title: Yeast Ndc80 OP ChIP-Seq

city: New Haven

first: Raymond

middle: K

last: Auerbach

state: CT

address: 266 Whitney Ave

institute: Yale University

description: Ndc80 ChIP-Seq, elevated Cse4 levels

data_processing: Raw sequencing data were first processed by the built-in Illumina analysis pipeline.  Prior to the actual sequence mapping, reads were parsed according to the 4-bp index, and barcodes were then removed. Those reads lacking an intact index were discarded. The remaining bases were aligned against the S. cerevisiae S288c reference genome version 2 (SGD/UCSC sacCer2, June 2008) by the ELAND algorithm (Illumina).  The peak scoring algorithm PeakSeq (Rozowsky et al., Nat Biotech (2009)) was used to identify statistically significant binding sites, changing only the following parameters to account for the compact S. cerevisiae genome: window size of 10 kb during the normalization step and bin size of 1 kb during the linear regression step (PeakSeqOutput files, with Q-value < 0.05).  ChIP-Seq data from epitope-tagged strains were scored against ChIP-Seq data from their matching untagged strains.  In both cases, anti-Myc antibodies were used during ChIP.  Scoring reference sets were created by pooling uniquely-mapping reads from biological replicates of untagged control strains.  As a reference sample marking open chromatin (Auerbach et al., PNAS (2009)), two lists of Sono-Seq significant regions were generated, obtained after scoring against either anti-Myc or anti-HA control sets.  To uncover Centromere-like Regions (CLRs), we took a conservative, stringent approach to minimize false positives lacking functional significance or failing qPCR validation.  For each biological replicate of a particular kinetochore component, only putative binding regions with Q-value < 10-5 were considered (Target bed files).  Several other criteria were then applied.  First, binding sites called in two biological replicates from Cse4, Mif2, Ndc10 and Ndc80 ChIP-Seq data, in either a Cse4 overexpression strain or in a wild-type strain, were overlapped with maxgap=150 (maximum gap between non-overlapping peaks).  Next, to identify a binding region as a CLR, 1) all four kinetochore proteins must be present at the putative site given the q-value threshold; and 2), for proteins in direct contact with DNA, mean PeakSeq ratios between both replicates at a particular target site should be above 2.00 for open chromatin marker Cse4 (same for PolII and Sono-Seq regions), and 1.50 for direct DNA binders Mif2 and Ndc10. Several other filters were used to distinguish between lower confidence regions and higher confidence regions for subsequent functional analyses, including comparison of PeakSeq experimental reads and PeakSeq background reads between CLRs and CENs, inspection of normalized signal tracks (good tagged/untagged signal ratio and low background in the appropriate untagged control desired), binding over a highly PolII-occupied ORF, and presence in a HOT region.  Regions that passed those criteria and filters were termed CLRs (Centromere-Like Regions).   Other binding sites that did not pass the aforementioned filters were referred to as LCNCRs (for low-confidence, negative control regions).

organism: Saccharomyces cerevisiae

source_name: Yeast cell

growth_protocol: Yeast cells were grown in 500 mL rich medium supplemented with adenine and uracil, in the presence of raffinose and galactose (YPAURG) to mid-log phase (OD = 0.5-0.7).

extract_protocol: ChIP-Seq experiments were performed at least in independent biological duplicates, as described previously (Aparicio et al., Curr Protoc Cell Biol (2004); LefranÃ§ois et al., BMC Genomics (2009); LefranÃ§ois et al., Methods Enzymology (2010)).  Yeast strains were grown in 500 mL YP media supplemented with adenine and uracil, in presence of glucose or galactose/raffinose, to mid-log phase (OD 600=0.5-0.7).  Proteins were crosslinked to DNA by treating cells with formaldehyde (1% final concentration) for 15 minutes, then quenched with glycine.  Cells were collected by filtration after two washes.  After cell lysis using a FastPrep machine (MP Biomedical), chromatin was sheared by sonication using a Branson Digital 450 sonifier (Branson).  Clarified, sonicated lysates were taken at this step for Sono-Seq, prior to immunopreciptation.  Immunoprecipitations of Myc-tagged and HA-tagged strains, as well as those of the respective control untagged strains, were carried out overnight with EZ-View anti-Myc or anti-HA affinity gels (Sigma).  For native RNA Polymerase II ChIP, cell lysates were incubated with Pol II 8WG16 mouse monoclonal antibody (Covance) and pulled down using Protein G agarose beads (Millipore).  After several washes and reversal of protein-DNA crosslinks, ChIP DNA was purified through a Qiagen MinElute PCR purification column (Qiagen). Illumina sequencing libraries were generated using adapters for multiplexing as described (LefranÃ§ois et al., BMC Genomics (2009); LefranÃ§ois et al., Methods Enzymology (2010)), and following Illuminaâs manufacturer recommendations.  Four barcoded libraries were mixed in equimolar ratios and processed on an Illumina Genome Analyzer II.  We have also used previously published Cse4 ChIP-Seq data, with normal Cse4 levels, which were deposited in the Gene Expression Omnibus (GEO) database under GSE13322.

