title: Data set A

description: Microarray data (GPR file )was extracted by Genepix (version 6.0)

source_name: D0 Female

growth_protocol: The mouse embryonic stem (ES) cell lines PGK12.1 (Norris et al. 1994), CCE/R (O'Neill et al. 2003) and 3F1 (Lee and Lu, 1999) were cultured as previously described (O'Neill et al. 2003).

treatment_protocol: Before differentiation

tag: D0 female ES cell cDNA

tag: D0 male ES cell cDNA


title: Mononucleosome_Control_After_replicate2

status: Public on Dec 01 2007

type: genomic

city: Salt Lake City

first: Timothy

middle: J

last: Parnell

state: UT

address: 2000 Circle of Hope

country: USA

institute: Huntsman Cancer Institute

description: Biological replicate 2 of 3 for mononucleosome DNA hybridization for the control strain after Sth1 degradation.

hyb_protocol: An aliquot of 250 ng of labeled DNA for each sample (channel) was used. Eight experiments were prepared for simultaneous hybridization to each array on the 8 pack slide using the 8 pack gasket coverslip. Hybridizations and washes were performed according to manufacturer's recommended procedures (Agilent Technologies).

scan_protocol: Slides were scanned using an Agilent Technologies Scanner G2505B.

data_processing: Data was extracted from the scanned images using Agilent Feature Extraction software version 8.5.1.1. The 22K 8 pack array format requires a minimum list of 50 pre-identified probes to use as median normalization controls. A list of 312 probes representing genomic loci previously reported as having low RSC occupancy was used as the normalization list in this experiment. Ratios between ChIP and Input channels were determined for probes that were not saturated or below background in either channel. Data from probes that had multiple genomic targets as determined by BLAST analysis were not included.

molecule: genomic DNA

organism: Saccharomyces cerevisiae

source_name: Genomic DNA derived from YBC2192

label_protocol: The template DNA was diluted in 25 ul volume containing 1.2X New England Biolab Buffer 2 and 1 ug of random primers (Invitrogen). The DNA was denatured by incubating at 94 C for 2 minutes and rapid cooled to 8 C. To the reaction 1 nM dATP, 1 nM dTTP, 1 nM dGTP, 0.5 nM dCTP, 1 nM labeled-dCTP, and 5 units of Exo-Klenow (New England Biolabs) was added for a final volume of 30 ul. The reaction was incubated at 37 C for 3 hours. The labeled products were purified using Qiagen PCR cleanup columns (not recommended) with an additional 35% guanidine HCl wash. DNA was eluted in 30 ul Elution Buffer. Incorporation and yield was checked using a Nano-Drop spectrophotometer.

growth_protocol: Cells were grown in liquid YP containing 2% raffinose and/or 2% galactose and 0.05 mM Cu(II)SO4.

extract_protocol: Cells were spheroplasted in spheroplastiing solution (1 M sorbitol, 10 mM Tris-HCl pH 7.4, 10 mM 2-Mercaptoethanol) using zymolyase at 37 C. Spheroplasting was stopped when the OD600 absorbance of an aliquot of cells in 1% SDS dropped to < 10% of the starting value. The cell suspension was adjusted to 0.5% SDS and 5 mM EDTA. Proteinase K was added to 50 ug/ml concentration and the suspension incubated at 65 C for overnight. The suspension was adjusted to 1.2 M potassium acetate and incubated in water/ice bath to precipitate proteins/SDS/salts. The samples were centrifuged at 20K x g for 10 minutes and the pellet discarded. Nucleic acids were then ethanol precipitated from the supernatant. The nucleic acids were resuspended in TE, RNase added, and incubated at 37 C for 10 minutes. The samples were then extracted once with phenol/chloroform, once with chloroform, and ethanol precipitated again. The yield of genomic DNA was measured by OD260. An aliquot of genomic DNA was then fragmented by three rounds of sonication for 25 seconds at 90% duty cycle at power setting 4. An aliquot of 2 ug of fragmented genomic DNA was used as template in the labeling reaction.

tag: Yeast strain

tag: relevant genotype

value: Sth1td ubr1(delta)

source_name: Mononucleosomal DNA from YBC2192 biological replicate 2 after Sth1 degradation.

label_protocol: refer to Sample_label_protocol_ch1.

growth_protocol: Cells were grown in liquid YP containing 2% raffinose and 0.05 mM Cu(II)SO4 at 27 C until OD600 of 0.4Ð0.6. The 'before' aliquot was removed from the culture and galactose was added to the remaining culture to 2% final and incubated for 60 minutes. The culture was then placed at 37 C for 120 minutes. An aliquot of cells were removed and crosslinked in 1% formaldehyde for 5 minutes.

extract_protocol: Cell pellets were resuspended in 1 ml of spheroplasting solution (1 M sorbitol, 10 mM Tris-HCl pH 7.4, 10 mM 2-Mercaptoethanol). The density of cell suspension was measured by absorbance at OD600 and an aliquot equal in cell amount to the other replicates (20-30 OD600 units) was taken. The aliquot of cells were pelleted and resuspended in 1 ml of fresh spheroplasting solution. Zymolyase was added to the samples and incubated at 37 C. Spheroplasting was stopped when the OD600 absorbance of an aliquot of cells in 1% SDS dropped to < 10% of the starting value. Spheroplasts were pelleted and resuspended in 500 ul MNase Digestion Buffer (1 M sorbitol, 50 mM Tris-HCl pH 7.4, 5 mM MgCl2, 1 mM CaCl2, 0.075% NP-40). To the suspension 3.75 units of Micrococcal Nuclease (USB) was added, followed by incubation at 37 C for 25 minutes. The digestion was stopped by adjusting to 0.5% SDS and 5 mM EDTA. Proteinase K was added to 50 ug/ml concentration and incubated at 65 C overnight. The suspension was adjusted to 1.2 M potassium acetate and incubated in water/ice bath to precipitate proteins/SDS/salts. The samples were centrifuged at 20K x g for 10 minutes and the pellet discarded. Nucleic acids were then ethanol precipitated from the supernatant. The nucleic acids were resuspended in TE, RNase added, and incubated at 37 C for 10 minutes. Samples were loaded on a 1.2% agarose gel and DNA fragments separated by electrophoresis. Mononucleosome DNA was excised from the gel and purified using the Qiagen Gel Extraction Kit. DNA yield was measured by OD260. An aliquot of 0.5-1.0 ug DNA was used as template in the labeling reaction.


title: Mononucleosome_Degron_Before_replicate3

description: Biological replicate 3 of 3 for mononucleosome DNA hybridization for the degron strain before Sth1 degradation.

source_name: Mononucleosomal DNA from YBC2191 biological replicate 3 before Sth1 degradation.

growth_protocol: Cells were grown in liquid YP containing 2% raffinose and 0.05 mM Cu(II)SO4 at 27 C until OD600 of 0.4Ð0.6. An aliquot of cells were removed and crosslinked in 1% formaldehyde for 5 minutes.

value: Sth1td pGAL1-UBR1


title: Data set M

type: RNA

city: Birmingham

first: Hong

last: Lin

address: Vincent Drive

department: Anatomy

country: United Kingdom

institute: Institute of Biomedical

hyb_protocol: The labelled samples were purified again and equal amounts (80~120pmol each) of Cy3- and Cy5-labelled cDNAs were combined and precipitated to a final vol. of 21.1ul. A hybridization reaction that contains Formamide (20ul), labelled probes (21.1ul), 10%SDS (0.8ul), 20xSSC (20ul), Cot1	(10ug) and Poly dA (8ug) are denatured at 95C for 2min and applied to arrays followed by overnight incubation at 42C.

scan_protocol: Slides were scanned using a GenePix 4000B scanner. PMT settings were set so as to balance overall signal in the Cy3 and Cy5 channels. Scans were automatically aligned using GenePix (version 6.0) software and then “cherry-picked” manually to eliminate abnormal spots.

data_processing: GPR file was normalised by Gepas software with global loess approach.

molecule: total RNA

organism: Mus musculus

source_name: D12 male

label_protocol: 200ng aliquots of cDNA were labelled with Cy3 fluorophore (Amersham) using Invitrogen Bioprime random primer labelling kits. Briefly, the cDNA was annealed with 20μl 2.5x random primers at 95C for 2 minutes. 1.2mM each of dATP, dGTP, dTTP and 0.6mMdCTP, 1ul of Cy3 dye and 1ul of Klenow fragment were then added to the reaction and incubated overnight at 37C.

extract_protocol: Total RNA was extracted using the RNeasy kit (Qiagen), according to the manufacturer’s protocol and treated with DNaseI (Qiagen).

treatment_protocol: cells of 12 days of differentiation

tag: D12 male ES cell cDNA

source_name: D12 Female

label_protocol: 200ng aliquots of cDNA were labelled with Cy5 fluorophore (Amersham) using Invitrogen Bioprime random primer labelling kits. Briefly, the cDNA was annealed with 20μl 2.5x random primers at 95C for 2 minutes. 1.2mM each of dATP, dGTP, dTTP and 0.6mMdCTP, 1ul of Cy5 dye and 1ul of Klenow fragment were then added to the reaction and incubated overnight at 37C.

tag: D12 Female ES cells


title: Rpc82-ChIP_Control_After_replicate3

description: Biological replicate 3 of 3 for RPC82 ChIP in the control strain after Sth1 degradation.

source_name: Chromatin precipitated from YBC2194 biological replicate 3 after Sth1 degradation using anti-Myc 9E11.

label_protocol:
Round A Amplification: The DNA sample was diluted in 10 ul reaction containing 1X of Sequenase Buffer and 40 pM ChIP-A primer (5'-GTTTCCCAGTCACGATCNNNNNNNNN-3'), denatured at 94 C for 2 min, and rapid cooled to 8 C. To the reaction 5 nM dNTP, 75 nM DTT, 0.5 ug BSA, and 4 units of Sequenase enzyme (USB) was added for a final volume of 15 ul. The temperature of the reaction was slowly raised to 37 C and incubated for 8 minutes. The DNA products were denatured again by incubating at 94 C for 2 minutes, followed by rapid cooling to 8 C. An additional 4 units of Sequenase enzyme was added, the temperature slowly raised to 37C and incubated for 8 minutes. The Round A products were diluted with water to 25 ul final volume.
Round B Amplification: A 7.5 ul aliquot of round A products was used as template in a PCR reaction using 2.5 units of Platinum Taq (Invitrogen), 2 mM MgCl2, and 5 uM ChIP-B primer (5'-GTTTCCCAGTCACGATC-3'). The amplification consisted of 27 rounds of 30 seconds at 94 C, 30 seconds at 40 C, 30 seconds at 50 C, and 60 seconds at 72 C. The Round B DNA products were purified using a Qiagen PCR cleanup column and eluted in 50 ul Elution Buffer. An aliquot was checked by agarose electrophoresis and the concentration determined by OD260.
Round C labeling: An aliquot of Round B product containing 2 ug of DNA was used as template in the labeling reaction. The template DNA was diluted in 25 ul volume containing 1.2X New England Biolab Buffer 2 and 10 pM of ChIP-B primer. The DNA was denatured by incubating at 94 C for 2 minutes and rapid cooled to 8 C. To the reaction 1 nM dATP, 1 nM dTTP, 1 nM dGTP, 0.5 nM dCTP, 1 nM labeled-dCTP, and 5 units of Exo-Klenow (New England Biolabs) was added for a final volume of 30 ul. The reaction was incubated at 37 C for 3 hours. The labeled products were purified using Qiagen PCR cleanup columns (not recommended) with an additional 35% guanidine HCl wash. DNA was eluted in 30 ul Elution Buffer. Incorporation and yield was checked using a Nano-Drop spectrophotometer.

growth_protocol: Cells were grown in liquid YP containing 2% raffinose and 0.05 mM Cu(II)SO4 at 27 C until OD600 of 0.4Ð0.6. The 'before' aliquot was removed from the culture and galactose was added to the remaining culture to 2% final and incubated for 60 minutes. The culture was then placed at 37 C for 120 minutes. An aliquot of cells were removed and crosslinked in 1% formaldehyde for 30 minutes.

