue

institute: Rockefeller University

hyb_protocol: 42 degrees overnight, Corning hybridization buffer

scan_protocol: 647 nm and 555 nm

data_processing: log2 of fold change (comparison of background-subtracted experimental intensity and LOWESS-normalized reference intensity)

organism: Taeniopygia guttata

source_name: green: whole brain

label_protocol: cDNA probe by reverse transcription

extract_protocol: Qiagen RNeasy Micro Kit (Cat. No. 74004)

tag: adult male zebra finch

source_name: red: HVC


title: S41


title: S39


title: S34


title: S40


title: Finch 3a; green: whole brain, red: HVC


title: S35


title: S38


title: S43


title: S44


title: S68


title: S17


title: S37


title: S77


title: S16


title: Finch 3b; green: HVC, red: whole brain

source_name: green: HVC

source_name: red: whole brain


title: S14


title: S15


title: Finch 2b; green: HVC, red: whole brain


title: Finch 2a; green: whole brain, red: HVC


title: Finch 2a; green: HVC, red: whole brain


title: S11


title: S13


title: Finch 1b; green: whole brain, red: HVC


title: Finch 1b; green: HVC, red: whole brain


title: S12


title: Finch 2b; green: brain, red HVC

source_name: green: brain


title: Finch 1a; green: whole brain, red: HVC


title: FOS vs. FOS+glucose Rep#2

city: Lincoln

first: Yong Jun

last: Goh

state: NE

address: 332 Food Industry Complex

department: Food Science and Technology

institute: University of Nebraska

description: Cells (2% inoculum from overnight culture in MRS broth) were grown in 100 ml of modified MRS supplemented with 2% FOS until an OD625nm 0.6. The culture was split into two 50-ml portions, and glucose was added to one of the portions at a 2% final concentration. Both cultures were grown for another 60 min (OD625nm ca. 1.0) before harvested for total RNA isolation.

data_processing: Hybridized arrays were scanned with ScanArray 5000 (Packard Instrument Co., Downers Grove, IL) at 10 micrometer per pixel resolution. The relative fluorescent intensity of Cy3 and Cy5 for each of the 7,680 features was quantified from TIFF image files (generated by ScanArray 5000) using ImaGene v.4.2 (BioDiscovery, Inc., Marina Del Rey, CA). The fluorescent signal intensities of the array spots for each channel were obtained by subtracting the median intensities of background pixels from the mean pixel intensities. Global mean normalization of the spot intensities from each channel on individual slides was performed using SNOMAD. All array data were composited using a Perl-based program, FormatALL (J. Wise and A. K. Benson, unpublished data), yielding the ratio representing the fold change in genes expression levels in FOS conditions (Cy5) with relative to FOS + glucose (Cy3).

organism: Lacticaseibacillus paracasei

source_name: FOS only

tag: strain 1195

source_name: FOS+glucose


title: Finch 3a; green: HVC, red: whole brain


title: S9


title: S10


title: Finch 1a; green: HVC, red: whole brain


title: FOS vs. FOS+glucose Rep#1

description: Cells (2% inoculum from overnight culture in MRS broth) were grown in 100 ml of modified MRS supplemented with 2% FOS until an OD625nm 0.6.  The culture was split into two 50-ml portions, and glucose was added to one of the portions at a 2% final concentration.  Both cultures were grown for another 60 min (OD625 ca. 1.0) before harvested for total RNA isolation.


title: S2


title: S6


title: S3


title: FOS vs. Fructose Rep#2

description: L. paracasei (2% [v/v] inoculum from overnight culture in MRS broth) were grown in 300 ml of modified MRS basal medium (with no added sugar source) until the cells reached a final OD625 nm of ca. 0.3. The culture was then divided into two 150-ml portions, where FOS or fructose was added to a 1% final concentration. Cells were collected for total RNA isolation after 30 min incubation (OD625 nm 0.3 – 0.4).

source_name: 1% FOS, 30 min

source_name: 1% fructose, 30 min


title: Canary 3b; green: whole brain, red: HVC

organism: Serinus canaria

tag: adult male canary


title: S4


title: FOS vs. Glucose Rep#1

description: L. paracasei (2% [v/v] inoculum from overnight culture in MRS broth) were grown in 300 ml of modified MRS basal medium (with no added sugar source) until the cells reached a final OD625 nm of ca. 0.3.  The culture was then divided into two 150-ml portions, where FOS or glucose was added to a 1% final concentration.  Cells were collected for total RNA isolation after 30 min incubation (OD625 nm 0.3 – 0.4).

