title: Mock DNase-Seq 20 min Cort

city: Bethesda

first: Songjoon

last: Baek

state: MD

address: 41 Library Drive

department: CCR

institute: NCI / NIH

description: DNase-seq in cells Mock treatment 600 nM Cort for 20 min

data_processing:
Illumina Casava 1.8 software was used for basecalling.
Areas of enrichment were identified using DNase2Hotspots software(htrceforge.net/projects/dnase2hotspots/) software with FDR 0%. Peaks with the maximum tag density lower than 10 were eliminated.
Raw signal density graphs were generated by counting tags at each genomic coordinate after extending the length of sequences to 150bp by strands.
Genome_build: mm9 (NCBI 37)
Supplementary_files_format_and_content: comma-delimited (CSV) text files include genomic locations (Chromosome, start, end ), z-score, p-value and the maximum density of hotspots. BedGraph is a file format that allows display of tag density profiles (signal maps) in a genome browser track

source_name: 3134 cells

growth_protocol: For maintenance cells were cultured in DMEM supplemented with 10% calf serum, sodium pyruvate, and penicillin-streptomycin cocktail.

extract_protocol:
Lysates were clarified from sonicated nuclei.
DHS-seq: Nuclei were isolated  and subjected to a limited DNase I digestion. Small DNA fragments where DNase I cut twice were isolated and prepared for sequencing according to the manufacturer (Illumina).

treatment_protocol: Cells were plated for experiments in DMEM growth medium supplemented with 10% charcoal-dextran treated serum 48 hours prior to conducting the experiment.  Cells were untreated or induced with 600nM Corticosterone, 100nM dexamethasone, or Mock treated.

tag: cell type

value: mouse mammary cells

value: 600 nM Cort for 20 min


title: Mock DNase-Seq 0 min Cort

status: Public on Feb 12 2015

type: SRA

country: USA

description: DNase-Seq in untreated cells

library_source: genomic

molecule: genomic DNA

organism: Mus musculus

tag: cell line

tag: treatment

value: none (untreated cells)


title: DNase-Seq 60 min Cort Pulsed

description: DNase-seq in cells treatment 600 nM Cort for 20 min + 40 min in hormone-free medium


title: DNase-Seq 0 min Cort

description: DNase-seq in untreated cells


title: Input Pol II

description: Combined Input DNA from treated and untreated 3134 cells

data_processing:
Illumina Casava 1.8 software was used for basecalling.
Areas of enrichment were identified using DNase2Hotspots software(htrceforge.net/projects/dnase2hotspots/) software with FDR 0%.   Peaks with the maximum tag density lower than 10 were eliminated.
Raw signal density graphs were generated by counting tags at each genomic coordinate after extending the length of sequences to 150bp by strands.
Genome_build: mm9 (NCBI 37)
Supplementary_files_format_and_content: comma-delimited (CSV)  text files include genomic locations (Chromosome, start,  end ), z-score, p-value and the maximum density of hotspots.

source_name: Input Pol II

extract_protocol:
Lysates were clarified from sonicated nuclei and GR-DNA complexes, Pol II-DNA complexes or CTCF-DNA complexes were isolated with antibody.
NEB Chip-Seq Prep Kit

tag: treated with

value: Combined (treated and untreated)


title: CTCF ChIP-Seq 0 min Cort

description: CTCF-ChIP in untreated cells

source_name: CTCF ChIP-Seq 0 min Cort

tag: chip antibody

value: anti-CTCF antibody

tag: chip antibody vendor

value: Millipore

tag: chip antibody cat. #


title: RpoN_RpoS_control_sample1

city: Braunschweig

first: Denitsa

last: Eckweiler

state: Lower Saxony

address: Spielmannstr. 7

department: Institute of Microbiology

country: Germany

institute: Technische Universität Braunschweig

description: ChIP-seq control rep1

data_processing:
Sequenced ChIP-seq reads were trimmed for adaptor sequence allowing for a minimal quality of 10 at their ends, then mapped to the PA14 genome using bowtie v0.12.7 with default parameters
ChIP-seq peaks were called using the SAM files and the MACS algorithm with the following options: -g 5200000 –nomodel --shiftsize=30 -p 0.05
Comparisons were performed in the following way: sample1 vs control1; sample2 vs control2
Genome_build: gi|116048575|ref|NC_008463.1|
Supplementary_files_format_and_content: The MACS output files contain the coordinates of the enriched genomic regions as well as their P-values, false discovery rate (FDR) and fold enrichment

organism: Pseudomonas aeruginosa

growth_protocol: PA14 wild type harboring a plasmid encoding for the polyhistidine-tagged sigma factor gene was cultivated under optimal conditions regarding sigma factor activity. In addition, sigma factor expression was strongly induced by adding L-arabinose (0.5%) to the culture for at least 45 minutes prior to formaldehyde-mediated cross-linking.

extract_protocol:
Polyhistidine-tagged sigma-factors were isolated by using anti6Xhis-antibodies (abcam) and magnetic beads coupled with protein-G (Invitrogen) on a rotator.
Immunoprecipitated DNA was linear amplified similar to the LinDA-ChIP-seq protocol published by Shankaranarayanan et al., Nature potocols, 2012. Library was constructed using the TruSeq DNA sample preparation kit according to the low-throughput protocol (Illumina).

treatment_protocol: DNA-protein crosslinking was performed by addition of formaldehyde to a final concentration of 0.5% for 5 minutes at ambient temperature. Reaction was quenched by addition of glycine to a final concentration of 137 mM. Cells were lysed in lysis buffer containing EDTA and lysozyme. Cells were disrupted and DNA was sheared by sonication.

tag: strain

tag: genotype/background


title: Mock DNase-seq 60 min Cort Constant

description: DNase-seq in cells Mock treatment 600 nM Cort for 60 min

value: 600 nM Cort for 60 min


title: Mock DNase-Seq 60 min Cort Pulsed

description: DNase-seq in cells Mock treatment 600 nM Cort for 20 min + 40 min in hormone-free medium


title: DNase-Seq 20 min Cort

description: DNase-seq in cells treatment 600 nM Cort for 20 min


title: Pol II ChIP-Seq Mock 60 min Cort Pulsed 2

description: Pol II-ChIP in cells Mock treated with 600 nM Cort for 20 min + 40 min in hormone-free medium

source_name: Pol II ChIP-Seq Mock 60 min Cort Pulsed

value: RNA polymerase II phosphoS5 antibody

value: Abcam


title: Pol II ChIP-Seq Mock 0 min Cort 2

description: Pol II-ChIP in Mock untreated cells

source_name: Pol II ChIP-Seq Mock 0 min Cort

value: Mock untreated cells


title: Pol II ChIP-Seq Mock 60 min Cort Constant 1

description: Pol II-ChIP in cells Mock treated with 600 nM Cort for 60 min


title: Pol II ChIP-Seq Mock 60 min Cort Pulsed 1

value: 600 nM Cort for 20 min + 40 min in hormone-free medium


title: Pol II ChIP-Seq Mock 0 min Cort 1

description: Pol II-ChIP in Mock untreated cells


title: FecI_knockout_mutant_sample2

description:
deletion mutant rep2
processed data file: FecI_knockout_vs_wild_type.csv

library_source: transcriptomic

data_processing:
The untrimmed RNA-seq reads were mapped to the PA14 genome using the stampy aligner with default settings.
Differential expression was estimated using the DESeq R library using a pre-filtering step. The overexpressing samples were used together with the empty vector controls for negative binomial test. Similarly, the relevant controls for the knockout mutant were the wildtype samples.
For detection of differentially expressed genes, both replicates for each condition were used, thus the corresponding data were generated on the basis of 4 samples for each comparison. Details can be found in the supplementary table and in the quality control file.
Quality control of the replicates and a summary of the samples used in the different comparisons are provided as supplementary information.
Genome_build: gi|116048575|ref|NC_008463.1|
Supplementary_files_format_and_content: DESeq output in CSV format presenting the differential gene expression after discarding rRNA/tRNA genes and the bottom 40% genes sorted by decreasing variance

molecule: total RNA

growth_protocol: Sigma factor hyper-expressing strains harboring the sigma factor gene in trans under control of the araBAD promoter and sigma factor deletion mutants were constructed. Under optimal conditions regarding sigma factor activity and optional induction of sigma factor expression, bacteria were harvested and total RNA was extracted.

extract_protocol:
Using Rneasy kit (Qiagen) plus Qiashredder columns.
Customized protocol (Dötsch et al., PloS ONE, 2012). Briefly, rRNA depletion using MICROBExpress kit with the Pseudomonas module (Ambion) and fragmentation by sonication using Covaris Adaptive Acoustics device (Covaris). Upon ligation of barcoded adapters, reverse transcription with SuperScript II kit (Invitrogen) followed. Amplification was done by PCR using Illumina platform compatible primers. Established cDNA library was normalized by double strand specific nuclease treatment (Evrogen) according DSN Normalization Sample Preparation Guide from Illumina.

treatment_protocol: Cells from 3 pooled cultures were harvested by addition of the same volume of RNA-protect buffer (Qiagen) and centrifugation.

tag: strain background

tag: genotype/variation

tag: sample type

value: PA14 bacterial culture


title: Pol II ChIP-Seq Mock 20 min Cort 2

description: Pol II-ChIP in cells Mock treated with 600 nM Cort for 20 min

source_name: Pol II ChIP-Seq Mock 20 min Cort


title: Pol II ChIP-Seq 60 min Cort Constant

description: Pol II-ChIP in cells treated with 600 nM Cort for 60 min

source_name: Pol II ChIP-Seq 60 min Cort Constant


title: Pol II ChIP-Seq Mock 20 min Cort 1


title: Pol II ChIP-Seq 20 min Cort

description: Pol II-ChIP in cells treated with 600 nM Cort for 20 min

source_name: Pol II ChIP-Seq 20 min Cort


title: GR ChIP-Seq input 60 min Cort Pulsed

description: Input DNA from cells treated with 600 nM Cort for 20 min + 40 min in hormone-free medium

source_name: GR ChIP-Seq input 60 min Cort Pulsed


title: GR ChIP-Seq input 20 min Cort

description: Input DNA from cells treated with 600 nM Cort for 20 min

source_name: GR ChIP-Seq input 20 min Cort


title: Pol II ChIP-Seq 0 min Cort

description: Pol II-ChIP in untreated cells

source_name: Pol II ChIP-Seq 0 min Cort


title: DNase-60 min Cort Constant

description: DNase-seq in cells treatment 600 nM Cort for 60 min


title: GR ChIP-Seq input 60 min Cort Constant

description: Input DNA from cells treated with 600 nM Cort for 60 min

source_name: GR ChIP-Seq input 60 min Cort Constant


title: GR ChIP-Seq 60 min Cort Pulsed

description:
cocktail of PA-510A and PA-511A from Affinity BioReagents and sc-1004 from Santa Cruz
GR-ChIP in cells treated with 600 nM Cort for 20 min + 40 min in hormone-free medium

source_name: GR ChIP-Seq 60 min Cort Pulsed

value: GR Antibody

value: Affinity BioReagents; Santa Cruz


title: GR ChIP-Seq 20 min Cort

description:
cocktail of PA-510A and PA-511A from Affinity BioReagents and sc-1004 from Santa Cruz
GR-ChIP in cells treated with 600 nM Cort for 20 min

source_name: GR ChIP-Seq 20 min Cort


title: GR ChIP-Seq 0 min Cort

description:
cocktail of PA-510A and PA-511A from Affinity BioReagents and sc-1004 from Santa Cruz
GR-ChIP in untreated cells

source_name: GR ChIP-Seq 0 min Cort


title: SigX_wildtype_sample1

description: deletion control rep1


title: SigX_knockout_mutant_new_sample2

description:
deletion mutant rep2
processed data file: SigX_knockout_vs_wild_type_new.csv


title: SigX_knockout_mutant_new_sample1

description:
deletion mutant rep1
processed data file: SigX_knockout_vs_wild_type_new.csv


title: Pol II ChIP-Seq Mock 60 min Cort Constant 2


title: RpoH_overexpressed_sample2

description:
overexpressing wildtype rep2
processed data file: RpoH_overexpression_vs_empty_vector.csv


title: RpoN_knockout_mutant_nitrogen_starvation_sample2

description:
deletion mutant rep2
processed data file: RpoN_knockout_vs_wild_type_nitrogen_starvation.csv


title: RpoN_knockout_mutant_nitrogen_starvation_sample1

description:
deletion mutant rep1
processed data file: RpoN_knockout_vs_wild_type_nitrogen_starvation.csv