extract_protocol:
Chromatin preparation: The washed cell pellets were resuspended in 500 ul of lysis buffer (50 mM Tris-HCl pH 7.5, 140 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% sodium deoxycholate, 0.4 mM DTT, 1X protease inhibitors) and mechanically lysed using 0.7 mm zirconia beads in a mechanical bead beater with four 2 minute rounds. The samples were centrifuged at 20K x g for 5 minutes and the supernatant discarded. The chromatin containing pellet was resuspended in 500 ul of fresh lysis buffer and sonicated five times for 25 seconds each at 90% duty cycle at power setting 4.5. The solubilized chromatin was separated from insoluble material by centrifugation at 20K x g for 10 minutes. The protein concentration was determined by Bradford assay.
Immunoprecipitation: An aliquot of 1000 ug was used for the immunoprecipitation. Extracts were incubated with ~40 ul of anti-mouse beads Dynabeads (Dynal Biotech) pre-bound with 1.7 ul of anti-Myc 9E11 in 500 ul of lysis buffer supplemented with 250 ug BSA Input chromatin overnight. Beads with captured immunocomplexes were washed sequentially with 1 ml of lysis buffer twice, lysis buffer adjusted to 500 mM NaCl once, LiCl wash buffer (10 mM Tris-HCl pH 8, 250 mM LiCl, 0.5% NP-40, 0.5% Sodium Deoxycholate, 1 mM EDTA) once, and TE (10 mM Tris-HCl pH 8, 1 mM EDTA) once. Immunocomplexes were released and cross links reversed by incubating the beads with 200 ul ChIP Elution Buffer (10 mM Tris-HCl pH 8, 1 mM EDTA, 1% SDS) at 65 C overnight. The DNA was purified using Qiagen PCR cleanup columns and eluted in 50 ul Elution Buffer. An aliquot of 7 ul was used in subsequent amplification and labeling procedures.

value: Sth1td ubr1(delta) RPC82-13myc

source_name: Chromatin Input derived from YBC2194 after Sth1 degradation.

growth_protocol: Refer to Sample_growth_protocol_ch1.

extract_protocol:
Chromatin preparation: Refer to Sample_extract_protocol_ch1.
Input: An aliquot of 1000 ug was reserved for input material. Input chromatin was incubated at 65 C overnight to reverse cross links. The DNA was purified using Qiagen PCR cleanup columns and eluted in 50 ul Elution Buffer. An aliquot of 2 ul was used in subsequent amplification and labeling procedures.


title: Mononucleosome_Degron_After_replicate2

description: Biological replicate 2 of 3 for mononucleosome DNA hybridization for the degron strain after Sth1 degradation.

source_name: Mononucleosomal DNA from YBC2191 biological replicate 2 after Sth1 degradation.


title: Mononucleosome_Degron_After_replicate3

description: Biological replicate 3 of 3 for mononucleosome DNA hybridization for the degron strain after Sth1 degradation.

source_name: Mononucleosomal DNA from YBC2191 biological replicate 3 after Sth1 degradation.


title: Mononucleosome_Degron_Before_replicate2

description: Biological replicate 2 of 3 for mononucleosome DNA hybridization for the degron strain before Sth1 degradation.

source_name: Mononucleosomal DNA from YBC2191 biological replicate 2 before Sth1 degradation.


title: Mononucleosome_Control_Before_replicate1

description: Biological replicate 1 of 3 for mononucleosome DNA hybridization for the control strain before Sth1 degradation.

source_name: Mononucleosomal DNA from YBC2192 biological replicate 1 before Sth1 degradation.


title: Mononucleosome_Degron_Before_replicate1

description: Biological replicate 1 of 3 for mononucleosome DNA hybridization for the degron strain before Sth1 degradation.

source_name: Mononucleosomal DNA from YBC2191 biological replicate 1 before Sth1 degradation.


title: Mononucleosome_Degron_After_replicate1

description: Biological replicate 1 of 3 for mononucleosome DNA hybridization for the degron strain after Sth1 degradation.

source_name: Mononucleosomal DNA from YBC2191 biological replicate 1 after Sth1 degradation.


title: Rpc82-ChIP_Degron_After_replicate2

description: Biological replicate 1 of 3 for RPC82 ChIP in the degron strain after Sth1 degradation.

source_name: Chromatin Input derived from YBC2193 after Sth1 degradation.

extract_protocol:
Chromatin preparation: The washed cell pellets were resuspended in 500 ul of lysis buffer (50 mM Tris-HCl pH 7.5, 140 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% sodium deoxycholate, 0.4 mM DTT, 1X protease inhibitors) and mechanically lysed using 0.7 mm zirconia beads in a mechanical bead beater with four 2 minute rounds. The samples were centrifuged at 20K x g for 5 minutes and the supernatant discarded. The chromatin containing pellet was resuspended in 500 ul of fresh lysis buffer and sonicated five times for 25 seconds each at 90% duty cycle at power setting 4.5. The solubilized chromatin was separated from insoluble material by centrifugation at 20K x g for 10 minutes. The protein concentration was determined by Bradford assay.
Input: An aliquot of 1000 ug was reserved for input material. Input chromatin was incubated at 65 C overnight to reverse cross links. The DNA was purified using Qiagen PCR cleanup columns and eluted in 50 ul Elution Buffer. An aliquot of 2 ul was used in subsequent amplification and labeling procedures.

value: Sth1td pGAL1-UBR1 RPC82-13myc

source_name: Chromatin precipitated from YBC2193 biological replicate 2 after Sth1 degradation using anti-Myc 9E11.

extract_protocol:
Chromatin preparation: Refer to Sample_extract_protocol_ch1.
Immunoprecipitation: An aliquot of 1000 ug was used for the immunoprecipitation. Extracts were incubated with ~40 ul of anti-mouse beads Dynabeads (Dynal Biotech) pre-bound with 1.7 ul of anti-Myc 9E11 in 500 ul of lysis buffer supplemented with 250 ug BSA Input chromatin overnight. Beads with captured immunocomplexes were washed sequentially with 1 ml of lysis buffer twice, lysis buffer adjusted to 500 mM NaCl once, LiCl wash buffer (10 mM Tris-HCl pH 8, 250 mM LiCl, 0.5% NP-40, 0.5% Sodium Deoxycholate, 1 mM EDTA) once, and TE (10 mM Tris-HCl pH 8, 1 mM EDTA) once. Immunocomplexes were released and cross links reversed by incubating the beads with 200 ul ChIP Elution Buffer (10 mM Tris-HCl pH 8, 1 mM EDTA, 1% SDS) at 65 C overnight. The DNA was purified using Qiagen PCR cleanup columns and eluted in 50 ul Elution Buffer. An aliquot of 7 ul was used in subsequent amplification and labeling procedures.


title: Rpc82-ChIP_Degron_After_replicate3

description: Biological replicate 3 of 3 for RPC82 ChIP in the degron strain after Sth1 degradation.

source_name: Chromatin precipitated from YBC2193 biological replicate 3 after Sth1 degradation using anti-Myc 9E11.


title: Rpc82-ChIP_Control_After_replicate1

description: Biological replicate 1 of 3 for RPC82 ChIP in the control strain after Sth1 degradation.

source_name: Chromatin precipitated from YBC2194 biological replicate 1 after Sth1 degradation using anti-Myc 9E11.


title: Rpc82-ChIP_Control_Before_replicate1

description: Biological replicate 1 of 3 for RPC82 ChIP in the control strain before Sth1 degradation.

growth_protocol: Cells were grown in liquid YP containing 2% raffinose and 0.05 mM Cu(II)SO4 at 27 C until OD600 of 0.4Ð0.6. An aliquot of cells were removed and crosslinked in 1% formaldehyde for 30 minutes.


title: Rpc82-ChIP_Control_Before_replicate3

description: Biological replicate 3 of 3 for RPC82 ChIP in the control strain before Sth1 degradation.


title: Rpc82-ChIP_Control_Before_replicate2

description: Biological replicate 2 of 3 for RPC82 ChIP in the control strain before Sth1 degradation.

source_name: Chromatin precipitated from YBC2194 biological replicate 2 after Sth1 degradation using anti-Myc 9E11.


title: Rpc82-ChIP_Degron_After_replicate1

source_name: Chromatin precipitated from YBC2193 biological replicate 1 after Sth1 degradation using anti-Myc 9E11.


title: Rpc82-ChIP_Control_After_replicate2

description: Biological replicate 2 of 3 for RPC82 ChIP in the control strain after Sth1 degradation.


title: Rpc82-ChIP_Degron_Before_replicate3

description: Biological replicate 3 of 3 for RPC82 ChIP in the degron strain before Sth1 degradation.

source_name: Chromatin Input derived from YBC2193 before Sth1 degradation.

source_name: Chromatin precipitated from YBC2193 biological replicate 3 before Sth1 degradation using anti-Myc 9E11.


title: Rpc82-ChIP_Degron_Before_replicate2

description: Biological replicate 2 of 3 for RPC82 ChIP in the degron strain before Sth1 degradation.

source_name: Chromatin precipitated from YBC2193 biological replicate 2 before Sth1 degradation using anti-Myc 9E11.


title: Rpc82-ChIP_Degron_Before_replicate1

description: Biological replicate 1 of 3 for RPC82 ChIP in the degron strain before Sth1 degradation.

source_name: Chromatin precipitated from YBC2193 biological replicate 1 before Sth1 degradation using anti-Myc 9E11.


title: H4myc-ChIP_Control_After_replicate2

description: Biological replicate 2 of 2 for H4-myc ChIP in the control strain after Sth1 degradation.

source_name: Chromatin Input derived from YBC2451 after Sth1 degradation.

growth_protocol: Cells were grown in liquid YP containing 2% raffinose and 0.05 mM Cu(II)SO4 at 27 C until OD600 of 0.4Ð0.6. The 'before' aliquot was removed from the culture and galactose was added to the remaining culture to 2% final and incubated for 60 minutes. The culture was then placed at 37 C for 120 minutes. An aliquot of cells were removed and crosslinked in 1% formaldehyde for 15 minutes.

value: Sth1td ubr1(delta) myc-HHF2

source_name: Chromatin precipitated from YBC2451 biological replicate 2 after Sth1 degradation using anti-Myc 9E11.


title: H4myc-ChIP_Control_After_replicate1

description: Biological replicate 1 of 2 for H4-myc ChIP in the control strain after Sth1 degradation.

source_name: Chromatin precipitated from YBC2451 biological replicate 1 after Sth1 degradation using anti-Myc 9E11.


title: H4myc-ChIP_Control_Before_replicate2

description: Biological replicate 2 of 2 for H4-myc ChIP in the control strain before Sth1 degradation.

source_name: Chromatin Input derived from YBC2451 before Sth1 degradation.

growth_protocol: Cells were grown in liquid YP containing 2% raffinose and 0.05 mM Cu(II)SO4 at 27 C until OD600 of 0.4Ð0.6. An aliquot of cells were removed and crosslinked in 1% formaldehyde for 15 minutes.

source_name: Chromatin precipitated from YBC2451 biological replicate 2 before Sth1 degradation using anti-Myc 9E11.


title: H4myc-ChIP_Degron_After_replicate2

description: Biological replicate 2 of 2 for H4-myc ChIP in the degron strain after Sth1 degradation.

source_name: Chromatin Input derived from YBC2449 after Sth1 degradation.

value: Sth1td pGAL1-UBR1 myc-HHF2

source_name: Chromatin precipitated from YBC2449 biological replicate 2 after Sth1 degradation using anti-Myc 9E11.


title: H4myc-ChIP_Degron_After_replicate1

description: Biological replicate 1 of 2 for H4-myc ChIP in the degron strain after Sth1 degradation.

source_name: Chromatin precipitated from YBC2449 biological replicate 1 after Sth1 degradation using anti-Myc 9E11.


title: H4myc-ChIP_Control_Before_replicate1

description: Biological replicate 1 of 2 for H4-myc ChIP in the control strain before Sth1 degradation.

source_name: Chromatin precipitated from YBC2451 biological replicate 1 before Sth1 degradation using anti-Myc 9E11.


title: H4myc-ChIP_Degron_Before_replicate2

description: Biological replicate 2 of 2 for H4-myc ChIP in the degron strain before Sth1 degradation.

source_name: Chromatin Input derived from YBC2449 before Sth1 degradation.

source_name: Chromatin precipitated from YBC2449 biological replicate 2 before Sth1 degradation using anti-Myc 9E11.


title: H4myc-ChIP_Degron_Before_replicate1

description: Biological replicate 1 of 2 for H4-myc ChIP in the degron strain before Sth1 degradation.

source_name: Chromatin precipitated from YBC2449 biological replicate 1 before Sth1 degradation using anti-Myc 9E11.


title: Rsc8-ChIP_Mock-After_replicate2

description: Biological replicate 2 of 3 for Rsc8-9myc ChIP at mock-after conditions.

