title: LN rep3

first: Jinkai

last: Wang

address: 650 Charles E. Young Drive South, CHS 33-228

institute: University of California, Los Angeles

data_processing:
Tophat (version 1.4.1) were used to map RNA-Seq reads to rat Ensembl transcripts first allowing up to 1 mismatch and the reads that were not fully mapped to the transcriptome will then be mapped to rat rn4 Ensembl genome allowing up to 2 mismatch.
Only the uniquely mapped read pairs were subjected to downstream analyses.
FPKMs (fragments per kb of exon per million sequence reads) were calculated using Cufflinks (version 2.0.0).
Genome_build: rn4
Supplementary_files_format_and_content: Supplemental excel file includes the FPKM values of all samples.

organism: Rattus norvegicus

source_name: Liver

growth_protocol: The rats were bred and maintained in an approved animal facility in microisolator caging on a 12-hour light-dark cycle.

extract_protocol:
Total RNA was isolated from liver tissue using standard TRIzol methods (Chomczynski and Sacchi 1987). RNA quality was determined via a BioAnalyzer 2100 (Agilent Technologies, Santa Clara, CA, USA), using an RIN threshold of 7.
RNA-Seq libraries were prepared with TruSeq RNA Sample Preparation Kits v2 (Illumina, San Diego, CA) according to manufacturer’s instructions.

tag: Sex

value: Male

value: liver

value: Lyon Normotensive


title: LN rep6

status: Public on Oct 31 2014

type: SRA

city: Los Angeles

state: CA

country: USA

library_source: transcriptomic

molecule: polyA RNA

tag: tissue

tag: genotype/variation


title: LN rep2


title: F2 ind34


title: F2 ind35


title: LN rep1


title: LN rep5


title: F2 ind36


title: LN rep4


title: F2 ind31


title: F2 ind33


title: F2 ind30


title: F2 ind32


title: F2 ind28


title: F2 ind29


title: WT 2

city: East Melbourne

first: Huiling

last: Xu

state: Victoria

address: St Andrew Place

department: Research Division

country: Australia

institute: Peter MacCallum Cancer Centre

data_processing:
RNA-seq reads were mapped using bowtie version 0.12.8 & TopHat v1.4.1. Transcripts assembled using Cufflink. Reads count was performed with htseq-count (version 1.0.0)
ChIP-seq reads were mapped with BWA for Illumina (version 1.2.3).
Genome_build: mm9
Supplementary_files_format_and_content: For RNA-seq, tab-delimited text files include limma-voom normalised log2 values for each Sample;
Supplementary_files_format_and_content: For ChIP-seq, peak calling was performed using MACS V3.1 with p value cutoff 1e-4 for peak detection and MFOLD 10 for enrichment ratio.

molecule: total RNA

organism: Mus musculus

source_name: WT small intestinal crypts

extract_protocol:
Small intestinal crypts were isolated in PBS containing 3 mM EDTA, 0.5 mM DTT and protease inhibitor (Roche). Adenomas were dissected from the small intestine. RNA was extracted using RNaEasy kit (Qiagene).For ChIP-seq, crypts and adenoma in 1% formaldehyde for 10 Min at room temperature. Chromatin was sheared using the Covaris S2 system following the manufacturer’s instructions. ChIP was performed using a ChIP validated rabbit anti-Rad21 antibody (Ab992, Abcam) and Magna-ChIP A kit (Millipore).
Library preparations were performed using the TruSeq RNA Sample Preparation protocol (Illumina, San Diego, CA) and correct size established using the DNA 1000 kit (Agilent technologies, Santa Clara, CA). Libraries were quantified with qPCR, normalised and pooled to 2nM before sequencing with paired-end 50 bp reads using standard protocols on the HiSeq2000 (Illumina, San Diego, CA).

tag: strain background

value: Wild type

value: Small intestinal crypts


title: F2 ind26


title: F2 ind25


title: F2 ind23


title: F2 ind24


title: F2 ind22


title: F2 ind27


title: F2 ind21


title: F2 ind20


title: F2 ind10


title: F2 ind19


title: F2 ind14


title: F2 ind16


title: F2 ind7


title: F2 ind11


title: F2 ind8


title: F2 ind18


title: F2 ind17


title: F2 ind13


title: F2 ind9


title: F2 ind3


title: LH rep4

value: Lyon Hypertensive


title: LH rep5


title: F2 ind5


title: LH rep6


title: F2 ind4


title: F2 ind2


title: F2 ind12


title: LH rep2


title: F2 ind1


title: LH rep3


title: LH rep1


title: N-10

status: Public on Oct 30 2014

city: Vancouver

first: Kelsie

middle: L

last: Thu

state: British Columbia

address: 675 West 10th Avenue

country: Canada

institute: BC Cancer Research Centre

data_processing:
Reads were filtered and aligned to NCBI GRCh37 reference genome and miRBase v18 using the BWA algorithm as previously described [TCGA: Comprehensive molecular portraits of human breast tumours. Nature 2012, 490(7418):61-70]
Genome_build: NCBI GRCh37 reference genome (hg19) and miRBase v18
Supplementary_files_format_and_content: Excel file containing normalized miRNA expression values (reads per kilobase per million (RPKM)) for 188 libraries
Supplementary_files_format_and_content: summarized miRNA expression data where expression values (in RPKM) for miRNA coding identical sequences from different genomic loci are summed, and expression values < 1 RPKM are converted to 0

organism: Homo sapiens

source_name: non-malignant lung parenchymal tissue from lung adenocarcinoma patient

extract_protocol:
Fresh-frozen lung adenocarcinoma (LUAC) tumor and patient matched non-malignant lung parenchymal tissues were collected from treatment naive patients; Tissue microdissection was guided by a lung pathologist to ensure >80% tumor cell or >80% non-malignant cell content. Total RNA was extracted using Trizol reagent
Full description of library construction, sequencing, read pre-processing, alignment and annotation are previously described [TCGA: Comprehensive molecular portraits of human breast tumours. Nature 2012, 490(7418):61-70]

tag: patient id

tag: disease

value: lung adenocarcinoma

value: CS

tag: gender

value: F

tag: tissue subtype

value: non-malignant lung parenchymal tissue

value: micro-dissected for >80% non-malignant cell content

tag: molecule subtype

value: microRNA


title: N-101


title: N-105

value: NS


title: F2 ind15


title: F2 ind6


title: N-102


title: N-1

value: M


title: N-107


title: N-113


title: N-11

value: FS


title: N-114


title: N-117


title: N-12


title: N-116


title: N-123


title: N-120


title: N-126


title: N-125


title: N-13


title: N-14


title: N-16


title: N-18


title: N-19


title: N-24


title: N-25


title: N-112


title: N-2


title: N-22


title: N-26


title: N-21


title: N-32


title: N-34


title: N-23


title: N-36


title: N-33


title: N-31


title: N-29


title: N-27


title: N-41


title: N-4


title: N-15


title: N-38


title: N-3


title: N-30


title: N-17


title: N-39


title: N-115


title: N-35


title: N-37


title: N-40


title: N-28


title: N-20


title: N-43


title: N-46


title: N-47


title: N-45


title: N-44


title: N-52


title: N-57


title: N-54


title: N-66


title: N-60


title: N-50


title: N-56


title: N-67


title: N-78


title: N-42


title: N-8


title: N-63


title: N-5


title: N-69


title: N-55


title: N-6


title: N-73


title: N-79


title: N-70


title: N-83


title: N-81


title: N-61


title: N-58


title: N-84


title: N-88


title: N-85


title: N-86


title: N-87


title: N-80


title: N-89


title: N-7


title: N-90


title: N-9


title: N-91


title: T-10

source_name: lung adenocarcinoma tumor

value: I

value: micro-dissected for >80% tumor cell content


title: T-1


title: N-95


title: N-96


title: T-101


title: T-11

value: II


title: N-99


title: N-92


title: T-112


title: T-102


title: T-113

value: III


title: T-115


title: T-116


title: T-114


title: T-107


title: T-13


title: T-123


title: T-125

value: IV


title: T-120


title: T-117


title: T-14


title: T-15


title: N-94


title: T-16


title: T-105


title: T-17


title: T-126


title: T-18


title: T-2


title: T-22


title: T-20


title: T-19


title: T-21


title: T-25


title: T-28


title: T-3


title: T-30


title: T-26


title: T-27


title: T-29


title: T-31


title: T-32


title: T-12


title: T-34


title: T-37


title: T-35


title: T-36


title: T-4


title: T-38


title: T-23


title: T-41


title: T-24


title: T-33


title: T-39


title: T-40


title: T-42


title: T-45


title: T-46


title: T-43


title: T-52


title: T-54


title: T-5


title: T-56


title: T-55


title: T-60


title: T-6


title: T-63


title: T-66


title: T-67


title: T-7


title: T-70


title: T-47


title: T-8


title: T-78


title: T-44


title: T-80


title: T-79


title: T-50


title: T-73


title: T-83


title: T-85


title: T-87


title: T-57


title: T-88


title: T-58


title: T-90


title: T-9


title: T-61


title: T-89


title: T-91


title: T-92


title: T-94


title: T-95


title: T-69


title: T-81


title: T-99


title: RNA-seq SKNMC shFLI1 48 hrs

city: Charlestown

first: Martin

last: Aryee

state: MA

address: 149 13th Street

department: Pathology

institute: Massachusetts General Hospital

data_processing:
Paired end reads were aligned to the reference genome using Tophat 1.4.1
Cufflinks 2.1.0 was used to generate read counts for Refseq genes (GTF downloaded from UCSC on 2013-09-11)
Genome_build: hg19
Supplementary_files_format_and_content: Read count tables generated by Cufflinks

source_name: SKNMC cells

growth_protocol: The human Ewing sarcoma cell lines A673 and SKMNC were grown in RPMI 1640 containing 10% FCS at 37°C with 5% CO2.

extract_protocol:
Total RNA was isolated using the RNeasy Kit (Qiagen).
5 ug of total RNA was used to isolate polyA mRNAs and construct Illumina sequencing libraries.

treatment_protocol: shRNAs from the RNAi Consortium were used for knockdown (pLKO.1): FLI1 (TRCN0000005322), GFP (GCAAGCTGACCCTGAAGTTCAT). Lentiviruses were produced using standard protocols. Briefly lentiviral shRNA plasmids were co-transfected with GAG/POL and VSV plasmids into 293T packaging cells using FugeneHD (Roche) to produce the virus. Viral supernatant was collected 72 hours after transfection and concentrated by ultracentrifugation using an SW41Ti rotor (Beckman Coulter) at 28,000 rpm for 120 min.

tag: cell line

value: Ewing sarcoma cell line SKMNC

tag: treatment

tag: time

value: 48 hrs


title: RNA-seq SKNMC shGFP 48 hrs

value: shGFP


title: RNA-seq SKNMC shGFP 96 hrs

value: 96 hrs


title: T-86

value: lung adenocarcinoma


title: RNA-seq A673 shGFP 48 hrs

source_name: A673 cells

value: Ewing sarcoma cell line A673


title: T-84


title: RNA-seq SKNMC shFLI1 96 hrs


title: T-96

value: F


title: H3K27ac ChIP-seq in A673 96hrs shFLI1

library_source: genomic

data_processing:
ChIP DNA and input controls were sequenced with the Hi-Seq Illumina Genome Analyzer.
ChIP-seq reads were aligned to the reference genome (hg19) using BWA or MAQ (http://maq.sourceforge.net/). Reads arising from plasmid DNA contamination were filtered for two libraries (SKNMC shFLI148.FLI1 and shFLI196.FLI1).
We extended aligned reads to 200 bp to approximate fragment sizes and derived 25-bp resolution density maps by counting the number of fragments overlapping each position with IGVtools. All density maps were normalized to 10M reads.
Genome_build: hg19
Supplementary_files_format_and_content: Wig files were generated in 25-bp step based on UCSC definition and converted into bigWig files using UCSC utilities.

molecule: genomic DNA

source_name: A673 after 96 hrs shFLI1

growth_protocol: A673 and SKMNC Ewing sarcoma cell lines were grown in RPMI 1640 containing 10% FCS at 37°C with 5% CO2. Cells were maintained at a density between 5 x 10^5 cells/ml and 2 x 10^6 cells/ml and split every 3 – 4 days. Primary pediatric mesenchymal stem cells were cultured in IMDM containing 10% FCS and 10ng/ml PDGF-BB (Peprotec).

extract_protocol:
Chromatin from formaldehyde-fixed cells (3-10 x 10^6 cells per epitope) was fragmented to a size range of 200 - 700 bases with a Branson 250 Sonifier. Solubilized chromatin was immunoprecipitated with the indicated antibodies and Protein G-Dynabeads (Life Technologies). Immunoprecipitated DNA was extracted with the Mini-Elute PCR purification kit (Qiagen) after crosslink reversal, RNAse A and Proteinase K treatment.
ChIP DNA samples were used to generate Illumina sequencing libraries following standard protocols.