title: RpoN_wildtype_nitrogen_starvation_sample2

description: deletion control rep2


title: RpoN_wildtype_nitrogen_starvation_sample1

description: deletion control rep1


title: Pol II ChIP-Seq 60 min Cort Pulsed

description: Pol II-ChIP in cells treated with 600 nM Cort for 20 min + 40 min in hormone-free medium

source_name: Pol II ChIP-Seq 60 min Cort Pulsed


title: SigX_chip_sample2

description: ChIP-seq treated rep2

value: harboring a plasmid encoding for the polyhistidine-tagged sigma factor gene

value: Anti-6X His tag  antibody - ChIP Grade

value: abcam

tag: chip antibody lot #


title: RpoH_overexpressed_sample1

description:
overexpressing wildtype rep1
processed data file: RpoH_overexpression_vs_empty_vector.csv


title: RpoN_RpoS_control_sample2

description: ChIP-seq control rep2


title: RpoS_chip_sample2

description: ChIP-seq treated rep2


title: SigX_control_sample2

description: ChIP-seq control rep2


title: RpoH_control_sample2

description: ChIP-seq control rep2


title: RpoH_empty_vector_sample2

description: overexpressing control rep2


title: RpoN_chip_sample2

description: ChIP-seq treated rep2


title: RpoH_chip_sample2

description: ChIP-seq treated rep2


title: RpoD_FliA_control_sample2

description: ChIP-seq control rep2


title: GR ChIP-Seq 60 min Cort Constant

description:
cocktail of PA-510A and PA-511A from Affinity BioReagents and sc-1004 from Santa Cruz
GR-ChIP in cells treated with 600 nM Cort for 60 min

source_name: GR ChIP-Seq 60 min Cort Constant

value: mouse mammary cells


title: RpoD_chip_sample2

description: ChIP-seq treated rep2


title: PvdS_chip_sample2

description: ChIP-seq treated rep2


title: GR ChIP-Seq input 0 min Cort

description: Input DNA from untreated cells

source_name: GR ChIP-Seq input 0 min Cort


title: FpvI_chip_sample2

description: ChIP-seq treated rep2


title: FpvI_chip_sample1

description: ChIP-seq treated rep1


title: FecI_chip_sample2

description: ChIP-seq treated rep2


title: FliA_chip_sample2

description: ChIP-seq treated rep2


title: AlgU_control_sample2

description: ChIP-seq control rep2


title: FecI2_chip_sample1

description: ChIP-seq treated rep1


title: AlgU_chip_sample2

description: ChIP-seq treated rep2


title: HeLa_CenpA_(20140605_2,7)

city: Seattle

first: Jorja

last: Henikoff

state: WA

address: 1100 Fairview AV N, A1-162

department: Basic Sciences

institute: Fred Hutchinson Cancer Research Center

description: HeLa cross-linked (X-ChIP) CENP-A ChIP

data_processing: Paired reads were merged using SeqPrep (https://github.com/jstjohn/SeqPrep) with parameters -q 30 (quality) and -L 25 (minimum merged pair length). SeqPrep also removed adaptors and low-quality reads.

organism: Homo sapiens

source_name: Immunoprecipitated DNA

extract_protocol: The HeLa cell line was grown in DMEM media using strandard protocols. Cross-linking ChIP was performed as described previously (PMID:24737864).

tag: cell

value: nuclei

value: Epithelial carcinoma

value: HeLa

value: Anti-CENP-A (Abcam Ab13939)


title: RpoH_empty_vector_sample1

description: overexpressing control rep1


title: HeLa_Input_(20140605_1,6)

description: HeLa cross-linked (X-ChIP) input

source_name: HeLa input

extract_protocol: The HeLa cell line was grown in DMEM media using strandard protocols. Input DNA was prepared from cross-linked cells as described previously (PMID:24737864).


title: HuRef_M2_B_(20140605_2,11)

description: HuRef native FLAG ChIP using KAPA polymerase.

extract_protocol: The HuRef lymphoblastoid cell line was grown in RPMI medium using standard protocols. Nuclei were prepared following a previously published protocol (PMID:23644596). Briefly, for each IP 40-60 million cells were collected and washed with 1X PBS. Cells were re-suspended in ice-cold buffer I (0.32 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) at a density equal to ~25 million cells/ml. An equal volume of ice-cold buffer I supplemented with 0.1% NP40 was added and samples were incubated on ice for 10 min. Nuclei solution was layered on ice-cold buffer III (1.2 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) and centrifuged at 10,000g for 20 min at 4 degrees C. The pellet was re-suspended in buffer A (0.34 M sucrose, 15 mM HEPES, pH 7.4, 15 mM NaCl, 60 mM KCl, 4 mM MgCl2, 1 mM DTT, 0.1 mM PMSF and protease inhibitor, 3mM Cacl2) at density equal to ~ 32.5 million cell/ml.  Micrococcal nuclease (MNase, Sigma, Cat # N3755) was added to ~2.5 units/ml, and digestion was carried out at 37 degrees C for 5 min. Reactions were stopped by addition of EGTA to a final concentration of 20 mM and EDTA to 10 mM. The final NaCl concentration was adjusted to 215 mM and needle extraction was performed as described previously (PMID:22184235) to enhance the solubility of kinetochore complex. The resulting solution was incubated overnight at 4 degrees C on nutator. Soluble chromatin was collected by centrifuging the mixture at 12000rpm at 4 degrees C for 8 min. Soluble chromatin was diluted 3X with 20 mM Tris, pH 8.0, 5 mM EDTA and 200 mM NaCl, and Triton-X was added at a final concentration of 0.1% (v/v). Next, the chromatin solution was pre-cleared using Protein A/G fast flow Sepharose beads for 20 min at 4 degrees C, and 15 mcg of antibody (Sigma anit-FLAG F1804) was added per ChIP sample and incubated overnight at 4 degrees C. Dynabeads were added to the samples and incubated at 4 degrees C for 2 hrs. Immunoprecipitated complexes were washed 6 times with 50mM Phosphate buffer pH 7.4, 5mM EDTA, 200mM NaCl, and DNA was extracted from Dynabeads as described (PMID:22184235). Solexa library construction was performed as described previously (PMID:22025700) and PCR-amplified using KAPA polymerase.

value: Lymphoblastoid

value: HuRef

value: Anti-FLAG (Sigma cat #F1804)


title: HuRef_H3_A_(20140605_1,10)

description: HuRef native H3 ChIP using Phusion polymerase.

extract_protocol: The HuRef lymphoblastoid cell line was grown in RPMI medium using standard protocols. Antibodies used in this study were purchased from Abcam (Cambridge MA). Nuclei were prepared following a previously published protocol (PMID:23644596). Briefly, for each IP 40-60 million cells were collected and washed with 1X PBS. Cells were re-suspended in ice-cold buffer I (0.32 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) at a density equal to ~25 million cells/ml. An equal volume of ice-cold buffer I supplemented with 0.1% NP40 was added and samples were incubated on ice for 10 min. Nuclei solution was layered on ice-cold buffer III (1.2 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) and centrifuged at 10,000g for 20 min at 4 degrees C. The pellet was re-suspended in buffer A (0.34 M sucrose, 15 mM HEPES, pH 7.4, 15 mM NaCl, 60 mM KCl, 4 mM MgCl2, 1 mM DTT, 0.1 mM PMSF and protease inhibitor, 3mM Cacl2) at density equal to ~ 32.5 million cell/ml.  Micrococcal nuclease (MNase, Sigma, Cat # N3755) was added to ~2.5 units/ml, and digestion was carried out at 37 degrees C for 5 min. Reactions were stopped by addition of EGTA to a final concentration of 20 mM and EDTA to 10 mM. The final NaCl concentration was adjusted to 215 mM and needle extraction was performed as described previously (PMID:22184235) to enhance the solubility of kinetochore complex. The resulting solution was incubated overnight at 4 degrees C on nutator. Soluble chromatin was collected by centrifuging the mixture at 12000rpm at 4 degrees C for 8 min. Soluble chromatin was diluted 3X with 20 mM Tris, pH 8.0, 5 mM EDTA and 200 mM NaCl, and Triton-X was added at a final concentration of 0.1% (v/v). Next, the chromatin solution was pre-cleared using Protein A/G fast flow Sepharose beads for 20 min at 4 degrees C, and 15 mcg of antibody (Abcam anti-H3 Ab1791) was added per ChIP sample and incubated overnight at 4 degrees C. Dynabeads were added to the samples and incubated at 4 degrees C for 2 hrs. Immunoprecipitated complexes were washed 6 times with 50mM Phosphate buffer pH 7.4, 5mM EDTA, 200mM NaCl, and DNA was extracted from Dynabeads as described (PMID:22184235). Solexa library construction was performed as described previously (PMID:22025700) and PCR-amplified using Phusion polymerase.

value: Anti-H3 (Abcam cat #Ab1791)


title: HuRef_M2_A_(20140605_1,11)

description: HuRef native FLAG ChIP using Phusion polymerase.

extract_protocol: The HuRef lymphoblastoid cell line was grown in RPMI medium using standard protocols. Nuclei were prepared following a previously published protocol (PMID:23644596). Briefly, for each IP 40-60 million cells were collected and washed with 1X PBS. Cells were re-suspended in ice-cold buffer I (0.32 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) at a density equal to ~25 million cells/ml. An equal volume of ice-cold buffer I supplemented with 0.1% NP40 was added and samples were incubated on ice for 10 min. Nuclei solution was layered on ice-cold buffer III (1.2 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) and centrifuged at 10,000g for 20 min at 4 degrees C. The pellet was re-suspended in buffer A (0.34 M sucrose, 15 mM HEPES, pH 7.4, 15 mM NaCl, 60 mM KCl, 4 mM MgCl2, 1 mM DTT, 0.1 mM PMSF and protease inhibitor, 3mM Cacl2) at density equal to ~ 32.5 million cell/ml.  Micrococcal nuclease (MNase, Sigma, Cat # N3755) was added to ~2.5 units/ml, and digestion was carried out at 37 degrees C for 5 min. Reactions were stopped by addition of EGTA to a final concentration of 20 mM and EDTA to 10 mM. The final NaCl concentration was adjusted to 215 mM and needle extraction was performed as described previously (PMID:22184235) to enhance the solubility of kinetochore complex. The resulting solution was incubated overnight at 4 degrees C on nutator. Soluble chromatin was collected by centrifuging the mixture at 12000rpm at 4 degrees C for 8 min. Soluble chromatin was diluted 3X with 20 mM Tris, pH 8.0, 5 mM EDTA and 200 mM NaCl, and Triton-X was added at a final concentration of 0.1% (v/v). Next, the chromatin solution was pre-cleared using Protein A/G fast flow Sepharose beads for 20 min at 4 degrees C, and 15 mcg of antibody (Sigma anit-FLAG F1804) was added per ChIP sample and incubated overnight at 4 degrees C. Dynabeads were added to the samples and incubated at 4 degrees C for 2 hrs. Immunoprecipitated complexes were washed 6 times with 50mM Phosphate buffer pH 7.4, 5mM EDTA, 200mM NaCl, and DNA was extracted from Dynabeads as described (PMID:22184235). Solexa library construction was performed as described previously (PMID:22025700) and PCR-amplified using Phusion polymerase.


title: HuRef_H3_B_(20140605_2,10)

description: HuRef native H3 ChIP using KAPA polymerase.

extract_protocol: The HuRef lymphoblastoid cell line was grown in RPMI medium using standard protocols. Antibodies used in this study were purchased from Abcam (Cambridge MA). Nuclei were prepared following a previously published protocol (PMID:23644596). Briefly, for each IP 40-60 million cells were collected and washed with 1X PBS. Cells were re-suspended in ice-cold buffer I (0.32 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) at a density equal to ~25 million cells/ml. An equal volume of ice-cold buffer I supplemented with 0.1% NP40 was added and samples were incubated on ice for 10 min. Nuclei solution was layered on ice-cold buffer III (1.2 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) and centrifuged at 10,000g for 20 min at 4 degrees C. The pellet was re-suspended in buffer A (0.34 M sucrose, 15 mM HEPES, pH 7.4, 15 mM NaCl, 60 mM KCl, 4 mM MgCl2, 1 mM DTT, 0.1 mM PMSF and protease inhibitor, 3mM Cacl2) at density equal to ~ 32.5 million cell/ml.  Micrococcal nuclease (MNase, Sigma, Cat # N3755) was added to ~2.5 units/ml, and digestion was carried out at 37 degrees C for 5 min. Reactions were stopped by addition of EGTA to a final concentration of 20 mM and EDTA to 10 mM. The final NaCl concentration was adjusted to 215 mM and needle extraction was performed as described previously (PMID:22184235) to enhance the solubility of kinetochore complex. The resulting solution was incubated overnight at 4 degrees C on nutator. Soluble chromatin was collected by centrifuging the mixture at 12000rpm at 4 degrees C for 8 min. Soluble chromatin was diluted 3X with 20 mM Tris, pH 8.0, 5 mM EDTA and 200 mM NaCl, and Triton-X was added at a final concentration of 0.1% (v/v). Next, the chromatin solution was pre-cleared using Protein A/G fast flow Sepharose beads for 20 min at 4 degrees C, and 15 mcg of antibody (Abcam anti-H3 Ab1791) was added per ChIP sample and incubated overnight at 4 degrees C. Dynabeads were added to the samples and incubated at 4 degrees C for 2 hrs. Immunoprecipitated complexes were washed 6 times with 50mM Phosphate buffer pH 7.4, 5mM EDTA, 200mM NaCl, and DNA was extracted from Dynabeads as described (PMID:22184235). Solexa library construction was performed as described previously (PMID:22025700) and PCR-amplified using KAPA polymerase.