source_name: 1% glucose, 30 min


title: FOS vs. Glucose Rep#2

description: L. paracasei (2% [v/v] inoculum from overnight culture in MRS broth) were grown in 300 ml of modified MRS basal medium (with no added sugar source) until the cells reached a final OD625 nm of ca. 0.3. The culture was then divided into two 150-ml portions, where FOS or glucose was added to a 1% final concentration. Cells were collected for total RNA isolation after 30 min incubation (OD625 nm 0.3 – 0.4).


title: S7


title: S5


title: slide25c-Milk-2970-3028-wtmallarddec03

city: Guelph

first: Wenjing

last: Tao

state: ON

address: Gordon street

department: Department of Pathobiology

institute: University of Guelph

description: Feature symbols and assigned values: Bad spot -100 X Absent -75 (absent spots are assigned based on gene list) Not found -50 | Not flagged 0 Good spot 100

hyb_protocol: Prehybridization: Combine 80 μl DIG Easy Hyb, 4 μl of blocking RNA/DNA, 65ºC for 2 min. Let the solution cool for 2 min at room temperature. Pipette 30-50μl prehyb buffer onto the chip, place the array chip into a slide box containing a small amount of DIG Easy Hyb, wrap the slide box with plastic wrap and place into a incubator, incubate at 42ºC for 45 min 2. Wash slides in room temperature DEPC-H2O, shaker for 10 min 3. Dip slides in room temperature isopropanol and spin dry in centrifuge for 5 min slides should be used immediately and should not dry for more than 1 hr (6) Hybridization: 1. Reused each labeled probe in 5μl of 2xHybridization buffer (make fresh) 2. Combine 5μl of each of purified Cy3- and Cy5-labeled probes, mix well 3. Add 4μl COT1-DNA (1μg/μl) 4. Add 5μl Poly(A)-DNA (5μg/μl) 5. Add 1μl tRNA (10μg/μl) 6. Heat probe mixture at 95ºC for 3 min to denature 7. Centrifuge probe for 1 min at maximum speed 8. Apply labeled probes to prehybridized microarray slide and coverslip (22 X 22 mm) 9. Place sealed slide box in a 42ºC incubator, and incubate for 16-20 hr (7) Washing: Prepare some pre-warmed (50ºC) 1X SSC + 0.1% SDS, also prepare some room temperature 1X SSC 1. Remove array from box and dip it a few times into 1X SSC. Once the cover slip has been removed, place the chip in the side holder and put the holder into the dish with 1X SSC + 0.1% SDS, leave the chip in 1X SSC + 0.1% SDS for 15 minutes, with a quick shake at 7.5 minutes. When 15 minutes is up, move the chip to a new dish of 1X SSC + 0.1% SDS, leave 15 minutes, with quick shake at 7.5 minutes. Repeat same wash in another new dish. 2. Wash slide in 1X SSC at room temperature to remove traces of SDS. 3. If necessary, do an additional wash in room temperature 0.1XSSC if you find that salt is precipitating on the chip. 4. Place slide into a clean slide box that has filter paper lining the bottom. Spin the slide box at 500 rpm for 5 min. 5. Scan slide if possible the same day