value: W303 Ndc80-9xMyc-KanMX4 Pgal1-Cse4

value: anti-Myc EZview affinity gel (Sigma)

tag: antibody manufacturer

value: Sigma


title: Yeast Ndc80 ChIP-Seq

status: Public on Jan 18 2013

type: SRA

country: USA

description: Ndc80 ChIP-Seq, normal Cse4 levels

library_source: genomic

molecule: genomic DNA

growth_protocol: Yeast cells were grown in 500 mL rich medium supplemented with adenine and uracil, in the presence of dextrose (YPADU) to mid-log phase (OD = 0.5-0.7).

tag: strain

value: W303 Ndc80-9xMyc-KanMX4

tag: antibody


title: Yeast Sono-Seq

description: Sono-Seq reference sample

tag: sample type

value: input sonicated DNA


title: Yeast Untagged Myc Control OP ChIP Seq

description: Untagged Myc control data set for scoring ChIP samples.

value: W303 Pgal1-Cse4


title: Yeast Ndc10 OP ChIP-Seq

description: Ndc10 ChIP-Seq, elevated Cse4 levels

value: W303 Ndc10-9xMyc-KanMX4 Pgal1-Cse4


title: Yeast Cse4 OP ChIP-Seq

description: Cse4 ChIP-Seq, elevated Cse4 levels

value: W303 Pgal1-Cse4-3xHA-KanMX4 (internal tag)

value: anti-HA EZview affinity gel (Sigma)


title: Yeast Cse4 scm3 ChIP-Seq

description: Cse4 ChIP-Seq, scm3 deletion

value: W303 Pgal1-Cse4-3xHA-KanMX4 (internal tag) scm3::HphMX4


title: Yeast PolII ChIP-Seq

description: Pol II ChIP-Seq

value: anti-RNA Polymerase II mouse monoclonal antibody, clone 8WG16 (Covance, MMS-126R)

value: Covance

tag: antibody catalog #


title: GREB1_XENO_REP1

city: Cambridge

first: Chandra

last: Chilamakuri

address: Robinson Way

country: United Kingdom

institute: Cancer Research UK Cambridge Institute

description: Chromatin IP against GREB1

data_processing:
Single end 36-bp ChIP-seq data were generated by the Illumina analysis pipeline version 1.6.1. For samples JC830, JC832, JC888, JC907 and JC945, version 1.7.0 was used.
Reads were aligned to the Human Reference Genome (assembly hg18, NCBI Build 36.6, March 2008 using BWA version 0.5.5; Li and Durbin, 2010). For samples JC830, JC832, JC888, JC907 and JC945, reads were aligned to the Human Reference Genome (assembly hg18, NCBI Build 36.6, March 2008 using BWA version 0.6.1-r104; Li and Durbin, 2010).
Reads were filtered by an alignment quality score removing all reads with a score less than 15. For samples JC830, JC832, JC888, JC907 and JC945, reads were filtered by an mapping quality score removing all reads with a score less than 5.
Peaks were called using MACS, version 1.3.7.1. For samples JC830, JC832, JC888, JC907 and JC945, peaks were called using MACS, version 1.4.1.
Genome_build: hg18
Supplementary_files_format_and_content: MACS peak files in .bed format

organism: Homo sapiens

growth_protocol: MCF7 cells were grown in DMEM with 10% FBS.

extract_protocol:
DNA extracted according to Schmidt D.(Methods,2009)
Library constructed according to Schmidt D.(Methods,2009)

treatment_protocol: Tamoxifen or ICI 182 drug treatments done at 100nM concentration for the indicated time

tag: cell line

tag: library id

tag: treatment

value: MCF7 xenograft tumour in mouse


title: Olig2_iOL_ChIPseq_Sample 7

status: Public on Jan 20 2013

city: Chengdu

first: Yang

last: Yu

state: Si Chuan

address: Ren Min Nan road

country: China

institute: Si Chuan University

data_processing:
ChIP-seq reads were aligned to the rn4 genome assembly
peaks were called using MACS version 1.4.0 with default settings.
peaks were further filtered using following criteria to define a more stringent protein-DNA interactome: 1) the p-value cutoff was set to <10-9; 2) an enrichment of 5 fold; 3) tag number > 20
Genome_build: rn4
Supplementary_files_format_and_content: bed files were generated using MACS 1.4.0

organism: Rattus norvegicus

source_name: immature oligodendrocyte cells

growth_protocol: Primary rat iOLs were isolated from cortices of pups at P2 using a differential detachment procedure as previously described (Chen et al., 2007). Isolated rat iOLs were grown in the iOL Growth Medium (Sato medium supplemented mitogens 10 ng/ml PDGF-AA and 20 ng/ml bFGF).

extract_protocol:
Lysates were clarified from sonicated nuclei and histone-DNA complexes were isolated with antibody.
The ChIP-seq libraries were prepared using 5500 SOLiD Fragment Library Core Kit (PN 4464412) as recommended by the manufacturer.

treatment_protocol: iOLs are differentiated in Oligodendrocyte Differentiation Medium (Sato medium supplemented with 15 nM thyroid hormone T3 and 10 ng/ml ciliary neurotrophic factor).