data_processing: Data was extracted from the scanned images using Agilent Feature Extraction software version 8.5.1.1. The 22K 8 pack array format requires a minimum list of 50 pre-identified probes to use as median normalization controls. A list of 200 probes which had the lowest raw (un-processed) ratio values was used as the normalization list in this experiment. The rationalization is that these probes and their corresponding values should represent the majority of the genome which is not enriched for the occupancy of RSC. Ratios between ChIP and Input channels were determined for probes that were not saturated or below background in either channel. Data from probes that had multiple genomic targets as determined by BLAST analysis were not included.

source_name: Chromatin Input derived from YBC1547 mock-after

growth_protocol: Cells were grown in liquid YP containing 2% raffinose at 27 C until OD600 of 0.4Ð0.6. The 'before' aliquot was removed from the culture and galactose was added to the remaining culture to 2% final and incubated for 60 minutes. The culture was then placed at 37 C for 120 minutes. An aliquot of cells were removed and crosslinked in 1% formaldehyde for 30 minutes.

extract_protocol:
Chromatin preparation: The washed cell pellets were resuspended in 500 ul of lysis buffer (50 mM Tris-HCl pH 7.5, 140 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% sodium deoxycholate, 0.4 mM DTT, 1X protease inhibitors) and mechanically lysed using 0.7 mm zirconia beads in a mechanical bead beater with four 2 minute rounds. The samples were centrifuged at 20K x g for 5 minutes and the supernatant discarded. The chromatin containing pellet was resuspended in 500 ul of fresh lysis buffer and sonicated five times for 25 seconds each at 90% duty cycle at power setting 4.5. The solubilized chromatin was separated from insoluble material by centrifugation at 20K x g for 10 minutes. The protein concentration was determined by Bradford assay.
Input: An aliquot of 400 ug was reserved for input material. Input chromatin was incubated at 65 C overnight to reverse cross links. The DNA was purified using Qiagen PCR cleanup columns and eluted in 50 ul Elution Buffer. An aliquot of 2 ul was used in subsequent amplification and labeling procedures.

source_name: Chromatin precipitated from YBC1547 mock-after replicate 2 using anti-Myc 9E11

extract_protocol:
Chromatin preparation: Refer to Sample_extract_protocol_ch1.
Immunoprecipitation: An aliquot of 400 ug was used for the immunoprecipitation. Extracts were incubated with ~40 ul of anti-mouse beads Dynabeads (Dynal Biotech) pre-bound with 1.7 ul of anti-Myc 9E11 in 500 ul of lysis buffer supplemented with 250 ug BSA Input chromatin overnight. Beads with captured immunocomplexes were washed sequentially with 1 ml of lysis buffer twice, lysis buffer adjusted to 500 mM NaCl once, LiCl wash buffer (10 mM Tris-HCl pH 8, 250 mM LiCl, 0.5% NP-40, 0.5% Sodium Deoxycholate, 1 mM EDTA) once, and TE (10 mM Tris-HCl pH 8, 1 mM EDTA) once. Immunocomplexes were released and cross links reversed by incubating the beads with 200 ul ChIP Elution Buffer (10 mM Tris-HCl pH 8, 1 mM EDTA, 1% SDS) at 65 C overnight. The DNA was purified using Qiagen PCR cleanup columns and eluted in 50 ul Elution Buffer. An aliquot of 7 ul was used in subsequent amplification and labeling procedures.


title: Rsc8-ChIP_Mock-After_replicate3

description: Biological replicate 3 of 3 for Rsc8-9myc ChIP at mock-after conditions.

source_name: Chromatin precipitated from YBC1547 mock-after replicate 3 using anti-Myc 9E11


title: gp91phox-/- 4hr_after mock inoculation, biological rep1

city: Bozeman

first: Kate

last: McInnerney

state: MT

address: 109 Lewis Hall

department: Microbiology

institute: Montana State University

description: Gene expression timecourse from AM following in vivo exposure to A. fumigatus conidia.

hyb_protocol: Following fragmentation, 10 micrograms of cRNA were hybridized for 16 hr_at 45C on GeneChip Mouse GeneChip 430A 2.0 Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 450.

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

data_processing: Data was analyzed with Agilent Genespring GX v. 7.3.1 using Affymetrix GCOS generated flag data from scaled CHP files, and RMA-normalized CEL files.

label: biotin

source_name: Alveolar Macrophage (AM)

label_protocol: Biotinylated cRNA were prepared according to the standard two-cycle Affymetrix protocol from 100 nanograms total RNA (Expression Analysis Technical Manual, 2004, Affymetrix).

extract_protocol: RNeasy RNA extraction kit (Quiagen) was performed on AM harvested fresh according to the manufacturer's instructions from mouse lungs.

treatment_protocol: Sample AM collected 4 hours after in vivo mock treatment

tag: AM collected from gp91phox-/-, 4 hour after mock treatment


title: gp91phox-/- 4hr_after mock inoculation, biological rep2


title: gp91phox-/- 4hr_after mock inoculation, biological rep3


title: Rsc3-ChIP_Mock-After_replicate2

description: Biological replicate 2 of 3 for Rsc3-9myc ChIP at mock-after conditions.

source_name: Chromatin Input derived from YBC1546 mock-after

source_name: Chromatin precipitated from YBC1546 mock-after replicate 2 using anti-Myc 9E11


title: gp91phox-/- 2hr_after mock inoculation, biological rep3

treatment_protocol: Sample AM collected 2 hours after in vivo mock treatment

tag: AM collected from gp91phox-/-, 2 hour after mock treatment


title: gp91phox-/- 2hr_after mock inoculation, biological rep1


title: Rsc8-ChIP_Mock-Before_replicate3

description: Biological replicate 3 of 3 for Rsc8-9myc ChIP at mock-before conditions.

source_name: Chromatin Input derived from YBC1547 mock-before

growth_protocol: Cells were grown in liquid YP containing 2% raffinose at 27 C until OD600 of 0.4Ð0.6. An aliquot of cells were removed and crosslinked in 1% formaldehyde for 30 minutes.

source_name: Chromatin precipitated from YBC1547 mock-before replicate 3 using anti-Myc 9E11


title: Rsc3-ChIP_Mock-After_replicate3

description: Biological replicate 3 of 3 for Rsc3-9myc ChIP at mock-after conditions.

source_name: Chromatin precipitated from YBC1546 mock-after replicate 3 using anti-Myc 9E11


title: Rsc8-ChIP_Mock-Before_replicate2

description: Biological replicate 2 of 3 for Rsc8-9myc ChIP at mock-before conditions.

source_name: Chromatin precipitated from YBC1547 mock-before replicate 2 using anti-Myc 9E11


title: Rsc3-ChIP_Mock-After_replicate1

description: Biological replicate 1 of 3 for Rsc3-9myc ChIP at mock-after conditions.

source_name: Chromatin precipitated from YBC1546 mock-after replicate 1 using anti-Myc 9E11


title: Rsc8-ChIP_Mock-Before_replicate1

description: Biological replicate 1 of 3 for Rsc8-9myc ChIP at mock-before conditions.

source_name: Chromatin precipitated from YBC1547 mock-before replicate 1 using anti-Myc 9E11


title: gp91phox-/- 4hr_after e7 conidia, biological rep1

treatment_protocol: Sample AM collected 4 hours after in vivo incubation with conidia

tag: AM collected from gp91phox-/-, 4 hour after exposure to Aspergillus fumigatus conidia


title: Rsc3-ChIP_Mock-Before_replicate1

description: Biological replicate 1 of 3 for Rsc3-9myc ChIP at mock-before conditions.

source_name: Chromatin Input derived from YBC1546 mock-before

source_name: Chromatin precipitated from YBC1546 mock-before replicate 1 using anti-Myc 9E11


title: Rsc3-ChIP_Mock-Before_replicate3

description: Biological replicate 3 of 3 for Rsc3-9myc ChIP at mock-before conditions.

source_name: Chromatin precipitated from YBC1546 mock-before replicate 3 using anti-Myc 9E11


title: Rsc8-ChIP_Mock-After_replicate1

description: Biological replicate 1 of 3 for Rsc8-9myc ChIP at mock-after conditions.

source_name: Chromatin precipitated from YBC1547 mock-after replicate 1 using anti-Myc 9E11


title: Rsc3-ChIP_Mock-Before_replicate2

description: Biological replicate 2 of 3 for Rsc3-9myc ChIP at mock-before conditions.

source_name: Chromatin precipitated from YBC1546 mock-before replicate 2 using anti-Myc 9E11


title: gp91phox-/- 4hr_after e7 conidia, biological rep2


title: gp91phox-/- 2hr_after mock inoculation, biological rep2


title: gp91phox-/- 4hr_after e6 conidia, biological rep3


title: liver_WT_rep2

city: Bethesda

first: Stephen

middle: John

last: Davies

state: MD

address: 4301 Jones Bridge Road

department: Microbiology and Immunology

institute: USUHS

description: Total RNA from whole liver of female wild type C57BL/6 mice

hyb_protocol: - Hold hybridization buffer (30 mM sodium citrate, 300 mM sodium chloride, 50% formamide, 0.4% SDS, pH 7.0) at 42°C. - Add twelve uL to each sample along with 3 uL of 1:1 salmon sperm/tRNA blocker, heat for 2 min at 98°C, centrifuge for 3 min at 14,000 x g. - Transfer 20 uL into a microarray hybridization chamber (MAUI Mixer, Biomicro Systems). - After overnight hybridization at 45°C, insert slides into a jig and submerge in Buffer A, comprised of 1X SSC and 0.05% SDS. - Remove adhesive MAUI Mixer lid and place slide in a rack in a jar containing fresh Buffer A. Once all slides are in the rack, agitate for 1 min, transfer to fresh Buffer A, agitate 2 min, transfer to Buffer B (0.1X SSC) for 2 min and then into fresh Buffer B. - Centrifuge slides for 5 min starting at 220 rpm and increasing to 580 in increments of approximately 100 rpm. Once dry, remove slides from the centrifuge and scan on a GenePix4000B Scanner (Axon Instruments/Molecular Devices).

scan_protocol: Images were scanned by GenePix4000B Scanner (Axon Instruments/Molecular Devices, Sunnyvale, CA)

data_processing:
Analyzed using the mAdb program (http://madb.niaid.nih.gov/).
Default Ratio: ChanB/ChanA (Cy5/Cy3)
Signal calculation: Mean Intensity minus Median Background
Any Features designated Control were excluded.
Normalization method: Precalculated 50th Percentile (Median)**

source_name: liver, wild type

label_protocol:
- Using RNase-free water, make up fresh Master Mix containing 1x RT buffer (Invitrogen), 1 mM DDT, and 1 mM each of dATP, dCTP, dGTP and dTTP.
- Prime the two RNA samples to be compared by adding 1 ul Oligo dT primer (1mg/ml) to 30 ug total RNA in 15 ul water.
- Heat samples to 70°C for 5 min., then snap cool in an ice-water bath. Centrifuge briefly to collect the contents to the bottom of the tube.
- Add these components in the order listed:
-   RNase inhibitor diluted 2X, 0.5 ul
-   Master Mix, 10.0 ul
-   dUTP-Cy3 OR dUTP Cy5, 2.0 ul
-   Superscript II, 1.5 ul
- Mix the components well with the RNA/primer sample and centrifuge briefly to collect the contents to the bottom of the tube.
- Incubate reverse transcription reaction mixture at 42°C for 90 min.
- After the reverse transcription reaction is complete, add 10ul of 1M NaOH and heat at 70oC for 15 min.
- Neutralize with 10ul 1M HCl.
- Combine the appropriate Cy3- and Cy5-labeled samples that will be co-hybridized onto the array.
- Add the combined probes to a Vivaspin 500 (10K cutoff) with 400 ul TE. Centrifuge at 12,000 X g (~10,000 rpm) for 8 min. Discard flow through.
- Repeat the wash step in the same Vivaspin filter at least three times.
- Remove the sample retained in the filter chamber with a pipette and transfer to a new tube. Adjust the sample volume to 20 ul.

extract_protocol:
- Homogenize tissue samples in 1ml of RNAzol reagent
- Add 0.2x volume of Chloroform. Shake tube well. Incubate 2 min.
- Centrifuge at 13,000 x g for 15 min at 4oC and collect upper clear layer.
- Add an equal volume of 70% Ethanol to the upper layer. Mix well and apply to RNeasy spin column. Check product information for capacity of columns.
- Centrifuge Discard flow through. Wash column 2x with RPE buffers supplied in the kit. Be certain ethanol has been added to RPE concentrate before use.
- After the last spin the column is dry so that the Ethanol is removed. Transfer the column to a fresh collection tube.
- Elute RNA with the designated volume of RNase free water. Allow to stand for 1 min then centrifuge collecting the eluted RNA. Repeat one time.
- Analyze for purity and concentration on a NanoDrop® ND-1000 Spectrophotometer (Wilmington, DE).