scan_protocol: Turn on the scanner and open the GenePix Pro software. Warm up the scanner for 15 min before image acquisition. Slide the scanner door open. Gently lift the tab on the front of slide holder to open the lid. Pull open the adjustment arm (lever) on the left of the slide holder with one finger, and insert the slide array side down (label out, face down) into the slide holder. Flick the lever slightly to ensure proper positioning of the slide. Gently close the slide holder lid until it snaps shut, and close the door. Click on the hardware button on the lower right, set the PMT ro 600V (default) for both channels (635 for Cy5 and Alexa 647, and 532 for Cy3 and Alexa 546). Perform a low resolution scan of the slide using the Preview Scan button (fast forward). Use the Histogram tab on the top to adjust the minimum intensity to 1000 and the maximum intensity to 65000. This is to eliminate background as most background is at an intensity of around 300-500. Observe the relative intensities of both channels (this is the ratio of the dyes, and making sure that the Log Axis box is checked). If necessary, adjust the PMT settings on the fly to achieve a color balance (ratio ~1.0). We routinely use a higher PMT setting for Cy5 (e.g., 670) than Cy3 (e.g., 600). Define the array location for high resolution scan using the View Scan Area button on the left. The box can be re-positioned and re-sized by clicking-and-dragging with the mouse. Click the Data Scan button to execute a high resolution scan of the specific area. Use the Histogram tab to monitor the intensities of both channels. If they do not overlap, click the Stop button and readjust the PMT settings. To save the image, click the File button and select Save Images. Save images as Multi-image TIFF Files and name your file. GenePix automatically adds the wavelength of the image as a suffix to the file name. Click the histogram tab, check the ratio. We want it to be around 1.0. Once the ratio of image is near 1.0, then select the elect button. After the scanner has stopped making noise, open the scanner and remove the array slide. Image Analysis After finish all scans, turn off the scanner and power supply. Close the current program. Click on the GenePix pro demo icon. Click the File… button and select Open Images to open previously saved images. Navigate to the directory that contains the previously saved TIFF images. Hold down the <Ctrl> key to select a pair of images from the wavelengths 635 and 532. Image display: Brightness/Contrast; Zoom Mode; Undo Zoom; Full Scale, etc Load the GAL (GenePix Array List) file containing gene list information by clicking the File button, and select the Load Array List command. Select the gal file and click open. This procedure automatically constructs blocks that match the number, size and positioning of printed features and assigns identifiers to each spot. Use the arrow keys to move the array grids to align with the spots as well as possible (no need to be perfect). Press F5 key to Find Features automatically. Right click to change between Block mode or Feature mode to fine-tune the alignment using the arrow keys (block-by-block). Press F5 to re-match the features. After the alignment is done, examine each grid. Using the Feature mode, flag spots that are bad (e.g., contaminated) or not found (e.g., weak) and make necessary adjustments. Select the spot and right click to apply the desired function (Good, Bad or Not found). To re-set feature positions, right click and select Clear Nudges. Flags can be re-set thesame way. Similar spots can be handled simultaneously by dragging a region around them. Select the “analyze” button, the columns of data should appear. Select the save as button and save the file. This will generate a gpr file (one per image) that can be used in the future (e.g., no need to redo the alignment), and 3 jpg (653, 532 and ratio) files.

data_processing: Genepix

organism: Bos taurus

source_name: healthy control

label_protocol: Indirect-labelling with cy3: (1) RT Reaction: After Trizol extraction and RNeasy purification, RNA is resuspended in H2O. After taking samples out from –80°C freezer, heat RNA samples to 50-65ºC (water bath) for 10 min and cool to RT for 10 min before starting. Bring water bath down to 42°C. Take 10μg total RNA and add DEPC H2O to get final amount of 14μl Add AncT primer (3μg/μl) 2μl Total volume 16μl Incubate at 70°C (heat block) for 10 min and chill on ice for 5 min Add: 5X First Strand Buffer 6μl 0.1M DTT 3μl 5X aa-dNTP mix 2.0μl SSII Reverse Transcriptase 1.5μl Rnase OUT 1.5μl Total volume 30μl Incubate at 42°C (water bath) for 3 hr. (Can be longer, e.g. over night) (2) Labeling Reaction: 1. to each labeling reaction add of 500mM EDTA (pH8.0) 10μl 2. vortex 3. add of 1M NaOH 10μl 4. vortex and spin briefly 5. incubate at 65°C (heat block) for 20 min to hydrolyze RNA, then cool to R/T 6. spin briefly 7. to neutralize, add 1M HCL 10μl vortex and spin briefly 8. perform probe purification with Qiagen PCR spin columns: a. add 300μl PB buffer to reaction, mix with pipette, and transfer to QIAquick column b. spin at maximum speed for 1 min, discard flow through c. wash with 740μl phosphate wash buffer (home-made buffer, NOT from kit) d. spin at maximum speed for 1 min e. empty collection tube and repeat wash steps c and d f. empty collection tube and spin at maximum speed for 1 min g. transfer column to a fresh tube (cut off caps), elute with 30μl elution buffer (home-made KPO4, NOT from kit) with 1 min incubation followed by spin at maximum speed for 1 min h. repeat elution step g, transfer to new tube i. speedvac dry (~ 40 minutes) (heater on middle) 9. Cy dye coupling a. resuspend the cDNA in 4.5μl 0.1M carbonate buffer, pH9.0 (< 1 month old) b. add 4.5μl NHS-Cy dyes c. incubate the reactions in dark for 2 hours at room temperature make prehyb. buffer and warm to 42°C Set heat block to 95°C (Alexa dye labeling: dissolve an aliquot Alexa 546 or 647 in 9 ul DMSO, pipette 5 ul Alexa dye to each cDNA in 4.5 ul of 0.1M carbonate buffer, pH9.0. Incubate both samples for 1-2 hours at room temperature in dark) (3) Purification: 1. Add 35μl 100mM NaOAc pH 5.2 to the reaction 2. Add 250μl PB buffer to reaction and follow QIAquick PCR purification protocol: a. Apply reaction volume to QIAquick spin column and centrifuge for 1 minute b. Discard flow-through, should be colored c. Add 0.74ml buffer PE (as provided with kit) to column and centrifuge for 1 minute d. Discard flow-through and centrifuge for an additional 1-minute at maximum speed e. Place QIAquick column in a clean 1.5ml microcentrifuge tube f. Elute twice with 30μl EB using 1-minute incubation time before centrifugation (eluted sample should look colored and filter should be white) g. Take 1μl of sample for dye incorporation analysis h. Speedvac dry sample (~ 40 minutes) (heat setting to middle) Alternative: precipitate DNA with 1/10 volume 3M sodium acetate, 1/100 volume glycogen, final conc. of 65% ethanol. (4) Dye Incorporation Analysis: 1. Take 1μl of sample for dye incorporation analysis 2. Measure absorbance at 260nm, 550nm (Cy3), for Cy3 incorporation: nucleotide/dye = 17.1*OD260/OD550 150pmole/slide is optimal for hybridization; nucleotide/dye should be < 50