value: Sprague-Dawley

tag: cell type

value: primary iOLs

value: T3 treatment

tag: genotype/variation

value: wild type

tag: chip antibody

value: Olig2 (Millipore, Ab9610)


title: Yeast Mif2 ChIP-Seq

description: Mif2 ChIP-Seq, normal Cse4 levels

value: W303 Mif2-9xMyc-KanMX4

value: anti-Myc EZview affinity gel (Sigma)


title: Prdm14WT_input

city: Kyoto

first: Yukihiro

last: Yabuta

state: Kyoto

address: Yoshida-Konoe-cho, Sakyo-ku

department: Anatomy and Cell Biology

country: Japan

institute: Kyoto University, Graduate school of medicine

data_processing:
For the ChIP-seq of PRDM14, Reads were summed in 10 bp bins along the chromosome for ChIP and WCE, respectively. The read number of each bin was normalized with the total number of aligned reads for each chromosome. Scanning the genome with a 300 bp (30 bin) sliding window, a one-sided Wilcoxon rank-sum test was performed to estimate the enrichment p-value for each window. The fold enrichment (ChIP/WCE) for each window was also calculated. To call peaks, we picked up the windows which satisfied the criteria of a fold enrichment > 3.0 and p-value < 1e-4 as candidates for binding sites (Langmead et al., 2009). To further eliminate uncertain sites, we removed the regions with low ChIP reads using either of two criteria: 1) The average number of ChIP reads per region/the average number of ChIP reads in the genome < 3.0; 2) The maximum read intensity in ChIP bins was less than 50. We derived the consensus sequence motif by using MEME (Hu et al., 2011).
Genome_build: mm9

organism: Mus musculus

source_name: ES cells

growth_protocol: ES cells were passaged in ESC maintenance medium [GMEM containing 2 mM L-glutamine (cat#11710-035; Invitrogen) with 1 × MEM non-essential amino acids (cat#11140-050; Invitrogen), 1 mM sodium pyruvate (cat#11360-070; Invitrogen), 100 M -mercaptoethanol (cat#21985-023; Invitrogen), 1 × 103 units/ml LIF (cat#ESG1107; Chemicon), and 20% FBS (Stem Cell Science)].

extract_protocol:
ESCs at 70–80% confluency (approximately 2×107 cells on 15 cm dish) were crosslinked with 1% formaldehyde for 5 min at RT and rinsed twice with 10 ml of cold PBS after quenching with 125mM glycine. The fixed cells were harvested by using a silicon scraper, collected by centrifugation at 700 g for 5 min at 4 ○C, resuspended in 5 ml of cold LB1 (20 mM Tris, 10 mM NaCl, 2.5 mM MgCl2, 0.2% NP-40, 1 mM PMSF, pH7.5) and pelleted by centrifugation at 1350 g for 5 min after the rotation for 10 min at 4 ○C. The pellet was resuspended in 5 ml of LB2 (20 mM Tris, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 1 mM PMSF, pH8.0) and rotated for 10 min at 4 ○C. The nuclei were pelleted by centrifugation at 1350 g for 5 min at 4 ○C and lysed in 2 ml of LB3 (20 mM Tris, 150 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 1% TritoneX-100, 0.1% sodium deoxycholate, 0.1% SDS, CompleteTM EDTA-free). The lysed nuclei were sonicated using the following conditions: Branson Sonifier 250D with standard tip; 15% output; 10 cycles (12sec ON, centrifugation at 13,200 g for 2 min at 4 ○C).
Protein-DNA complexes were immunoprecipitated for overnight at 4 ○C using 10 μg of antibodies bound to 250 μl of Dynabeads Protein G (Invitrogen). Immunoprecipitates were washed five times with 1 ml of RIRA buffer (50 mM HEPES-KOH, 0.5 M LiCl, 1 mM EDTA, 0.5% Na-Deoxycholate, 1% NP-40, pH 7.4), and once with 250 μl of TE50 (50 mM Tris, 10 mM EDTA, pH8.0). Beads were resuspended with 200 μl of EB (1% SDS, TE50). The complexes were eluted at 65 ○C for 20 min. The crosslinks for both the immunoprecipitated and input DNA fragments were reversed by incubating at 65 ○C for at least 6 hrs. After RNase A and Proteinase K digestion, the DNA is purified by phenol/chloroform extraction and EtOH precipitation with 30 μg glycogen and dissolved with TE.
DNA from whole cell extracts (WCE) and ChIP fractions was further sheared to an average size of approximately 150 bp by ultrasonication (Covaris, Woburn, MA), end-repaired, ligated to sequencing adapters and amplified according to the manufacturer’s instructions (Applied Biosystems SOLiD Library Preparation Protocol).
For the ChIP of PRDM14 using AGP14 ESC-like cells, gel-purified amplified DNA of between 100 and 150 bp was sequenced on Life Technologies SOLiD 4 platforms to generate single-end 50-bp reads. For SUZ12 and histone H3K27me3, amplified DNA of between 200 and 350 bp, purified using an Agencourt AMPureXP Kit, was sequenced on a Life Technologies SOLiD 5500xl platform to generate single-end 50 bp reads.