treatment_protocol: shRNAs from the RNAi Consortium were used for knockdown (pLKO.1 backbone): FLI1 (TRCN0000005322), GFP (GCAAGCTGACCCTGAAGTTCAT). EWS-FLI1 was expressed using a lentiviral vector (pLIV). Lentiviruses were produced using standard protocols. Briefly, lentiviral shRNA or expression plasmids were co-transfected with GAG/POL and VSV plasmids into 293T packaging cells using FugeneHD (Roche) to produce the virus. Viral supernatant was collected 72 hours after transfection and concentrated by ultracentrifugation.

tag: chip antibody

value: H3K27ac (Abcam, ab4729)


title: H3K27ac ChIP-seq in A673 96hrs shGFP

source_name: A673 after 96 hrs shGFP


title: H3K27ac ChIP-seq in A673 48hrs shFLI1

source_name: A673 after 48 hrs shFLI1

value: H3K27ac (Active Motif, 39133)


title: H3K27ac ChIP-seq in A673 48hrs shGFP

source_name: A673 after 48 hrs shGFP


title: FLI1 ChIP-seq in A673 48hrs shFLI1

value: FLI1 (Santa Cruz, sc-356)


title: FLI1 ChIP-seq in A673 48hrs shGFP


title: A673 whole cell extract

value: WCE


title: H3K4me1 ChIP-Seq in A673

value: H3K4me1 (Abcam, ab8895)


title: H3K4me3 ChIP-Seq in A673

value: H3K4me3 (Millipore, 07-473)


title: p300 ChIP-seq in SKMNC 96hrs shFLI1

source_name: SKMNC after 96 hrs shFLI1

value: p300 (Santa Cruz, sc-585)


title: ELF1 ChIP-seq in SKMNC 96hrs shFLI1

value: ELF1 (Santa Cruz, sc-631)


title: FLI1 ChIP-Seq in A673


title: H3K27ac ChIP-Seq in A673


title: H3K27me3 ChIP-Seq in A673

value: H3K27me3 (Millipore, 07-449)


title: GABPA ChIP-seq in SKMNC 96hrs shFLI1

value: GABPA (Santa Cruz, sc-22810)


title: WDR5 ChIP-seq in A673

value: WDR5 (Bethyl, A302-429A)


title: RNA-seq A673 shFLI1 48 hrs

value: 48 hrs


title: H3K27ac ChIP-seq in SKMNC 48hrs shFLI1

source_name: SKMNC after 48 hrs shFLI1

value: Ewing sarcoma cell line SKMNC


title: H3K4me1 ChIP-seq in SKMNC 96hrs shFLI1


title: FLI1 ChIP-seq in SKMNC 96hrs shFLI1


title: FLI1 ChIP-seq in SKMNC 48hrs shFLI1


title: p300 ChIP-seq in SKMNC 96 hrs shGFP

source_name: SKMNC after 96 hrs shGFP

value: shGFP


title: ELF1 ChIP-seq in SKMNC 96 hrs shGFP


title: H3K4me1 ChIP-seq in SKMNC 96hrs shGFP


title: GABPA ChIP-seq in SKMNC 96hrs shGFP


title: H3K27ac ChIP-seq in SKMNC 96hrs shGFP


title: H3K27me3 ChIP-seq in SKMNC 96hrs shGFP


title: FLI1 ChIP-seq in SKMNC 96hrs shGFP96


title: H3K27ac ChIP-seq in SKMNC 48hrs shGFP

source_name: SKMNC after 48 hrs shGFP


title: FLI1 ChIP-seq in SKMNC 48hrs shGFP


title: H3K4me3 ChIP-seq in SKMNC

source_name: SKMNC


title: SKMNC whole cell extract

description: SKNMC.WCE


title: H3K27me3 ChIP-seq in SKMNC


title: WDR5 ChIP-seq in SKMNC


title: H3K4me1 ChIP-seq in SKMNC


title: FLI1 ChIP-seq in SKMNC


title: H3K27ac ChIP-seq in SKMNC


title: MV4;11_WCE_0nM

city: Watertown

first: Eric

last: Olson

address: 480 Arsenal Street

institute: Syros Pharmaceuticals

data_processing:
Samples 20140404_1218 and 20140404_1219 were aligned to hg19 using Bowtie2 with default parameters
All other samples were aligned to a combined hg19/dm3 genome and reads were separated into each organism post-alignment. See manuscript for details.
Supplementary_files_format_and_content: TDF files(s) represent counts of aligned reads within 1 bp bins with each read being extended 200 in the direction of alignment. Counts are in reads-per-million and floored at 0.1

source_name: AML

extract_protocol:
Whole cell extracts were sonicated to solubilize the chromatin. The chromatin extracts containing DNA fragments with an average size of 500 bp were immunoprecipitated using different antibodies. Purified immunoprecipitated DNA were prepared for sequencing according to a modified version of the Solexa Genomic DNA protocol. Fragmented DNA was end repaired and subjected to 18 cycles of LM-PCR using oligos provided by Illumina. Amplified fragments between 150 and 300bp (representing shear fragments between 50 and 200nt in length and ~100bp of primer sequence) were isolated by agarose gel electrophoresis and purified.
Lysates were clarified from sonicated cells and target bound fragments were isolated with antibody.

treatment_protocol: MV4-11 cells were treated with DMSO for 4 days.

tag: spike-in reference

value: Drosophila melanogaster

tag: drug

value: DMSO

tag: duration

value: 4 days


title: Jurkat_WCE_75%_R2

source_name: Peripheral blood cells

growth_protocol: Jurkat cells were treated in RPMI, 10% FBS and 1% penstrep media with regular cell care maintenance.    S2 cells were grown in SFX medium (Thermo/HyClone #SH30278.02) with 1% Penicillin/Streptomycin (Invitrogen #15140) at room temperature. S2 cells acquired from NCCC and crosslinked following Syros-specified conditions.

treatment_protocol: Jurkat cells were treated with either DMSO or EPZ (Dot1L inhibitor), then collected individually and mixed according to the following composition by cell number: 25% DMSO to 75% EPZ.

value: Jurkat

value: DMSO / EPZ

tag: concentration

value: 20 uM


title: MV4;11_WCE_0.5nM

treatment_protocol: MV4-11 cells were treated with 0.5nM EPZ-5676 for 4 days.


title: Jurkat_WCE_100%_R2

treatment_protocol: Jurkat cells were treated with either DMSO or EPZ (Dot1L inhibitor), then collected individually and mixed according to the following composition by cell number: 0% DMSO to 100% EPZ.


title: H3K27me3 ChIP-seq in SKMNC 96hrs shFLI1


title: MV4;11_WCE_2nM

treatment_protocol: MV4-11 cells were treated with 2nM EPZ-5676 for 4 days.


title: Jurkat_WCE_50%_R2

treatment_protocol: Jurkat cells were treated with either DMSO or EPZ (Dot1L inhibitor), then collected individually and mixed according to the following composition by cell number: 50% DMSO to 50% EPZ.


title: MV4;11_WCE_5nM

treatment_protocol: MV4-11 cells were treated with 5nM EPZ-5676 for 4 days.

value: 5 nM


title: Jurkat_WCE_25%_R2

treatment_protocol: Jurkat cells were treated with either DMSO or EPZ (Dot1L inhibitor), then collected individually and mixed according to the following composition by cell number: 75% DMSO to 25% EPZ.


title: Jurkat_WCE_0%_R2

treatment_protocol: Jurkat cells were treated with either DMSO or EPZ (Dot1L inhibitor), then collected individually and mixed according to the following composition by cell number: 100% DMSO to 0% EPZ.


title: Jurkat_WCE_100%_R1

treatment_protocol: Jurkat cells were treated with either DMSO or 20 uM EPZ5676 (Dot1L inhibitor), then collected individually and mixed according to the following composition by cell number: 0% DMSO to 100% EPZ.


title: Jurkat_WCE_75%_R1

treatment_protocol: Jurkat cells were treated with either DMSO or 20 uM EPZ5676 (Dot1L inhibitor), then collected individually and mixed according to the following composition by cell number: 25% DMSO to 75% EPZ.


title: Jurkat_WCE_25%_R1

treatment_protocol: Jurkat cells were treated with either DMSO or 20 uM EPZ5676 (Dot1L inhibitor), then collected individually and mixed according to the following composition by cell number: 75% DMSO to 25% EPZ.


title: Jurkat_WCE_50%_R1

treatment_protocol: Jurkat cells were treated with either DMSO or 20 uM EPZ5676 (Dot1L inhibitor), then collected individually and mixed according to the following composition by cell number: 50% DMSO to 50% EPZ.


title: Jurkat_WCE_NoSpike_50%

growth_protocol: Jurkat cells were treated in RPMI, 10% FBS and 1% penstrep media with regular cell care maintenance.


title: Jurkat_K79_NoSpike_50%

tag: antibody manu.

value: Abcam

tag: antibody catalog


title: hs_CFU_2

city: New York

first: John

last: Hale

state: NY

address: 310 East 67th Street

institute: New York Blood Center

description: Cultured CD34+ cells

data_processing:
Illumina Casava 1.8.2 software was used for basecalling.
FASTQ format sequencing reads were aligned to the hg19 using TopHat Version 2.0.9 software with default parameters to reference annotation.
Differentially expressed and regulated genes with were computed using Cuffdiff 2.1.1.
Genome_build: hg19
Supplementary_files_format_and_content: diff, count_tracking and fpkm_tracking text files

source_name: CFU-E cells

extract_protocol:
Total RNAs were extracted by RNeasy Plus Mini Kit (Cat# 74134, QIAGEN). Illumina TruSeq RNA Sample Prep Kit (Part # 15008136) was used with 1 ug of total RNA for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: cell type

value: colony-forming unit-erythroid cells (CFU-E)

tag: tissue

value: Cord blood


title: hs_BFU_3

description: Cultured CD34+ cells

source_name: BFU-E cells

value: burst-forming unit-erythroid cells (BFU-E)


title: hs_CFU_3

description: Cultured CD34+ cells


title: hs_BFU_2

description: Cultured CD34+ cells


title: Jurkat_WCE_0%_R1

treatment_protocol: Jurkat cells were treated with either DMSO or 20 uM EPZ5676 (Dot1L inhibitor), then collected individually and mixed according to the following composition by cell number: 100% DMSO to 0% EPZ.


title: hs_CFU_1

description: Cultured CD34+ cells


title: hs_CD34_3

description: Cultured CD34+ cells

source_name: CD34 positive cells

value: CD34 positive cells (CD34)


title: Jurkat_K4_100%_R2

data_processing:
Sample 20140404_1216 was aligned to hg19 using Bowtie2 with default parameters
Sample 20131220_425 was aligned to dm3 using Bowtie2 with default parameters
All other samples were aligned to a combined hg19/dm3 genome and reads were separated into each organism post-alignment. See manuscript for details.
Supplementary_files_format_and_content: IGV compatible TDF files represent spike-in normalized (.spikein) counts for reads aligning to human (.hg19) or drosophila (.dm3). See manuscript for details. Except for 20140404_1216 and 20131220_425 which did not contain spike-in and were normalized using RPM.

tag: spike-in reference organism

tag: spike-in cell line

value: Millipore


title: hs_CD34_1

description: Cultured CD34+ cells


title: hs_BFU_1

description: Cultured CD34+ cells


title: Jurkat_K79_NoSpike


title: MV4;11_K79_5nM

treatment_protocol: MV4-11 cells were treated with 5 nM EPZ for 4 days.


title: MV4;11_K4_2nM

treatment_protocol: MV4-11 cells were treated with 2nM EPZ 5676 for 4 days.


title: MV4;11_K79_2nM

treatment_protocol: MV4-11 cells were treated with 2 nM EPZ for 4 days.


title: MV4;11_K4_0nM

treatment_protocol: MV4-11 cells were treated with DMSO for 4 days.


title: H3K27ac ChIP-seq in SKMNC 96hrs shFLI1

treatment_protocol: shRNAs from the RNAi Consortium were used for knockdown (pLKO.1 backbone): FLI1 (TRCN0000005322), GFP (GCAAGCTGACCCTGAAGTTCAT). EWS-FLI1 was expressed using a lentiviral vector (pLIV). Lentiviruses were produced using standard protocols. Briefly, lentiviral shRNA or expression plasmids were co-transfected with GAG/POL and VSV plasmids into 293T packaging cells using FugeneHD (Roche) to produce the virus. Viral supernatant was collected 72 hours after transfection and concentrated by ultracentrifugation.

value: H3K27ac (Active Motif, 39133)


title: Jurkat_K79_100%_R1


title: Jurkat_K79_0nM

treatment_protocol: Jurkat cells were treated with DMSO for 4 days.


title: MV4;11_K79_0.5nM

treatment_protocol: MV4-11 cells were treated with 0.5 nM EPZ for 4 days.


title: Jurkat_K79_75%_R1

treatment_protocol: Jurkat cells were treated with either DMSO or EPZ (Dot1L inhibitor), then collected individually and mixed according to the following composition by cell number: 25% DMSO to 75% EPZ.