extract_protocol: TRIzol reagent

tag: no evidence of S. aureus mastitis

source_name: mastitis

label_protocol: Indirect-labelled with cy5: (1) RT Reaction: After Trizol extraction and RNeasy purification, RNA is resuspended in H2O. After taking samples out from –80°C freezer, heat RNA samples to 50-65ºC (water bath) for 10 min and cool to RT for 10 min before starting. Bring water bath down to 42°C. Take 10μg total RNA and add DEPC H2O to get final amount of 14μl Add AncT primer (3μg/μl) 2μl Total volume 16μl Incubate at 70°C (heat block) for 10 min and chill on ice for 5 min Add: 5X First Strand Buffer 6μl 0.1M DTT 3μl 5X aa-dNTP mix 2.0μl SSII Reverse Transcriptase 1.5μl Rnase OUT 1.5μl Total volume 30μl Incubate at 42°C (water bath) for 3 hr. (Can be longer, e.g. over night) (2) Labeling Reaction: 1. to each labeling reaction add of 500mM EDTA (pH8.0) 10μl 2. vortex 3. add of 1M NaOH 10μl 4. vortex and spin briefly 5. incubate at 65°C (heat block) for 20 min to hydrolyze RNA, then cool to R/T 6. spin briefly 7. to neutralize, add 1M HCL 10μl vortex and spin briefly 8. perform probe purification with Qiagen PCR spin columns: a. add 300μl PB buffer to reaction, mix with pipette, and transfer to QIAquick column b. spin at maximum speed for 1 min, discard flow through c. wash with 740μl phosphate wash buffer (home-made buffer, NOT from kit) d. spin at maximum speed for 1 min e. empty collection tube and repeat wash steps c and d f. empty collection tube and spin at maximum speed for 1 min g. transfer column to a fresh tube (cut off caps), elute with 30μl elution buffer (home-made KPO4, NOT from kit) with 1 min incubation followed by spin at maximum speed for 1 min h. repeat elution step g, transfer to new tube i. speedvac dry (~ 40 minutes) (heater on middle) 9. Cy dye coupling a. resuspend the cDNA in 4.5μl 0.1M carbonate buffer, pH9.0 (< 1 month old) b. add 4.5μl NHS-Cy dyes c. incubate the reactions in dark for 2 hours at room temperature make prehyb. buffer and warm to 42°C Set heat block to 95°C (Alexa dye labeling: dissolve an aliquot Alexa 546 or 647 in 9 ul DMSO, pipette 5 ul Alexa dye to each cDNA in 4.5 ul of 0.1M carbonate buffer, pH9.0. Incubate both samples for 1-2 hours at room temperature in dark) (3) Purification: 1. Add 35μl 100mM NaOAc pH 5.2 to the reaction 2. Add 250μl PB buffer to reaction and follow QIAquick PCR purification protocol: a. Apply reaction volume to QIAquick spin column and centrifuge for 1 minute b. Discard flow-through, should be colored c. Add 0.74ml buffer PE (as provided with kit) to column and centrifuge for 1 minute d. Discard flow-through and centrifuge for an additional 1-minute at maximum speed e. Place QIAquick column in a clean 1.5ml microcentrifuge tube f. Elute twice with 30μl EB using 1-minute incubation time before centrifugation (eluted sample should look colored and filter should be white) g. Take 1μl of sample for dye incorporation analysis h. Speedvac dry sample (~ 40 minutes) (heat setting to middle) Alternative: precipitate DNA with 1/10 volume 3M sodium acetate, 1/100 volume glycogen, final conc. of 65% ethanol. (4) Dye Incorporation Analysis: 1. Take 1μl of sample for dye incorporation analysis 2. Measure absorbance at 260nm, 650nm (Cy5) for Cy5 incorporation: nucleotide/dye = 28.5*OD260/OD650 150pmole/slide is optimal for hybridization; nucleotide/dye should be < 50