title: PvdS_control_sample2

description: ChIP-seq control rep2


title: HuRef_CenpC_2A_(20140605_1,9)

description: HuRef native CENP-C ChIP. Biological replicate 2 using Phusion polymerase.

extract_protocol: The HuRef lymphoblastoid cell line was grown in RPMI medium using standard protocols. Antibodies used in this study were purchased from Abcam (Cambridge MA). Nuclei were prepared following a previously published protocol (PMID:23644596). Briefly, for each IP 40-60 million cells were collected and washed with 1X PBS. Cells were re-suspended in ice-cold buffer I (0.32 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) at a density equal to ~25 million cells/ml. An equal volume of ice-cold buffer I supplemented with 0.1% NP40 was added and samples were incubated on ice for 10 min. Nuclei solution was layered on ice-cold buffer III (1.2 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) and centrifuged at 10,000g for 20 min at 4 degrees C. The pellet was re-suspended in buffer A (0.34 M sucrose, 15 mM HEPES, pH 7.4, 15 mM NaCl, 60 mM KCl, 4 mM MgCl2, 1 mM DTT, 0.1 mM PMSF and protease inhibitor, 3mM Cacl2) at density equal to ~ 32.5 million cell/ml.  Micrococcal nuclease (MNase, Sigma, Cat # N3755) was added to ~2.5 units/ml, and digestion was carried out at 37 degrees C for 5 min. Reactions were stopped by addition of EGTA to a final concentration of 20 mM and EDTA to 10 mM. The final NaCl concentration was adjusted to 215 mM and needle extraction was performed as described previously (PMID:22184235) to enhance the solubility of kinetochore complex. The resulting solution was incubated overnight at 4 degrees C on nutator. Soluble chromatin was collected by centrifuging the mixture at 12000rpm at 4 degrees C for 8 min. Soluble chromatin was diluted 3X with 20 mM Tris, pH 8.0, 5 mM EDTA and 200 mM NaCl, and Triton-X was added at a final concentration of 0.1% (v/v). Next, the chromatin solution was pre-cleared using Protein A/G fast flow Sepharose beads for 20 min at 4 degrees C, and 15 mcg of antibody (Abcam anti-CENP-C Cat # 33034) was added per ChIP sample and incubated overnight at 4 degrees C. Dynabeads were added to the samples and incubated at 4 degrees C for 2 hrs. Immunoprecipitated complexes were washed 6 times with 50mM Phosphate buffer pH 7.4, 5mM EDTA, 200mM NaCl, and DNA was extracted from Dynabeads as described (PMID:22184235). Solexa library construction was performed as described previously (PMID:22025700) and PCR-amplified using Phusion polymerase.

value: Anti-CENP-A (Abcam cat #Ab13939)


title: HuRef_CenpC_1B_(20140605_2,4)

description: HuRef native CENP-C ChIP. Biological replicate 1 using KAPA polymerase.

extract_protocol: The HuRef lymphoblastoid cell line was grown in RPMI medium using standard protocols. Antibodies used in this study were purchased from Abcam (Cambridge MA). Nuclei were prepared following a previously published protocol (PMID:23644596). Briefly, for each IP 40-60 million cells were collected and washed with 1X PBS. Cells were re-suspended in ice-cold buffer I (0.32 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) at a density equal to ~25 million cells/ml. An equal volume of ice-cold buffer I supplemented with 0.1% NP40 was added and samples were incubated on ice for 10 min. Nuclei solution was layered on ice-cold buffer III (1.2 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) and centrifuged at 10,000g for 20 min at 4 degrees C. The pellet was re-suspended in buffer A (0.34 M sucrose, 15 mM HEPES, pH 7.4, 15 mM NaCl, 60 mM KCl, 4 mM MgCl2, 1 mM DTT, 0.1 mM PMSF and protease inhibitor, 3mM Cacl2) at density equal to ~ 32.5 million cell/ml.  Micrococcal nuclease (MNase, Sigma, Cat # N3755) was added to ~2.5 units/ml, and digestion was carried out at 37 degrees C for 5 min. Reactions were stopped by addition of EGTA to a final concentration of 20 mM and EDTA to 10 mM. The final NaCl concentration was adjusted to 215 mM and needle extraction was performed as described previously (PMID:22184235) to enhance the solubility of kinetochore complex. The resulting solution was incubated overnight at 4 degrees C on nutator. Soluble chromatin was collected by centrifuging the mixture at 12000rpm at 4 degrees C for 8 min. Soluble chromatin was diluted 3X with 20 mM Tris, pH 8.0, 5 mM EDTA and 200 mM NaCl, and Triton-X was added at a final concentration of 0.1% (v/v). Next, the chromatin solution was pre-cleared using Protein A/G fast flow Sepharose beads for 20 min at 4 degrees C, and 15 mcg of antibody (Abcam anti-CENP-C Cat # 33034) was added per ChIP sample and incubated overnight at 4 degrees C. Dynabeads were added to the samples and incubated at 4 degrees C for 2 hrs. Immunoprecipitated complexes were washed 6 times with 50mM Phosphate buffer pH 7.4, 5mM EDTA, 200mM NaCl, and DNA was extracted from Dynabeads as described (PMID:22184235). Solexa library construction was performed as described previously (PMID:22025700) and PCR-amplified using KAPA polymerase.


title: HuRef_CenpA_2A_(20140605_1,8)

description: HuRef native CENP-A ChIP. Biological replicate 2 using Phusion polymerase.

extract_protocol: The HuRef lymphoblastoid cell line was grown in RPMI medium using standard protocols. Antibodies used in this study were purchased from Abcam (Cambridge MA). Nuclei were prepared following a previously published protocol (PMID:23644596). Briefly, for each IP 40-60 million cells were collected and washed with 1X PBS. Cells were re-suspended in ice-cold buffer I (0.32 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) at a density equal to ~25 million cells/ml. An equal volume of ice-cold buffer I supplemented with 0.1% NP40 was added and samples were incubated on ice for 10 min. Nuclei solution was layered on ice-cold buffer III (1.2 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) and centrifuged at 10,000g for 20 min at 4 degrees C. The pellet was re-suspended in buffer A (0.34 M sucrose, 15 mM HEPES, pH 7.4, 15 mM NaCl, 60 mM KCl, 4 mM MgCl2, 1 mM DTT, 0.1 mM PMSF and protease inhibitor, 3mM Cacl2) at density equal to ~ 32.5 million cell/ml.  Micrococcal nuclease (MNase, Sigma, Cat # N3755) was added to ~2.5 units/ml, and digestion was carried out at 37 degrees C for 5 min. Reactions were stopped by addition of EGTA to a final concentration of 20 mM and EDTA to 10 mM. The final NaCl concentration was adjusted to 215 mM and needle extraction was performed as described previously (PMID:22184235) to enhance the solubility of kinetochore complex. The resulting solution was incubated overnight at 4 degrees C on nutator. Soluble chromatin was collected by centrifuging the mixture at 12000rpm at 4 degrees C for 8 min. Soluble chromatin was diluted 3X with 20 mM Tris, pH 8.0, 5 mM EDTA and 200 mM NaCl, and Triton-X was added at a final concentration of 0.1% (v/v). Next, the chromatin solution was pre-cleared using Protein A/G fast flow Sepharose beads for 20 min at 4 degrees C, and 15 mcg of antibody (Abcam anti-CENP-A Ab13939) was added per ChIP sample and incubated overnight at 4 degrees C. Dynabeads were added to the samples and incubated at 4 degrees C for 2 hrs. Immunoprecipitated complexes were washed 6 times with 50mM Phosphate buffer pH 7.4, 5mM EDTA, 200mM NaCl, and DNA was extracted from Dynabeads as described (PMID:22184235). Solexa library construction was performed as described previously (PMID:22025700) and PCR-amplified using Phusion polymerase.


title: HuRef_CenpC_1A_(20140605_1,4)

description: HuRef native CENP-C ChIP. Biological replicate 1 using Phusion polymerase.


title: HuRef_CenpC_2B_(20140605_2,9)

description: HuRef native CENP-C ChIP. Biological replicate 2 using KAPA polymerase.


title: HuRef_CenpA_1A_(20140605_1,2)

description: HuRef native CENP-A ChIP. Biological replicate 1 using Phusion polymerase.


title: HuRef_Input_2B_(20140605_2,5)

description: HuRef native ChIP input. Biological replicate 2 using KAPA polymerase.

source_name: HuRef input

extract_protocol: The HuRef lymphoblastoid cell line was grown in RPMI medium using standard protocols. Antibodies used in this study were purchased from Abcam (Cambridge MA). Nuclei were prepared following a previously published protocol (PMID:23644596). Briefly, for each IP 40-60 million cells were collected and washed with 1X PBS. Cells were re-suspended in ice-cold buffer I (0.32 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) at a density equal to ~25 million cells/ml. An equal volume of ice-cold buffer I supplemented with 0.1% NP40 was added and samples were incubated on ice for 10 min. Nuclei solution was layered on ice-cold buffer III (1.2 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) and centrifuged at 10,000g for 20 min at 4 degrees C. The pellet was re-suspended in buffer A (0.34 M sucrose, 15 mM HEPES, pH 7.4, 15 mM NaCl, 60 mM KCl, 4 mM MgCl2, 1 mM DTT, 0.1 mM PMSF and protease inhibitor, 3mM Cacl2) at density equal to ~ 32.5 million cell/ml.  Micrococcal nuclease (MNase, Sigma, Cat # N3755) was added to ~2.5 units/ml, and digestion was carried out at 37 degrees C for 5 min. Reactions were stopped by addition of EGTA to a final concentration of 20 mM and EDTA to 10 mM. The final NaCl concentration was adjusted to 215 mM and needle extraction was performed as described previously (PMID:22184235) to enhance the solubility of kinetochore complex. The resulting solution was incubated overnight at 4 degrees C on nutator. Soluble chromatin was collected by centrifuging the mixture at 12000rpm at 4 degrees C for 8 min. An aliquot containing 1/20th of the entire chromatin solution was subjected to DNA extraction. Solexa library construction was performed as described previously (PMID:22025200) and PCR-amplified using KAPA polymerase.


title: WT EB CapSeq

city: Pittsburgh

first: Sarah

middle: J

last: Hainer

state: PA

address: 4249 Fifth Ave

department: Department of Biological Sciecnes

institute: University of Pittsburgh

data_processing:
Library strategy: CAP-seq
Remove 5' barcode and 3' barcode (or called linker)
Map the reads to mm9 genome and annotation
A custom PERL pipeline is used to summarize reads and generate histogram at single nt level.
Generic genome browse is used to visualize the data
Genome_build: mm9

source_name: murine embryoid bodies (day 12)

growth_protocol: E14 mouse embryonic stem cells (ESCs) were cultured on gelatin-coated dishes without feeder cells in high-glucose DMEM medium (Sigma) containing 10% serum (Corning) and LIF at 37°C with 5% CO2. Embryoid bodies (EBs) were grown by removing LIF from ESC medium and plating on uncoated petri dishes. Cells were maintained in Medium devoid of LIF for 12 days and harvested.