title: MV4;11_K4_0.5nM

treatment_protocol: MV4-11 cells were treated with 0.5nM EPZ 5676 for 4 days.


title: Jurkat_K79_0%_R1


title: Jurkat_K79_50%_R1

treatment_protocol: Jurkat cells were treated with either DMSO or EPZ (Dot1L inhibitor), then collected individually and mixed according to the following composition by cell number: 50% DMSO to 50% EPZ.


title: hs_CD34_2

description: Cultured CD34+ cells


title: Jurkat_K4_100%_R1

treatment_protocol: Jurkat cells were treated with either DMSO or EPZ (Dot1L inhibitor), then collected individually and mixed according to the following composition by cell number: 0% DMSO to 100% EPZ.

tag: antibody lot


title: Jurkat_K79_25%_R1

treatment_protocol: Jurkat cells were treated with either DMSO or EPZ (Dot1L inhibitor), then collected individually and mixed according to the following composition by cell number: 75% DMSO to 25% EPZ.


title: Jurkat_K79_75%_R2


title: Jurkat_K4_75%_R1


title: Jurkat_K4_0%_R1

treatment_protocol: Jurkat cells were treated with either DMSO or EPZ (Dot1L inhibitor), then collected individually and mixed according to the following composition by cell number: 100% DMSO to 0% EPZ.


title: Jurkat_K4_25%_R1


title: Jurkat_K4_50%_R1


title: Jurkat_K79_50%_R2


title: Jurkat_K79_100%_R2


title: Jurkat_K79_5nM

treatment_protocol: Jurkat cells were treated with 5 nM EPZ-5676 for 4 days.


title: Jurkat_K79_25%_R2


title: Jurkat_K4_50%_R2


title: Jurkat_K79_0%_R2


title: Jurkat_K4_0%_R2

growth_protocol: Jurkat cells were treated in RPMI, 10% FBS and 1% penstrep media with regular cell care maintenance.    S2 cells were grown in SFX medium (Thermo/HyClone #SH30278.02) with 1% Penicillin/Streptomycin (Invitrogen #15140) at room temperature. Acquired from NCCC, with Syros-specified crosslinking conditions.


title: Jurkat_K4_25%_R2


title: S2_input

city: Bethesda

first: Brian

last: Oliver

state: MD

address: 50 South Drive

department: LBG

institute: NIDDK, NIH

description: Two biological replicates were produced. Each fastq file for S2_input represents an independent biological replicate. All fastq files for S2_input were pooled before alignment to the genome.

data_processing:
Sequencing was done on a GAI instrument and base-calls were performed with Illumina software pipeline version 0.3.0
All reads were trimmed to 25 bp before alignment to the genome using Bowtie version 0.12.7
Peaks were called using SPP version 1.11 WTD method with FDR of 0.01.  Duplicate reads were removed from the libraries before peak calling with the Picard tool MarkDuplicates (version 1.95)
wig files  for S2_DSXF and S2_DsxM (input subtracted) were made using SPP's smoothed tag density function with bandwidth=200 and step=100 and tag.shift=0. The score represents the smoothed input subtracted tag density.
Genome_build: dm3, Flybase release 5 with no Uextra

organism: Drosophila melanogaster

source_name: S2 cells without transfection

growth_protocol: Schneider's Drosophila line 2 cells (abbreviated S2) derived from Drosophila embryos were maintained in Schneider's Drosophila medium (Invitrogen) containing 10% heat inactivated Fetal Bovine Serum (JRH Biosciences) and antibiotics (0.5 U/ml penicillin and 0.5 μg/ml streptomycin (Invitrogen). The cells were maintained in monolayers at 25 °C.

extract_protocol:
Formaldehyde was added to a final concentration of 1 % to cross-link proteins to DNA. The fixation reaction was incubated on a shaker for 10 min at room temperature. The reaction was quenched by adding glycine to a final concentration of 125 mM and incubation for 5 min on a shaker at room temperature. Subsequently the cells were washed twice with ice-cold PBS. After centrifugation at 500 x g (1680 rpm) for 5 min at 4 °C the cell pellet was resuspended in 10 ml ice-cold cell lysis buffer (5 mM pH8.0 PIPES buffer, 85 mM Potassium chloride, 0.5% Nonidet P40 and protease inhibitors (cOmplete, EDTA-free, Roche, indianapolis, IN)) for 10 minutes at 4 °C. Nuclei were released by douncing with a Wheaton homogenizer (pestle B). The crude nuclear extract was collected by centrifugation at 500 x g (1680 rpm) for 5 min at 4 °C, resuspended in 2 ml ice–cold nuclear lysis buffer (50 mM pH8.1 Tris.HCl, 10 mM EDTA, 1 % SDS and protease inhibitors) and incubated for 20 minutes at 4 °C. After adding 1 ml ice-cold IP dilution buffer (0.01 % SDS, 1.1 % TritonX-100, 1.2 mM pH 8 EDTA.Na2, 16.7 mM pH8 Tris.HCl, 167 mM NaCl and protease inhibitors) and 0.3 g acid-washed glass beads (Sigma-Aldrich, St. Louis, MO) to the nuclear extract, the chromatin was sheared to 200-1000 bp using a Misonix Sonicator 3000 (Misonix, Inc. Farmingdale, NY). Sonication was performed on ice water with 8 pulses of 30 seconds and 30 second intervals. Thereafter the cell debris was removed by centrifugation at 16000 x g (13000 rpm) for 10 min at 4 °C. Aliquots of sonicated chromatin were stored at -80 °C. To assess chromatin quality, aliquots of the chromatin (100 μl) were treated with 40 µg proteinase K (Invitrogen, Carlsbad, CA) for 2 h at 50 °C after the DNA-protein complexes were de-crosslinked at 65°C overnight. After purification of the DNA by phenol-chloroform extraction and ethanol precipitation 3 μl aliquots were subjected to electrophoresis and the chromatin fractions with optimal size (200-1000 bp) range were subjected to immunoprecipitation experiments.
Libraries were made using the Genomic DNA sample preparation kit (Illumina). Adapter-ligated DNA of 200 ± 25 bp range was excised from the gel before PCR amplification.

value: no transfection

tag: amount of dna used to make library


title: heat30.parent30_1

city: Eugene

first: Patrick

middle: C.

last: Phillips

state: OR

address: 5289 University of Oregon

department: Institute of Ecology and Evolution

institute: University of Oregon

data_processing:
Base-calls were performed using Illumina software
Initial processing was performed using the process_shortreads component of Stacks. In short, reads were demultiplexed, and the barcodes were removed. Reads with ambiguous barcodes were rescued if there were fewer than 2 mismatches, and discarded otherwise. Reads failing the Illumina chastity filter were also removed.
Filtered reads were aligned to the C. remanei genome using GSNAP version 2013-03-31 with parameters -N 1 -s -w 10000 --pairmax-RNA 10000
Reads aligning to genes were counted using htseq-count with parameters -m union -s no
Genome_build: C. remanei (strain 356) v1_4 (Fierst et al, in preparation; NCBI BioProject ID PRJNA248909)
Supplementary_files_format_and_content: tab-delimited text files with read counts per gene (note: a fasta file including all transcript sequences for this dataset is linked as a Series supplementary file).

organism: Caenorhabditis remanei

source_name: pooled L1 tissue

growth_protocol: Worms were maintained on NGM-lite seeded with E. coli strain OP50 at 20°C

extract_protocol:
Approximately 100,000 individuals per replicate were flash frozen in Trizol. Total RNA was extracted using a standard Trizol extraction protocol. Poly-A mRNA was isolated from total RNA using the Dynabeads mRNA purification kit (Ambion).
mRNA was sheared to 200-600nt fragments using a buffered zinc solution (RNA fragmentation kit; Ambion). cDNA was synthesized from fragments using Superscript III, and barcoded sequencing adaptors were added to enable multiplexing of samples.

treatment_protocol: Populations were age-synchronized during the embryonic stage via a standard bleaching procedure. Embryos were incubated at either 20°C or 30°C in liquid buffer without food for 20 hours. Pooled L1 larvae were passed through a 20 micron filter immediately prior to tissue collection.

tag: strain

value: Heat-selected, derived from PX443

tag: developmental stage

tag: parental temperature

tag: larval rearing temperature

tag: batch

value: E


title: ATAC-Seq SKNMC

description: SKNMC.ATACseq

data_processing:
Library strategy: ATAC-seq
Paired end reads were aligned to the reference genome using BWA 0.6.2
Read start sites were adjusted to represent the center of the transposon binding event (+4 bp in the plus strand and -5 bp in the minus strand). Cut site density was calculated over 150bp-wide sliding windows at 20bp steps using Bedtools. Each experiment was normalized to 10M reads.
Genome_build: hg19
Supplementary_files_format_and_content: Bigwig files of ATAC-Seq cut site density

source_name: SKNMC cells

growth_protocol: The human Ewing sarcoma cell lines A673 and SKMNC were grown in RPMI 1640 containing 10% FCS at 37°C with 5% CO2. Cells were maintained between a density of 5 x 10^5 cells/ml and 2 x 10^6 cells/ml and split every 3 – 4 days. Primary pediatric mesenchymal stem cells were cultured in IMDM containing 10% FCS and 10ng/ml PDGF-BB (Peprotec).

extract_protocol:
1x10^5 cells for each condition were resuspended in lysis buffer for 10 minutes, washed, and incubated in Transposase reaction mix for 30 minutes at 37 °C (Nextera DNA sample Prep Kit, Illumina).
DNA was purified and adaptor sequences were added to the fragmented DNA by PCR to construct Illumina sequencing libraries.

treatment_protocol: EWS-FLI1 was expressed using a lentiviral vector (pLIV). Lentiviruses were produced using standard protocols. Briefly, lentiviral shRNA or expression plasmids were co-transfected with GAG/POL and VSV plasmids into 293T packaging cells using FugeneHD (Roche) to produce the virus. Viral supernatant was collected 72 hours after transfection and concentrated by ultracentrifugation using an SW41Ti rotor (Beckman Coulter) at 28,000 rpm for 120 min.

tag: cell line

value: Ewing sarcoma cell line SKMNC


title: S2_DsxF

description: Three biological replicates were produced.  The fastq files R4_s6_S2_DsxFIP_rep1.txt and  R5_s7_S2_DsxFIP_rep1.txt represent two sequence runs of the same library, from one biological replicate.  R6_s1_S2_DsxFIP_rep2.txt and R6_s6_S2_DsxFIP_rep3.txt represent sequence runs of two more independent biological replicates.  All fastq files  for S2_DsxF  were pooled before alignment to the genome.

data_processing:
Sequencing was done on a GAI instrument and base-calls were performed with Illumina software pipeline version 0.3.0
All reads were trimmed to 25 bp before alignment to the genome using Bowtie version 0.12.7
Peaks were called using SPP version 1.11 WTD method with FDR of 0.01.  Duplicate reads were removed from the libraries before peak calling with the Picard tool MarkDuplicates (version 1.95)
Bedgraph files  for S2_DSXF and S2_DsxM (input subtracted) were made using SPP's smoothed tag density function with bandwidth=200 and step=100 and tag.shift=0. The score represents the smoothed input subtracted tag density.
Genome_build: dm3, Flybase release 5 with no Uextra

source_name: S2 cells transfected with pMT5.1- DSXF -V5-His B

tag: chip antibody

value: anti-V5 (Invitrogen)

tag: treatment

value: Transfection of S2 cells was performed using Effectene Transfection Reagent (Qiagen, Valencia, CA).  pCoBlast (Invitrogen, Carlsbad, CA) was used as the selection plasmid. For the transfection procedure, cells were diluted to a density of 10^6 cells/ml and grew afterwards for about 20 hours. 1 µg expression plasmid, 50 ng pCoBlast, Effectene Transfection components and 840 µl complete Schneider’s Drosophila medium were mixed according to the manufacturer’s protocol and immediately added drop-wise onto the cells. The cells were subsequently grown in complete Schneider’s Drosophila medium for 60 hours prior to selection with 30 µg/ml blasticidin (Invitrogen, Carlsbad, CA). After 5 weeks of selection, the blasticidin-resistant cells were maintained in complete Schneider’s Drosophila medium containing 25 µg/ml of blasticidin. Expression of the recombinant proteins from the MT promoter was induced by adding copper sulfate to the medium to a final concentration of 500 μM. Before induction of expression cells were seeded to a density of 1*106 cells/ ml. Expression of the recombinant proteins was induced at a cell density of 3*10^6 cells/ ml. 60 ml of induced transfected Drosophila S2 cells expressing the recombinant proteins DSXM -V5-His or DSXF -V5-His respectively were grown in cell culture bottles for 36 hours reaching a cell density of 4.5*10^6 cells per ml.


title: S2_DsxM

description: Three biological replicates were produced. Each fastq file for S2_DSM represents an independent biological replicate. All fastq files for  S2_DsxM were pooled before alignment to the genome.