value: Mouse embryonic stem cells (mESCs)


title: Yeast Untagged Myc Control ChIP Seq

value: Sigma


title: Prdm14KO_SUZ12

value: SUZ12 (Abcam, ab12073)


title: Yeast Ndc10 ChIP-Seq

description: Ndc10 ChIP-Seq, normal Cse4 levels

value: W303 Ndc10-9xMyc-KanMX4


title: ER_XENO_REP1

description: Chromatin IP against ER

value: ER; sc-534 (santa cruz) Lot-(HC-20)


title: AGP14_IP

value: Prdm14-/-, rescued by AVI-EGFP-Prdm14 (AGP14)

value: GFP (Abcam, ab290)


title: nGFP_IP

value: Prdm14-/-, rescued by EGFP


title: Yeast Untagged HA Control OP ChIP Seq

description: Untagged HA control data set for scoring ChIP samples.

value: W303 Pgal1-Cse4

value: anti-HA EZview affinity gel (Sigma)


title: Prdm14WT_SUZ12


title: GREB1_ICI_REP2

description: Chromatin IP against GREB1

value: MCF7 cells in Complete DMEM media and 4 hours of 100nM ICI 182780(rep2)


title: ER_REP2

description: Chromatin IP against ER

value: MCF7 cells in Complete DMEM media


title: GREB1_REP2

description: Chromatin IP against GREB1

value: MCF7 cells in Complete DMEM media (rep2)


title: GREB1_ICI_REP1

description: Chromatin IP against GREB1

value: MCF7 cells in Complete DMEM media and 4 hours of 100nM ICI 182780(rep1)


title: Yeast Mif2 OP ChIP-Seq

description: Mif2 ChIP-Seq, elevated Cse4 levels

value: W303 Mif2-9xMyc-KanMX4 Pgal1-Cse4


title: Prdm14WT_H3K27me3

value: H3K27me3 (Upstate/Millipore, 07-4492)


title: GREB1_REP1

description: Chromatin IP against GREB1

value: MCF7 cells in Complete DMEM media (rep1)


title: Control

city: Amsterdam

first: Ching Yong

last: Yick

address: Meibergdreef 9

department: Respiratory Medicine

country: Netherlands

institute: Academic Medical Center

description: pool of 5 non-atopic controls

library_source: transcriptomic

data_processing:
The general quality of the sequencing run was checked using the GS Run Browser version 2.6 (Roche).
Sequence reads were mapped against the human genome (hg19; UCSC) using the GS Reference Mapper version 2.6 (Roche).
Comparison of the numbers of reads per gene between asthma and healthy controls was based on the Poisson distribution and carried out with SPSS including correction for multiple testing.
With 9 subjects in total, 4 biopsy specimens per subject, and a significance level of 0.0001, we expected to find 48 true and 2 false significant genes. This resulted in a false discovery rate of approximately 4%.
Genome_build: hg19
Supplementary_files_format_and_content: Comma-separated files contain mapped genes and the following information: number of unique matching reads, percent of  all unique matches, percent of all reads

molecule: total RNA

source_name: Endobronchial biopsy

extract_protocol: 4 endobronchial biopsies per subject were collected during a bronchoscopy. Total RNA was isolated with TRIzol. cDNA was prepared using the Ovation RNA-Seq System (NuGEN). cDNA libraries of >350bp were constructed using SPRIworks Fragment Library System II (Beckman Coulter) with the addition of GS FLX Titanium Rapid Library multiplex identifier (MID) adaptors (Roche). Emulsion-based clonal amplification was performed using the GS FLX Titanium emPCR Kit (Lib-L; Roche). RNA-Seq was performed using the GS FLX+ instrument (454/Roche).

tag: tissue

value: tissue from airway wall

tag: disease state

value: Healthy non-atopic control


title: INPUT

description: Input DNA

source_name: INPUT

value: Input


title: E2F4_TAM

description: Chromatin IP against E2F4

value: E2F4; sc-866 (santa cruz) Lot-(A2804)

value: MCF7 cells in complete DMEM media treated with 100nm Tamoxifen for 48hrs


title: Asthma

description: pool of 4 patients

value: Atopic asthma


title: ER_REP1

description: Chromatin IP against ER

value: ER; sc-534 (santa cruz) Lot-(HC-20)


title: E2F4_ICI

description: Chromatin IP against E2F4

value: MCF7 cells in complete DMEM media treated with 100nm ICI 182780 for 48hrs


title: TLE3_REP1

description: Chromatin IP against TLE3


title: JU-A10-T_G DGE

city: Qingdao

first: Guodong

last: Liu

state: Shandong

address: Binhai Road 72

department: State Key Laboratory of Microbial Technology

institute: Shandong University

description: Penicillium decumbens strain JU-A10-T grown in glucose medium.

data_processing: 3' adaptor sequences were first removed from the sequencing reads to produce 21-nt long tags. Then low quality tags, one copy tags and tags which are not 21-nt long were removed to generate clean tags. Reference 21-nt tag databases were created from predicted P. decumbens transcripts plus downstream 300-nt sequences of two strains. All clean tags were mapped to corresponding reference tag databases and no mismatch was allowed. The copy number of unambiguous tags (tags mapped to one single gene) for each gene was normalized to TPM (number of transcripts per million clean tags) for differential expression analysis. Linked .txt files include sequence, copynumber, and normalized tags per million of clean tags.