tag: Staph aureus mastitis


title: slide26b-Milk-2875-2869-wtAug10


title: Canary 3a; green: whole brain, red: HVC


title: FOS vs. Fructose Rep#1


title: S8

source_name: Mref

tag: single male reference genomic DNA


title: slide31-Milk-2834-2883-wtmallarddec03

source_name: Healthy control


title: slide30-Milk-2929-2914-wtmallarddec03


title: slide26a-Milk-2970-3028-wtmallarddec03


title: Canary 3b; green: HVC, red: whole brain


title: Canary 2b; green: whole brain; red: HVC


title: S1


title: slide28-Milk-2848-2916-wtmallarddec03


title: slide23b-Milk-3011-3072-wtmallarddec03


title: S69

tag: single female genomic DNA


title: F11

tag: B-lymphocyte cell line DNA, a son of pedigree XC01423


title: Canary 2a; green: whole brain, red: HVC


title: F12

tag: B-lymphocyte cell line DNA, a daughter of pedigree XC01423


title: slide44-milk-2916-2848-wtAug10

hyb_protocol: (5) Prehybridization: Combine 80 μl DIG Easy Hyb, 4 μl of blocking RNA/DNA, 65ºC for 2 min. Let the solution cool for 2 min at room temperature. Pipette 30-50μl prehyb buffer onto the chip, place the array chip into a slide box containing a small amount of DIG Easy Hyb, wrap the slide box with plastic wrap and place into a incubator, incubate at 42ºC for 45 min 2. Wash slides in room temperature DEPC-H2O, shaker for 10 min 3. Dip slides in room temperature isopropanol and spin dry in centrifuge for 5 min slides should be used immediately and should not dry for more than 1 hr (6) Hybridization: 1. Reused each labeled probe in 5μl of 2xHybridization buffer (make fresh) 2. Combine 5μl of each of purified Cy3- and Cy5-labeled probes, mix well 3. Add 4μl COT1-DNA (1μg/μl) 4. Add 5μl Poly(A)-DNA (5μg/μl) 5. Add 1μl tRNA (10μg/μl) 6. Heat probe mixture at 95ºC for 3 min to denature 7. Centrifuge probe for 1 min at maximum speed 8. Apply labeled probes to prehybridized microarray slide and coverslip (22 X 22 mm) 9. Place sealed slide box in a 42ºC incubator, and incubate for 16-20 hr (7) Washing: Prepare some pre-warmed (50ºC) 1X SSC + 0.1% SDS, also prepare some room temperature 1X SSC 1. Remove array from box and dip it a few times into 1X SSC. Once the cover slip has been removed, place the chip in the side holder and put the holder into the dish with 1X SSC + 0.1% SDS, leave the chip in 1X SSC + 0.1% SDS for 15 minutes, with a quick shake at 7.5 minutes. When 15 minutes is up, move the chip to a new dish of 1X SSC + 0.1% SDS, leave 15 minutes, with quick shake at 7.5 minutes. Repeat same wash in another new dish. 2. Wash slide in 1X SSC at room temperature to remove traces of SDS. 3. If necessary, do an additional wash in room temperature 0.1XSSC if you find that salt is precipitating on the chip. 4. Place slide into a clean slide box that has filter paper lining the bottom. Spin the slide box at 500 rpm for 5 min. 5. Scan slide if possible the same day