source_name: S2 cells transfected with pMT5.1- DSXM-V5-His B


title: heat20.parent30_1


title: control30.parent30_2

value: Control-selected, derived from PX443

value: C


title: ancestor30.parent30_1

value: A


title: control20.parent30_1


title: heat30.parent30_2


title: heat20.parent30_2


title: FatBody_RNAseq_TraF_12hr_R2

description:
replicate 2
EC90

data_processing:
Reads passing the Illumina chastity filter were mapped to the D. melanogaster genome and assigned to gene models using Tophat 1.4.1 (Trapnell et al., 2009) and default settings except for the following; -G=true, minimum intron length was set to 42bp (-i 42), the maximum multihits was set to 1 (-g 1).
Transcript abundance was determined using Cufflinks 2.1.1 (Trapnell et al., 2013) with maximum bundle fragments set to 10,000,000 (--max-bundle-frags 10000000) due to high read density at the Yp loci, and upper quartile normalization was used (-N).
Genome_build: dm3, Flybase release 5 with no Uextra
Supplementary_files_format_and_content: Text files with abundance measurements in FPKM

source_name: fat body

growth_protocol: 15 males and 30 virgin females were used to set up  6-8 crosses in bottles. Parent flies were flipped to new bottles after 5 days. All flies were raised and maintained at 18°C. Experimental and control flies were collected within 24 hours of eclosion and aged 5 days.

extract_protocol:
Fat body tissue was dissected from age-matched adult flies of the genotypes w1118; tra2ts2/tra2ts1 (experimental) or w1118 (control for dsxF->dsxM experiments); for dsxM->dsxF experiments the genotypes were: y1 w*; P{w+mc=UAS-Tra.F}20J7; P{w+mc=tubP-GAL80ts}7/P{w+mc=tubP-GAL4}LL7 (experimental) or P{w+mc=tubP-GAL80ts}7/P{w+mc=tubP-GAL4}LL7 (control). All samples were raised at 18°C until 5 days after eclosion when adults were shifted to either 29°C (for tra2ts) or 30°C (for UAS-TraF) for 0, 12, or 24 hours.  Total RNA was extracted from fat body dissected at room temperature (placed on ice after 30 minutes) using TRIzol Reagent following manufacturer’s protocol (Ambion Life Technologies, Carlsbad, CA, USA). Purified RNA was treated with DNAse I following manufacturer's protocol (New England Biolabs, Ipswich, MA, USA) and purified again using phenol:chloroform extraction followed by ethanol precipitation.  Duplicate RNA-seq libraries were constructed from 200 ng total RNA from independent dissection of each sample using the TruSeq RNA Sample Preparation v2 high-throughput (HT) protocol (Illumina, San Diego, CA, USA, 2011).  Libraries were sequenced on the HiSeq 2000 machine following a 76 bp single-end protocol (Illumina, San Diego, CA, USA).
True-Seq v2 protocol starting with 200 ng of total RNA.

treatment_protocol: On the 5th day, 15-20 time matched flies were separated into 3 vials designated as t=0, t=12, and t=24hrs. Flies were maintained at 29°C for 0, 12, and 24hrs and dissected at 25°C.

tag: Stage

value: adult

value: fat body

tag: genotype

value: XY; UAS-tra[F]; tub-Gal4/tub-Gal80[ts]

tag: hours at restrictive temperature


title: control20.parent30_2


title: FatBody_RNAseq_TraF_0hr_R2

description:
replicate 2
EC89


title: control30.parent30_1


title: FatBody_RNAseq_Control_24hr_R2

description:
replicate 2
EC94

value: XY; tub-Gal4/tub-Gal80[ts]


title: FatBody_RNAseq_Control_12hr_R1

description:
replicate 1
EC87


title: ancestor20.parent30_2


title: ancestor20.parent30_1


title: ancestor30.parent30_2


title: FatBody_RNAseq_Control_0hr_R2

description:
replicate 2
EC92


title: Drosophila_K79_NoSpike

growth_protocol: S2 cells were grown in SFX medium (Thermo/HyClone #SH30278.02) with 1% Penicillin/Streptomycin (Invitrogen #15140) at room temperature.  Mouse embryonic fibroblasts were grown in DMEM with 10% FBS supplemented with glutamine.

tag: antibody manu.

value: Abcam

tag: antibody catalog


title: FatBody_RNAseq_Tra2TS_24hr_R1

description:
replicate 1
EC61

tag: tissue

value: XX; tra2[ts1]/tra2[ts2]


title: FatBody_RNAseq_Control_0hr_R1

description:
replicate 1
EC86


title: FatBody_RNAseq_TraF_24hr_R2

description:
replicate 2
EC91


title: FatBody_RNAseq_Tra2TS_24hr_R2

description:
replicate 2
EC67


title: FatBody_RNAseq_Tra2TS_0hr_R2

description:
replicate 2
EC65


title: FatBody_RNAseq_Tra2TS_12hr_R1

description:
replicate 1
EC60


title: FatBody_RNAseq_TraF_24hr_R1

description:
replicate 1
EC85


title: FatBody_RNAseq_TraF_12hr_R1

description:
replicate 1
EC84


title: FatBody_RNAseq_TraF_0hr_R1

description:
replicate 1
EC83


title: FatBody_RNAseq_Control_24hr_R1

description:
replicate 1
EC107


title: FatBody_RNAseq_Control_12hr_R2

description:
replicate 2
EC93


title: FatBody_RNAseq_Tra2TS_0hr_R1

description:
replicate 1
EC59


title: FatBody_RNAseq_w1118_24hr_R1

description:
replicate 1
EC64


title: FatBody_RNAseq_w1118_24hr_R2

description:
replicate 2
EC70


title: MLL-AF9_UNC1999_SUZ12

city: Chapel Hill

first: Bowen

state: NC

address: 450 West Drive, CB7295, Lineberger Cancer Center, GREG WANG Lab, RM 31-331

department: Biochemistry and Biophysics

institute: UNC-Chapel Hill

description: Chromatin IP

data_processing:
All reads were mapped to the mouse genome (mm9) using the BWA alignment software. Unique reads mapped to a single best-matching location with no more than two mismatches were kept, which were then subject to removal of reads generated by PCR- caused duplicates using the Picard and Samtools.
Profiles of ChIP-Seq read densities were displayed in the Integrative Genomics Viewer (IGV, Broad Institute) and  the Integrative Genomics Browser (IGB, Affymetrix).
The MACS software was used for peak identification with data from input as controls and default parameters.
Genome_build: mm9
Supplementary_files_format_and_content: wig files; enrichment values in 25bp windows

source_name: MLL-AF9-puro transformed murine leukemia cell line

growth_protocol: Transformed murine leukemia progenitor cell lines were cultured in Opti-MEM base medium supplemented with 10% FBS, 1% antibiotics, 50 μM mercaptoethanol, and stem cell factor (SCF)

extract_protocol:
Lysates were clarified from sonicated nuclei and histone (or PRC2)-DNA complexes were isolated with antibody.
Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit (Part# 0801-0303). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 15 cycles and library fragments of ~250 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Genome Analyzer following the manufacturer's protocols.

treatment_protocol: Cells were treated with 3μM of the indicated compounds for 4 days, followed by ChIP-seq preparation.


title: MLL-AF9_UNC1999_H3K27ac

description: Chromatin IP

value: H3K27ac (Abcam 4729)


title: MLL-AF9_UNC2400_SUZ12

description: Chromatin IP


title: MLL-AF9_UNC1999_H3K27me3

description: Chromatin IP

value: H3K27me3 (Millipore 07-449)


title: FatBody_RNAseq_w1118_0hr_R2

description:
replicate 2
EC68


title: MV4;11_K79_0nM

tag: spike-in reference organism

value: Drosophila melanogaster

tag: spike-in cell line

tag: drug

value: DMSO

tag: duration

value: 4 days


title: MLL-AF9_UNC1999_Input_DNA

description: input DNA


title: FatBody_RNAseq_w1118_0hr_R1

description:
replicate 1
EC62


title: MLL-AF9_UNC2400_H3K27ac

description: Chromatin IP


title: MLL-AF9_UNC2400_Input_DNA

description: input DNA


title: MLL-AF9_mock_H3K27me3

description: Chromatin IP

value: mock


title: Female_Ovary_Dam_rep1

description: Two biological replicates were collected and sequenced for the treatment condition (UAS-DamDsxF) and control condition (UAS-Dam). Peaks were called by incorporating data from both replicates thus there is one peak file per tissue and sex.

data_processing:
Sequencing was done on a HiSeq2000 instrument and base-calls were performed with Illumina software pipeline CASAVA-1.8.0/RTAVersion-1.12.4/HiSeqControl-1.4.5
17 bp were trimmed from either side of the reads before alignment to the genome using Bowtie version 0.12.7 with the parameters -m1 -v2 -5 17 -3 17 --best.  Trimming was done to remove primer sequence present in many of the reads and improve % alignment to the genome.
Duplicate reads were removed from the aligned reads using with the Picard tool MarkDuplicates version 1.95.
Following duplicate-removal, reads were counted in 500 bp bins across the genome (GTF file = GSE27269_Dmel_500bp_Tiled.GTF; file available as supplementary data file on the GSE27269 series accession) using HTSeq-count version 0.5.1p2 with parameters -m union -s no
DESeq version 1.12 was used to identify 500 bp bins with statistically significant (adjusted P<0.01) enrichment (log2 Fold Change DamDsx/Dam Only > 0) of DamDsx reads compared to DamOnly reads. Both biological replicates for DamDsx and DamOnly were used to run DESeq with the parameter sharingMode = "fit-only". This step enabled incorporation of both replicates into the process of peak calling. Bins with zero reads in both DamDsx and DamOnly control were removed from the analysis. Peaks were formed by merging adjacent signficantly DamDSX-enriched bins into single units using BEDTools version 2.16.2. We included as peaks all significantly enriched isolated 500 bp bins.
BedGraph files were made using the smoothed tag density function of SPP (version 1.11) with the parameters bandwidth=200, step=100 and tag.shift=0. The score represents the smoothed DamOnly control-subtracted DamDSX tag density.
*_occupancysignal_pvalues.txt files contain the depth-normalized read count averaged between replicates for each 500 bp bin and the associated pvalue and Benjamini-Hochberg adjusted pvalue.BaseMeanA = DamOnly control. BaseMeanB = Dam-Dsx treatment. fold change and log2 fold change represent DamDsx Treatment/DamOnly Control. This file was produced with DESeq v.1.12 as described above.
genome build: dm3, Flybase release 5 with no Uextra

source_name: Adult Drosophila ovary

growth_protocol: Flies were raised and maintained at constant light, temperature and humidity (25 ˚C and 60% relative humidity) before DNA extraction. Flies were grown on a cornmeal medium supplemented with Baker's Yeast pellets.

extract_protocol:
Genomic DNA was extracted using components of Qiagen's DNEasy Blood and Tissue kit (Qiagen, Valencia, CA, USA).  Samples processed for DamID-seq were homogenized in 175 µl PBS and incubated with 200 mg of RNAse A for 2 minutes at room temperature.  Tissue was lysed with 20 µl of proteinase K and 200 µl of buffer AL for ten minutes at 70 °C. 200 µl of ethanol were added to each sample and they were transferred to the spin columns after which genomic DNA extraction continued following manufacturer's instructions with the exception of a 30 minute incubation prior to first elution and a second elution step after a 10 minute incubation. Genomic DNA extracted for DamID-chip was extracted following manufacturer's protocol except for the following modifications: a 1.5 hr incubation with lysis buffer prior to the addition of proteinase K, addition of 400 µl of Buffer AL and 300 µl of 100% ethanol, two rounds of both AW1 and AW2 and an incubation with the elution buffer for 30 minutes prior to two rounds of elution. 2.5 - 3 µg of fatbody genomic DNA and 0.3 µg of ovary genomic DNA was used for selective PCR amplification of methylated DNA. DNA was incubated with 10-30 units of DpnI in 50-100 µl of Buffer 4 (New England Biolabs , Ipswich, MA, USA). DpnI was inactivated at 80 °C for 20 minutes and digested DNA was purified through a Qiagen PCR Purification Qiaquick column following manufacturer's protocol and eluted in 30 µl ddH20. One-half of the DpnI reaction products were ligated to 40 pmol of the doublestranded DamID adaptors (top strand :  5'-CTAATACGACTCACTATAGGGCAGCGTGGTCGCGGCCGAGGA-3'; bottom strand: 5'-TCCTCGGCCG-3') for 2 hours at 16 °C with 400 units of T4 ligase (New England Biolabs, Ipswich, MA, USA) or 5 units of T4 ligase (Roche, Indianapolis, IN, USA) in a 20 µl reaction volume.  All 20 µl of the adapter-ligated DNA were then subjected to DpnII digestion with 10 units of DpnII in a 80 µl reaction volume for at least one hour. PCR amplification was performed with 20 µl of the DpnII digested DNA in an 80 µl volume with 100 pmol PCR primer (5'-TCCTCGGCCG-3'), 16 nmol of each dNTP and 1.6 µl PCR Advantage enzyme mix in 1X PCR Advantage Reaction Buffer (Clontech, Mountain View, CA, USA) or 62.5 pmol PCR primer (5'-TCCTCGGCCG-3'), 16 nmol of each dNTP, 80 nmol MgCl2 in 1X buffer with 8 units of taq polymerase (Fermentas, Pittsburgh, PA, USA). DNA was amplified with the following program: 10 minutes at 68 °C, 1 minute at 94 °C, 5 minutes at 65 °C and 15 minutes at 68 °C, followed by 3 cycles of 1 minute at 94 °C, 1 minute at 65 °C and 10 minutes at 68 °C and then 17 cycles of 1 minute at 94 °C, 1 minute at 65 °C and 2 minutes at 68 °C.  DNA was purified through a Qiaquick column (Qiagen, Valencia, CA, USA).
PCR-amplified DNA was sonicated in a 200 µl volume of Qiagen's EB buffer in a BioRuptor Sonicator (Diagenode, Denville, NJ, USA) set on high for 3 X 15 minutes in a 4 °C water bath.  Following sonication, DNA was purified through a Qiaquick column (Qiagen, Valencia, CA, USA).
20 ng of sonicated DamID-prepared DNA were used to make libraries following the protocol in the Illumina ChIP-seq Sample Preparation Kit (Illumina, San Diego, CA, USA). A gel slice of 250-350 bp was excised from the gel prior to PCR amplification.