organism: Penicillium decumbens

source_name: Fungal mycelia

growth_protocol: Conidial suspension was inoculated to the medium at a final concentration of 1e6 per ml, and the strain was grown at 30 degrees C, 200 rpm.

extract_protocol: Total RNA was extracted, and oligo (dT) beads were used to purify mRNA and guide double-stranded cDNA synthesis. The enzyme NlaIII was used to cut the cDNA at CATG sites. After Illumina adapter 1 was linked to the CATG site, MmeI was used to cut at 17-bp downstream of the CATG site. Then Illumina adapter 2 was linked at 3' end and the fragments were PCR amplified. Finally, the 85-bp strips were gel purified and digested for Solexa sequencing. Each tunnel generated millions of raw reads with sequencing length of 35bp.

tag: medium

value: 2% (wt/vol) glucose

tag: time


title: RXRA_REP1

description: Chromatin IP against RXRA

value: RXRA;  sc-553 (santa cruz)-Lot(E0411)


title: SF3B1 mutant patient #2

city: Cleveland

first: John

last: Barnard

state: Ohio

address: 9500 Euclid Ave.

department: Quantitative Health Sciences

institute: Cleveland Clinic Foundation

data_processing:
100-bp paired-end RNA-Seq reads were mapped to the hg19 RefSeq human transcriptome and spliceosome by DNAnexus (http://dnanexus.com) using a Bayesian method, in which a read was mapped when its posterior probability of mapping exceeded 0.9.
These filtered posterior probabilities were summed to generate fractional read counts per gene and per exon, with probabilities from splice-junction spanning reads counted for each relevant exon.
Genome_build: hg19
Supplementary_files_format_and_content: Mapped exon counts; Alternative Splicing format: type The type of element – either an exon or a junction;  gene_id The unique identifier for the gene from the reference database; gene_name The reference gene name in the database; chromosome_name Chromosome name for the gene; strand Genomic strand for the gene; prev_exon_left If the element is a splice site, this represents the left boundary of the donor exon. If the element is an exon, this represents the left boundary; prev_exon_right If the element is a splice site, this represents the right boundary of the donor exon. If the element is an exon, this represents the right boundary; next_exon_left If the element is a splice site, this represents the left boundary of the acceptor exon. If the element is an exon, this field is not used; next_exon_right If the element is a splice site, this represents the right boundary of the acceptor exon. If the element is an exon, this field is not used; read_count The sum of the posterior probabilities for all confident reads that map to the element.

source_name: bone marrow mononuclear cells

extract_protocol:
Isolated total RNA from bone marrow mononuclear cells
RNA libraries were prepared for sequencing using standard Illumina protocols

value: bone marrow

tag: genotype

value: SF3B1 somatic heterozygote, K700E

tag: disease

value: RARS, 50% ring sideroblasts


title: JU-A10-T_C DGE

description: Penicillium decumbens strain JU-A10-T grown in cellulose medium.

growth_protocol: Conidial suspension was inoculated to 2% (wt/vol) glucose medium at a final concentration of 1e6 per ml, and the strain was grown at 30 degrees C, 200 rpm for 39 h as the pre-culture. Then mycelia were collected by vacuum filtration, washed with sterile water and grown in 2% (wt/vol) cellulose medium for 2 h.

value: 2% (wt/vol) cellulose

value: first grown in 2% (wt/vol) glucose medium for 39 h and then transferred into 2% (wt/vol) cellulose medium for another 2-h growth


title: KLF4_REP1

description: Chromatin IP against KLF4

value: KLF4; R&D systems AF3640-Lot(xvh0310101)

value: MCF7 cells in Complete DMEM media


title: 114-2_C DGE

description: Penicillium decumbens strain 114-2 grown in cellulose medium.


title: 114-2_G DGE

description: Penicillium decumbens strain 114-2 grown in glucose medium.


title: Prdm14KO_H3K27me3

value: Mouse embryonic stem cells (mESCs)

value: H3K27me3 (Upstate/Millipore, 07-4492)


title: JU-A10-T_CW DGE

description: Penicillium decumbens strain JU-A10-T grown in cellulose-wheat bran medium.