tag: Strain

tag: Gender

value: Female

source_name: Reference pool (liver, RAG, OT-II/RAG and wild type)


title: gp91phox-/- 4hr_after e6 conidia, biological rep2


title: Megakaryocyte CD41+_39C_second replicate

city: Gothenburg

first: Lars

last: Palmqvist

address: Sahlgrenska University Hospital

department: Institute of Biomedicine

country: Sweden

institute: University of Gothenburg

description: The gene array data CEL-files were imported into GeneSpring® software version 7.3 (Silicon Genetics, Redwood City, CA). The GC-RMA method (Irizarry et al., 2003) was used for normalization and data was processed as follows: all values below 0.01 were set to 0.01: all of the genes in each sample were divided by the median of the genes present on the HG133A gene chip. All samples were then normalized against the 37ºC control samples. Each measurement for each gene in those specific samples was divided by the median of that gene's measurements in the corresponding control samples. We judged genes to be differentially expressed when the difference was of at least 1.5 fold; and the extent of difference in expression was significantly different in an ANOVA t-test (p<0.05).

hyb_protocol: Protocol	EukGE-WS2v4 Wash A1 Recovery Mixes	0 Wash A1 Temperature (C)	25 Number of Wash A1 Cycles	10 Mixes per Wash A1 Cycle	2 Wash B Recovery Mixes	0 Wash B Temperature (C)	50 Number of Wash B Cycles	4 Mixes per Wash B Cycle	15 Stain Temperature (C)	25 First Stain Time (seconds)	600 Wash A2 Recovery Mixes	0 Wash A2 Temperature (C)	25 Number of Wash A2 Cycles	10 Mixes per Wash A2 Cycle	4 Second Stain Time (seconds)	600 Third Stain Time (seconds)	600 Wash A3 Recovery Mixes	0 Wash A3 Temperature (C)	30 Number of Wash A3 Cycles	15 Mixes per Wash A3 Cycle	4 Holding Temperature (C)	25 Station	1 Module	3

scan_protocol: Pixel Size	3 Filter	570 Scan Temperature Scanner ID Number of Scans	2 Scanner Type	HP

data_processing: MAS 5.0

label: Biotin

organism: Homo sapiens

source_name: Cord blood

label_protocol: The labeling was done as previously described (Perez-Iratxeta et al., 2005) and performed at the StemCore Laboratories (www.stemcore.ca, Ottawa, Canada)

growth_protocol: CD41a+ Mk issued from CB CD34+ cells were cultured for 7-days with TPO at 100 ng/ml and purified by positive magnetic selection using a CD41a-FITC monoclonal antibody (Immunotech, Marseille, France) and MACS columns according to manufacturer’s instructions (Miltenyi, Auburn, CA, USA) with a purity >95%.

extract_protocol: RNA was then extracted following the manufacturer’s instructions (RNeasy kit, Qiagen, Mississauga, Ontario, Canada).

treatment_protocol: To define potential differences between megakaryocytes (Mk) derived at 37°C or 39°C, we compared their gene expression repertoire by micro-gene chip technology (Affymetrix GeneChip HG133A). Two independent experiments were carried out on Mk at 37 or 39°C.

tag: Human CB cell collection, cryopreservation and CD34+ cell enrichment were previously described (Proulx et al., 2003). CD34+ cells were enriched by negative selection according to manufacturer’s instructions (Human Progenitor Enrichment Cocktail and StemSep column, StemCell Technologies, Vancouver, BC, Canada).


title: gp91phox-/- 4hr_after e6 conidia, biological rep1

source_name: Alveolar Macrophage (AM)


title: Megakaryocyte CD41+_39C_first replicate


title: Megakaryocyte CD41+_37C_second replicate


title: gp91phox-/- 4hr_after e7 conidia, biological rep3


title: liver_RAG_rep2

description: Total RNA from whole liver of female RAG KO C57BL/6 mice

source_name: liver, RAG KO

label_protocol:
- Using RNase-free water, make up fresh Master Mix containing 1x RT buffer (Invitrogen), 1 mM DDT, and 1 mM each of dATP, dCTP, dGTP and dTTP.
- Prime the two RNA samples to be compared by adding 1 ul Oligo dT primer (1mg/ml) to 30 ug total RNA in 15 ul water.
- Heat samples to 70°C for 5 min., then snap cool in an ice-water bath. Centrifuge briefly to collect the contents to the bottom of the tube.
- Add these components in the order listed:
- RNase inhibitor diluted 2X, 0.5 ul
- Master Mix, 10.0 ul
- dUTP-Cy3 OR dUTP Cy5, 2.0 ul
- Superscript II, 1.5 ul
- Mix the components well with the RNA/primer sample and centrifuge briefly to collect the contents to the bottom of the tube.
- Incubate reverse transcription reaction mixture at 42°C for 90 min.
- After the reverse transcription reaction is complete, add 10ul of 1M NaOH and heat at 70oC for 15 min.
- Neutralize with 10ul 1M HCl.
- Combine the appropriate Cy3- and Cy5-labeled samples that will be co-hybridized onto the array.
- Add the combined probes to a Vivaspin 500 (10K cutoff) with 400 ul TE. Centrifuge at 12,000 X g (~10,000 rpm) for 8 min. Discard flow through.
- Repeat the wash step in the same Vivaspin filter at least three times.
- Remove the sample retained in the filter chamber with a pipette and transfer to a new tube. Adjust the sample volume to 20 ul.


title: gp91phox-/- 2hrs post e6 conidia, biological rep2

treatment_protocol: Sample AM collected 2 hours after in vivo incubation with conidia

tag: AM collected from gp91phox-/-, 2 hour after exposure to Aspergillus fumigatus conidia


title: liver_OT-I/RAG_rep2

description: Total RNA from whole liver of female OT-II/RAG C57BL/6 mice

source_name: liver, OT-II/RAG


title: gp91phox-/- 2hrs post e6 conidia, biological rep3


title: gp91phox-/- 2hr_after e7 conidia, biological rep3


title: Data Set S

description: Scanned image were automatically aligned using GenePix (version 6.0) software and then “cherry-picked” manually to eliminate abnormal spots. Microarray data (GPR file) were then extracted by Genepix (version 6.0)

scan_protocol: Slides were scanned using a GenePix 4000B scanner. PMT settings were set so as to balance overall signal in the Cy3 and Cy5 channels.

data_processing: The data in GPR file were normalised by Gepas software with global loess approach.

source_name: D21 Female

growth_protocol: The mouse embryonic stem (ES) cell lines PGK12.1 (Norris et al. 1994), CCE/R (O'Neill et al. 2003) and 3F1 (Lee and Lu, 1999) were cultured as previously described (O'Neill et al. 2003). Differentiation was induced by replating on non-adherent plastic dishes in the absence of LIF. Adult control cells were thymic lymphocytes from 4 week old Balb/c mice.

extract_protocol: Total RNA was extracted using the RNeasy kit (Qiagen), according to the manufacturer’s protocol and treated with DNaseI (Qiagen). First strand cDNAs were reverse transcribed from 3μg total RNA in a reaction containing 4μl 5x first-strand buffer, 1 μl 0.1M DTT, 1μl oligo dT primer (50 μM), 200unit RT-Superscript III transcriptase (RT-SSIII, Invitrogen), 40unit RNAase inhibitor (Boehringer Mannheim) made up to 20μl. The cDNA was purified with a Qiagen PCR purification kit.

treatment_protocol: Cells after 21 days of differentiation

tag: D21 female ES cells

source_name: D21 male

treatment_protocol: cells after 21 days of differentiation

tag: D21 male ES cells


title: gp91phox-/- 2hr_after e7 conidia, biological rep1


title: liver_WT_rep3


title: HUVEC vs M199 8hr exposure 2nd expt

city: Memphis

first: Katherine

middle: S

last: Barker

state: TN

address: 881 Madison Avenue, Room 305

department: Clinical Pharmacy

institute: University of Tennessee Health Science Center

hyb_protocol: Hybridized microarrays were washed according to manufacturer’s protocols and subjected to a signal enhancement protocol consisting of an initial incubation with streptavidin-phycoerythrin conjugate followed by goat anti-streptavidin biotinylated antibody and a final staining with the streptavidin-phycoerythrin conjugate.

scan_protocol: The microarrays were scanned using the GeneArray™ scanner with an argon ion laser excitation source and emission detected by a photomultiplier tube through a 570-nm long pass filter.  Digitized image data were processed using GeneChip® Operating Software.

data_processing: normalized to all data points

source_name: human umbilical vein endothelial cells

label_protocol: Ten micrograms of total RNA were subjected to first- and second-strand cRNA synthesis incorporating biotin-labeled nucleotides.  cRNA was then fragmented and subsequently hybridized overnight with microarray chips using the manufacturer’s hybridization buffer.

growth_protocol: grown in M199 culture medium supplemented with human serum at 37 degrees Celsius for 8 hours in 5% CO2

extract_protocol: Trizol reagent

treatment_protocol: 8hr confluent culture of HUVECs in M199 culture medium

tag: HUVECs cultured with M199 culture medium for 8hrs


title: Megakaryocyte CD41+_37C_first replicate

tag: Human cord blood (CB) cell collection, cryopreservation and CD34+ cell enrichment has been previously described (Proulx et al., 2003). CD34+ cells were enriched by negative selection according to manufacturer’s instructions (Human Progenitor Enrichment Cocktail and StemSep column, StemCell Technologies, Vancouver, BC, Canada).


title: gp91phox-/- 2hrs post e6 conidia, biological rep1


title: gp91phox-/- 2hr_after e7 conidia, biological rep2


title: HUVEC vs M199 8hr exposure 1st expt

tag: HUVECs cultured in M199 culture medium 8hrs


title: gp91phox-/- AM 0hr_control, biological rep2

treatment_protocol: Sample AM collected at t=0

tag: AM collected from gp91phox-/-, at 0 hours (no exposure to Aspergillus fumigatus conidia)


title: liver_OT-II/RAG_rep3


title: liver_RAG_rep1

description: Total RNA from whole liver of female RAG-/--C57BL/6 mice

source_name: liver, RAG


title: gp91phox-/- AM 0hr_control, biological rep3

description: Gene expression timecourse from AM following in vivo exposure to A. fumigatus conidia.


title: liver_OT-II/RAG_rep1

description: Total RNA from whole liver of female OT-II/RAG-/--C57BL/6 mice


title: HUVEC vs M199 3hr 2nd expt

growth_protocol: grown in M199 culture medium supplemented with human serum at 37 degrees Celsius for 3 hours in 5% CO2

treatment_protocol: 3hr confluent culture of HUVECs in M199 culture medium

tag: HUVECs cultured with M199 culture medium 3hrs


title: C57Bl/6 AM 4hrs after mock txmt, biological rep3

tag: AM collected from C57Bl/6 and gp91phox-/-, 4 hour after mock inoculation (no conidia)


title: HUVEC vs SC5314 8hr exposure 1st expt

treatment_protocol: 8hr co-culture of confluent culture of HUVECs with SC5314 strain of Candida albicans in M199 culture medium

tag: HUVECs co-cultured with SC5314 strain (Candida albicans) for 8hrs in M199 culture medium


title: liver_WT_rep1


title: HUVEC vs M199 3hr exposure 1st expt

tag: HUVECs cultured for 3hrs in M199 culture medium


title: HUVEC vs SC5314 8hr 2nd expt


title: C57Bl/6 AM 4hrs after mock txmt, biological rep2


title: C57Bl/6 AM 4hrs after mock txmt, biological rep4


title: liver_RAG_rep3

description: Total RNA from whole liver of female RAG C57BL/6 mice


title: HUVEC vs SC5314 3hr exposure 2nd expt

treatment_protocol: 3hr co-culture of confluent culture of HUVECs with SC5314 strain of Candida albicans in M199 culture medium

tag: HUVECs co-cultured with SC5314 strain (Candida albicans) for 3hrs in M199 culture medium


title: HUVEC vs SC5314 3hr exposure 1st expt


title: gp91phox-/- AM 0hr_control, biological rep1


title: C57Bl/6 AM 4hrs after e7 conidia, biological rep4

tag: AM collected from C57Bl/6, 4 hour after exposure to Aspergillus fumigatus conidia


title: HUVEC vs CAN34 8hr exposure 2nd expt

treatment_protocol: 8hr co-culture of confluent culture of HUVECs with CAN34 strain of Candida albicans in M199 culture medium

tag: HUVECs co-cultured with CAN34 strain (Candida albicans) for 8hrs in M199 culture medium


title: Zmet2 study 11-day seedling aerial tissue genotype B73 zmet2-m1 Rep1

city: Saint Paul

first: Nathan

middle: M

last: Springer

state: MN

address: 1445 Gortner Ave

department: Plant Biology

institute: University of Minnesota

description: No other relevant details

hyb_protocol: Hybridization was performed according to the recommended Affymetrix protocols at the University of Minnesota Microarray facility.