treatment_protocol: We collected replicates for experiments by crossing the male flies bearing UAS-DamDsxF, UAS-DamDsxM  or UAS-Dam (serves as control) to female virgin w1118 flies.  In order to generate the pUAST-att-NDamMyc-dsxM/F DNA constructs, dsx male (dsxM) and dsx female (dsxF) cDNA sequences were PCR amplified using using sex-specific dsx cDNA plasmid templates (a gift from G. Lee). The forward PCR primer (common to dsx male and female) was Dsx-Dam-F (GTC GAC ATG GTT TCG GAG GAG AAC TGG) and contains a SalI site. The sex-specific reverse primers were DsxM-Dam-R2 (GGT ACC CGG GGA TTA CAC TTT GAT AGT C) and DsxF-Dam-R3 (GGT ACC CAG TTT TGA TAC CCA GAC CC) used to amplify dsxM and dsxF respectively.  dsxM and dsxF PCR products were cloned and sequenced in full prior to ligation as SalI/KpnI fragments into the XhoI and KpnI sites of pUAST-att-NDamMyc (a gift from T. Southall) generating plasmids pUAST-att-NDamMyc-dsxM and pUAST-att-NDamMyc-dsxF , respectively.  The UAS-Dam, UAS-DamDsxF, or UAS-DamDsxM constructs were inserted independently into the attP2 site on chromosome 3L band 681A-B2 between CG6310 and Mocs1 (Groth et al., 2004 Genetics 166: 1775-1782) using the PhiC31 site-directed integration method. Crosses were set up with 5 virgin females and 3 males.  Progeny that were UAS-Dam/+, UAS-DamDsxF/+ or UAS-DamDsxM/+ were collected within 24 hours of eclosion, sexed and aged for 5 days at 25  °C with w1118 flies of the opposite sex.  At 5 days of age, flies were dissected in PBS and fatbody or ovaries were collected.

value: ovary

tag: Sex

value: Female

tag: age

value: 5-day old adult


title: Female_Ovary_Dam_rep2


title: FatBody_RNAseq_w1118_12hr_R2

description:
replicate 2
EC69


title: Female_Ovary_DamDsxF_rep2

value: w1118; UAS-DamDsxF/+


title: FatBody_RNAseq_w1118_12hr_R1

description:
replicate 1
EC63


title: MLL-AF9_mock_Input_DNA

description: input DNA


title: Male_Fatbody_Dam_rep2

description: Two biological replicates were collected and sequenced for the treatment condition (UAS-DamDsxM) and control condition (UAS-Dam). Peaks were called by incorporating data from both replicates thus there is one peak file per tissue and sex.

data_processing:
Sequencing was done on a GAIIx instrument
Base calling was performed with the Illumina software pipeline 1.8.70.0
17 bp were trimmed from either side of the reads before alignment to the genome using Bowtie version 0.12.7 with the parameters -m1 -v2 -5 17 -3 17 --best.  Trimming was done to remove primer sequence present in many of the reads and improve % alignment to the genome.
Duplicate reads were removed from the aligned reads using with the Picard tool MarkDuplicates version 1.95.
Following duplicate-removal, reads were counted in 500 bp bins across the genome (GTF file = GSE27269_Dmel_500bp_Tiled.GTF; file available as supplementary data file on the GSE27269 series accession) using HTSeq-count version 0.5.1p2 with parameters -m union -s no
DESeq version 1.12 was used to identify 500 bp bins with statistically significant (adjusted P<0.01) enrichment (log2 Fold Change DamDsx/Dam Only > 0) of DamDsx reads compared to DamOnly reads. Both biological replicates for DamDsx and DamOnly were used to run DESeq with the parameter sharingMode = "fit-only". This step enabled incorporation of both replicates into the process of peak calling. Bins with zero reads in both DamDsx and DamOnly control were removed from the analysis. Peaks were formed by merging adjacent signficantly DamDSX-enriched bins into single units using BEDTools version 2.16.2. We included as peaks all significantly enriched isolated 500 bp bins.
BedGraph files were made using the smoothed tag density function of SPP (version 1.11) with the parameters bandwidth=200, step=100 and tag.shift=0. The score represents the smoothed DamOnly control-subtracted DamDSX tag density.
*_occupancysignal_pvalues.txt files contain the depth-normalized read count averaged between replicates for each 500 bp bin and the associated pvalue and Benjamini-Hochberg adjusted pvalue.BaseMeanA = DamOnly control. BaseMeanB = Dam-Dsx treatment. fold change and log2 fold change represent DamDsx Treatment/DamOnly Control. This file was produced with DESeq v.1.12 as described above.
genome build: dm3, Flybase release 5 with no Uextra

source_name: Adult male fatbody

value: Male

value: Adult abdominal fatbody with associated abdominal cuticle, oenocytes and cuticular muscles


title: Female_Fatbody_DamDsxF_rep2

source_name: Adult female fatbody

value: w1118; UAS-Dam-DsxF/+


title: Female_Fatbody_DamDsxF_rep1


title: Male_Fatbody_DamDsxM_rep2

value: w1118; UAS-DamDsxM/+


title: Female_Fatbody_Dam_rep1


title: MLL-AF9_UNC2400_H3K27me3

description: Chromatin IP


title: Male_Fatbody_DamDsxM_rep1

data_processing:
Sequencing was done on a GAIIx instrument
Base calling was performed with the Illumina software pipeline 1.6.47.1
17 bp were trimmed from either side of the reads before alignment to the genome using Bowtie version 0.12.7 with the parameters -m1 -v2 -5 17 -3 17 --best.  Trimming was done to remove primer sequence present in many of the reads and improve % alignment to the genome.
Duplicate reads were removed from the aligned reads using with the Picard tool MarkDuplicates version 1.95.
Following duplicate-removal, reads were counted in 500 bp bins across the genome (GTF file = GSE27269_Dmel_500bp_Tiled.GTF; file available as supplementary data file on the GSE27269 series accession) using HTSeq-count version 0.5.1p2 with parameters -m union -s no
DESeq version 1.12 was used to identify 500 bp bins with statistically significant (adjusted P<0.01) enrichment (log2 Fold Change DamDsx/Dam Only > 0) of DamDsx reads compared to DamOnly reads. Both biological replicates for DamDsx and DamOnly were used to run DESeq with the parameter sharingMode = "fit-only". This step enabled incorporation of both replicates into the process of peak calling. Bins with zero reads in both DamDsx and DamOnly control were removed from the analysis. Peaks were formed by merging adjacent signficantly DamDSX-enriched bins into single units using BEDTools version 2.16.2. We included as peaks all significantly enriched isolated 500 bp bins.
BedGraph files were made using the smoothed tag density function of SPP (version 1.11) with the parameters bandwidth=200, step=100 and tag.shift=0. The score represents the smoothed DamOnly control-subtracted DamDSX tag density.
*_occupancysignal_pvalues.txt files contain the depth-normalized read count averaged between replicates for each 500 bp bin and the associated pvalue and Benjamini-Hochberg adjusted pvalue.BaseMeanA = DamOnly control. BaseMeanB = Dam-Dsx treatment. fold change and log2 fold change represent DamDsx Treatment/DamOnly Control. This file was produced with DESeq v.1.12 as described above.
genome build: dm3, Flybase release 5 with no Uextra

value: w1118; UAS-Dam-DsxM/+


title: A9430196

status: Public on Oct 29 2014

city: Newark

first: Shawn

middle: R

last: Thatcher

state: DE

address: 15 Innovation Way

institute: University of Delaware

data_processing:
Illumina Casava software used for basecalling.
Tophat2 was used to map reads to genome and maize transcriptome (version 5a)
Cufflinks was used to discover novel transcripts
Cuffdiff was used to quantify known and novel transcripts
Genome_build: Maize transcriptome version 5a
Supplementary_files_format_and_content: Tab-delimited FPKM expression

organism: Zea mays

source_name: maize leaf

growth_protocol: B73, Mo17 and 88 Syn10 IBM DH lines were grown hydroponically, under long day conditions (16/8 hour day/night cycle at 26°C:/22°C) and harvested at 5 weeks of age

extract_protocol:
Total RNA was isolated from frozen maize leaf tissues via Qiagen RNeasy kit for total RNA isolation (Qiagen, Valencia, California USA).
Libraries were prepared using the TruSeq paired-end mRNA-Seq kit and  protocol from Illumina, Inc. and sequenced on the Illumina HiSeq 2500  system with Illumina TruSeq SBS v3 reagents. The resulting sequences were  trimmed based on quality scores and mapped to the maize B73 reference  genome sequence V2 and maize working gene set V5a with Tophat2.


title: A9430197


title: Male_Fatbody_Dam_rep1

treatment_protocol: We collected replicates for experiments by crossing the male flies bearing UAS-DamDsxF , UAS-DamDsxM  or UAS-Dam (serves as control) to female virgin w1118 flies.  In order to generate the pUAST-att-NDamMyc-dsxM/F DNA constructs, dsx male (dsxM) and dsx female (dsxF) cDNA sequences were PCR amplified using using sex-specific dsx cDNA plasmid templates (a gift from G. Lee). The forward PCR primer (common to dsx male and female) was Dsx-Dam-F (GTC GAC ATG GTT TCG GAG GAG AAC TGG) and contains a SalI site. The sex-specific reverse primers were DsxM-Dam-R2 (GGT ACC CGG GGA TTA CAC TTT GAT AGT C) and DsxF-Dam-R3 (GGT ACC CAG TTT TGA TAC CCA GAC CC) used to amplify dsxM and dsxF respectively.  dsxM and dsxF PCR products were cloned and sequenced in full prior to ligation as SalI/KpnI fragments into the XhoI and KpnI sites of pUAST-att-NDamMyc (a gift from T. Southall) generating plasmids pUAST-att-NDamMyc-dsxM and pUAST-att-NDamMyc-dsxF , respectively.  The UAS-Dam, UAS-DamDsxF, or UAS-DamDsxM constructs were inserted independently into the attP2 site on chromosome 3L band 681A-B2 between CG6310 and Mocs1 (Groth et al., 2004 Genetics 166: 1775-1782) using the PhiC31 site-directed integration method. Crosses were set up with 5 virgin females and 3 males.  Progeny that were UAS-Dam/+, UAS-DamDsxF/+ or UAS-DamDsxM/+ were collected within 24 hours of eclosion, sexed and aged for 5 days at 25  °C with w1118 flies of the opposite sex.  At 5 days of age, flies were dissected in PBS and fatbody or ovaries were collected.

value: 5 day old adult


title: MLL-AF9_mock_SUZ12

description: Chromatin IP


title: A9430194


title: A9430195


title: Female_Ovary_DamDsxF_rep1


title: A9430193


title: A9430192


title: A9430191


title: A9430189


title: MV4;11_K4_5nM

treatment_protocol: MV4-11 cells were treated with 5nM EPZ 5676 for 4 days.