extract_protocol:
Small RNAs of from total RNAs were resolved on a 15% polyacrylamide 7M Urea Gel, along with 2 picomoles of RNA standard (18- and 40-nt) in separate lanes. SYBR gold staining was used to visualize the RNA standards. A gel fragment was excised from the sample lanes in the migration range between the two standards. RNA was eluted from the gel fragment in (0.3M NaCl-TE pH7.5) solution overnight and isopropanol-precipitated with 20mg of glycogen as the carrier. The gel purified RNA was incubated with 0.25 U/µl TAP (Epicentre) and 1U/µl SuperRNaseIN (Ambion) in 10µl reaction containing 1X buffer at 37 oC for 1 hr, followed by phenol-chloroform extraction and isopropanol precipitation. The TAP-treated samples were then incubated with 5µM of 3’-end linker (1, 2, 3 from IDT DNA Technologies), 1 Unit/µl of SuperRNaseIN, 10% DMSO and 1 U/µl T4 RNA ligase (Takara) in 10µl ligation buffer (50mM Tris-Cl pH7.5, 10mM MgCl2,10mg/mL BSA, 10mM DTT). The 3’ ligated products were gel purified, eluted, precipitated and then incubated with 5 µM of 5’ adapter oligonucleotide with 4nt barcode at the 5' end, 1 Unit/µl SuperRNaseIN (Ambion) and 2 U/µl of T4 RNA ligase in ligation buffer (50mM Tris-HCI pH7.5), 10mM MgCI2, 10mM DTT, 1mM ATP) and 10% DMSO. The ligated products were gel purified and reverse transcribed in a standard 20µl reaction (SuperScript III, Invitrogen). The cDNA was amplified by PCR and purified in a 8% acrylamide gel. PCR products generated for all the samples were sequenced on a Solexa sequencing HiSeq 2000 platform (Illumina, Inc.) and single-end 100 nt reads were obtained.
small RNA cloning, 5' and 3' ligation dependent

value: Embryoid bodies


title: HuRef_Input_1A_(20140605_1,1)

description: HuRef native ChIP input. Biological replicate 1 using Phusion polymerase.

extract_protocol: The HuRef lymphoblastoid cell line was grown in RPMI medium using standard protocols. Antibodies used in this study were purchased from Abcam (Cambridge MA). Nuclei were prepared following a previously published protocol (PMID:23644596). Briefly, for each IP 40-60 million cells were collected and washed with 1X PBS. Cells were re-suspended in ice-cold buffer I (0.32 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) at a density equal to ~25 million cells/ml. An equal volume of ice-cold buffer I supplemented with 0.1% NP40 was added and samples were incubated on ice for 10 min. Nuclei solution was layered on ice-cold buffer III (1.2 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) and centrifuged at 10,000g for 20 min at 4 degrees C. The pellet was re-suspended in buffer A (0.34 M sucrose, 15 mM HEPES, pH 7.4, 15 mM NaCl, 60 mM KCl, 4 mM MgCl2, 1 mM DTT, 0.1 mM PMSF and protease inhibitor, 3mM Cacl2) at density equal to ~ 32.5 million cell/ml.  Micrococcal nuclease (MNase, Sigma, Cat # N3755) was added to ~2.5 units/ml, and digestion was carried out at 37 degrees C for 5 min. Reactions were stopped by addition of EGTA to a final concentration of 20 mM and EDTA to 10 mM. The final NaCl concentration was adjusted to 215 mM and needle extraction was performed as described previously (PMID:22184235) to enhance the solubility of kinetochore complex. The resulting solution was incubated overnight at 4 degrees C on nutator. Soluble chromatin was collected by centrifuging the mixture at 12000rpm at 4 degrees C for 8 min. An aliquot containing 1/20th of the entire chromatin solution was subjected to DNA extraction. Solexa library construction was performed as described previously (PMID:22025200) and PCR-amplified using Phusion polymerase.


title: HuRef_Input_2A_(20140605_1,5)

description: HuRef native ChIP input. Biological replicate 2 using Phusion polymerase.


title: HuRef_CenpA_1B_(20140605_2,2)

description: HuRef native CENP-A ChIP. Biological replicate 1 using KAPA polymerase.

extract_protocol: The HuRef lymphoblastoid cell line was grown in RPMI medium using standard protocols. Antibodies used in this study were purchased from Abcam (Cambridge MA). Nuclei were prepared following a previously published protocol (PMID:23644596). Briefly, for each IP 40-60 million cells were collected and washed with 1X PBS. Cells were re-suspended in ice-cold buffer I (0.32 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) at a density equal to ~25 million cells/ml. An equal volume of ice-cold buffer I supplemented with 0.1% NP40 was added and samples were incubated on ice for 10 min. Nuclei solution was layered on ice-cold buffer III (1.2 M sucrose, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 15 mM Tris, pH 7.5, 0.5 mM DTT, 0.1 mM PMSF and protease inhibitor) and centrifuged at 10,000g for 20 min at 4 degrees C. The pellet was re-suspended in buffer A (0.34 M sucrose, 15 mM HEPES, pH 7.4, 15 mM NaCl, 60 mM KCl, 4 mM MgCl2, 1 mM DTT, 0.1 mM PMSF and protease inhibitor, 3mM Cacl2) at density equal to ~ 32.5 million cell/ml.  Micrococcal nuclease (MNase, Sigma, Cat # N3755) was added to ~2.5 units/ml, and digestion was carried out at 37 degrees C for 5 min. Reactions were stopped by addition of EGTA to a final concentration of 20 mM and EDTA to 10 mM. The final NaCl concentration was adjusted to 215 mM and needle extraction was performed as described previously (PMID:22184235) to enhance the solubility of kinetochore complex. The resulting solution was incubated overnight at 4 degrees C on nutator. Soluble chromatin was collected by centrifuging the mixture at 12000rpm at 4 degrees C for 8 min. Soluble chromatin was diluted 3X with 20 mM Tris, pH 8.0, 5 mM EDTA and 200 mM NaCl, and Triton-X was added at a final concentration of 0.1% (v/v). Next, the chromatin solution was pre-cleared using Protein A/G fast flow Sepharose beads for 20 min at 4 degrees C, and 15 mcg of antibody (Abcam anti-CENP-A Ab13939) was added per ChIP sample and incubated overnight at 4 degrees C. Dynabeads were added to the samples and incubated at 4 degrees C for 2 hrs. Immunoprecipitated complexes were washed 6 times with 50mM Phosphate buffer pH 7.4, 5mM EDTA, 200mM NaCl, and DNA was extracted from Dynabeads as described (PMID:22184235). Solexa library construction was performed as described previously (PMID:22025700) and PCR-amplified using KAPA polymerase.


title: FecI_chip_sample1

description: ChIP-seq treated rep1


title: HuRef_Input_1B_(20140605_2,1)

description: HuRef native ChIP input. Biological replicate 1 using KAPA polymerase.


title: WT ESC CapSeq

source_name: murine embryonic stem cells

growth_protocol: E14 mouse embryonic stem cells (ESCs) were cultured on gelatin-coated dishes without feeder cells in high-glucose DMEM medium (Sigma) containing 10% serum (Corning) and LIF at 37°C with 5% CO2.

value: embryonic stem cells


title: HuRef_CenpA_2B_(20140605_2,8)

description: HuRef native CENP-A ChIP. Biological replicate 2 using KAPA polymerase.


title: Mbd3 KD ESCs_MNase-seq

data_processing:
Paired end reads were collapsed and adapter sequences were removed from fastq files. Reads were mapped to the mouse mm9 genome using Bowtie2 and only uniquely mapped reads with zero, one, or two mismatches were used for further analysis.  The read size distribution was determined for each library and reads were separated into nucleosome sized fragments (135-165). To calculate occupancy around TSS, DHS, or transcription factor sites, seqMINER was used to sum read occurrence either 2000 bp upstream and downstream, 1000 bp upstream and downstream, or 500 bp upstream and downstream of the reference sequence. Occurrences were binned in 20 bp intervals and reads were normalized to the average genome-wide coverage. TSS reference sites were used based on mm9 TSS coordinates. DHS reference sites were based on mouse ENCODE data (GSM1014154). Coordinates of DHSs outside of TSSs were obtained by subtraction of called TSSs from ENCODE DHS coordinates (mm9).
Genome_build: mm9

growth_protocol: E14 mouse embryonic stem cells (ESCs) were cultured on gelatin-coated dishes without feeder cells in high-glucose DMEM medium (Sigma) containing 10% serum (Corning) and LIF at 37°C with 5% CO2

extract_protocol:
ES cells were crosslinked with formaldehyde (Sigma) to a final concentration of 1%, incubated at room temperature for 10 min and quenched with glycine to a final concentration of 125 mM. Cells were washed with ice cold PBS and pelleted. Cell pellets were resuspended in lysis buffer (10 mM Tris [pH 7.5], 10 mM NaCl, 2 mM MgCl2, 0.5% NP-40, 1 mM CaCl2) with protease inhibitors (Roche) and incubated for 15 min at 4°C. Permeabilized cells were treated with ~10 Units/106 cells of micrococcal nuclease (Roche) for 5 min at 37°C. Reactions were stopped with the addition of EDTA. Control samples with no MNase addition were treated similarly. Samples were incubated for 4 hr at 4°C with rotation with RNaseA/T1 (Ambion) then incubated overnight at 65°C with 0.01% SDS and proteinase K (Ambion). DNA was extracted with PCI using phase-lock tubes, ethanol precipitated, and resuspended in 30 μL TE.
Samples were treated with CIP, end-repaired, A-tailed, and adaptor-ligated as described (Illumina). Between each step, DNA was cleaned with PCI extraction and ethanol precipitation. After adaptor ligation, DNA was selected and purified with Agencourt Ampure beads and then PCR amplified with KAPA HiFi polymerase using 16 cycles of PCR.

treatment_protocol: KD with siRNA

tag: knockdown

value: Mbd3 KD by siRNA


title: HeLa_CenpC_(20140605_2,12)

description: HeLa cross-linked (X-ChIP) CENP-C ChIP

value: Anti-CENP-C (Abcam Ab33034)


title: Smarca4 KD ESCs_MNase-seq

value: Smarca4 KD by siRNA


title: Smarca4 KD MEFs_CAP-seq_R2

data_processing:
Fastq files from single read libraries were collapsed and split based on the 5  barcode in the linker sequence. The 5  barcode and 3  linker sequences were then trimmed from reads and reads were mapped to the mouse mm9 genome using Bowtie1 with reads at least 19 nt long (although most were 60 nt   80 nt long). The mutation rate allowed for the alignment was 0 for reads 19-24 nt, 1 for 25-29 nt, 2 for 30-39 nt, 3 for 40-49 nt, 4 for 50-59 nt, and 5 for 60-69 nt, and 6 for 70 nt or bigger. Reads were separated into RNA classes including: coding, lincRNAs, miRNAs, piRNAs, and  structural  RNAs (tRNAs, rRNAs, snRNAs, and snoRNAs) based on Ensembl data, miRBase 18 and non-coding RNA database fRNAdb 2.4. Read counts were normalized to parts per million non-structural RNA reads and a gff2 file was generated to visualize alignments on a Genome Browser Gbrowse 1.70.
To map reads based on DHS location, a custom PERL script was used to search an interval of 500 nt upstream to 500 nt downstream of a given read for the nearest gene, using the DHS locations obtained from the ENCODE data set (GSM1014154) with TSS locations removed. Using all reads called in this interval, similar analysis was performed as described above for antisense reads initiating from annotated TSSs.
To map reads within coding genes, a custom PERL script was used to search an interval of 500 nt downstream to 2000 nt downstream of annotated TSS locations for a given read for the nearest gene. Based on the orientation of the 3  end of the CapSeq read, the read was assigned as either sense or antisense to the coding gene. Using the sense and antisense reads as separate groups, similar analysis was performed as described above.
To map reads based on TTS location, a custom PERL script was used to search an interval of 300 nt upstream to 300 nt downstream of a given read for the nearest TTS. Based on the orientation of the 3  end of the CapSeq read, the read was assigned as either sense or antisense to the TTS. Using only the antisense reads, similar analysis was performed to that described above.
To analyze changes in transcription start site usage, only uniquely mapped, 5' perfectly matched, CapSeq reads of at least 30 nt were included in this analysis. RNA reads were normalized to 10 million sense protein coding reads, and a histogram for the 5  ends of mapped reads was generated for each sample. Sites with less than 5 reads were removed and sites were also removed if they were at least 10-fold less than the upstream or downstream neighboring gene as these sites could be generated by sequencing errors of the nearby abundant reads.
To compare antisense TSS CapSeq reads with MNase-Seq reads upstream of the NDR, CapSeq reads arising in the antisense direction from 500 nt upstream to 100 nt downstream of annotated TSSs were called if there were 5 or more reads in the averaged EGFP KD or Smarca4 KD ESC CapSeq datasets for which there were also MNase-Seq reads. MNase-Seq reads were averaged -500 bp to -200 bp upstream of the annotated TSS for either EGFP KD or Smarca4 KD. For both the CapSeq and MNase-Seq datasets, the Smarca4 KD data was divided by the EGFP KD data, and the log2 (ratio) was calculated. The data was clustered using Cluster 3.0 and visualized by Java TreeView.
Genome_build: mm9
Note: Fastq file contains 5' barcode sequence 'GCTG'

source_name: mouse embryonic fibroblasts

growth_protocol: T-antigen immortalized mouse fibroblasts (MEFs) were cultured in high-glucose DMEM containing 10% serum at 37°C with 5% CO2.