scan_protocol: The Genechip 3000 scanner was used to scan each array

data_processing: The MAS5.0 values are available in the submitted file; RMA processing of the .cel files was also used.

organism: Zea mays

source_name: B73 zmet2-m1 seedling

label_protocol: Eight μg of total RNA was labeled for each hybridization using the One-Cycle cDNA Synthesis Kit, according to the manufacturer’s instructions (Affymetrix, Santa Clara CA).

growth_protocol: The zmet2-m1 allele was generated by Mu transposon mutagenesis and contains a Mu insertion in exon 18 that results in a loss-of-function mutation.  This allele was backcrossed into the Mo17 and B73 maize inbred lines for six generations and then self-pollinated for two generations to produce homozygous near-isogenic lines of Mo17 and B73 that are homozygous for the zmet2-m1 allele.  Three biological replicates of the B73 and B73 zmet2-m1 genotypes were grown sequentially over a 5 week period during May and June 2005.  Three additional biological replicates involving the B73, B73 zmet2-m1, Mo17 and Mo17 zmet2-m1 genotypes were grown during January and February 2006.  The plants were grown using standard greenhouse conditions (1:1 mix of autocalaved field soil and MetroMix; 16 hours light and 8 hours dark; daytime temperature of 30 C and night temperature of 22 C) and were sampled for gene expression on the 11th day after planting between 8 am and 9 am.  The plants were cut immediately above the highest root, thus all above-ground tissues and meristems were collected.  For each biological replicate, the seeds were planted such that one seed of each genotype was present in every pot.  Eight pots were selected for tissue collection resulting in sampling of eight plants per genotype in each replicate.  The sampled tissues were flash frozen in liquid nitrogen and stored at -80C prior to RNA isolation.

extract_protocol: For RNA isolation tissues from 8 seedlings per genotype per biological replicate were pooled and ground in liquid nitrogen.  RNAs were extracted using Trizol reagent according to the manufacturer’s instructions (Invitrogen Corp., Carlsbad CA) and purified using the RNeasy kit, according to the manufactures instructions (Qiagen Corp., Valencia, CA).  The quality and quantity of all purified RNA samples were assessed using agarose gel electrophoresis and the Nanodrop spectrophotometer (Nanodrop Technologies, Montchanin, DE).

tag: Stage

value: 11-day seedling all aerial tissue; Genotype:B73 zmet2-m1


title: Zmet2 study 11-day seedling aerial tissue genotype B73 zmet2-m1 Rep3

description: No other relevant details


title: C57Bl/6 AM 4hrs after e7 conidia, biological rep3


title: Zmet2 study 11-day seedling aerial tissue genotype B73 zmet2-m1 Rep2

description: No other relevant details


title: C57Bl/6 AM 4hrs after mock txmt, biological rep1


title: Zmet2 study 11-day seedling aerial tissue genotype Mo17 Rep3

description: No other relevant details

source_name: Mo17 seedling

value: 11-day seedling all aerial tissue; Genotype:Mo17


title: HUVEC vs CAN34 8hr exposure 1st expt


title: HUVEC vs CAN34 3hr exposure 2nd expt

treatment_protocol: 3hr co-culture of confluent culture of HUVECs with CAN34 strain of Candida albicans in M199 culture medium

tag: HUVECs co-cultured with CAN34 strain (Candida albicans) for 3 hrs in M199 culture medium


title: HUVEC vs CAN34 3hr exposure 1st expt

tag: HUVECs co-cultured with CAN34 strain (Candida albicans) for 3hrs in M199 culture medium


title: Zmet2 study 11-day seedling aerial tissue genotype Mo17 zmet2-m1 Rep3

description: No other relevant details

source_name: Mo17 zmet2-m1 seedling

value: 11-day seedling all aerial tissue; Genotype:Mo17 zmet2-m1


title: C57Bl/6 AM 4hrs post e6 conidia, biological rep3

source_name: Alveolar Macrophage (AM) and control untreated


title: prostatic cell line NB26

city: Princeton

first: Xi-De

last: Wang

state: NJ

institute: Bristol-Myers Squibb

description: Gene expression data from cultured prostatic cell line

hyb_protocol: Following fragmentation, 10 ug of cRNA were hybridized, washed and stained using the Affymetrix Fluidics Station 400 according to Affymetrix standard procedures

scan_protocol: GeneChips were scanned using the Affymetrix GeneChip Scanner 3000

data_processing: The data were normalized with RMA.

source_name: cultured human prostatic cells

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

extract_protocol: Total RNA isolated from cell culture using RNeasy kit (Qiagen) according to the manufacturer's instructions.

tag: cell line


title: C57Bl/6 AM 4hrs after e7 conidia, biological rep2


title: Zmet2 study 11-day seedling aerial tissue genotype B73 zmet2-m1 Rep6

description: No other relevant details


title: prostatic cell line PWR1E


title: prostatic cell line DUCaP


title: C57Bl/6 AM 4hrs post e6 conidia, biological rep4


title: C57Bl/6 AM 4hrs after e7 conidia, biological rep1


title: Zmet2 study 11-day seedling aerial tissue genotype B73 Rep6

description: No other relevant details

source_name: B73 seedling

value: 11-day seedling all aerial tissue; Genotype:B73


title: prostatic cell line W99


title: Zmet2 study 11-day seedling aerial tissue genotype B73 zmet2-m1 Rep5

description: No other relevant details


title: Zmet2 study 11-day seedling aerial tissue genotype Mo17 Rep2

description: No other relevant details


title: prostatic cell line NB11


title: Zmet2 study 11-day seedling aerial tissue genotype Mo17 zmet2-m1 Rep2

description: No other relevant details


title: C57Bl/6 AM 4hrs post inoc with beads, biological rep2

treatment_protocol: Sample AM collected 4 hours after in vivo incubation with polystyrene beads

tag: AM collected from C57Bl/6, 4 hour after exposure to polystyrene beads


title: C57Bl/6 AM 4hrs post e6 conidia, biological rep2


title: prostatic cell line RWPE2


title: C57Bl/6 AM 4hrs post inoc with beads, biological rep1


title: C57Bl/6 2hr_mock txmt, biological rep4

treatment_protocol: Sample AM collected 2 hours after in vivo mock inoculation

tag: AM collected from C57Bl/6, 2 hour after mock inoculation


title: Zmet2 study 11-day seedling aerial tissue genotype Mo17 Rep1

description: No other relevant details


title: prostatic cell line RWPE1


title: Zmet2 study 11-day seedling aerial tissue genotype Mo17 zmet2-m1 Rep1

description: No other relevant details


title: C57Bl/6 2hr_mock txmt, biological rep1


title: prostatic cell line HPV10


title: Zmet2 study 11-day seedling aerial tissue genotype B73 zmet2-m1 Rep4

description: No other relevant details


title: Zmet2 study 11-day seedling aerial tissue genotype B73 Rep4

description: No other relevant details


title: prostatic cell line MDAPCa2b


title: C57Bl/6 AM 4hrs post e6 conidia, biological rep1


title: C57Bl/6 2hr_mock txmt, biological rep2


title: Zmet2 study 11-day seedling aerial tissue genotype B73 Rep5

description: No other relevant details


title: prostatic cell line HPV7


title: C57Bl/6 2hr_mock txmt, biological rep3


title: C57Bl/6 AM 2hr_post e7 conidia, biological rep3

tag: AM collected from C57Bl/6, 2 hours after exposure to Aspergillus fumigatus conidia


title: C57Bl/6 AM 2hr_post e7 conidia, biological rep1


title: C57Bl/6 AM 2hr_post e7 conidia, biological rep4


title: C57Bl/6 AM 2hrs post e6 conidia, biological rep4


title: prostatic cell line LNCaP


title: C57Bl/6 AM 2hrs post e6 conidia, biological rep3


title: prostatic cell line WPMY1


title: prostatic cell line DU145


title: C57Bl/6 AM 2hr_post e7 conidia, biological rep2


title: prostatic cell line VCaP


title: C57Bl/6 AM time 0 control, biological rep4

treatment_protocol: Sample AM collected at 0 hours (no in vivo incubation with conidia)

tag: AM collected time 0 control


title: prostatic cell line 22Rv


title: C57Bl/6 AM time 0 control, biological rep3

source_name: Alveolar Macrophage (AM) control untreated


title: prostatic cell line PC3


title: C57Bl/6 AM 2hrs post e6 conidia, biological rep1


title: C57Bl/6 AM 2hrs post e6 conidia, biological rep2


title: C57Bl/6 AM time 0 control, biological rep2


title: Skeletal_muscle_control_rep2

status: Public on Nov 30 2007

city: Tampere

first: Alejandra

last: Rodriguez-Martinez

address: Biokatu 6

department: Institute of Biomedical Technology

country: Finland

institute: University of Tampere

description:
Strain: C57BL/6.
Gender: male.
Age: 18 weeks.
Tissue: skeletal muscle.
Second biological replicate of control group

hyb_protocol: Using Illumina reagents and protocols. Hybridization: 18h at 55 degrees C.

scan_protocol: Standard  scan factor in Illumina BeadArray reader.

data_processing: Array data were normalized with Inforsense KDE version 2.0.4 using quantile normalization method.

source_name: Skeletal muscle, control diet

label_protocol: Using Illumina TotalPrep RNA Amplification kit (#IL1791, Ambion), following the menufacturer's protocol. Input amount: 200ng of total RNA. Incubation of labeling reaction: 14h.

growth_protocol: control diet containing 0.02% of iron

extract_protocol: RNeasy RNA isolation kit including RNase-free DNase treatment (Qiagen), according to the manufacturer's protocol (Qiagen)

tag: Strain

tag: Gender

value: male.

tag: Age

value: 18 weeks.

tag: Tissue

value: skeletal muscle.


title: Skeletal_muscle_iron_rep3

description:
Strain: C57BL/6.
Gender: male.
Age: 18 weeks.
Tissue: skeletal muscle.
Third biological replicate of iron group

source_name: Skeletal muscle, iron diet

growth_protocol: iron diet: control diet supplemented with 2% carbonyl iron


title: Skeletal_muscle_control_rep1

description:
Strain: C57BL/6.
Gender: male.
Age: 18 weeks.
Tissue: skeletal muscle.
First biological replicate of control group

source_name: skeletal muscle, control diet


title: Skeletal_muscle_iron_rep2

description:
Strain: C57BL/6.
Gender: male.
Age: 18 weeks.
Tissue: skeletal muscle.
Second biological replicate of iron group


title: C57Bl/6 AM time 0 control, biological rep1


title: Sedentary vs. Exercise 4

first: Joshua

last: Hunsberger

address: 35 Convent Drive MSC3711

institute: NIH

hyb_protocol: A two-step hybridization and labeling protocol was used where the chip was hybridized to cDNA overnight, washed stringently to remove nonspecifically bound probe, and then poststained with fluorescent dendrimers.

scan_protocol: Slides were scanned using a GenePix scanner (Axon Instruments).

data_processing: Image analysis was performed using GenePix Pro 4.0 software. Resulting files from GenePix 4.0 (Axon Instruments) analysis were imported into Genespring 5.0 (Silicon Genetics) for additional visualization and data mining. Hybridization spots were considered positive only if there was a signal intensity of twice the background or more in at least one channel of half of the replicates. Per-chip normalization was performed by dividing the expressed genes by the median of two housekeeping control genes, β-tubulin and cyclophilin, that were not regulated.

source_name: Total RNA from murine hippocampal RNA labeled with Cyanine-5 (red).

label_protocol: Five micrograms of total RNA from 1 week exercise and sedentary mice (n=4 each group) was reverse-transcribed into cDNA and indirectly labeled using a sensitive fluorescent labeling procedure (Genisphere).