value: Millipore

tag: concentration

value: 5 nM


title: A9430187


title: A9430186


title: A9430184


title: A9430185


title: A9430181


title: A9430179


title: A9430183


title: A9430180


title: A9430182


title: A9430190


title: Female_Fatbody_Dam_rep2


title: A9430188


title: A9430198


title: A9430178


title: A9430177


title: A9430174


title: A9430175


title: A9430172


title: A9430169


title: A9430173


title: A9430168


title: A9430170


title: A9430171


title: A9430166


title: A9430167


title: A9430163


title: A9430162


title: A9430161


title: A9430160


title: A9430157


title: A9430158


title: A9430155


title: A9430176


title: A9430154


title: A9430153


title: A9430150


title: A9430152


title: A9430148


title: A9430147


title: A9430146


title: A9430144


title: A9430142


title: A9430145


title: A9430138


title: A9430165


title: A9430141


title: A9430139


title: A9430137


title: A9430132


title: A9430159


title: A9430143


title: A9430136


title: A9430156


title: A9430135


title: A9430151


title: A9430128


title: A9430131


title: A9430149


title: A9430130


title: A9430123


title: A9430127


title: A9430121


title: A9430125


title: A9430124


title: A9430119


title: A9430122


title: A9430120


title: A9430117


title: A9430115


title: A9430114


title: A9430111


title: A9430110


title: A9430129


title: A9430108


title: A9430106


title: A9430112


title: A9430109


title: A9430107


title: A9430105


title: A9430104


title: A9430126


title: A9430102


title: A9430101


title: A9430100


title: A9430118


title: A9430113


title: Delg1 delta PCNA protein ChIP

first: zhiguo

last: zhang

address: 1130 St. Nicholas Avenue

department: Pediatric and Genetics and Development

institute: Columbia University

description: Sample name: y217

data_processing:
Reads were aligned to the yeast genome (sacCer3) using the Bowtie2 software. Only the consist pair reads were used for the further analysis.
The genome-wide reads coverage of Watson and Crick strand was calculated by BEDTools and  in-house Perl  programs.
Genome_build: sacCer3
Supplementary_files_format_and_content: bigwig files were generated for Watson and Crick strand, scores represent the normalization reads density.

organism: Saccharomyces cerevisiae

source_name: whole cell

growth_protocol: For G1 cell synchronization, alpha factor was added at cell density OD600=0.4-0.5 to finial concentration at 5ug/ml for wild type Bar1 and 50ng/ml for bar1 mutants.  After 3hr arrest at 30°C, the cell was released to fresh YPD medium with or without 200mM hydroxyurea.

extract_protocol:
the collected samples were incubated with 1% fresh-made formaldehyde at 25°C for 20mins, and quenched with 0.125M Glysine for 5mins.  The harvested cells were lysed with glass beads, and the chromatin pellet were washed and sonicated. The average DNA fragment size is about 200-400bp.  Sheared chromatin was immunoprecipitated with an protein-specific antibody. The chromatin was reverse-cross-linked and immunoprecipitated.  DNA was exacted with the Chelex-100 method
ssDNA library preparation method was modified from previous one (Meyer et al., 2012). To preserve the strand information, we use a specific RNA ligase, which also can ligated the ssDNAs in a sequence-independent way. The ssDNA library strand information was kept by sequential 3’ end and  5’ end ligation.

tag: media

value: YPD+HU

tag: cell cycle stage

value: S

value: Anti-FLAG (SIGMA,F1804)


title: 84min Pol δ protein ChIP

description: Sample name: y223

value: YPD

tag: chip antibody


title: 84min Pol δ ChIP-eSPAN

description: Sample name: y226


title: 84min Pol δ BrdU IP

description: Sample name: y220

value: Anti-BrdU (BD BioScience, 347580)


title: Delg1 delta PCNA ChIP-eSPAN

description: Sample name: y218


title: 96min Pol δ BrdU IP

description: Sample name: y221


title: 96min Pol δ protein ChIP

description: Sample name: y224


title: A9430103


title: 96min Pol δ ChIP-eSPAN

description: Sample name: y227


title: 72min Pol δ protein ChIP

description: Sample name: y222


title: 72min Pol δ ChIP-eSPAN

description: Sample name: y225


title: 72min Pol δ BrdU IP

description: Sample name: y219


title: 84min PCNA ChIP-eSPAN

description: Sample name: y186


title: 96min PCNA BrdU IP

description: Sample name: y181


title: 84min PCNA protein ChIP

description: Sample name: y183


title: 96min PCNA protein ChIP

description: Sample name: y184


title: 72min PCNA protein ChIP

description: Sample name: y182


title: 72min PCNA ChIP-eSPAN

description: Sample name: y185


title: 96min Pol ɛ ChIP-eSPAN

description: Sample name: y85


title: 96min Pol ɛ BrdU IP

description: Sample name: y83


title: 72min PCNA BrdU IP

description: Sample name: y179


title: 84min PCNA BrdU IP

description: Sample name: y180


title: 96min Pol ɛ protein ChIP

description: Sample name: y84


title: Delg1 delta PCNA BrdU IP

description: Sample name: y216


title: 96min PCNA ChIP-eSPAN

description: Sample name: y187


title: 96min Pol ɛ input

description: Sample name: y82


title: 84min Pol ɛ ChIP-eSPAN

description: Sample name: y81


title: 84min Pol ɛ protein ChIP

description: Sample name: y80


title: 84min Pol ɛ BrdU IP

description: Sample name: y79


title: 72min Pol ɛ protein ChIP

description: Sample name: y76


title: 72min Pol ɛ BrdU IP

description: Sample name: y75


title: 72min Pol ɛ input

description: Sample name: y74


title: 72min Pol ɛ ChIP-eSPAN

description: Sample name: y77


title: Rfc1 protein ChIP repeat

description: Sample name: y209


title: Rfc1 ChIP-eSPAN repeat

description: Sample name: y210


title: Rfc1 ChIP-eSPAN

description: Sample name: y137


title: Rfc1 BrdU IP repeat

description: Sample name: y208


title: FatBody_RNAseq_Tra2TS_12hr_R2

description:
replicate 2
EC66

value: adult

value: fat body

value: XX; tra2[ts1]/tra2[ts2]


title: Rfa1 protein ChIP repeat

description: Sample name: y205


title: Rfc1 input

description: Sample name: y134


title: Rfa1 ChIP-eSPAN repeat

description: Sample name: y206


title: Rfa1 protein ChIP

description: Sample name: y122


title: Rfa1 BrdU IP repeat

description: Sample name: y204


title: Pol32 ChIP-eSPAN

description: Sample name: y28


title: Jurkat_K4_75%_R2

value: Drosophila melanogaster

tag: cell line

value: Jurkat

value: DMSO / EPZ

value: 4 days

value: 20 uM


title: Rfa1 BrdU IP

description: Sample name: y121


title: Rfa1 input

description: Sample name: y120


title: Pol32 protein ChIP

description: Sample name: y27


title: Pol32 BrdU IP

description: Sample name: y26


title: Pol δ ChIP-eSPAN

description: Sample name: y89


title: Pol δ protein ChIP

description: Sample name: y88


title: Pol δ BrdU IP

description: Sample name: y87


title: Pol32 input

description: Sample name: y25


title: Pol δ input

description: Sample name: y86


title: 84min Pol ɛ input

description: Sample name: y78


title: Pol ɛ BrdU IP

description: Sample name: y18


title: Rfc1 BrdU IP

description: Sample name: y135


title: Pol α ChIP-eSPAN repeat

description: Sample name: y41


title: Pol ɛ input

description: Sample name: y17


title: Pol ɛ protein ChIP

description: Sample name: y19


title: Rfc1 protein ChIP

description: Sample name: y136


title: Rfa1 ChIP-eSPAN

description: Sample name: y123


title: Pol α protein ChIP repeat

description: Sample name: y40


title: Pol α input repeat

description: Sample name: y38


title: Pol α protein ChIP

description: Sample name: y23


title: Pol α ChIP-eSPAN

description: Sample name: y24


title: PCNA protein ChIP repeat

description: Sample name: y128


title: PCNA BrdU IP repeat

description: Sample name: y127


title: PCNA input repeat

description: Sample name: y126


title: Pol α input

description: Sample name: y21


title: Pol α BrdU IP

description: Sample name: y22


title: Mcm10 ChIP-eSPAN

description: Sample name: y69

value: Anti-HA (abcam,ab16918)


title: PCNA ChIP-eSPAN repeat

description: Sample name: y129


title: PCNA protein ChIP

description: Sample name: y48


title: PCNA BrdU IP

description: Sample name: y47


title: Mcm10 input

description: Sample name: y66


title: Mcm6 ChIP-eSPAN

description: Sample name: y65


title: Mcm10 protein ChIP

description: Sample name: y68


title: 12-106 positive

city: Coventry

first: Jan

middle: J

last: Brosens

address: University Hospital Coventry and Warwickshire

department: Warwick Medical School

country: United Kingdom

institute: University of Warwick

data_processing:
Transcriptomic maps were created by aligning single-end reads with tophat-2.0.4 against the University of California Santa Cruz (UCSC) hg19 reference transcriptome from the Illumina iGenomes resource.
Gene counts were estimated using HTSeq-0.5.3p3.
Genome_build: hg19
Supplementary_files_format_and_content: Table containing Transcripts per million (TPM) for each sample.  TPM were calculated according to Wagner et al. (2012) http://www.ncbi.nlm.nih.gov/pubmed/22872506

source_name: Endometrial Stromal Cell

growth_protocol: Single cell suspensions of human endometrial stromal cells (HESCs) were isolated from mid-luteal biopsies. Stromal single cell suspensions were layered over Ficoll-Paque PLUS and centrifuged to remove erythrocytes. The medium / Ficoll-Paque PLUS interface, containing stromal cells, was carefully aspirated, washed with DMEM/F-12 medium, and then subjected to magnetic bead separation to isolate W5C5+ and W5C5- cells using anti-W5C5 (SUSD2) antibody. Freshly isolated W5C5+ and W5C5- cells were expanded in growth medium consisting of DMEM/F12 with 10% dextran-coated charcoal-treated fetal bovine serum , 1% L-glutamine , 1% antibiotic-antimycotic solution , insulin (2 µg / ml), estradiol (1 nM) and basic fibroblast growth factor (10 ng / ml).  Confluent cultures were harvested by lysis in RNA STAT-60.

extract_protocol:
Total RNA was purified from RNA STAT-60 lysate according to the manufacturer's protocol.  Quantity and purity were determined using the Nanodrop ND1000 spectrophotometer.  3 μg each sample was treated with Amplification Grade DNase I (Invitrogen) according to manufacturer's instructions.  Samples were cleaned up by sodium acetate/ethanol precipitation and resolubilised in nuclease-free water.  From this point onwards, all steps were performed by Source Bioscience (Nottingham, UK).  Sample quality control was performed using the Agilent 2100 Bioanalyser.  All samples had RIN ≥ 9.1
The mRNA libraries were prepared in accordance to the Illumina TruSeq RNA sample preparation v2 guide (September 2012, Rev. D) for Illumina Paired-End Multiplexed Sequencing. According to the Illumina mRNA libraries preparation protocol, poly-A mRNA in the tRNA samples were first purified using Illumina poly-T oligo-attached magnetic beads and two rounds of purification. During the second elution of the poly-A RNA, the mRNA were also fragmented and primed with random hexamers for cDNA synthesis. Cleaved mRNAs were reverse transcribed into first strand cDNA using reverse transcriptase and random primers. The RNA template was then removed and a replacement strand synthesized to generate double-stranded cDNA. Following the standard protocol, after the first and second strands cDNA synthesis, ends were repaired, dA base added, and Illumina Indexing adapters were ligated. Finally, cDNA fragments that have adapter molecules on both ends underwent 15 cycles of PCR to amplify the amount of prepared material.

tag: cell type

value: Endometrial Stromal Cell

tag: culture progenitor w5c5 status

value: W5C5 negative


title: Mcm6 protein ChIP

description: Sample name: y64


title: Mcm6 BrdU IP

description: Sample name: y63


title: 12-99 positive


title: Mcm6 input

description: Sample name: y62


title: 12-101 negative

value: W5C5 positive


title: Mcm4 protein chip

description: Sample name: y55

value: YPD+HU


title: Mcm4 input

description: Sample name: y54

value: S


title: 12-106 negative


title: PCNA ChIP-eSPAN

description: Sample name: y49


title: 12-91 positive


title: 12-91 negative


title: Cdc45 BrdU IP

description: Sample name: y51


title: Cdc45 protein ChIP

description: Sample name: y52


title: G1 Mcm6 input

description: Sample name: y60


title: G1 Mcm4 input

description: Sample name: y56


title: G1 Mcm4 protein chip

description: Sample name: y58

value: Anti-FLAG (SIGMA,F1804)


title: Rpb3U_Input-2

status: Public on Oct 28 2014

first: David

middle: Johannes

last: Clark

address: 6 Center Drive Bldg 6A Rm 2A02

department: DDB

institute: NICHD, NIH

description: Biological replicate 2

data_processing:
Nucleosome sequences were aligned to the yeast genome (SacCer2) using Bowtie with default settings.
Nucleosome occupancy maps were created using programs described by Cole et al. (2011) Nucleic Acids Res. 39, 9521-9535.
The processed data were converted to the bigwig files supplied here: different DNA length exclusion limits were used in the processing (listed with Processed Data Files)
The .bw files for the Rpb3 samples include only unique read pairs.
Genome_build: SacCer2

source_name: Input: Rpb3 (no 3AT) Experiment 3

growth_protocol: All strains were grown to mid-log phase (OD600 about 0.8) at 30 degC in synthetic complete medium (untreated), or in synthetic complete medium without histidine followed by addition of 10 mM 3AT for 20 min (3AT-induced).