value: 1% (wt/vol) cellulose plus 1% (wt/vol) wheat bran


title: HeLa_BAC_STARRseq_input

status: Public on Jan 17 2013

city: Vienna

first: Daniel

last: Gerlach

address: Dr.-Boehringer-Gasse 5-11

department: Global Computational Biology and Digital Sciences

country: Austria

institute: Boehringer Ingelheim RCV GmbH & Co KG

description: PCR amplified Plasmid library, used for HeLa STARR-seq

data_processing:
Basecall where performed using Real-Time Analysis (RTA) version > 1.12.4.2 or CASAVA 1.9.1.
Reads were mapped to the hg19 genome using bowtie (bowtie -p 12 -f -v 3 -m 1 --best --strata --quiet INDEX reads.fa) and an index on chr1 - chr22, chrX, and chrY. For all subsequently analysis we subsampled data overlapping the BAC regions. Significantly enriched regions (peaks) were called with an in-house pipeline using read density profiles of cDNA and input and an hypergeometric test to assign a p-value to each peak. Enrichment values were corrected within a 95% confidence interval taking into account the number of independent fragments at a single peak summit position.
Genome_build: hg19

source_name: BAC library (in STARR-seq vector)

extract_protocol: Genomic DNA (source: RPCI 11 human BAC library) was isolated, sheared and size selected (~500bp). Following the instructions of NEBNext® DNA Library Prep Reagent Set for Illumina® (NEB; cat. no. E6000L), Illumina Multiplexing Adapters (Illumina Inc; cat. no. PE-400-1001) were ligated and homology arms for In-Fusion® recombination were added by PCR, followed by recombination into the STARR-seq vector (pGL4-Promotor backbone (Promega; cat. no. E6651) with the sequence between BgIlI and FseI replaced with the following sequence, containing a Super Core Promoter 1 (SCP1) (1), an ORF (sgGFP, Qbiogene, Inc), a ccdB suicide gene flanked by homology arms (used for cloning the genomic enhancer candidates during library generation), and the pGL3’s SV40 late polyA-signal). The In-Fusion® reactions were transformed (MegaX DH10B; Invitrogen), grown in liquid culture and plasmids were isolated.

value: RPCI human BAC library 11 (6 BACS)

tag: experimental procedure

value: STARR-Seq


title: HeLa_BAC_STARRseq

description: PCR amplified cDNA (STARR-seq transcript)

data_processing:
Basecall where performed using Real-Time Analysis (RTA) version > 1.12.4.2 or CASAVA 1.9.1.
Reads were mapped to the hg19 genome using bowtie (bowtie -p 12 -f -v 3 -m 1 --best --strata --quiet INDEX reads.fa) and an index on chr1 - chr22, chrX, and chrY. For all subsequently analysis we subsampled data overlapping the BAC regions. Significantly enriched regions (peaks) were called with an in-house pipeline using read density profiles of cDNA and input and an hypergeometric test to assign a p-value to each peak. Enrichment values were corrected within a 95% confidence interval taking into account the number of independent fragments at a single peak summit position.
Genome_build: hg19
Supplementary_files_format_and_content: All processed data files are in plain text. For STARR-seq we report for each peak the chromosome, the summit position, the enrichment over input (log2-scale) at the summit, and the p-value.

molecule: polyA RNA

source_name: HeLa Kyoto cells

value: HeLa Kyoto


title: COT2_REP1

description: Chromatin IP against COT2

value: COT2; R&D systems PP-H7147-00-Lot(A2)


title: input DNA

city: San Francisco

first: Nevan

last: Krogan

address: 1700 4th Street

department: Cellular & Molecular Pharmacology

institute: University of California, San Francisco

description: Sample 2

data_processing:
reads were mapped to hg19 using bowtie version 0.12.7
peaks were called for each replicate separately using the same input control with PeakRanger 1.15
Consensus peaks of each replicate were identified using the bedtools 2.17.0 utility intersectBed with a minimum overlap of 200.  The start position of the consensus region is the minimum position of the two replicates and the end position is the maximum of the two replicates.
Consensus peaks were filtered such that both replicate peaks had a PeakRanger FDR value of 1e-7 or better.
FDR-filtered consensus peak regions were used to discover the top scoring motif with CUDA-MEME 2.7.3, and these peaks were filtered such that there was one instance of the motif TGYGGYY.
Genes were assigned to consensus peaks using GREAT version 2.0.2.
Genome_build: hg19
Supplementary_files_format_and_content: one bed file for each replicate that includes their PeakRanger FDR value; one tab file representing the consensus peaks and the identity of the replicates making them up

growth_protocol: cells were maintained in RPMI-1640 media containing 10% FBS, penicillin, streptomycin, and 10 ug/mL Blasticidin

extract_protocol:
Approximately 1e8 cells in 100ml of media (approx 1e6 cells/ml) were treated with 1% formaldehyde for 15 minutes.  Solubilized chromatin was prepared by incubation with lysis buffers, a few cycles of sonication and MNase treatment to obtain an average DNA size of ~1kb.  ChIP was performed with anti-RUNX1 polyclonals.
Purified ChIP or input DNA was further sheared to an average size of ~200bp using a BioRuptor.  The ends of the DNA fragments were polished using an Epicentre End-IT kit, polyadenylated with dATP and Klenow exo- and ligated to adapters, followed by 10 rounds of PCR with high fidelty Taq.

treatment_protocol: cells were induced with 1 ug/mL doxycycline for ~16 hours to allow for use as an appropriate comparison control with any other Jurkat Trex lines that carry a Tet-inducible construct.