extract_protocol:
Small RNAs of from total RNAs were resolved on a 15% polyacrylamide 7M Urea Gel, along with 2 picomoles of RNA standard (18- and 40-nt) in separate lanes. SYBR gold staining was used to visualize the RNA standards. A gel fragment was excised from the sample lanes in the migration range between the two standards. RNA was eluted from the gel fragment in (0.3M NaCl-TE pH7.5) solution overnight and isopropanol-precipitated with 20mg of glycogen as the carrier. The gel purified RNA was incubated with 0.25 U/µl TAP (Epicentre) and  1U/µl SuperRNaseIN (Ambion) in 10µl reaction containing 1X buffer at 37 oC for 1 hr, followed by phenol-chloroform extraction and isopropanol precipitation. The TAP-treated samples were then incubated with 5µM of 3’-end linker (1, 2, 3 from IDT DNA Technologies), 1 Unit/µl of SuperRNaseIN, 10% DMSO and 1 U/µl T4 RNA ligase (Takara) in 10µl ligation buffer (50mM Tris-Cl pH7.5, 10mM MgCl2,10mg/mL BSA, 10mM DTT).  The 3’ ligated products were gel purified, eluted, precipitated and then incubated with 5 µM of 5’ adapter oligonucleotide with 4nt barcode at the 5' end, 1 Unit/µl SuperRNaseIN (Ambion) and 2 U/µl of T4 RNA ligase in ligation buffer (50mM Tris-HCI pH7.5), 10mM MgCI2, 10mM DTT, 1mM ATP) and 10% DMSO. The ligated products were gel purified and reverse transcribed in a standard 20µl reaction (SuperScript III, Invitrogen). The cDNA was amplified by PCR and purified in a 8% acrylamide gel. PCR products generated for all the samples were sequenced on a Solexa sequencing HiSeq 2000 platform (Illumina, Inc.) and single-end 100 nt reads were obtained.
small RNA cloning, 5' and 3' ligation dependent

treatment_protocol: KD with esiRNA

value: Smarca4 KD by esiRNA

value: T-antigen immortalized from primary MEFs

value: embryonic fibroblasts


title: EGFP KD ESCs_MNase-seq

value: EGFP KD by siRNA


title: FecI2_chip_sample2

description: ChIP-seq treated rep2

extract_protocol:
Polyhistidine-tagged sigma-factors were isolated by using anti6Xhis-antibodies (abcam) and magnetic beads coupled with protein-G (Invitrogen) on a rotator.
Immunoprecipitated DNA was linear amplified similar to the LinDA-ChIP-seq protocol published by Shankaranarayanan et al., Nature potocols, 2012. Library was constructed using the TruSeq DNA sample preparation kit according to the low-throughput protocol (Illumina).

value: harboring a plasmid encoding for the polyhistidine-tagged sigma factor gene

value: Anti-6X His tag  antibody - ChIP Grade

value: abcam


title: Smarca4 KD MEFs_CAP-seq_R1

data_processing:
Fastq files from single read libraries were collapsed and split based on the 5  barcode in the linker sequence. The 5  barcode and 3  linker sequences were then trimmed from reads and reads were mapped to the mouse mm9 genome using Bowtie1 with reads at least 19 nt long (although most were 60 nt   80 nt long). The mutation rate allowed for the alignment was 0 for reads 19-24 nt, 1 for 25-29 nt, 2 for 30-39 nt, 3 for 40-49 nt, 4 for 50-59 nt, and 5 for 60-69 nt, and 6 for 70 nt or bigger. Reads were separated into RNA classes including: coding, lincRNAs, miRNAs, piRNAs, and  structural  RNAs (tRNAs, rRNAs, snRNAs, and snoRNAs) based on Ensembl data, miRBase 18 and non-coding RNA database fRNAdb 2.4. Read counts were normalized to parts per million non-structural RNA reads and a gff2 file was generated to visualize alignments on a Genome Browser Gbrowse 1.70.
To map reads based on DHS location, a custom PERL script was used to search an interval of 500 nt upstream to 500 nt downstream of a given read for the nearest gene, using the DHS locations obtained from the ENCODE data set (GSM1014154) with TSS locations removed. Using all reads called in this interval, similar analysis was performed as described above for antisense reads initiating from annotated TSSs.
To map reads within coding genes, a custom PERL script was used to search an interval of 500 nt downstream to 2000 nt downstream of annotated TSS locations for a given read for the nearest gene. Based on the orientation of the 3  end of the CapSeq read, the read was assigned as either sense or antisense to the coding gene. Using the sense and antisense reads as separate groups, similar analysis was performed as described above.
To map reads based on TTS location, a custom PERL script was used to search an interval of 300 nt upstream to 300 nt downstream of a given read for the nearest TTS. Based on the orientation of the 3  end of the CapSeq read, the read was assigned as either sense or antisense to the TTS. Using only the antisense reads, similar analysis was performed to that described above.
To analyze changes in transcription start site usage, only uniquely mapped, 5' perfectly matched, CapSeq reads of at least 30 nt were included in this analysis. RNA reads were normalized to 10 million sense protein coding reads, and a histogram for the 5  ends of mapped reads was generated for each sample. Sites with less than 5 reads were removed and sites were also removed if they were at least 10-fold less than the upstream or downstream neighboring gene as these sites could be generated by sequencing errors of the nearby abundant reads.
To compare antisense TSS CapSeq reads with MNase-Seq reads upstream of the NDR, CapSeq reads arising in the antisense direction from 500 nt upstream to 100 nt downstream of annotated TSSs were called if there were 5 or more reads in the averaged EGFP KD or Smarca4 KD ESC CapSeq datasets for which there were also MNase-Seq reads. MNase-Seq reads were averaged -500 bp to -200 bp upstream of the annotated TSS for either EGFP KD or Smarca4 KD. For both the CapSeq and MNase-Seq datasets, the Smarca4 KD data was divided by the EGFP KD data, and the log2 (ratio) was calculated. The data was clustered using Cluster 3.0 and visualized by Java TreeView.
Genome_build: mm9
Note: Fastq file contains 5' barcode sequence 'ATCA'


title: EGFP KD MEF_CAP-seq_R2

data_processing:
Fastq files from single read libraries were collapsed and split based on the 5  barcode in the linker sequence. The 5  barcode and 3  linker sequences were then trimmed from reads and reads were mapped to the mouse mm9 genome using Bowtie1 with reads at least 19 nt long (although most were 60 nt   80 nt long). The mutation rate allowed for the alignment was 0 for reads 19-24 nt, 1 for 25-29 nt, 2 for 30-39 nt, 3 for 40-49 nt, 4 for 50-59 nt, and 5 for 60-69 nt, and 6 for 70 nt or bigger. Reads were separated into RNA classes including: coding, lincRNAs, miRNAs, piRNAs, and  structural  RNAs (tRNAs, rRNAs, snRNAs, and snoRNAs) based on Ensembl data, miRBase 18 and non-coding RNA database fRNAdb 2.4. Read counts were normalized to parts per million non-structural RNA reads and a gff2 file was generated to visualize alignments on a Genome Browser Gbrowse 1.70.
To map reads based on DHS location, a custom PERL script was used to search an interval of 500 nt upstream to 500 nt downstream of a given read for the nearest gene, using the DHS locations obtained from the ENCODE data set (GSM1014154) with TSS locations removed. Using all reads called in this interval, similar analysis was performed as described above for antisense reads initiating from annotated TSSs.
To map reads within coding genes, a custom PERL script was used to search an interval of 500 nt downstream to 2000 nt downstream of annotated TSS locations for a given read for the nearest gene. Based on the orientation of the 3  end of the CapSeq read, the read was assigned as either sense or antisense to the coding gene. Using the sense and antisense reads as separate groups, similar analysis was performed as described above.
To map reads based on TTS location, a custom PERL script was used to search an interval of 300 nt upstream to 300 nt downstream of a given read for the nearest TTS. Based on the orientation of the 3  end of the CapSeq read, the read was assigned as either sense or antisense to the TTS. Using only the antisense reads, similar analysis was performed to that described above.
To analyze changes in transcription start site usage, only uniquely mapped, 5' perfectly matched, CapSeq reads of at least 30 nt were included in this analysis. RNA reads were normalized to 10 million sense protein coding reads, and a histogram for the 5  ends of mapped reads was generated for each sample. Sites with less than 5 reads were removed and sites were also removed if they were at least 10-fold less than the upstream or downstream neighboring gene as these sites could be generated by sequencing errors of the nearby abundant reads.
To compare antisense TSS CapSeq reads with MNase-Seq reads upstream of the NDR, CapSeq reads arising in the antisense direction from 500 nt upstream to 100 nt downstream of annotated TSSs were called if there were 5 or more reads in the averaged EGFP KD or Smarca4 KD ESC CapSeq datasets for which there were also MNase-Seq reads. MNase-Seq reads were averaged -500 bp to -200 bp upstream of the annotated TSS for either EGFP KD or Smarca4 KD. For both the CapSeq and MNase-Seq datasets, the Smarca4 KD data was divided by the EGFP KD data, and the log2 (ratio) was calculated. The data was clustered using Cluster 3.0 and visualized by Java TreeView.
Genome_build: mm9
Note: Fastq file contains 5' barcode sequence 'TGAC'

value: EGFP KD by esiRNA


title: EGFP KD MEF_CAP-seq_R1


title: Smarca4 KD ESCs_CAP-seq_R2

treatment_protocol: KD with esiRNAs

value: Smarca4 KD by esiRNAs


title: Smarca4 KD ESCs_CAP-seq_R1


title: EGFP KD ESCs_CAP-seq_R1

value: EGFP KD by esiRNAs


title: EGFP KD ESCs_RNA-seq_R1

data_processing:
Fastq files from single read libraries were collapsed and split based on the 5  barcode in the linker sequence. The 5  barcode sequences were then trimmed from reads and reads were mapped to the mouse mm9 genome using Bowtie1 with reads at least 19 nt long. Read counts were normalized to parts per million reads.
To map reads based on TSS location, a custom PERL script was used to search an interval of 2000 nt upstream to 500 nt downstream of a given read for the nearest gene. Antisense reads were then binned, averaged for the two RNA-Seq libraries, and called if there were 5 reads or more for one of the KD datasets. The reads from these libraries were then sorted from lowest to highest in Smarca4 KD and visualized through Java TreeView. To map reads based on DHS location, a custom PERL script was used to search an interval of 1000 nt upstream to 1000 nt downstream of a given read for the nearest gene. All reads were then binned, averaged for the two RNA-Seq libraries, and called if there were 5 reads or more for one of the KD datasets. The reads from these libraries were clustered using using Cluster 3.0 and visualized through Java TreeView.
Genome_build: mm9
Note: Fastq file contains 5' barcode sequence 'TCGT'

extract_protocol:
RNA was isolated with TRIzol reagent, as per the manufacturers instructions.
5 μg of total RNA was depleted of rRNA using a Ribo-Zero Gold kit (Epicentre) and first strand cDNA was made using 200 ng RNA, 3 ug random hexamers, 1 μg Superscript III, and standard dNTPs in the presence of Actinomycin D. First strand cDNA was purified by PCI extraction, ethanol precipitated, and resuspended in water. Second strand cDNA was synthesized with E. coli DNA polymerase, in the presence of E. coli DNA ligase and RNase H (all from NEB), using dNTPs in which dUTP was substituted for dTTP, at 16°C for 2 hrs. After purification on a Clean and Concentrate column (Zymo Research), cDNA was fragmented using Fragmentase (NEB), PCI extracted and precipitated as above. End repair, A-tailing, and barcoded adapter ligation were performed as described (Illumina), and the second strand cDNA was digested with USER enzyme (NEB). Libraries were amplified from the resulting adapter-ligated first-strand cDNA using Phusion (NEB) and 16 cycles of amplification. Libraries were size-selected on 1.5% agarose gels to a size range of ~180-250 bp.