extract_protocol: The DNA was repaired using the DNA repair kit from New England Biolabs. The repaired DNA was processed for paired-end sequencing according to the Illumina protocol.

treatment_protocol: Formaldehyde-fixed and sonicated

tag: strain

tag: genotype

tag: treatment

value: no 3-aminotriazole


title: 443A_Input-1

description: Biological replicate 1

source_name: Input: HA-H2B (+3AT) Experiment 2

value: MATa can1-100 his3-11 leu2-3,112 lys2delta trp1-1 ura3-1 hta2-htb2delta::TRP1 HA-6His-HTB1::HIS3


title: Rpb3A_IP-2

description: Biological replicate 2

source_name: IP: Rpb3 (+3AT) Experiment 3


title: Rpb3U_Input-1

description: Biological replicate 1

source_name: Input:Rpb3 (no 3AT) Experiment 2


title: 443A_Input-2

description: Biological replicate 2

source_name: Input: HA-H2B (+3AT) Experiment 3


title: Rpb3A_Input-1

description: Biological replicate 1

source_name: Input: Rpb3 (+3AT) Experiment 2


title: Rpb3U_IP-1

description: Biological replicate 1

source_name: IP: Rpb3 (no 3AT) Experiment 2


title: 443A_IP-1

description: Biological replicate 1

source_name: IP: HA-H2B (+3AT) Experiment 2


title: 443U_IP-2

description: Biological replicate 2

source_name: IP: HA-H2B (No 3AT) Experiment 3


title: 443U_Input-2

description: Biological replicate 2

source_name: Input: HA-H2B (no 3AT) Experiment 3


title: 443U_Input-1

description: Biological replicate 1

source_name: Input: HA-H2B (no 3AT) Experiment 2


title: Cdc45 ChIP-eSPAN

description: Sample name: y53


title: Rpb3A_IP-1

description: Biological replicate 1

source_name: IP: Rpb3 (+3AT) Experiment 2


title: Rpb3U_IP-2

description: Biological replicate 2

source_name: IP: Rpb3 (No 3AT) Experiment 3


title: Rpb3A_Input-2

description: Biological replicate 2

source_name: Input: Rpb3 (+3AT) Experiment 3


title: 12-99 negative


title: G1 Mcm6 protein chip

description: Sample name: y61

value: YPD


title: Pol ɛ ChIP-eSPAN

description: Sample name: y20


title: PCNA input

description: Sample name: y46


title: 439A_IP-1

description: Biological replicate 1

source_name: IP: HA-H4 (+3AT) Experiment 2

value: MATa ade2-1 leu2-3,112 trp1-1 ura3-1 hht1-hhf1delta::LEU2 URA3::HHT2-HA-HHF2


title: 439A_IP-2

description: Biological replicate 1

source_name: IP: HA-H4 (+3AT) Experiment 3


title: 439A_Input-2

description: Biological replicate 2

source_name: Input: HA-H4 (+3AT) Experiment 3


title: 439A_Input-1

description: Biological replicate 2

source_name: Input: HA-H4 (+3AT) Experiment 2


title: Mcm10 BrdU IP

description: Sample name: y67

source_name: whole cell

value: Anti-BrdU (BD BioScience, 347580)


title: 443A_IP-2

description: Biological replicate 2

source_name: IP: HA-H2B (+3AT) Experiment 3


title: 111J-2_iNCP-1

description: For replicates, see GSE 26943

source_name: nucleosome core particles (+3AT)

treatment_protocol: MNase limit digest


title: 439U_Input-1

description: Biological replicate 1

source_name: Input: HA-H4 (no 3AT) Experiment 2


title: 439U_IP-1

description: Biological replicate 1

source_name: IP: HA-H4 (no 3AT) Experiment 2


title: OP9_DLL1_donor4

status: Public on Oct 27 2014

city: Gent

first: Annelynn

last: Wallaert

address: De Pintelaan 185

country: Belgium

institute: UGent

data_processing:
Illumina Casava1.7 software used for basecalling.
after QC with FastQC v0.10.1, reads were mapped to hg19 whole genome using tophat v2.0.10
A count file for each sample was generated using HTSeq-0.6.1 and Ensembl igenomes GTF file as reference.
Count files were merged into a count matrix and normalized in R using DESeq2
Differential expression analysis was done with DESeq2
Genome_build: hg19
Supplementary_files_format_and_content: tab-delimited text files include normailzed count values for each Sample

source_name: CD34+, OP9-DLL1, donor4

extract_protocol:
Total RNA was harvested with the miRNeasy mini kit (Qiagen) with Dnase treatment on-column.
RNA libraries were prepared for sequencing using standard Illumina protocols with poly-A enrichment

treatment_protocol: CD34+ thymocytes were purified out of pediatric thymus samples using MACS and seeded onto confluent OP9-GFP (control) or OP9-DLL1 plates for 48h in α-MEM media supplemented with 20% heat-inactivated FCS plus 100U penicilin, 100 µg/ml streptomycin, 2 mM L-glutamine and cytokines SCF, Flt-3 and IL-7 at 5 ng/ml each.

tag: co-culture


title: 111H_cArray-2

description: Biological replicate 2

source_name: under-digested chromatin (no 3AT)

treatment_protocol: MNase under-digest


title: 111J-2_cArray-1

description: Biological replicate 1


title: 439U_IP-2

description: Biological replicate 2

source_name: IP: HA-H4 (No 3AT) Experiment 3


title: OP9_GFP_donor4

source_name: CD34+, OP9-GFP, donor4


title: CUTLL1_GSI_48h_3

growth_protocol: Cells were grown in RPMI supplemented with 20% fetal bovine serum, 1% glutamin and 1% penicilin/streptomycin

treatment_protocol: Cells were seeded at 1*10^6 cells/ml and treated with either DMSO or 1 µM of Compound E. Cells were harvested 12 and 48h after treatment.

value: GSI

tag: time


title: OP9_GFP_donor3

source_name: CD34+, OP9-GFP, donor3


title: 111J-2_cNCP-1

description: For replicates, see GSE 26943

source_name: nucleosome core particles (no 3AT)


title: ChIP FOXL2

city: GIF SUR YVETTE

first: Adrien

last: Georges

address: CEA/SACLAY

country: France

institute: CNRS

description: DNA ChiPed with anti-FOXL2 C-terminal antibody (from lab)

data_processing:
Base calling, CASAVA-1.8.2
Read alignment, Bowtie Galaxy version v1.1.2, Full list of parameters : Will you select a reference genome from your history or use a built-in index?:indexed;Select a reference genome:mm9female;Is this library mate-paired?:single;FASTQ file:1: Foxl2;Bowtie settings to use:full;Skip the first n reads (-s):0;Only align the first n reads (-u):-1;Trim n bases from high-quality (left) end of each read before alignment (-5):0;Trim n bases from low-quality (right) end of each read before alignment (-3):0;Maximum number of mismatches permitted in the seed (-n):2;Maximum permitted total of quality values at mismatched read positions (-e):70;Seed length (-l):28;Whether or not to round to the nearest 10 and saturating at 30 (--nomaqround):Round to nearest 10;Number of mismatches for SOAP-like alignment policy (-v):-1;Whether or not to try as hard as possible to find valid alignments when they exist (-y):Do not try hard;Report up to n valid alignments per read (-k):1;Whether or not to report all valid alignments per read (-a):Do not report all valid alignments;Suppress all alignments for a read if more than n reportable alignments exist (-m):1;Write all reads with a number of valid alignments exceeding the limit set with the -m option to a file (--max):False;Write all reads that could not be aligned to a file (--un):False;Whether or not to make Bowtie guarantee that reported singleton alignments are 'best' in terms of stratum and in terms of the quality values at the mismatched positions (--best):noBest;Maximum number of backtracks permitted when aligning a read (--maxbts):125;Override the offrate of the index to n (-o):-1;Seed for pseudo-random number generator (--seed):-1;Suppress the header in the output SAM file:False;
Unmapped reads removal, Filter SAM galaxy version 1.0.0
Peak calling, MACS Galaxy version 1.0.1, Full list of parameters : Tag size:50;Band width:160;Pvalue cutoff for peak detection:0,00001;Select the regions with MFOLD high-confidence enrichment ratio against background to build model:20;Parse xls files into into distinct interval files:False;Save shifted raw tag count at every bp into a wiggle file:wig;Extend tag from its middle point to a wigextend size fragment.:-1;Resolution for saving wiggle files:20;Use fixed background lambda as local lambda for every peak region:False;3 levels of regions around the peak region to calculate the maximum lambda as local lambda:1000,5000,10000;Build Model:create_model;Diagnosis report:no_diag;Perform the new peak detection method (futurefdr):True;
Genome_build: mm9
Supplementary_files_format_and_content: .bed file contain 5 columns : chr, peakStart, peakEnd, peakID and score
Supplementary_files_format_and_content: .wig file contain unnormalized read mapping abundancies (20bp digits)

source_name: cells from preantral follicles maintained for 9 days in vitro

growth_protocol: The ovaries of 20 8-week old female Swiss mice were dissected and digested in DMEM/F-12 medium (Gibco) containing 1mg/mL collagenase (Sigma-Aldrich), 0,5mg/mL BSA (Sidma-Aldrich) and 2000 U/mL DNase I (Thermo-Fisher), in a ratio of 500µL per ovary, at 37°C for 30 min. Ovaries were progressively dispersed during digestion using pipette tips of decreasing size to flush the organ debris. The mix was then diluted in 10 volumes of DMEM/F-12 medium, filtered using a 500 µm membrane, spinned at 300 g. The pellet was rinsed in cold PBS and spinned again, and the pelleted debris were resuspended in a PBS/Ficoll mix of density 1.10. A PBS/Ficoll gradient was made by depositing PBS/Ficoll layers of density 1.08, 1.07, 1.06, 1.05, 1.03 and 1. The gradient was spinned at 1500 g for 20min, and 1 mL fractions were collected and diluted in 5 mL culture medium (i.e. DMEM/F-12 supplemented with 4% FBS, 1% Insulin-Transferrin-Selenium-X supplement, 1% Penicillin-Streptomycine (Gibco) and 10-7M androstenedione (Sigma-Aldrich). We then manually collected intact preantral follicles, characterized by the presence of several layers of granulosa cells around the oocyte and the absence of antrum. They mainly migrated to the 1.07/1.06 and 1.06/1.05 interfaces. Follicles were then rinsed, transferred to culture plates and grown overnight. On day 1, they were dispersed using trypsin and then cultured in monolayers. Culture medium was changed on day 3, and cells were passed on days 5 and 7.

extract_protocol:
Cells grown to confluence in three 150-mm plates were fixed in situ with 1% Formaldehyde for 10 min at room temperature, fixation was then blocked with 125 mM glycine, cells were rinsed and scrapped in PBS. Cells were first lysed in Lysis buffer A (50 mM Hepes pH 7.5, NaCl 140 mM, EDTA 1 mM, glycerol 10% NP-40 0.5% and Triton X-100 0.25%), then spinned and rinsed in lysis buffer B (Tris 10 mM pH 7.5, NaCl 200 mM, EDTA 1 mM, EGTA 0.5 mM), centrifugated again and finally resuspended in lysis buffer C (Tris 10 mM pH 7.5, NaCl 100 mM, EDTA 1 mM, EGTA 0.5 mM, Na–deoxycholate 0.5%, N-lauryl sarcosine 0.5%). Chromatin was sheared in a Bioruptor sonicator, used at high power with 30/30 s cycles for 20 min, then 1% Triton X-100 was added to lysates, and cell debris were centrifugated. In parallel, 30 μg anti-FOXL2 were incubated with 200 μl ProteinG Dynabeads for 4 h in PBS containing 0.1% BSA. Lysates were incubated with the beads overnight, beads were then washed five times in wash buffer (50 mM Hepes pH 7.6, 500 mM LiCl, 1 mM EDTA, 0.7% Na–deoxycholate, 1% NP-40) and once in TBS. DNA–protein complexes were then eluted in 10 mM Tris pH 8, 1 mM EDTA, 1% SDS at 65 °C for 15 min. Crosslinks were reversed at 65 °C overnight, then recovered DNA was treated with RNaseA, proteinase K and purified using phenol–chloroform. Input samples underwent to the same procedure.
Adapters were ligated using a SPRI-TE Nucleic Acid Extractor Robot with a SPRIworks Fragment Library Kit I and adapters from TruSeq DNA LT Sample Prep kit (Illumina). Ligated fragments were amplified with C and D primers(C primer :5’-AATGATACGGCGACCACCGAGATCTACAC-3’b and D primer : 5’-CAAGCAGAAGACGGCATACGAGAT-3’) using Phusion HF polymerase (Thermo Fisher Scientific) for 14 PCR cycles. 200 to 400 bp fragments were selected on gel and purified on a Qiagen column.