extract_protocol: Total hippocampal RNA from individual animals was isolated (RNA Aqueous, Ambion). Optical density values (260/280) were consistently at or over 1.9.

tag: Exercise; strain

source_name: Total RNA from murine hippocampal RNA labeled with Cyanine-3 (green).

tag: Sedentary; strain


title: Skeletal_muscle_iron_rep1

description:
Strain: C57BL/6.
Gender: male.
Age: 18 weeks.
Tissue: skeletal muscle.
First biological replicate of iron group


title: Sedentary vs. Exercise 2


title: Heart_control_rep1

description:
Strain: C57BL/6.
Gender: male.
Age: 18 weeks.
Tissue: cardiac muscle.
First biological replicate of control group

source_name: cardiac muscle, control diet

value: cardiac muscle.


title: Heart_iron_rep2

description:
Strain: C57BL/6.
Gender: male.
Age: 18 weeks.
Tissue: cardiac muscle.
Second biological replicate of iron group

source_name: cardiac muscle, iron diet


title: Sedentary vs. Exercise 3


title: Sedentary vs. Exercise 1


title: Heart_control_rep2

description:
Strain: C57BL/6.
Gender: male.
Age: 18 weeks.
Tissue: cardiac muscle.
Second biological replicate of control group


title: Heart_iron_rep3

description:
Strain: C57BL/6.
Gender: male.
Age: 18 weeks.
Tissue: cardiac muscle.
Third biological replicate of iron group

growth_protocol: iron diet:control diet supplemented with 2% carbonyl iron


title: Heart_iron_rep1

description:
Strain: C57BL/6.
Gender: male.
Age: 18 weeks.
Tissue: cardiac muscle.
First biological replicate of iron group


title: muscle_LF_ d3_biorep1

city: Maastricht

first: Janneke

last: de Wilde

address: Universiteitssingel 50

department: Human Biology

country: Netherlands

institute: Maastricht University

description: Before the diet intervention mice (age: 9 weeks) were switched to the LF run-in diet for 3 weeks

hyb_protocol: Hybridization was performed according to the manufactures instructions as described in the eukaryotic section of the GeneChip Expression Analysis Technical Manual (Affymetrix, 2004)

scan_protocol: Scanning was performed according to the manufactures instructions as described in the eukaryotic section of the GeneChip Expression Analysis Technical Manual (Affymetrix, 2004)

data_processing: Expression levels of probe sets were normalized using GC-RMA(slow) (unlogged scale). Probe sets were redefined according to Dai et al (Nucleic Acids Research, 2005). In this method probes are annotated using up to date databases and assigned to unique gene identifiers, in this case Entrez ID's, instead of the classic GeneChip probe sets. As selection criteria we used: fold change > 1.3 and raw p value < 0.05

source_name: mouse 2010, quadriceps muscle, low fat diet, day 3

label_protocol: Labeling was performed according to the manufactures instructions as described in the eukaryotic section of the GeneChip Expression Analysis Technical Manual (Affymetrix, 2004)

growth_protocol: Mice were fed a standard growth diet for 2 weeks prior to switching to the LF run-in diet for 3 weeks. Following the run-in period mice were randomly assigned to the LF or the HF group for 3 or 28 days.

extract_protocol: According the manufactures instruction, total RNA was extracted from the quadriceps muscle with Trizol reagent (Invitrogen, Breda, the Netherlands) and purified with the Rneasy Mini Kit

treatment_protocol: After 3 or 28 days of diet intervention, mice were sacrificed, the quadriceps muscle was dissected and snap-frozen in liquid nitrogen

value: C56BL/6J, Gender:Male, Age:12 weeks


title: Heart_control_rep3

description:
Strain: C57BL/6.
Gender: male.
Age: 18 weeks.
Tissue: cardiac muscle.
Third biological replicate of control group


title: muscle_HF_ d28_biorep4

source_name: mouse 1754, quadriceps muscle, high fat diet, day 28

extract_protocol:
According the manufactures instruction, total RNA was extracted from the quadriceps muscle with Trizol reagent (Invitrogen, Breda, the Netherlands) and purified with the Rneasy Mini Kit
(Qiagen, Venlo, the Netherlands). Contaminating genomic DNA was removed with the RNase-free DNase set (Qiagen, Venlo, the Netherlands) according the manufactures instructions.

value: C56BL/6J, Gender:Male, Age:16 weeks


title: muscle_HF_ d28_biorep5

source_name: mouse 1757, quadriceps muscle, high fat diet, day 28


title: muscle_HF_ d28_biorep1

source_name: mouse 1751, quadriceps muscle, high fat diet, day 28


title: muscle_HF_ d28_biorep2

source_name: mouse 1752, quadriceps muscle, high fat diet, day 28


title: muscle_HF_ d28_biorep3

source_name: mouse 1753, quadriceps muscle, high fat diet, day 28


title: muscle_LF_ d28_biorep4

source_name: mouse 2023, quadriceps muscle, low fat diet, day 28


title: muscle_LF_ d28_biorep6

source_name: mouse 2025, quadriceps muscle, low fat diet, day 28


title: muscle_LF_ d28_biorep5

source_name: mouse 2024, quadriceps muscle, low fat diet, day 28


title: non-branched in vitro-formed UBs and MM, rep2

city: La Jolla

first: Sanjay

last: Nigam

state: CA

address: 9500 Gilman Dr

department: Pediatrics

institute: University of California, San Diego

hyb_protocol: Following fragmentation, 10 ug of cRNA were hybridized for 16 hr at 45C on GeneChip Rat 230 2.0 Genome Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 450.

data_processing: The data were analyzed with GCOS using Affymetrix default analysis settings

organism: Rattus norvegicus

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

growth_protocol: Embryonic kidneys were immediately processed after dissection. Recombination was cultured for 6 days.

extract_protocol: total RNA was extracted using the Ambion Acqueous micro kit


title: Skeletal_muscle_control_rep3

description:
Strain: C57BL/6.
Gender: male.
Age: 18 weeks.
Tissue: skeletal muscle.
Third biological replicate of control group


title: muscle_LF_ d28_biorep3

source_name: mouse 2022, quadriceps muscle, low fat diet, day 28


title: muscle_LF_ d28_biorep1

source_name: mouse 2020, quadriceps muscle, low fat diet, day 28


title: muscle_HF_ d3_biorep6

source_name: mouse 1772, quadriceps muscle, high fat diet, day 3


title: HeLa cells with siRNAs specific for human hnRNPL and hnRNPLL, biological rep1

city: Giessen

first: Lee-Hsueh

last: Hung

state: Hessen

address: Heinrich-Buff-Ring 58

department: Biochemistry

country: Germany

institute: University of Giessen

description: Exon level expression

hyb_protocol: Approximately 5.5ug of labeled DNA target was hybridized to the Affymetrix GeneChip Human Exon 1.0 ST Array at 45oC for 16 hours per manufacturers recommendation (see http://www.affymetrix.com/products/arrays/exon_application.affx for additional information). Hybridized arrays were washed and stained on a GeneChip Fluidics Station 450.

scan_protocol: Arrays were scanned using the Affymetrix GCS 3000 7G and Gene-Chip Operating Software v. 1.4 to produce .CEL intensity files.

data_processing: .CEL files were background adjusted, quantile normalized netween all 12 samples and summarized in probeset level using RMAExpress. (http://rmaexpress.bmbolstad.com)

source_name: Human cervical carcinoma cell line

label_protocol: Ribosomal RNA was depleted from 1µg of total RNA using the RiboMinus Human/Mouse Transcriptome Isolation kit (Invitrogen Corp., Carlsbad, CA). cDNA was generated using the GeneChip WT cDNA Synthesis and Amplification Kit (Affymetrix, Inc., Santa Clara, CA) per manufacturer's instructions. cDNA was fragmented and end labeled using the GeneChip® WT Terminal Labeling Kit (Affymetrix, Inc.).

extract_protocol: One day before transfection, HeLa cells were seeded in 6-well culture plates (1.5 x 10e5 cells per well) or 10-cm culture dishes (4.3 x 10e5 cells per dish). siRNA duplex (at a final concentration in culture medium of 30 nM) was transfected with Lipofectamine 2000 (Invitrogen) according to the manufacturer's instructions. siRNA duplices were from MWG Biotech (Ebersberg, Germany). Four days after siRNA transfection, total RNA was isolated using Trizol (Invitrogen) and RNeasy kit (QIAGEN).

tag: siRNA, cell line


title: muscle_HF_ d3_biorep5

source_name: mouse 1771, quadriceps muscle, high fat diet, day 3


title: HeLa cells with siRNAs specific for human hnRNPL and hnRNPLL, biological rep2

description: Exon level expression


title: rat kidney at embryonic day 18, rep1

source_name: rat kidney at embryonic day 18, rep1

tag: rat kidney at embryonic day 18, rep1


title: rat kidney at week 4 post birth, rep3

source_name: rat kidney at week 4 post birth, rep3

tag: rat kidney at week 4 post birth, rep3


title: rat kidney at week 4 post birth, rep2

source_name: rat kidney at week 4 post birth, rep2

tag: rat kidney at week 4 post birth, rep2


title: HeLa cells with siRNA specific for human hnRNPLL, biological rep3

description: Exon level expression


title: muscle_HF_ d28_biorep6

source_name: mouse 1758, quadriceps muscle, high fat diet, day 28


title: HeLa cells with siRNAs specific for human hnRNPL and hnRNPLL, biological rep3

description: Exon level expression


title: muscle_HF_ d3_biorep4

source_name: mouse 1770 quadriceps muscle, high fat diet, day 3


title: rat kidney at embryonic day 18, rep3

source_name: rat kidney at embryonic day 18, rep3

tag: rat kidney at embryonic day 18, rep3


title: rat kidney at embryonic day 18, rep4

source_name: rat kidney at embryonic day 18, rep4

tag: rat kidney at embryonic day 18, rep4


title: HeLa cells with siRNA specific for human hnRNPL, biological rep2

description: Exon level expression


title: HeLa cells with siRNA specific for human hnRNPL, biological rep1

description: Exon level expression


title: HeLa cells with siRNA specific for human hnRNPLL, biological rep1

description: Exon level expression


title: Yeast replication isw2 nhp10 MMS 150min b

city: Seattle

first: Jack

middle: A.

last: Vincent

state: WA

address: 1100 Fairview Ave N

department: Basic Sciences

institute: Fred Hutchinson CRC

description:
Unreplicated vs Replicated DNA during MMS treatment
isw2 nhp10 double mutant strain
150 min time-point

hyb_protocol: • hybridization buffer: 3X SSC + 10%SDS + 1 mg/mL poly(dA) • blocking agent: no prehybridization • slide blocking: no prehybridization • probe blocking: 1mg/mL polydA during hybridization • wash procedure: wash1: 1X SSC, 0.03% SDS wash2: 1X SSC dip slides 15 times wash3: 0.2X SSC: shake slides 75rpm for 20minutes wash4: 0.05X SSC: shake slides 75rpm for 10 minutes spin-dry slides in centrifuge 50g for 5 minutes. • quantity of labelled target used: all material generated from 0.5 ug of EcoRI digested genomic DNA • time, concentration, volume, temperature: 16h, 26 microliters at 63°C • Hybridization instrument: Manual, TelChem hybridization chambers in waterbath

scan_protocol: Scanning hardware GenePix 4000B scanner (Molecular Devices) Image analysis GenePix Pro v6.0 (Molecular Devices)

data_processing:
Percent replication for each spot was determined as described (Alvino, GM et al, Mol Cell Bio, 2007). The calculation of percent replication at each time point is dependent on two measurements; the percentage of total genomic replication in the population at each time point and the number of cycling cells over the entire time course. • Total genomic % replication: The total percent replication for the genome was determined after fractionation of each CsCl gradient. Fractions were blotted to a nylon membrane and then hybridized to a radiolabeled genomic probe (as described above). Percent replication was determined by quantifying the signal from each fraction (by phosphorimager), plotting the value for each fraction, and then comparing the area under the curve corresponding to replicated and unreplicated fractions (see http://fangman-brewer.genetics.washington.edu/density_transfer.html). The total % replication for this sample was as follows: 150min 35%
• Ratio of cycling cells: The ratio of cycling cells to non-cycling cells was estimated from the maximum percentage of budded cells obtained during the time course. The maximum percent budded was 86% for isw2 nhp10 experiments.