title: Smarca4 KD ESCs_RNA-seq_R1

data_processing:
Fastq files from single read libraries were collapsed and split based on the 5  barcode in the linker sequence. The 5  barcode sequences were then trimmed from reads and reads were mapped to the mouse mm9 genome using Bowtie1 with reads at least 19 nt long. Read counts were normalized to parts per million reads.
To map reads based on TSS location, a custom PERL script was used to search an interval of 2000 nt upstream to 500 nt downstream of a given read for the nearest gene. Antisense reads were then binned, averaged for the two RNA-Seq libraries, and called if there were 5 reads or more for one of the KD datasets. The reads from these libraries were then sorted and visualized through Java TreeView. To map reads based on DHS location, a custom PERL script was used to search an interval of 1000 nt upstream to 1000 nt downstream of a given read for the nearest gene. All reads were then binned, averaged for the two RNA-Seq libraries, and called if there were 5 reads or more for one of the KD datasets. The reads from these libraries were then sorted and visualized through Java TreeView.
Genome_build: mm9
Note: Fastq file contains 5' barcode sequence 'ACGT'


title: Smc1a Input

city: London

first: Gopuraja

last: Dharmalingam

address: Faculty of Medicine, Imperial College, Hammersmith Hospital Campus

department: Epigenetics section

country: United Kingdom

institute: MRC London Institute of Medical Sciences

data_processing:
Raw reads were aligned to the mm9 mouse genome assembly using Bowtie version 0.12.8 (Langmead et al., 2008) with parameters “–m 1 –v 2”.
Peaks were called using MACS with default parameters. Wiggle files were produced by MACS.
Genome_build: mm9
Supplementary_files_format_and_content: MACS-generated bed and wig files.

source_name: CD4+CD8+ double positive thymocytes

growth_protocol: 6 weeks old mice

extract_protocol:
DP thymocytes from control and Rad21-/- mice were FACS-sorted, fixed and subsequently lysed and sonicated to extract soluble chromatin. H3K27Ac was immunoprecipitated using polyclonal antibodies. Smc1a was immunoprecipitated from WT samples only.
Library was constructed using NEBNext ChIP-Seq Library Prep Kit (NEB, catalog number E6200S), according to the manufacturer's instructions

treatment_protocol: Conditional deletion of Rad21 in DP thymocytes

tag: age

value: 6 weeks old

value: CD4+ CD8+ double positive (DP) thymocyte

value: Wild type

tag: chip antibody target

value: Control

value: none (input)


title: Smc1a ChIP-seq Rep2

tag: cell type

value: Anti-Smc1a (Bethyl Laboratories, catalog number A300-055A)


title: Smarca4 KD ESC_RNA-seq_R2

data_processing:
Fastq files from single read libraries were collapsed and split based on the 5  barcode in the linker sequence. The 5  barcode sequences were then trimmed from reads and reads were mapped to the mouse mm9 genome using Bowtie1 with reads at least 19 nt long. Read counts were normalized to parts per million reads.
To map reads based on TSS location, a custom PERL script was used to search an interval of 2000 nt upstream to 500 nt downstream of a given read for the nearest gene. Antisense reads were then binned, averaged for the two RNA-Seq libraries, and called if there were 5 reads or more for one of the KD datasets. The reads from these libraries were then sorted from lowest to highest in Smarca4 KD and visualized through Java TreeView. To map reads based on DHS location, a custom PERL script was used to search an interval of 1000 nt upstream to 1000 nt downstream of a given read for the nearest gene. All reads were then binned, averaged for the two RNA-Seq libraries, and called if there were 5 reads or more for one of the KD datasets. The reads from these libraries were clustered using using Cluster 3.0 and visualized through Java TreeView.
Genome_build: mm9
Note: Fastq file contains 5' barcode sequence of GCT

tag: cell line

value: embryonic stem cells


title: H3K27Ac KO 1

value: Anti-acetyl lysine 27 of histone H3 (Active Motif, catalog number 39133)


title: H3K27Ac WT 2


title: Smc1a ChIP-seq Rep1


title: Input WT 1


title: Input WT 2


title: H3K27Ac WT 1


title: Input KO 2


title: PARE library from Anther 4.0mm and pollen

status: Public on Feb 11 2015

city: St. Louis

first: Blake

middle: C.

last: Meyers

state: MO

address: 975 N Warson Road

institute: Donald Danforth Plant Science Center

data_processing:
All reads from small RNA, PARE or RNA-seq libraries were trimmed to remove adapter sequences using a Perl script. After trimming off adapters, small RNA reads with the length between 18 and 34 were kept; PARE reads that are longer than 21-nt are discarded, and 21-nt PARE reads are chopped to 20-nt by removing the last nucleotide. Small RNA and PARE reads were mapped to the reference genome using bowtie, and RNA-seq reads were mapped using Tophat. We developed three websites and databases (for sRNA, PARE and RNA-seq, respectively) using the Maize AGPv2 genome, to store and analyze all libraries; we used these visualization tools extensively for this analysis. These Web sites are available at http://mpss.udel.edu/maize_sRNA2/ , http://mpss.udel.edu/maize_RNAseq2/ and http://mpss.udel.edu/maize_pare/
Genome_build: MAIZE_AGPv2_genome

organism: Zea mays subsp. mays

source_name: anther

growth_protocol: Fertile anthers of the W23 inbred line, mac1-Y211 and msca1 introgressed five times into W23, and ocl4 in the A188 inbred were grown in Stanford, CA under greenhouse conditions.

extract_protocol:
Total RNA was isolated using Tri reagent (Molecular Research Center, Cincinnati, OH) or Plant RNA Reagent (Invitrogen, Carlsbad, CA).
Small RNA and RNA-seq libraries were constructed using TruSeq™ Small RNA Sample Prep Kit and TruSeq™ RNA Sample Prep Kit (Illumina, San Diego, CA). PARE libraries were constructed as previously described. All libraries were sequenced on an Illumina Hi-Seq 2000 instrument at the Delaware Biotechnology Institute, Newark, DE.

value: Wildtype (W23)


title: PARE library from Anther 2.0mm, 2.5mm and 3.0mm


title: PARE library from Maize B73, pollen

value: Wildtype (B73)


title: PARE library from Maize mop1 mutant, pollen


title: EGFP KD ESCs_CAP-seq_R2


title: ocl4, 0.7mm Anther, rep2

data_processing:
All reads from small RNA, PARE or RNA-seq libraries were trimmed to remove adapter sequences using a Perl script. After trimming off adapters, small RNA reads with the length between 18 and 34 were kept; PARE reads that are longer than 21-nt are discarded, and 21-nt PARE reads are chopped to 20-nt by removing the last nucleotide. Small RNA and PARE reads were mapped to the reference genome using bowtie, and RNA-seq reads were mapped using Tophat. We developed three websites and databases (for sRNA, PARE and RNA-seq, respectively) using the Maize AGPv2 genome, to store and analyze all libraries; we used these visualization tools extensively for this analysis. These Web sites are available at http://mpss.udel.edu/maize_sRNA2/ , http://mpss.udel.edu/maize_RNAseq2/ and http://mpss.udel.edu/maize_pare/ .  Total RNA was isolated using Tri reagent (Molecular Research Center, Cincinnati, OH) or Plant RNA Reagent (Invitrogen, Carlsbad, CA). Small RNA and RNA-seq libraries were constructed using TruSeq™ Small RNA Sample Prep Kit and TruSeq™ RNA Sample Prep Kit (Illumina, San Diego, CA). PARE libraries were constructed as previously described (Zhai et al., Methods, 2013). All libraries were sequenced on an Illumina Hi-Seq 2000 instrument at the Delaware Biotechnology Institute, Newark, DE.
Genome_build: MAIZE_AGPv2_genome

treatment_protocol: no treatment


title: ocl4, 1.5mm Anther, rep2


title: ocl4, 1.0mm Anther, rep2


title: ocl4, 2.0mm Anther, rep2


title: ocl4, 2.0mm Anther, rep1


title: ocl4, 1.0mm Anther, rep1


title: EGFP KD ESC_RNA-seq_R2

data_processing:
Fastq files from single read libraries were collapsed and split based on the 5  barcode in the linker sequence. The 5  barcode sequences were then trimmed from reads and reads were mapped to the mouse mm9 genome using Bowtie1 with reads at least 19 nt long. Read counts were normalized to parts per million reads.
To map reads based on TSS location, a custom PERL script was used to search an interval of 2000 nt upstream to 500 nt downstream of a given read for the nearest gene. Antisense reads were then binned, averaged for the two RNA-Seq libraries, and called if there were 5 reads or more for one of the KD datasets. The reads from these libraries were then sorted from lowest to highest in Smarca4 KD and visualized through Java TreeView. To map reads based on DHS location, a custom PERL script was used to search an interval of 1000 nt upstream to 1000 nt downstream of a given read for the nearest gene. All reads were then binned, averaged for the two RNA-Seq libraries, and called if there were 5 reads or more for one of the KD datasets. The reads from these libraries were clustered using using Cluster 3.0 and visualized through Java TreeView.
Genome_build: mm9
Note: Fastq file contains 5' barcode sequence 'CATT'


title: ocl4, 0.4mm Anther, rep2


title: ocl4, 1.5mm Anther, rep1


title: ocl4, 0.4mm Anther, rep1


title: ocl4, 0.7mm Anther, rep1


title: H3K27Ac KO 2


title: mac1, 2.0mm Anther, rep2


title: Input KO 1


title: mac1, 1.5mm Anther, rep2


title: mac1, 0.4mm Anther, rep2


title: PARE library from Anther 1.0 and 1.5 mm


title: mac1, 1.0mm Anther, rep2


title: mac1, 0.7mm Anther, rep2


title: mac1, 0.7mm Anther, rep1


title: mac1, 1.0mm Anther, rep1


title: mac1, 2.0mm Anther, rep1


title: mac1, 1.5mm Anther, rep1


title: msca1, 2.0mm Anther, rep2


title: mac1, 0.4mm Anther, rep1


title: msca1, 1.5mm Anther, rep2


title: msca1, 0.4mm Anther, rep2


title: msca1, 1.0mm Anther, rep2


title: msca1, 0.7mm Anther, rep2


title: msca1, 2.0mm Anther, rep1


title: msca1, 0.7mm Anther, rep1


title: 1.5mm Anther, rep3


title: msca1, 1.0mm Anther, rep1


title: 1.0mm Anther, rep3


title: msca1, 0.4mm Anther, rep1


title: mac1, 0.7mm Anther


title: 0.7mm Anther, rep3


title: msca1, 1.5mm Anther, rep1


title: wildtype, 0.4mm Anther


title: wildtype, 0.7mm Anther


title: ocl4, 0.7mm Anther


title: 0.4mm Anther, rep3


title: mac1, 0.4mm Anther


title: ocl4, 0.4mm Anther


title: HCC

city: San Diego

first: Shicheng

last: Guo

state: CA

address: 8730 Costa Verde

institute: MCRI, SHSCI, GHH, UCSD

description: Sample 4

data_processing:
Illumina Casava1.7 software used for base-calling.
sequence were mapped to the hg19 human genome reference assembly by BWA alignment tools with the default settings.
The peaks (hypermethylatged regions) were identified using MACS software.
human CpG islands (CGIs) were downloaded from the UCSC database. The genomic methylation profile generated.
Genome_build: hg19
Supplementary_files_format_and_content: genomic methylation profile wig file

source_name: HCC

extract_protocol:
Peripheral venous blood sample was drawn for each subject, and the serum sample was separated within 2 hours. CfDNA was extracted from 800-μl aliquots of serum using the blood DNA kit (TianGen, Shanghai, PRC).
MBD enriched cf DNA libraries were prepared for sequencing using standard Illumina protocols

tag: etiology

value: HBV positive HCC

tag: tissue

value: Peripheral venous blood

tag: molecule subtype

value: serum cell-free DNA


title: HBV Cirrhosis

description: Sample 3

source_name: HBV Cirrhosis

value: HBV related liver cirrhosis


title: HBV Carrier

description: Sample 2

source_name: HBV Carrier

value: Chronic HBV infection


title: Ark_null_0hr [HS_B7]

city: La Jolla

first: Huaiyu

last: Sun

address: 10010 North Torrey Pines Road

department: Molecular and Cell Biology Laboratory

institute: Salk Institute for Biological Studies

description: subject to poly(A) RNA enrichment

data_processing:
Base-calling was done by Hiseq Control Software (HCS) 2.0.12 and Real Time Analysis (RTA) v1.17.21.3, with demultiplexing  by CASAVA 1.8.2.
Sequence reads were aligned to annotated mouse genome (mm9) by STAR (version 2.3.0e), with parameters shown in the following sample commande line:   STAR   --runThreadN 24   --genomeDir /genomics/software/STAR/indexes/Genome_mm9  --genomeLoad LoadAndKeep   --readFilesIn HS1-1-3-ATCACG_ATCACG_L003_R1_001.fastq    --outFileNamePrefix ./HS1_STAR_   --outSAMstrandField intronMotif   --sjdbGTFfile mm9.refseq.gtf
Conversion of SAM files to sorted BAM files was done with samtools (version 0.1.19-44428c).
The gene expression level, represented by fragments per kilo base per million reads (FPKM), was calculated using Cufflinks (v2.2.1).
Extraction of FPKM values, data cleanup, and gene name annotation were performed with Python scripts. For each batch of 8 sequenced samples, only genes with an average FPKM greater than 0.25 were preserved for further analysis.
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited text file format, include FPKM values for each sample.