value: Jurkat Trex (Invitrogen)


title: Hela_PTB_CLIP_2nd

city: La Jolla, San Diego

first: Jie

last: Huang

state: CA

address: 9500 Gilman Drive, Room 231

department: Department of Cellular and Molecular Medicine

institute: UC San Diego

data_processing:
CLIP-seq: After removing 3' adaptor sequence, the tags longer than 20 nt were mapped to the reference human genome (hg18) with Bowtie 0.12.7 by allowing maximum 2 mismatches in the first 25 nt, and only tags with less than or equal to 10 mapped locations were kept.
RNA-seq: Paired-end reads were separately mapped into a custom Bowtie index, which was built from UCSC knownGenes (hg18) not including introns. Under the constraint of the insert size, only the paired-end reads whose two ends are about 50 nt apart on the same gene were kept. For each pair, the coordinates of the two end reads were mapped back to the genome based on its gene's genomic location, and then joined together. Only the pair-end reads with one unique genomic location were kept.
3' end RNA-seq: After removing 3' adaptor sequence, the tags longer than 20 nt were mapped to the reference human genome (hg18) with Bowtie 0.12.7 by allowing maximum 2 mismatches in the first 25 nt, and only tags with less than or equal to 10 mapped locations were kept. Then scanned the genome for polyA-stretch defined as consecutive 8As, removed them; adaptively clustered tags within a specific distance 30 nt along transcripts and sorted cluster intervals by the length and tag in a decreasing order. If a cluster contains a known 3’end within a 300 nt window, we used the end and then counted the number of reads in each cluster.
Genome_build: UCSC hg18
Supplementary_files_format_and_content: bed format

source_name: Hela cells

growth_protocol: HeLa cells were maintained in Dulbecco’s Modified Eagle Medium (DMEM) containing 10% FBS (Omega Scientific) and 100U of penicillin/streptomycin (Life Technology).

extract_protocol: CLIP-seq library was prepared according to previously published standard protocol. RNA-seq was typically done after shRNA treatment for 72 hrs. Trizol-isolated  RNA was enriched in two rounds for Poly(A+) RNA with paramagnetic oligo(dT), fragmented into ~200 nt in length, converted to cDNA with Superscript III (Invitrogen), and subjected to deep sequencing.

treatment_protocol: For CLIP-seq samples, Hela cells were UV-irradiated at 400mj. For pair-end RNA-seq, Hela cells were treated with control shRNA or PTB specific shRNA. For 3' end RNA-seq, Hela cells were further treated with actinomycin D (2 μg/ml) for 4 hours.

value: Hela

tag: clip antibody

value: Anti-PTB mAb BB7


title: RUNX1

description: Sample 1

tag: chip antibody vendor

value: Santa Cruz

tag: chip antibody cat. #


title: OSC DHS-seq biological replicate 2

description: PCR amplified DNaseI digested genomic DNA

data_processing:
Basecall where performed using Real-Time Analysis (RTA) version > 1.12.4.2 or CASAVA 1.9.1
Reads were mapped to the dm3 genome using bowtie (bowtie -f -v 3 -m 1 --best --strata --quiet -S INDEX -) excluding chrU, chrUextra, and chrM. We estimated the enrichment and p-values for specific genomic regions using the cDNA and the input together with a hypergeometric test. To call peaks we also used MACS14 with default parameters and an FDR cutoff of 5%.
Genome_build: dm3
Supplementary_files_format_and_content: All processed data files are in plain text. The DHS-seq peaks are directly taken from the MACS output.

organism: Drosophila melanogaster

source_name: OSC cells

extract_protocol: 5-7 10^7 cells were harvested. DNase treatment was done as in (L. Cappabianca, H. Thomassin, R. Pictet, T. Grange, Genomic footprinting using nucleases., Methods in molecular biology (Clifton, N.J.) 119, 427-42 (1999)). For size selection (~120bp) a sucrose gradient was emplyed (SW TI40, 30000rpm, 16h). Library was constructed following instructions of NEBNext® DNA Library Prep Reagent Set for Illumina® (NEB; cat. no. E6000L). Library amplifcation was done using KAPA Library Amp Real Time (KAPA biosystems: cat.no. KK2701).

value: OSC cells

tag: sample type


title: OSC DHS-seq biological replicate 1

value: PCR amplified DNaseI digested genomic DNA


title: OSC DHS-seq Input

data_processing:
Basecall where performed using Real-Time Analysis (RTA) version > 1.12.4.2 or CASAVA 1.9.1
Reads were mapped to the dm3 genome using bowtie (bowtie -f -v 3 -m 1 --best --strata --quiet -S INDEX -) excluding chrU, chrUextra, and chrM. We estimated the enrichment and p-values for specific genomic regions using the cDNA and the input together with a hypergeometric test. To call peaks we also used MACS14 with default parameters and an FDR cutoff of 5%.
Genome_build: dm3


title: 114-2_CW DGE

description: Penicillium decumbens strain 114-2 grown in cellulose-wheat bran medium.