source_name: S.cerevisiae, 150min post alpha factor, 0.015% MMS, replicated DNA

label_protocol: 0.5ug of each sample was labeled with either Cy3 or Cy5-dUTP (GE Biosciences). 21ul DNA solution was heat denatured at 100°C for 5 min then quick chilled in an ice slurry. The following was added while samples were still on ice: 5ul 10x dNTP Mix (1.2mM each dATP, dCTP, dGTP; 0.6mM dTTP; 10mM Tris 8.0), 3ul Cy-dUTP, 1ul exo- Klenow (50000 U/ml, New England Biolabs). Samples were incubated at 37°C for 1hr. Reaction was stopped by adding 5ul 0.5M EDTA. 60ul of 1mg/ml tRNA was added to each reaction and unincorporated nucleotides were removed using a Biospin 30 column (BioRad). Oppositely labeled samples (i.e. Cy3-replicated and Cy5-unreplicated) from the same timepoint were combined for cohybridization. 20ul polydA::dT (1mg/ml; Sigma) was added to each probe mix, then DNA was precipitated after addition of 1/10 volume 3M NaOAc and 2 volumes 100% ethanol.

growth_protocol: Density transfer experiments were performed essentially as described to isotopically label newly replicated DNA (see the Fangman-Brewer lab website at UW). Cells were grown a minimum of 7 generations in minimal medium containing 13C and 15N as the sole carbon and nitrogen sources (dense media). Cells were synchronized with α-factor for 105 minutes. Cells were then filtered and transferred to complete media (YC) containing 12C and 14N (light media) in the continued presence of α-factor for 75 minutes prior to release (termed conditioning phase). MMS was added to 0.015% 15 minutes into the conditioning phase resulting in a 60-minute exposure during G1 arrest. After the conditioning phase, cells were filtered, washed, and released in light media in the presence or absence of 0.015% MMS. This sample was collected at 150 min after release.

extract_protocol:
Standard Yeast genomic prep: Bead beating + phenol:chloroform extraction.
Unreplicated DNA was separated from replicated in a CsCl gradient (see the Fangman-Brewer lab website at UW.) The CsCl gradient was drip-fractionated. Fractions were then applied to slot blots and probed for DNA content. Fractions from the bottom of the gradient (the most dense fractions) contain unreplicated DNA, while fractions from the top of the gradient (least dense fractions) contain newly replicated DNA. Fractions containing unreplicated DNA (or replicated DNA) were pooled and EtOH precipitated.

tag: Collected 150 minutes post alpha factor release. Treated with MMS at 0.015%

tag: replicated DNA

source_name: S.cerevisiae, 150min post alpha factor, 0.015% MMS, unreplicated DNA

label_protocol: same as channel 1

growth_protocol: same as channel 1

extract_protocol: same as channel 1

tag: unreplicated DNA


title: muscle_HF_ d3_biorep3

source_name: mouse 1769, quadriceps muscle, high fat diet, day 3


title: Yeast replication isw2 nhp10 MMS 150min a

data_processing:
Percent replication for each spot was determined as described (Alvino, GM et al, Mol Cell Bio, 2007). The calculation of percent replication at each time point is dependent on two measurements; the percentage of total genomic replication in the population at each time point and the number of cycling cells over the entire time course. • Total genomic % replication: The total percent replication for the genome was determined after fractionation of each CsCl gradient. Fractions were blotted to a nylon membrane and then hybridized to a radiolabeled genomic probe (as described above). Percent replication was determined by quantifying the signal from each fraction (by phosphorimager), plotting the value for each fraction, and then comparing the area under the curve corresponding to replicated and unreplicated fractions (see http://fangman-brewer.genetics.washington.edu/density_transfer.html).
The total % replication for this sample was as follows: 150min 35%
• Ratio of cycling cells: The ratio of cycling cells to non-cycling cells was estimated from the maximum percentage of budded cells obtained during the time course. The maximum percent budded was 86% for isw2 nhp10 experiments.

tag: Unreplicated DNA


title: rat kidney at embryonic day 13, rep1

source_name: rat kidney at embryonic day 13, rep1

tag: rat kidney at embryonic day 13, rep1


title: HeLa cells with siRNA specific for human hnRNPL, biological rep3

description: Exon level expression


title: HeLa cells with siRNA specific for luciferase GL2, biological rep1

description: Exon level expression


title: HeLa cells with siRNA specific for human hnRNPLL, biological rep2

description: Exon level expression


title: muscle_HF_ d3_biorep1

source_name: mouse 1767, quadriceps muscle, high fat diet, day 3

tag: Strain


title: rat kidney at embryonic day 13, rep3

source_name: rat kidney at embryonic day 13, rep3

tag: rat kidney at embryonic day 13, rep3


title: rat kidney at week 4 post birth, rep1

source_name: rat kidney at week 4 post birth, rep1

tag: rat kidney at week 4 post birth, rep1


title: muscle_LF_ d28_biorep2

source_name: mouse 2021, quadriceps muscle, low fat diet, day 28


title: muscle_HF_ d3_biorep2

source_name: mouse 1768, quadriceps muscle, high fat diet, day 3


title: Yeast replication isw2 nhp10 MMS 60min b

description:
Unreplicated vs Replicated DNA during MMS treatment
isw2 nhp10 double mutant strain
60 min time-point

data_processing:
Percent replication for each spot was determined as described (Alvino, GM et al, Mol Cell Bio, 2007). The calculation of percent replication at each time point is dependent on two measurements; the percentage of total genomic replication in the population at each time point and the number of cycling cells over the entire time course. • Total genomic % replication: The total percent replication for the genome was determined after fractionation of each CsCl gradient. Fractions were blotted to a nylon membrane and then hybridized to a radiolabeled genomic probe (as described above). Percent replication was determined by quantifying the signal from each fraction (by phosphorimager), plotting the value for each fraction, and then comparing the area under the curve corresponding to replicated and unreplicated fractions (see http://fangman-brewer.genetics.washington.edu/density_transfer.html).
The total % replication for this sample was as follows: 60min 20%
• Ratio of cycling cells: The ratio of cycling cells to non-cycling cells was estimated from the maximum percentage of budded cells obtained during the time course. The maximum percent budded was 86% for isw2 nhp10 experiments.

source_name: S.cerevisiae, 60min post alpha factor, 0.015% MMS, replicated DNA

growth_protocol: Density transfer experiments were performed essentially as described to isotopically label newly replicated DNA (see the Fangman-Brewer lab website at UW). Cells were grown a minimum of 7 generations in minimal medium containing 13C and 15N as the sole carbon and nitrogen sources (dense media). Cells were synchronized with α-factor for 105 minutes. Cells were then filtered and transferred to complete media (YC) containing 12C and 14N (light media) in the continued presence of α-factor for 75 minutes prior to release (termed conditioning phase). MMS was added to 0.015% 15 minutes into the conditioning phase resulting in a 60-minute exposure during G1 arrest. After the conditioning phase, cells were filtered, washed, and released in light media in the presence or absence of 0.015% MMS. This sample was collected at 60 min after release.

tag: Collected 60 minutes post alpha factor release.

tag: Replicated DNA

source_name: S.cerevisiae, 60min post alpha factor, 0.015% MMS, unreplicated DNA


title: rat kidney at embryonic day 18, rep2

source_name: rat kidney at embryonic day 18, rep2

tag: rat kidney at embryonic day 18, rep2


title: rat kidney at embryonic day 13, rep2

source_name: rat kidney at embryonic day 13, rep2

tag: rat kidney at embryonic day 13, rep2


title: Yeast replication isw2 nhp10 MMS 120min b

description:
Unreplicated vs Replicated DNA during MMS treatment
isw2 nhp10 double mutant strain
120 min time-point

data_processing:
Percent replication for each spot was determined as described (Alvino, GM et al, Mol Cell Bio, 2007). The calculation of percent replication at each time point is dependent on two measurements; the percentage of total genomic replication in the population at each time point and the number of cycling cells over the entire time course. • Total genomic % replication: The total percent replication for the genome was determined after fractionation of each CsCl gradient. Fractions were blotted to a nylon membrane and then hybridized to a radiolabeled genomic probe (as described above). Percent replication was determined by quantifying the signal from each fraction (by phosphorimager), plotting the value for each fraction, and then comparing the area under the curve corresponding to replicated and unreplicated fractions (see http://fangman-brewer.genetics.washington.edu/density_transfer.html).
The total % replication for this sample was as follows: 120min 32%
• Ratio of cycling cells: The ratio of cycling cells to non-cycling cells was estimated from the maximum percentage of budded cells obtained during the time course. The maximum percent budded was 86% for isw2 nhp10 experiments.

source_name: S.cerevisiae, 120min post alpha factor, 0.015% MMS, replicated DNA

growth_protocol: Density transfer experiments were performed essentially as described to isotopically label newly replicated DNA (see the Fangman-Brewer lab website at UW). Cells were grown a minimum of 7 generations in minimal medium containing 13C and 15N as the sole carbon and nitrogen sources (dense media). Cells were synchronized with α-factor for 105 minutes. Cells were then filtered and transferred to complete media (YC) containing 12C and 14N (light media) in the continued presence of α-factor for 75 minutes prior to release (termed conditioning phase). MMS was added to 0.015% 15 minutes into the conditioning phase resulting in a 60-minute exposure during G1 arrest. After the conditioning phase, cells were filtered, washed, and released in light media in the presence or absence of 0.015% MMS. This sample was collected at 120 min after release.

tag: Collected 120 minutes post alpha factor release. Treated with MMS at 0.015%

source_name: S.cerevisiae, 120min post alpha factor, 0.015% MMS, unreplicated DNA


title: HeLa cells with siRNA specific for luciferase GL2, biological rep3

description: Exon level expression

source_name: Human cervical carcinoma cell line

tag: siRNA, cell line


title: HeLa cells with siRNA specific for luciferase GL2, biological rep2

description: Exon level expression


title: Yeast replication isw2 nhp10 MMS 90min b

description:
Unreplicated vs Replicated DNA during MMS treatment
isw2 nhp10 double mutant strain
90 min time-point

data_processing:
Percent replication for each spot was determined as described (Alvino, GM et al, Mol Cell Bio, 2007). The calculation of percent replication at each time point is dependent on two measurements; the percentage of total genomic replication in the population at each time point and the number of cycling cells over the entire time course. • Total genomic % replication: The total percent replication for the genome was determined after fractionation of each CsCl gradient. Fractions were blotted to a nylon membrane and then hybridized to a radiolabeled genomic probe (as described above). Percent replication was determined by quantifying the signal from each fraction (by phosphorimager), plotting the value for each fraction, and then comparing the area under the curve corresponding to replicated and unreplicated fractions (see http://fangman-brewer.genetics.washington.edu/density_transfer.html).
The total % replication for this sample was as follows: 90min 27%
• Ratio of cycling cells: The ratio of cycling cells to non-cycling cells was estimated from the maximum percentage of budded cells obtained during the time course. The maximum percent budded was 86% for isw2 nhp10 experiments.

source_name: S.cerevisiae, 90min post alpha factor, 0.015% MMS, replicated DNA

growth_protocol: Density transfer experiments were performed essentially as described to isotopically label newly replicated DNA (see the Fangman-Brewer lab website at UW). Cells were grown a minimum of 7 generations in minimal medium containing 13C and 15N as the sole carbon and nitrogen sources (dense media). Cells were synchronized with α-factor for 105 minutes. Cells were then filtered and transferred to complete media (YC) containing 12C and 14N (light media) in the continued presence of α-factor for 75 minutes prior to release (termed conditioning phase). MMS was added to 0.015% 15 minutes into the conditioning phase resulting in a 60-minute exposure during G1 arrest. After the conditioning phase, cells were filtered, washed, and released in light media in the presence or absence of 0.015% MMS. This sample was collected at 90 min after release.