growth_protocol: Mouse embryo fibroblasts (Ark WT and Ark-/-) were gifts of Vasso Episkopou (Imperial College London) and were maintained in Dulbecco’s modified Eagle’s medium (DMEM) with 10% fetal bovine serum.

extract_protocol:
Total RNA was extracted using a Nucleospin RNA kit (Clontech) with in-column DNase digestion. The RNA quality was verified to have an RNA integrity number (RIN) greater than 7.5 through using a Bioanalyzer (Agilent).
Stranded mRNA-seq (mRNA sequencing) libraries were prepared using the TruSeq Stranded mRNA Sample Prep Kit (Illumina). mRNA was selected by poly(dT) magnetic beads and then fragmented and reverse-transcribed; cDNA was end-repaired, adenylated, adapter-ligated and PCR amplified.

treatment_protocol: Cells were grown in the 60-mm dish for 2 days to reach ~40% confluence and were treated with TGFβ (PeproTech) at 2.5 ng/ml for 1h, 4h, or 16h as indicated.

value: embryo fibroblasts

value: Ark-/-; Ark_null

tag: treated with

value: none (0hr control)


title: Ark_null_TGFb_1hr [HS_B8]

description: subject to poly(A) RNA enrichment

value: TGFβ at 2.5 ng/ml for 1h


title: Ark_null+rvArksim_TGFb_4hr [HS_B6]

description: subject to poly(A) RNA enrichment

treatment_protocol:
retrovirus protocol: Recombinant retrovirus was used to re-introduce the expression of the SUMO-interacting motif (SIM) mutant (Arksim, with core hydrophobic residues, V298/V299/V300 in SIM1 and V380/V381/L383 in SIM3, changed to A) in Ark-/- MEFs. The  retroviral vector CAG-GFP was derived from murine Moloney leukemia virus (MMLV), with the synthetic CAG promoter driving the simultaneous expression of Arkadia and enhanced green fluorescent protein (EGFP) via an internal ribosomal entry site. Virus was packaged in 293T cells and was concentrated through ultraspeed centrifugation. For infection, Ark-/- MEFs were freshly plated in a 60-mm dish for 4 h and infected for 24 h with virus preparations of ~50,000 CFU to a ~95% efficiency. Early passages of the infected MEFs were used for experiments to avoid retroviral silencing after cell divisions.
Cells were grown in the 60-mm dish for 2 days to reach ~40% confluence and were treated with TGFβ (PeproTech) at 2.5 ng/ml for 1h, 4h, or 16h as indicated.

value: Ark-/-; rvCAG-Arksim

value: Ark-sim mutant-reconstituted Ark-/-

value: TGFβ at 2.5 ng/ml for 4h


title: Ark_null+rvArkwt_TGFb_4hr [HS_B3]

description: subject to poly(A) RNA enrichment

treatment_protocol:
retrovirus protocol: Recombinant retrovirus was used to re-introduce the expression of wild-type Arkadia (ArkWT) in Ark-/- MEFs. The retroviral vector CAG-GFP was derived from murine Moloney leukemia virus (MMLV), with the synthetic CAG promoter driving the simultaneous expression of Arkadia and enhanced green fluorescent protein (EGFP) via an internal ribosomal entry site. Virus was packaged in 293T cells and was concentrated through ultraspeed centrifugation. For infection, Ark-/- MEFs were freshly plated in a 60-mm dish for 4 h and infected for 24 h with virus preparations of ~50,000 CFU to a ~95% efficiency. Early passages of the infected MEFs were used for experiments to avoid retroviral silencing after cell divisions.
Cells were grown in the 60-mm dish for 2 days to reach ~40% confluence and were treated with TGFβ (PeproTech) at 2.5 ng/ml for 1h, 4h, or 16h as indicated.

value: Ark-/-; rvCAG-ArkWT

value: Ark-WT-reconstituted Ark-/-


title: Ark_null+rvArksim_TGFb_1hr [HS_B5]

description: subject to poly(A) RNA enrichment


title: Ark_null+rvArkwt_TGFb_1hr [HS_B2]

description: subject to poly(A) RNA enrichment


title: Ark_null_0hr [HS_A5]

description: subject to poly(A) RNA enrichment


title: Ark_null_TGFb_16hr [HS_A8]

description: subject to poly(A) RNA enrichment

value: TGFβ at 2.5 ng/ml for 16h


title: Ark_nul_TGFb_1hr [HS_A6]

description: subject to poly(A) RNA enrichment


title: Ark_null_TGFb_4hr [HS_A7]

description: subject to poly(A) RNA enrichment


title: Ark_WT_TGFb_ 16hr [HS_A4]

description: subject to poly(A) RNA enrichment

source_name: Ark_WT_TGFb_ 16hr

value: Ark+/+; Ark_wild type


title: Ark_WT_0hr [HS_A1]

description: subject to poly(A) RNA enrichment


title: Ark_WT_TGFb_1hr [HS_A2]

description: subject to poly(A) RNA enrichment


title: Ark_WT_TGFb_ 4hr [HS_A3]

description: subject to poly(A) RNA enrichment

source_name: Ark_WT_TGFb_ 4hr


title: Col-0 ozone rep2 (control genotype for tga2 tag5 tga6)

status: Public on Feb 10 2015

city: Beijing

first: Yuxin

last: Hu

address: No.20 Nanxincun, Xiangshan

department: Key Laboratory of Plant Molecular Physiology

country: China

institute: Institute of Botany, CAS

description: control genotypel for tga2 tga5 tga6

data_processing:
RNA-seq data analysis was performed with the JAVA-based client-server system Chipster
Quality of sequences was checked with FastQC package
First 13 reads were trimmed FRINSEQ package, then mapped to wild type Col-0 whole genome using TopHat2 with following settings:Do not ignore novel junctions, hits allowed=2, allowing 2 nucleotide mismatches, minimum anchor length=8, Maximum number of mismatches allowed in the anchor=0, minimum intron length=20, Maximu intron length=2000
Generated BAM files were sorted and indexed with SAMtools package
Number of Reads of each gene was counted and calculated with HTSeq package
Genome_build: Col-0 TAIR10.23
Supplementary_files_format_and_content: Tab-separated values files including AGI code, chromsomes, start, end, length and RPKM values for each Sample

organism: Arabidopsis thaliana

source_name: whole rosette

growth_protocol: Seeds were sown on 1:1 peat: vermiculite, stratified for three days, and then grown at 22°C/19°C for a week. Subsequently the geminated seedlings were transplanted into new 1:1 peat: vermiculite mixture, one plants per pot (6cm x 6cm), and eight plants per genotype. All plants were randomly placed, grown in controlled chamber (Weiss Bio1300; Weiss Gallenkamp), at 22°C/19°C, and relative humidity of 70%/90%, under a 12-h light/12-h dark cycle for two weeks.

extract_protocol:
Ten to 15 rosettes per genotype (Col-0 set1, coi1-16 ein2 sid2, Col-0 set2, tga2 tga5 tga6 ) from control and ozone treatment were pooled and frozen into liquid nitrogen. Three biological replicates of RNA were isolated with GeneJet Plant RNA purification Mini Kit (Fermentas, now part of Thermo Scientific). Quality and quantity of RNA was assessed using Agilent 2100 Bioanalyzer and NanoDrop.
Libraries were prepared from 100 ng of total RNA with Illumina TruSeq RNA Sample Prep Kit v2 according to the kit manual.

treatment_protocol: Plants were exposed with 350 nL L-1 ozone for two hours.

tag: genotype

tag: developmental stage

value: 3 weeks old plants

tag: treatment

value: 350 ppb ozone exposure for 2hr


title: tga2 tga5 tga6 ctrl rep1

value: tga2 tga5 tga6

value: control


title: tga2 tga5 tga6 ozone rep2


title: tga2 tga5 tga6 ozone rep1


title: tga2 tga5 tga6 ctrl rep2


title: tga2 tga5 tga6 ctrl rep3


title: Healthy Control

description: Sample 1

source_name: Healthy Control

value: Healthy Control

value: Peripheral venous blood

value: serum cell-free DNA


title: Col-0 ozone rep3 (control genotype for tga2 tag5 tga6)

description: control genotypel for tga2 tga5 tga6


title: Col-0 ozone rep1 (control genotype for tga2 tag5 tga6)

description: control genotypel for tga2 tga5 tga6


title: Col-0 ctrl rep2 (control genotype for tga2 tag5 tga6)

description: control genotypel for tga2 tga5 tga6


title: Col-0 ctrl rep3 (control genotype for tga2 tag5 tga6)

description: control genotypel for tga2 tga5 tga6


title: Col-0 ctrl rep1 (control genotype for tga2 tag5 tga6)

description: control genotypel for tga2 tga5 tga6


title: coi1-16 ein2 sid2 ozone rep3

value: coi1-16 ein2 sid2


title: coi1-16 ein2 sid2 ozone rep2


title: Ark_null+rvArksim_0hr [HS_B4]

description: subject to poly(A) RNA enrichment

value: embryo fibroblasts

value: none (0hr control)


title: Ark_null+rvArkwt_0hr [HS_B1]

description: subject to poly(A) RNA enrichment


title: coi1-16 ein2 sid2 ozone rep1


title: coi1-16 ein2 sid2 ctrl rep1


title: coi1-16 ein2 sid2 ctrl rep3


title: Col-0 ozone rep2 (control genotype for coi1-16 ein2 sid2)

description: control genotype for coi1-16 ein2 sid2


title: Col-0 ctrl rep2 (control genotype for coi1-16 ein2 sid2)

description: control genotype for coi1-16 ein2 sid2


title: Col-0 ozone rep3 (control genotype for coi1-16 ein2 sid2)

description: control genotype for coi1-16 ein2 sid2


title: Col-0 ozone rep1 (control genotype for coi1-16 ein2 sid2)

description: control genotype for coi1-16 ein2 sid2


title: tga2 tga5 tga6 ozone rep3


title: OsglHAT1-OE Rep1

city: Nagoya

first: Tomoyuki

last: Furuta

state: Aichi

address: Furo-cho

country: Japan

institute: Nagoya university

description: Transgenic plant in Nipponbare

data_processing:
Sequencing was performed on an Illumina Genome Analyzer IIx Sequencer.
A total of 33 base pair single-end reads were aligned to the transcript sequence of the Nipp genome from IRGSP (http://rapdb.dna.affrc.go.jp/download/archive/irgsp1/) using Bowtie.
The read mapped to each reference was counted
Reads Per Million of library size (RPKM) were calculated
Pairwise comparisons of RNA-Seqs were performed by using TbT normalization with EdgeR (Kadota et al., 2012) to detect significant differences in gene expression with a false discovery rate <0.05
Genome_build: IRGSP-1.0
Supplementary_files_format_and_content: tab-delimited text files include RPM values for each Sample ...

organism: Oryza sativa

source_name: Shoot apical meristem

extract_protocol:
Total RNA was isolated by using the RNeasy Plant Mini Kit (Qiagene) and then digested by recombinant DNase I (RNase-free, Takara) to remove possible genomic DNA contamination, following the manufacturer’s instructions.
Single-end libraries were constructed using the Tru-seq RNA library construction kit (Illumina).