label_protocol: Indirect-labelling with cy3: (1) RT Reaction: After Trizol extraction and RNeasy purification, RNA is resuspended in H2O. After taking samples out from –80°C freezer, heat RNA samples to 50-65ºC (water bath) for 10 min and cool to RT for 10 min before starting. Bring water bath down to 42°C. Take 10μg total RNA and add DEPC H2O to get final amount of 14μl Add AncT primer (3μg/μl) 2μl Total volume 16μl Incubate at 70°C (heat block) for 10 min and chill on ice for 5 min Add: 5X First Strand Buffer 6μl 0.1M DTT 3μl 5X aa-dNTP mix 2.0μl SSII Reverse Transcriptase 1.5μl Rnase OUT 1.5μl Total volume 30μl Incubate at 42°C (water bath) for 3 hr. (Can be longer, e.g. over night) (2) Labeling Reaction: 1. to each labeling reaction add of 500mM EDTA (pH8.0) 10μl 2. vortex 3. add of 1M NaOH 10μl 4. vortex and spin briefly 5. incubate at 65°C (heat block) for 20 min to hydrolyze RNA, then cool to R/T 6. spin briefly 7. to neutralize, add 1M HCL 10μl vortex and spin briefly 8. perform probe purification with Qiagen PCR spin columns: a. add 300μl PB buffer to reaction, mix with pipette, and transfer to QIAquick column b. spin at maximum speed for 1 min, discard flow through c. wash with 740μl phosphate wash buffer (home-made buffer, NOT from kit) d. spin at maximum speed for 1 min e. empty collection tube and repeat wash steps c and d f. empty collection tube and spin at maximum speed for 1 min g. transfer column to a fresh tube (cut off caps), elute with 30μl elution buffer (home-made KPO4, NOT from kit) with 1 min incubation followed by spin at maximum speed for 1 min h. repeat elution step g, transfer to new tube i. speedvac dry (~ 40 minutes) (heater on middle) 9. Cy dye coupling a. resuspend the cDNA in 4.5μl 0.1M carbonate buffer, pH9.0 (< 1 month old) b. add 4.5μl NHS-Cy dyes c. incubate thereactions in dark for 2 hours at room temperature make prehyb. buffer and warm to 42°C Set heat block to 95°C (Alexa dye labeling: dissolve an aliquot Alexa 546 or 647 in 9 ul DMSO, pipette 5 ul Alexa dye to each cDNA in 4.5 ul of 0.1M carbonate buffer, pH9.0. Incubate both samples for 1-2 hours at room temperature in dark) (3) Purification: 1. Add 35μl 100mM NaOAc pH 5.2 to the reaction 2. Add 250μl PB buffer to reaction and follow QIAquick PCR purification protocol: a. Apply reaction volume to QIAquick spin column and centrifuge for 1 minute b. Discard flow-through, should be colored c. Add 0.74ml buffer PE (as provided with kit) to column and centrifuge for 1 minute d. Discard flow-through and centrifuge for an additional 1-minute at maximum speed e. Place QIAquick column in a clean 1.5ml microcentrifuge tube f. Elute twice with 30μl EB using 1-minute incubation time before centrifugation (eluted sample should look colored and filter should be white) g. Take 1μl of sample for dye incorporation analysis h. Speedvac dry sample (~ 40 minutes) (heat setting to middle) Alternative: precipitate DNA with 1/10 volume 3M sodium acetate, 1/100 volume glycogen, final conc. of 65% ethanol. (4) Dye Incorporation Analysis: 1. Take 1μl of sample for dye incorporation analysis 2. Measure absorbance at 260nm, 550nm (Cy3), for Cy3 incorporation: nucleotide/dye = 17.1*OD260/OD550 150pmole/slide is optimal for hybridization; nucleotide/dye should be < 50