tag: Collected 90 minutes post alpha factor release. Treated with MMS at 0.015%

source_name: S.cerevisiae, 90min post alpha factor, 0.015% MMS, unreplicated DNA


title: Yeast replication isw2 nhp10 MMS 120min a


title: non-treated

city: Genève

first: Jon

last: Reinders

state: Genève

address: 30 quai Ernest-Ansermet

department: Department of Plant Biology

country: Switzerland

institute: Universite de Geneve

description: one sample analysis (non-treated gDNA1, non-treated gDNA2, non-treated gDNA3)

hyb_protocol: Following a 2 hour fragmentation, 9 ug of DNA were hybridized for 16 hr at 45C on Arabidopsis Tiling 1.0R arrays . GeneChips were washed and stained as directed by Affymetrix

scan_protocol: GeneChips were scanned using the GeneChip® Scanner 3000 7G

data_processing: Arabidopsis data (CEL) files were analyzed using the Tiling Analysis Software (TAS, Affymetrix) using the following parameters. The normalization parameters were scaled to an intensity of 100 on a log2 scale and the p-value was -10log10. The probe analysis was set to a bandwidth of 80 and perfect match/mismatch intensities were analyzed using a one-sided upper test. The interval analysis on intensity values was set to a threshold of 1-e5 with a maximum gap of 80 and a minimum run to 40, resulting in a “sliding window” analysis at a 161 bp length. For analyzing methylation polymorphisms, hypo- and hypermethylated intervals were calculated using TAS. Per entry, the three biological replicate bisulfite-treated DNA files and the three biological replicate non treated DNA files were selected as the treatment and control group files, respectively, in a two sample comparison analysis using quantile normalization of both groups together. The resulting ColBS+ and met1-3BS+ BAR files, for the above two-sample comparisons, were loaded into IGB (Affymetrix). Since IGB performs only a one-sided statistical test, hypomethylated and hypermethylated comparisons were made separately. The hypomethylation graph was the difference in the log2 scaled (Col_BS+) from (met1-3_BS+) results and the hypermethylation was the difference in the log2 scaled (met1-3_BS+) from (Col_BS+). The graphs of these comparisons were adjusted to zero and the positive cutoff of 4.0 was applied. Additionally, the three CEL files per entry were analyzed in TAS using the one sample detection analysis to create a single BAR files for each entry alone. The one sample parameter were as above and allowed further visual inspection and comparison.

organism: Arabidopsis thaliana

source_name: see description for corresponding files

label_protocol: DNA was amplified using our BiMP amplification method. DNA labeling reaction with terminal deoxynucleotidyl transferase and DNA labeling reagent (Affymetrix) were added to each DNA sample

growth_protocol: Plants were grown in soil under under 8 hr light/16 hr dark. Leaf tissue from 9-12 individuals was pooled per DNA sample.  Thre replicate samples per entry were collected.

extract_protocol: For the bisulfite treatment, the EpiTect Bisulfite modification kit (Qiagen) was used as directed by the manufacturer.

treatment_protocol: CTAB extracted DNA samples were Rnase A and Proteinase K treated (Warnecke et al. 2002), phenol extracted (Sigma) and then ethanol precipitated. DNA samples (4.0μg) were digested with a 2-fold excess of DraI restriction enzyme (Promega), as recommended by the manufacturer, and phenol extracted (Sigma) and ethanol precipitated.

tag: Genotype

value: M/M; treatment:bisulfite +


title: BiMP met1-3 BS+

description: one sample analysis (met_bs+1, met_bs+2, met_bs+3)

source_name: BiMP met1-3 BS+


title: Yeast replication isw2 nhp10 MMS 90min a


title: muscle_LF_ d3_biorep5

source_name: mouse 2017, quadriceps muscle, low fat diet, day 3


title: Yeast replication isw2 nhp10 MMS 60min a

tag: Collected 60 minutes post alpha factor release. Treated with MMS at 0.015%


title: muscle_LF_ d3_biorep6

source_name: mouse 2019, quadriceps muscle, low fat diet, day 3


title: BiMP Col-0 BS+

description: one sample analysis (Col_bs+1, Col_bs+2, Col_bs+3)

source_name: BiMP Col-0 BS+


title: muscle_LF_ d3_biorep3

source_name: mouse 2014, quadriceps muscle, low fat diet, day 3


title: BiMP met1-3BS+ normalized

description: two sample analysis ((met_bs+1, met_bs+2, met_bs+3) and (non-treated gDNA1, non-treated gDNA2, non-treated gDNA3)), as experimental and control, respectively.


title: muscle_LF_ d3_biorep4

source_name: mouse 2015, quadriceps muscle, low fat diet, day 3


title: Yeast replication MMS 90min b

description:
Unreplicated vs Replicated DNA during MMS treatment
90 min time-point

data_processing:
Percent replication for each spot was determined as described (Alvino, GM et al, Mol Cell Bio, 2007). The calculation of percent replication at each time point is dependent on two measurements; the percentage of total genomic replication in the population at each time point and the number of cycling cells over the entire time course.
• Total genomic % replication:
The total percent replication for the genome was determined after fractionation of each CsCl gradient. Fractions were blotted to a nylon membrane and then hybridized to a radiolabeled genomic probe (as described above). Percent replication was determined by quantifying the signal from each fraction (by phosphorimager), plotting the value for each fraction, and then comparing the area under the curve corresponding to replicated and unreplicated fractions (see http://fangman-brewer.genetics.washington.edu/density_transfer.html). The total % replication for this sample was as follows:
90min 66%
• Ratio of cycling cells:
The ratio of cycling cells to non-cycling cells was estimated from the maximum percentage of budded cells obtained during the time course. The maximum percent budded was 94% for WT experiments.

extract_protocol:
Standard Yeast genomic prep: Bead beating + phenol:chloroform extraction.
Unreplicated DNA was separated from replicated in a CsCl gradient (see the Fangman-Brewer lab website at UW.)
The CsCl gradient was drip-fractionated. Fractions were then applied to slot blots and probed for DNA content. Fractions from the bottom of the gradient (the most dense fractions) contain unreplicated DNA, while fractions from the top of the gradient (least dense fractions) contain newly replicated DNA. Fractions containing unreplicated DNA (or replicated DNA) were pooled and EtOH precipitated.

tag: Collected 90 minutes post alpha factor release.

tag: Treated with MMS at 0.015%


title: Yeast replication MMS 120min a

description:
Unreplicated vs Replicated DNA during MMS treatment
120 min time-point

data_processing:
Percent replication for each spot was determined as described (Alvino, GM et al, Mol Cell Bio, 2007). The calculation of percent replication at each time point is dependent on two measurements; the percentage of total genomic replication in the population at each time point and the number of cycling cells over the entire time course.
• Total genomic % replication:
The total percent replication for the genome was determined after fractionation of each CsCl gradient. Fractions were blotted to a nylon membrane and then hybridized to a radiolabeled genomic probe (as described above). Percent replication was determined by quantifying the signal from each fraction (by phosphorimager), plotting the value for each fraction, and then comparing the area under the curve corresponding to replicated and unreplicated fractions (see http://fangman-brewer.genetics.washington.edu/density_transfer.html). The total % replication for this sample was as follows:
120min 81%
• Ratio of cycling cells:
The ratio of cycling cells to non-cycling cells was estimated from the maximum percentage of budded cells obtained during the time course. The maximum percent budded was 94% for WT experiments.

tag: Collected 120 minutes post alpha factor release.

tag: replicated DNA


title: Yeast replication isw2 nhp10 MMS 45min a

description:
Unreplicated vs Replicated DNA during MMS treatment
isw2 nhp10 double mutant strain
45 min time-point

data_processing:
Percent replication for each spot was determined as described (Alvino, GM et al, Mol Cell Bio, 2007). The calculation of percent replication at each time point is dependent on two measurements; the percentage of total genomic replication in the population at each time point and the number of cycling cells over the entire time course.
• Total genomic % replication:
The total percent replication for the genome was determined after fractionation of each CsCl gradient. Fractions were blotted to a nylon membrane and then hybridized to a radiolabeled genomic probe (as described above). Percent replication was determined by quantifying the signal from each fraction (by phosphorimager), plotting the value for each fraction, and then comparing the area under the curve corresponding to replicated and unreplicated fractions (see http://fangman-brewer.genetics.washington.edu/density_transfer.html). The total % replication for this sample was as follows:
45min 14%
• Ratio of cycling cells:
The ratio of cycling cells to non-cycling cells was estimated from the maximum percentage of budded cells obtained during the time course. The maximum percent budded was 86% for isw2 nhp10 experiments.

source_name: S.cerevisiae, 45min post alpha factor, 0.015% MMS, unreplicated DNA

growth_protocol: Density transfer experiments were performed essentially as described to isotopically label newly replicated DNA (see the Fangman-Brewer lab website at UW). Cells were grown a minimum of 7 generations in minimal medium containing 13C and 15N as the sole carbon and nitrogen sources (dense media). Cells were synchronized with α-factor for 105 minutes. Cells were then filtered and transferred to complete media (YC) containing 12C and 14N (light media) in the continued presence of α-factor for 75 minutes prior to release (termed conditioning phase). MMS was added to 0.015% 15 minutes into the conditioning phase resulting in a 60-minute exposure during G1 arrest. After the conditioning phase, cells were filtered, washed, and released in light media in the presence or absence of 0.015% MMS. This sample was collected at 45 min after release.

tag: Collected 45 minutes post alpha factor release.

tag: Unreplicated DNA

source_name: S.cerevisiae, 45min post alpha factor, 0.015% MMS, replicated DNA


title: Yeast replication MMS 120min b

tag: Strain

tag: unreplicated DNA


title: non-branched in vitro-formed UBs and MM, rep1


title: Yeast replication MMS 60min b

description:
Unreplicated vs Replicated DNA during MMS treatment
60 min time-point

hyb_protocol: Hybridization protocol 	• hybridization buffer: 3X SSC + 10%SDS + 1 mg/mL poly(dA) • blocking agent: no prehybridization • slide blocking: no prehybridization • probe blocking: 1mg/mL polydA during hybridization • wash procedure: wash1: 1X SSC, 0.03% SDS wash2: 1X SSC dip slides 15 times wash3: 0.2X SSC: shake slides 75rpm for 20minutes wash4: 0.05X SSC: shake slides 75rpm for 10 minutes spin-dry slides in centrifuge 50g for 5 minutes. • quantity of labelled target used: all material generated from 0.5 ug of EcoRI digested genomic DNA • time, concentration, volume, temperature: 16h, 26 microliters at 63°C • Hybridization instrument: Manual, TelChem hybridization chambers in waterbath

data_processing:
Percent replication for each spot was determined as described (Alvino, GM et al, Mol Cell Bio, 2007). The calculation of percent replication at each time point is dependent on two measurements; the percentage of total genomic replication in the population at each time point and the number of cycling cells over the entire time course.
• Total genomic % replication:
The total percent replication for the genome was determined after fractionation of each CsCl gradient. Fractions were blotted to a nylon membrane and then hybridized to a radiolabeled genomic probe (as described above). Percent replication was determined by quantifying the signal from each fraction (by phosphorimager), plotting the value for each fraction, and then comparing the area under the curve corresponding to replicated and unreplicated fractions (see http://fangman-brewer.genetics.washington.edu/density_transfer.html). The total % replication for this sample was as follows:
60min 45%
• Ratio of cycling cells:
The ratio of cycling cells to non-cycling cells was estimated from the maximum percentage of budded cells obtained during the time course. The maximum percent budded was 94% for WT experiments.

source_name: S.cerevisiae, 60min post alpha factor, 0.015% MMS, replicated DNA

growth_protocol: Growth protocol 	Density transfer experiments were performed essentially as described to isotopically label newly replicated DNA (see the Fangman-Brewer lab website at UW). Cells were grown a minimum of 7 generations in minimal medium containing 13C and 15N as the sole carbon and nitrogen sources (dense media). Cells were synchronized with α-factor for 105 minutes. Cells were then filtered and transferred to complete media (YC) containing 12C and 14N (light media) in the continued presence of α-factor for 75 minutes prior to release (termed conditioning phase). MMS was added to 0.015% 15 minutes into the conditioning phase resulting in a 60-minute exposure during G1 arrest. After the conditioning phase, cells were filtered, washed, and released in light media in the presence or absence of 0.015% MMS. This sample was collected at 60 min after release.

extract_protocol:
Standard Yeast genomic prep: Bead beating + phenol:chloroform extraction.
Unreplicated DNA was separated from replicated DNA in a CsCl gradient (see the Fangman-Brewer lab website at UW.)
The CsCl gradient was drip-fractionated. Fractions were then applied to slot blots and probed for DNA content. Fractions from the bottom of the gradient (the most dense fractions) contain unreplicated DNA, while fractions from the top of the gradient (least dense fractions) contain newly replicated DNA. Fractions containing unreplicated DNA (or replicated DNA) were pooled and EtOH precipitated.

tag: Collected 60 minutes post alpha factor release.

source_name: S.cerevisiae, 60min post alpha factor, 0.015% MMS, unreplicated DNA


title: Yeast replication MMS 60min a


title: muscle_LF_ d3_biorep2

source_name: mouse 2012, quadriceps muscle, low fat diet, day 3