value: Swiss

tag: gender

value: female

tag: tissue

value: ovary


title: CUTLL1_DMSO_48h_3

value: DMSO


title: OKF6-TERT1R

first: Katarzyna

middle: Maria

last: Marcinkiewicz

address: 1300 York Ave. Room E405

department: Pharmacology

institute: Weill Cornell Graduate School of Medical Sciences

data_processing:
Sequence data (base call files or bcl files) generated from the sequencer are  converted to FASTQ files using the Illumina CASAVA 1.8.2 pipeline.
The ERRBS pipeline used has been described (Akalin et. al., 2012).
Genome_build: hg19

source_name: immortalized oral keratinocytes

growth_protocol: OKF6-TERT1R cells were cultured in Keratinocyte-SFM medium (# 10744019, Gibco, CA, USA), supplemented with 0.3 mM CaCl2, 0.2 ng/ml EGF, penicillin/streptomycin/L-glutamine (#10378-016, Gibco, CA, USA) and bovine pituitary extract (BPE).  SCC-9 cells were cultured in Dulbecco's Modified Eagle’s Medium: Nutrient Mixture F-12 (DMEM/F-12; Gibco, CA, USA), supplemented with 10% fetal calf serum and 400 ng/ml hydrocortisone.

extract_protocol:
Total genomic DNA was extracted from OKF6-TERT1R and SCC-9 cells using DNeasy Blood & Tissue Kit (Qiagen, Germantown, MD, USA), including an RNAse treatment step.
as decribed: Akalin A, Garrett-Bakelman FE, Kormaksson M, Busuttil J, Zhang L, Khrebtukova I, Milne TA, Huang Y, Biswas D, Hess JL, et al. (2012). Base-pair resolution DNA methylation sequencing reveals profoundly divergent epigenetic landscapes in acute myeloid leukemia PLoS Genet 8, e1002781.

treatment_protocol: The cells were plated in 10 cm2 tissue culture plates at the density of 2 × 106 cells/plate.

tag: cell


title: SCC-9

source_name: tongue squamous cell carcinoma cell line


title: Cdc45 input

description: Sample name: y50


title: 111J-2_iArray-1

description: Biological replicate 1

source_name: under-digested chromatin (+3AT)


title: CUTLL1_DMSO_12h_3


title: CUTLL1_GSI_12h_3


title: Input

description: Input DNA submitted to same decrosslink and preparation as ChIPped DNA

value: cells from preantral follicles maintained for 9 days in vitro


title: CUTLL1_GSI_48h_2


title: 439U_Input-2

description: Biological replicate 2

source_name: Input: HA-H4 (no 3AT) Experiment 3


title: CUTLL1_DMSO_48h_2


title: 443U_IP-1

description: Biological replicate 1

source_name: IP: HA-H2B (no 3AT) Experiment 2


title: OP9_DLL1_donor3

source_name: CD34+, OP9-DLL1, donor3


title: CUTLL1_GSI_48h_1


title: 111H_iArray-2

description: Biological replicate 2


title: CUTLL1_DMSO_48h_1


title: CUTLL1_DMSO_12h_2


title: CUTLL1_DMSO_12h_1


title: CUTLL1_GSI_12h_1


title: NAIVE_1

city: New Haven

first: Vince

last: Schulz

state: CT

address: 333 Cedar St. LMP 4085

department: Department of Pediatrics

institute: Yale University

data_processing:
Illumina Casava 1.8.2 software was used for basecalling.
FASTQ format sequencing reads were aligned to the hg19 genome using TopHat Version 2.0.4 software with default parameters
Reads in every gene were counted using htseq-count (http://www-huber.embl.de/users/anders/HTSeq/doc/overview.html). Genes with less than 3 samples with >1 read counts/million (cpm) were called not expressed and were not used in further analysis. The read count data was normalized using the TMM method. Differentially expressed genes were identified using the edgeR exact test.
Genome_build: hg19
Supplementary_files_format_and_content: Counts per gene and log2 RPKM

source_name: Naïve T cell

extract_protocol:
To obtain primary human Tfh and Teff cells, human tonsils were cut into small sections and homogenized by crushing followed by straining through a 40mM nylon filter. CD4+ T lymphocytes were enriched using a negative selection biotin-based magnetic separation kit (EasySep; StemCell Technologies) prior to cell surface staining with the antibodies to:  CD4 (clone RPA-T4), CD45RA (clone HI-100), TCRb (clone IP26), PD-1 (clone EH12.1), CXCR5 (clone RF8B2), and PSGL-1 (clone KPL-1)(all from BD Biosciences). Staining of CXCR5 was performed at room temperature (25°C) with 1 hour incubation. Stained and rinsed cells were analyzed using an LSRII Multilaser Cytometer (BD Biosciences) or specific populations sorted using a FACSAria (BD Bioscience). RNA was isolated from lymphocytes and prepared for RNA-seq analyses as described (Su MY, Steiner LA, Bogardus H, et al. Identification of biologically relevant enhancers in human erythroid cells. J Biol Chem. 2013;288(12):8433-8444).
Paired end RNA libraries were prepared for sequencing using standard Illumina protocols at the Yale Center for Genome Analysis.

tag: t cell type

value: Naïve

value: Human tonsil

tag: cell surface markers


title: NAIVE_3


title: TEFF_2

source_name: TEFF T cell

value: TEFF


title: NAIVE_4


title: TFH_2

source_name: TFH T cell

value: TFH


title: TEFF_1


title: TEFF_3


title: TFH_H3K27me3

data_processing:
Illumina Casava 1.8.2 software was used for basecalling.
FASTQ format sequencing reads were mapped to the human genome (hg19 NCBI Build 37) using the BWA version 0.6.1 alignment program with default parameters.
The Model-based Alignment of ChIP-Seq (MACS) program version 1.4.0rc2 was used to identify H3K4me1 and H3K27Ac peaks using default parameters.
The MACS2 program version 2.0.10.20131216 was used to identify broad regions bound by H3K27me3
Genome_build: hg19
Supplementary_files_format_and_content: The H3K4me1 and H3K27Ac macs1.4 peaks.bed output files have a fifth field that is -10*log10(pvalue).  H3K27me3 MACS2 output is in broadPeak format.

extract_protocol:
To obtain primary human Tfh and Teff cells, human tonsils were cut into small sections and homogenized by crushing followed by straining through a 40mM nylon filter. CD4+ T lymphocytes were enriched using a negative selection biotin-based magnetic separation kit (EasySep; StemCell Technologies) prior to cell surface staining with the antibodies to:  CD4 (clone RPA-T4), CD45RA (clone HI-100), TCRb (clone IP26), PD-1 (clone EH12.1), CXCR5 (clone RF8B2), and PSGL-1 (clone KPL-1)(all from BD Biosciences). Staining of CXCR5 was performed at room temperature (25°C) with 1 hour incubation. Stained and rinsed cells were analyzed using an LSRII Multilaser Cytometer (BD Biosciences) or specific populations sorted using a FACSAria (BD Bioscience).  ChIP assays were performed as previously described with minor variations (Steiner, L. A., Maksimova, Y., Schulz, V., Wong, C., Raha, D., Mahajan, M. C., Weissman, S. M., and Gallagher, P. G. (2009) Mol Cell Biol). 1 × 107 cells were cross-linked with 1% formaldehyde for 10 min at room temperature, followed by Dounce homogenization. Cross-linked nuclei were isolated, followed by sonication with a Covaris S2 shearing device (duty cycle 10%, intensity 5, 15 minutes), to obtain chromatin-containing DNA fragments with an average size of ~200-400bp. For each ChIP, 4 µg of antibody was used. Samples were immunoprecipitated with antibodies against monomethyl histone 3 lysine 4 (H3k4me1, Abcam ab8895), trimethyl histone 3 lysine 27 (Abcam ab6002), and acetyl histone 3 lysine 27 (H3K27Ac, Abcam ab4729).  The antigen-antibody complex was captured on protein G beads, washed four times with radioimmunoprecipitation assay buffer, and then washed with phosphate-buffered saline. The DNA-protein complex was eluted from the protein G beads with 0.1 NaHCO3/1% SDS at 65°C, and cross-linking of DNA-protein adducts was reversed by 4 hours of incubation at 65°C. After proteinase K and RNase digestion of the reverse-cross-linked sample, DNA was cleaned with the QIAquick PCR Purification Kit (Qiagen) according to manufacturer instructions.
ChIP libraries were prepared using the KAPA Library Preparation Kit (KAPA Biosystems) per the manufacturer's protocol. Briefly, ChIP DNA was end repaired, followed by addition of an adenine base at the 3' and NEXTflex ChIP-Seq Barcodes (BiooScientific) adaptor ligation. The modified DNA was then size selected on a 2% agarose and purified by using a gel extraction kit (Qiagen).  One microliter of size-selected adaptor ligated DNA was initially analyzed by SYBR Green Q-PCR to determine the number of cycles required to reach 50% of complete amplification.  Adaptor ligated DNA was PCR amplified with one initial heating step of 98°C for 45 second, followed by the pre-determined number of cycles with a melting temperature of 98°C for 15 s, an annealing temperature of 65°C for 30 s, and a product extension at 72°C for 30 s. At the end of the amplification, a final extension at 72°C for 1 min was performed. Amplified ChIP DNA was subjected to deep sequencing using standard Yale Center for Genome analysis protocols.


title: TEFF_H3K4me1


title: TFH_H3K4me1


title: TFH_1


title: NAIVE_2


title: TEFF_H3K27Ac


title: TEFF_TI

value: TI


title: TFH_H3K27Ac


title: TEFF_H3K27me3


title: HAEC-Input-ChIP-Seq

city: Kuopio

first: Minna

middle: U

last: Kaikkonen

department: A.I. Virtanen Institute, Department of Biotechnology and Molecular Medicine

country: Finland

institute: University of Eastern Finland

description: chromatin immunoprecipitation

data_processing:
Libraries were sequenced for 36 or 50 cycles on an Illumina Genome Analyzer II or HiSeq 2000, respectively, according to the manufacturer’s instructions.
For ChIP-Seq and GRO-Seq samples (single end reads), reads were aligned to the mm9 genome (NCBI build using bowtie (v0.12.7), keeping only reads that mapped best to a single, unique location. RNA-Seq samples were aligned using Tophat.
GRO-Seq and RNA-Seq results were trimmed to remove A-stretches originating from the library preparation. Data analysis was performed using HOMER and the detailed instructions for analysis can be found at http://biowhat.ucsd.edu/homer/
For ChIP-Seq, aligned read files were analyzed with HOMER (http://biowhat.ucsd.edu/homer/) to find enriched H3K4me2 regions. Each ChIP-Seq experiment was normalized to a total of 10^7 uniquely mapped tags by adjusting the number of tags at each position in the genome to the correct fractional amount given the total tags mapped.
For TCC analysis, reads from paired-end sequencing were separately filtered and aligned to the genome using bowtie. Each read was trimmed from the 3’ end after GATC site and minimum read length was set to 25 bp. Paired-end reads were connected and additional filtering steps were performed using HOMER 4.3. Read pairs with exact same ends were only considered once and read pairs were removed if they were separated by less than 1.5x the estimated sequencing insert length to remove likely continuous genomic fragments or re-ligation events. Paired-end reads originating from regions of unusually high tag density were left out by removing reads from 10 kb regions that contain more than 5 times the average number of reads. Reads were filtered according to their distance from a restriction site keeping only those read pairs that had both of their reads aligning at most 1.5x fragment length estimate from a restriction site. Reads were removed if their ends form a self-ligation with adjacent restriction sites.
Genome_build: hg19
Supplementary_files_format_and_content: Positions for H3K4me2 regions are provided for ChIP-Seq.

source_name: human aortic endothelial cells

growth_protocol: HUVECs were isolated from umbilical cords obtained from the maternity ward of the Kuopio University Hospital by the approval of the Kuopio University Hospital Ethics Committee. HUVECs were maintained in Endothelial Cell Growth medium (EGM; 0.1% human epidermal growth factor, 0.1% hydrocortisone, 0.1% Gentamicin - Amphotericin-B, 0.4% bovine brain extract, 2% FBS; Lonza) on cell culture flasks coated with 10 g/ml fibronectin (Sigma, St. Louis, MO) and 0.05 % gelatin in PBS. Human aortic endothelial cells (HAECs) were purchased from Life Technologies.

extract_protocol: Lysates were clarified from sonicated nuclei and protein-DNA complexes were isolated with antibody. Libraries were prepared according to Illumina's instructions. Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (minus exo) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation, ChIP DNA was PCR-amplified with Illumina Genomic adaptors or with NEXTflex DNA barcodes adaptors and library fragments were size-selected (150-250bp) from a 2% agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on Illumina HiSeq 2000 according to the manufacturer’s instructions.

treatment_protocol: Cells were seeded on 10-cm dishes and allowed to adhere for 24 h after which the cells were washed with HBSS and lipoprotein deficient medium (2% LPDS) was added to the wells. After 16 h cells were treated with 50 ng/ml rhVEGF-A165 (R&D Systems, Minneapolis, MN) for 2-8h.

value: human aortic endothelial cells


title: HAEC-H3K4me2-ChIP-Seq

description: chromatin immunoprecipitation