label_protocol: indirect-labelled with cy5: (1) RT Reaction: After Trizol extraction and RNeasy purification, RNA is resuspended in H2O. After taking samples out from –80°C freezer, heat RNA samples to 50-65ºC (water bath) for 10 min and cool to RT for 10 min before starting. Bring water bath down to 42°C. Take 10μg total RNA and add DEPC H2O to get final amount of 14μl Add AncT primer (3μg/μl) 2μl Total volume 16μl Incubate at 70°C (heat block) for 10 min and chill on ice for 5 min Add: 5X First Strand Buffer 6μl 0.1M DTT 3μl 5X aa-dNTP mix 2.0μl SSII Reverse Transcriptase 1.5μl Rnase OUT 1.5μl Total volume 30μl Incubate at 42°C (water bath) for 3 hr. (Can be longer, e.g. over night) (2) Labeling Reaction: 1. to each labeling reaction add of 500mM EDTA (pH8.0) 10μl 2. vortex 3. add of 1M NaOH 10μl 4. vortex and spin briefly 5. incubate at 65°C (heat block) for 20 min to hydrolyze RNA, then cool to R/T 6. spin briefly 7. to neutralize, add 1M HCL 10μl vortex and spin briefly 8. perform probe purification with Qiagen PCR spin columns: a. add 300μl PB buffer to reaction, mix with pipette, and transfer to QIAquick column b. spin at maximum speed for 1 min, discard flow through c. wash with 740μl phosphate wash buffer (home-made buffer, NOT from kit) d. spin at maximum speed for 1 min e. empty collection tube and repeat wash steps c and d f. empty collection tube and spin at maximum speed for 1 min g. transfer column to a fresh tube (cut off caps), elute with 30μl elution buffer (home-made KPO4, NOT from kit) with 1 min incubation followed by spin at maximum speed for 1 min h. repeat elution step g, transfer to new tube i. speedvac dry (~ 40 minutes) (heater on middle) 9. Cy dye coupling a. resuspend the cDNA in 4.5μl 0.1M carbonate buffer, pH9.0 (< 1 month old) b. add 4.5μl NHS-Cy dyes c. incubate the reactions in dark for 2 hours at room temperature make prehyb. buffer and warm to 42°C Set heat block to 95°C (Alexa dye labeling: dissolve an aliquot Alexa 546 or 647 in 9 ul DMSO, pipette 5 ul Alexa dye to each cDNA in 4.5 ul of 0.1M carbonate buffer, pH9.0. Incubate both samples for 1-2 hours at room temperature in dark) (3) Purification: 1. Add 35μl 100mM NaOAc pH 5.2 to the reaction 2. Add 250μl PB buffer to reaction and follow QIAquick PCR purification protocol: a. Apply reaction volume to QIAquick spin column and centrifuge for 1 minute b. Discard flow-through, should be colored c. Add 0.74ml buffer PE (as provided with kit) to column and centrifuge for 1 minute d. Discard flow-through and centrifuge for an additional 1-minute at maximum speed e. Place QIAquick column in a clean 1.5ml microcentrifuge tube f. Elute twice with 30μl EB using 1-minute incubation time before centrifugation (eluted sample should look colored and filter should be white) g. Take 1μl of sample for dye incorporation analysis h. Speedvac dry sample (~ 40 minutes) (heat setting to middle) Alternative: precipitate DNA with 1/10 volume 3M sodium acetate, 1/100 volume glycogen, final conc. of 65% ethanol. (4) Dye Incorporation Analysis: 1. Take 1μl of sample for dye incorporation analysis 2. Measure absorbance at 260nm, 650nm (Cy5) for Cy5 incorporation: nucleotide/dye = 28.5*OD260/OD650 150pmole/slide is optimal for hybridization; nucleotide/dye should be < 50


title: slide24b-milk-2947-2924-wtmallarddec03


title: slide27-milk-2869-2875-wtmallarddec03


title: Canary 2a; green: HVC, red: whole brain


title: slide41-milk-2883-2834-wtAug10


title: slide43-milk-3072-3011-wtAug10


title: slide29b-milk-2929-2914-wtmallarddec03


title: Canary 2b; green: HVC, red: whole brain


title: F13


title: Canary 1a; green: HVC, red: whole brain

source_name: Green: HVC

source_name: Red: whole brain


title: F14


title: Canary 1b; green: HVC, red: whole brain


title: slide20b-2875-2869-wtmallarddec03

source_name: Mastitis


title: slide33b-milk-2947-2924-wtmallarddec03


title: Canary 3a; green: HVC, red: whole brain


title: F10


title: F9


title: F6

tag: B-lymphocyte cell line DNA, mother of pedigree XC01423


title: slide22-2869-2875-wtmallarddec03


title: slide9b-2929-2914-wtmallarddec03