tag: cultivar

value: Nipponbare

value: Shoot apical meristem


title: Col-0 ctrl rep1 (control genotype for coi1-16 ein2 sid2)

description: control genotype for coi1-16 ein2 sid2


title: OsglHAT1-OE Rep3

description: Transgenic plant in Nipponbare


title: OsglHAT1-OE Rep2

description: Transgenic plant in Nipponbare


title: GW6a-4.6 Rep1

description: CSSL having a substituted genomic fragment from kasalath genome in the background of Nipponbare


title: Nipponbare Rep1

description: Nipponbare


title: GW6a-4.6 Rep3


title: GW6a-4.6 Rep2


title: BAT Input

city: Cincinnati

first: Hee-Woong

last: Lim

state: Ohio

address: 3333 Burnet Ave. MLC 7024

department: Division of Biomedical Informatics

institute: Cincinnati Children's Hospital Medical Center

data_processing:
All the ChIP-seq libraries were sequenced by the Functional Genomics Core at the University of Pennsylvania using HiSeq 2000 or Illumina Genome Analyzer IIx.
ChIP-seq reads were aligned to mouse genome (mm9) using Bowtie with options, ‘-k 1 -m 1 --best --strata’. All redundant reads were discarded except one per genomic position. Peak-calling for PRDM16 and MED1 was performed using Homer. After initial calling, all peaks were resized to 200 bp, and a 1 RPM cut-off was applied. PRDM16 binding sites were more rigorously defined as follows. After initial peak-calling, replicates were compared to check reproducibility then pooled. Final peak-calling was performed for a pooled WT sample using a pooled KO sample as a background control. Any genomic regions that overlapped with ENCODE blacklist regions were eliminated.
BigWig files for RPM-normalized stack-height profiles were generated using Homer and bedGraphToBigWig command.
Genome_build: mm9
Supplementary_files_format_and_content: RPM-normalized bigWig files

source_name: Wildtype BAT

growth_protocol: C57BL6 male mice were maintained in a 12 hour dark /light cycle. Mice used for PRDM16 ChIP seq were sacrificed at 8 weeks of age. Mice used for Pol2, Med1 and H3K27ac were sacrificed at 9 months of age or older for tissue collection.

extract_protocol:
Interscapular BAT was isolated and processed for each sacrificed mouse. The tissues were immediately minced, crosslinked in 1 % formaldehyde/PBS. MED1 ChIPs were performed using EGS as an additional crosslinker. For each fat pad, material from 3-4 mice was used for sequencing. To prepare ChIP extracts, crosslinked tissue was suspended in a ChIP buffer containing 20mM Tris, 0.1% SDS, 1% Triton-100, 150 mM NaCl, 1 mM EDTA, protease inhibitors. Samples were incubated at 4 C for 10 min, and subjected to microtip probe sonication under conditions optimized for IP efficiency. Extracts were cleared of insoluble lipid by microfuge centrifugation at full speed followed by extract transfer to a new tube. The process was repeated until minimal lipid accumulated at the top of the extract after centrifugation.
ChIP-Seq libraries were produced and sequenced according to Illumina protocols. Libraries were multiplexed for sequencing.

tag: strain

value: Prdm16(fl/fl) C57Bl6

value: Brown adipose tissue


title: coi1-16 ein2 sid2 ctrl rep2


title: Nipponbare Rep2

description: Nipponbare


title: Fibroblasts_Ctrl-1_sham-siRNA_1

first: Xiang-Dong

last: Fu

address: 9500 Gilman Drive, Room 231

department: CMM

institute: UC San Diego

description: control human fibroblasts, treated with sham siRNA

data_processing:
The sequenced 40 bases were mapped to the oligo pool sequences with Bowtie by allowing 1 mismatch in each of the donor and acceptor sequences.
Genome_build: hg18
Supplementary_files_format_and_content: The processed files contain both the genomic location (the first 6 columns) and counts for 2 isoforms (last 2 columns) of each alternative splicing event.

source_name: Primary skin fibroblast

growth_protocol: DMEM with 10% FBS for HeLa cells. DMEM with 20% FBS and 1x NEAA for primary fibroblasts.

extract_protocol:
Total RNA was extracted from cells using Trizol reagent (Life technologies).
RASL library was constructed according to Li H, et al protocol published Current Protocols in Molecular Biology.

treatment_protocol: Transgene expression was induced by 4μg/ml tetracycline. siRNA was transfected using Lipofectmine RNAiMAX (Life technologies)

value: Skin biopsy from healthy control

tag: strategy

value: RASL-seq

value: HeLa

tag: sample type


title: Fibroblasts_Ctrl-1_sham-siRNA_2

value: control human fibroblasts, treated with sham siRNA


title: Fibroblasts_Ctrl-1_FUS-siRNA_1

description: control human fibroblasts, treated with FUS siRNA


title: Fibroblasts_Ctrl-1_SMN-siRNA_2

description: control human fibroblasts, treated with SMN siRNA


title: Fibroblasts_Ctrl-1_SMN-siRNA_1

value: control human fibroblasts, treated with SMN siRNA


title: Fibroblasts_Ctrl-1_FUS-siRNA_2

value: control human fibroblasts, treated with FUS siRNA


title: Fibroblasts_Ctrl-1_notreat_2

description: control human fibroblasts, no treatment

value: control human fibroblasts, no treatment


title: Fibroblasts_Ctrl-1_notreat_1

description: control human fibroblasts, no treatment


title: Col-0 ctrl rep3 (control genotype for coi1-16 ein2 sid2)

description: control genotype for coi1-16 ein2 sid2

value: 3 weeks old plants

value: control


title: Fibroblasts_Ctrl-2_SMN-siRNA_1


title: Fibroblasts_Ctrl-2_SMN-siRNA_2


title: Fibroblasts_Ctrl-2_FUS-siRNA_2


title: Fibroblasts_Ctrl-2_sham-siRNA_2


title: Fibroblasts_Ctrl-2_notreat_2

description: control human fibroblasts, no treatment


title: A375

city: Bejing

first: Shaobin

last: Wang

address: No.7 Beitucheng West Road, Chaoyang District,

institute: Beijing institute of Genomics

data_processing:
The reads whose quality were lower than 10 were removed firstly, then the rest high quality reads were mapped against to the human genome using the Corona_Lite software (Applied Biosystems, USA). To be specific, a three-step strategy was adopted. Firstly, the 50nt reads were mapped to the genome allowing 5 mismatches. Secondly, the no-hit reads were trimmed 5 nucleotides from the end and repeated mapping. Thirdly, the no-hit reads from the second step were trimmed 5 nucleotides again and repeated the third times mapping. Given that introns were spliced in mature mRNA, three junction datasets were constructed which composed of 5' exons downstream sequences and the 3' exons upstream sequences. The sequences length of 3 junction datasets were 40, 35 and 30nt respectively. The reads not mapped to the genome were also searched against to these junction datasets. Removing the reads mapped to the ribosomal RNA, the last hit reads were assigned to individual genes according to the mapping site information. The number of hit reads of each gene represents its relative expression value. In order to comparation between different samples, the relative expression value was further normalized to RPKM (the reads per kilobase of exon model per million mapped reads) value.
Genome Build:
A375.wig: hg18

molecule: polyA RNA

source_name: skin

extract_protocol: SOLiD whole transcriptome analysis kit

value: human primary melanoma


title: A2058

data_processing:
The reads whose quality were lower than 10 were removed firstly, then the rest high quality reads were mapped against to the human genome using the Corona_Lite software (Applied Biosystems, USA). To be specific, a three-step strategy was adopted. Firstly, the 50nt reads were mapped to the genome allowing 5 mismatches. Secondly, the no-hit reads were trimmed 5 nucleotides from the end and repeated mapping. Thirdly, the no-hit reads from the second step were trimmed 5 nucleotides again and repeated the third times mapping. Given that introns were spliced in mature mRNA, three junction datasets were constructed which composed of 5' exons downstream sequences and the 3' exons upstream sequences. The sequences length of 3 junction datasets were 40, 35 and 30nt respectively. The reads not mapped to the genome were also searched against to these junction datasets. Removing the reads mapped to the ribosomal RNA, the last hit reads were assigned to individual genes according to the mapping site information. The number of hit reads of each gene represents its relative expression value. In order to comparation between different samples, the relative expression value was further normalized to RPKM (the reads per kilobase of exon model per million mapped reads) value.
Genome Build:
A2058.wig: hg18

value: human metastatic melanoma


title: HEMn

data_processing:
The reads whose quality were lower than 10 were removed firstly, then the rest high quality reads were mapped against to the human genome using the Corona_Lite software (Applied Biosystems, USA). To be specific, a three-step strategy was adopted. Firstly, the 50nt reads were mapped to the genome allowing 5 mismatches. Secondly, the no-hit reads were trimmed 5 nucleotides from the end and repeated mapping. Thirdly, the no-hit reads from the second step were trimmed 5 nucleotides again and repeated the third times mapping. Given that introns were spliced in mature mRNA, three junction datasets were constructed which composed of 5' exons downstream sequences and the 3' exons upstream sequences. The sequences length of 3 junction datasets were 40, 35 and 30nt respectively. The reads not mapped to the genome were also searched against to these junction datasets. Removing the reads mapped to the ribosomal RNA, the last hit reads were assigned to individual genes according to the mapping site information. The number of hit reads of each gene represents its relative expression value. In order to comparation between different samples, the relative expression value was further normalized to RPKM (the reads per kilobase of exon model per million mapped reads) value.
Genome Build:
HEMn.wig: hg18

value: HEMn

value: human normal melanocyte


title: Fibroblasts_Ctrl-2_FUS-siRNA_1


title: Fibroblasts_Ctrl-2_sham-siRNA_1


title: Fibroblast_M511nfs_2

description: patient fibroblasts with M511nfs*6 mutation

value: Skin biopsy from ALS patient


title: Fibroblast_H517Q_1

description: patient fibroblasts with H517Q mutation

value: patient fibroblasts with H517Q mutation


title: Fibroblast_M511nfs_1

value: patient fibroblasts with M511nfs*6 mutation


title: Fibroblast_H517Q_2

description: patient fibroblasts with H517Q mutation


title: Fibroblast_R521G_2

description: patient fibroblasts with R521G mutation

value: patient fibroblasts with R521G mutation


title: Fibroblasts_Ctrl-2_notreat_1

description: control human fibroblasts, no treatment

value: Skin biopsy from healthy control


title: Fibroblast_R521G_1

description: patient fibroblasts with R521G mutation

value: RASL-seq


title: Fibroblasts_Ctrl-3_1

description: control human fibroblasts, Kin2ALS6

value: HeLa

value: control human fibroblasts, Kin2ALS6


title: H3K27ac ChIP-seq, Prdm16 KO BAT

source_name: Prdm16 Knockout BAT

value: Myf5Cre-Prdm16(fl/fl) C57Bl6

value: Brown adipose tissue

tag: antibody

value: H3K27ac (Abcam ab4729)


title: Fibroblasts_Ctrl-2_2

description: control human fibroblasts, Kin1ALS17

value: control human fibroblasts, Kin1ALS17


title: Fibroblasts_Ctrl-1_2

description: control human fibroblasts, con1

value: control human fibroblasts, con1


title: H3K27ac ChIP-seq, WT BAT

value: Prdm16(fl/fl) C57Bl6


title: Fibroblasts_Ctrl-2_1

description: control human fibroblasts, Kin1ALS17


title: FLP-IN_P525L_FUS-RNAi_2

description: P525L + FUS-RNAi

source_name: HeLa

value: human cervical cancer patient imortalized

value: P525L + FUS-RNAi


title: Fibroblasts_Ctrl-1_1

description: control human fibroblasts, con1


title: Fibroblasts_Ctrl-3_2

description: control human fibroblasts, Kin2ALS6


title: FLP-IN_R521G_FUS-RNAi_2

description: R521G + FUS-RNAi

value: R521G + FUS-RNAi


title: FLP-IN_R521G_FUS-RNAi_3

description: R521G + FUS-RNAi


title: Med1 ChIP-seq, WT BAT

value: Med1(Bethyl A300-793A)


title: Med1 ChIP-seq, Prdm16 KO BAT


title: PolII ChIP-seq, Prdm16 KO BAT

value: PolII (Santa Cruz sc-899 or sc-9001)


title: FLP-IN_R521G_FUS-RNAi_1

description: R521G + FUS-RNAi


title: PolII ChIP-seq, WT BAT


title: FLP-IN_R514G_FUS-RNAi_2

description: R514G + FUS-RNAi

value: R514G + FUS-RNAi


title: Prdm16 ChIP-seq, Prdm16 KO BAT, rep1

value: Made in Lab (Harms et al. 2014. Cell Metab.)


title: FLP-IN_R514G_FUS-RNAi_1

description: R514G + FUS-RNAi


title: Prdm16 ChIP-seq, Prdm16 KO BAT, rep2


title: Prdm16 ChIP-seq, WT BAT, rep2


title: FLP-IN_R495X_FUS-RNAi_2

description: R495X + FUS-RNAi

value: R495X + FUS-RNAi