tag: strain


title: S2 DHS-seq Input

source_name: S2 cells

value: S2 cells


title: Healthy patient #1

value: bone marrow

value: SF3B1 wildtype

value: normal


title: STARR-seq Input library 500

description: PCR amplified Plasmid library, used for OSC STARR-seq

data_processing:
Basecall where performed using Real-Time Analysis (RTA) version > 1.12.4.2 or CASAVA 1.9.1
Reads were mapped to the dm3 genome using bowtie (bowtie -p 12 -f -X 2000 -v 3 -m 1 --best --strata --quiet INDEX -1 reads_1.fa -2 reads_2.fa) excluding chrU, chrUextra, and chrM. For all subsequently analysis we used only reads collapsed on chromosome, start, end, strand (fragments). Significantly enriched regions (peaks) were called with an in-house pipeline using read density profiles of cDNA and input and an hypergeometric test to assign a p-value to each peak. Enrichment values were corrected within a 95% confidence interval taking into account the number of independent fragments at a single peak summit position.
Genome_build: dm3

source_name: whole genome library (in STARR-seq vector)

extract_protocol: Genomic DNA (source: embryos of the sequenced strain: y; cn bw sp) was isolated, sheared and size selected (~500bp). Following the instructions of NEBNext® DNA Library Prep Reagent Set for Illumina® (NEB; cat. no. E6000L), Illumina Multiplexing Adapters (Illumina Inc; cat. no. PE-400-1001) were ligated and homology arms for In-Fusion® recombination were added by PCR, followed by recombination into the STARR-seq vector (pGL3-Promotor backbone (Promega; cat. no. E1751) with the sequence between BglII and FseI replaced with the following sequence, containing a Drosophila Synthetic Core Promoter (DSCP) (1), an ORF (sgGFP, Qbiogene, Inc), a ccdB suicide gene flanked by homology arms (used for cloning the genomic enhancer candidates during library generation), and the pGL3s SV40 late polyA-signal.). The In-Fusion® reactions were transformed (MegaX DH10B; Invitrogen), grown in liquid culture and plasmids were isolated.

value: y; cn bw sp - sequenced strain (Adams et al. 2000)


title: SF3B1 mutant patient #1

value: SF3B1 somatic heterozygote, H666Q

value: RARS-T, >15% ring sideroblasts


title: STARR-seq Input library 5.2

description: PCR amplified Plasmid library, used for S2 STARR-seq


title: S2 DHS-seq biological replicate 2


title: OSC STARR-seq biological replicate 2

data_processing:
Basecall where performed using Real-Time Analysis (RTA) version > 1.12.4.2 or CASAVA 1.9.1
Reads were mapped to the dm3 genome using bowtie (bowtie -p 12 -f -X 2000 -v 3 -m 1 --best --strata --quiet INDEX -1 reads_1.fa -2 reads_2.fa) excluding chrU, chrUextra, and chrM. For all subsequently analysis we used only reads collapsed on chromosome, start, end, strand (fragments). Significantly enriched regions (peaks) were called with an in-house pipeline using read density profiles of cDNA and input and an hypergeometric test to assign a p-value to each peak. Enrichment values were corrected within a 95% confidence interval taking into account the number of independent fragments at a single peak summit position.
Genome_build: dm3
Supplementary_files_format_and_content: All processed data files are in plain text. For STARR-seq we report for each peak the chromosome, the summit position, the enrichment over input (log2-scale) at the summit, and the p-value.

value: PCR amplified cDNA (STARR-seq transcript)


title: S2 DHS-seq biological replicate 1


title: S2 STARR-seq biological replicate 2


title: OSC STARR-seq biological replicate 1


title: S2 STARR-seq biological replicate 1


title: S2 RNA-seq biological replicate 1

description: PCR amplified cDNA (cellular polyA+ RNA)

data_processing:
Basecall where performed using Real-Time Analysis (RTA) version > 1.12.4.2 or CASAVA 1.9.1
Reads were mapped to the dm3 genome and transcriptome using bowtie (bowtie -f -v 3 -m 1 --best --strata --quiet -S INDEX -) and bfast reconciling the outputs. We estimated RPKM values for each gene based on the number of reads falling over each exon merging exons from a all genes' isoforms into one meta-transcript. Reads counts were normalized by the total reads per library and the merged meta-gene exons lengths.
Genome_build: dm3
Supplementary_files_format_and_content: Processed data files are in plain text. For the RNA-seq data we report 3 different gene identifiers and the RPKM value for each gene.

extract_protocol: RNA was isolated using QIAShredder (QIAGEN; cat.no. 79654) and RNeasy Mini Kit (QIAGEN; cat.no. 74106; including DNase I on column digestion) from 5-10 x 10^6 cells. 5ug total RNA was then processed as described before (S. Zhong et al., High-throughput illumina strand-specific RNA sequencing library preparation., Cold Spring Harbor protocols 2011, 940-9 (2011)) with minor adjustments. End-repair, dA-tailing and adapter ligation were performed with the NEBNext DNA Library Prep Reagent Set for Illumina (NEB, cat.no. E6000L) with proportionally reduced enzyme amounts to account for low RNA concentrations. Indexed library amplification was carried out using KAPA Library Amp Real Time (KAPA biosystems: cat.no. KK2701).


title: S2 RNA-seq biological replicate 2

value: PCR amplified cDNA (cellular polyA+ RNA)


title: OSC RNA-seq biological replicate 1


title: Saccharomyces_cerevisiae_Glucose_GCC_Reps1and2

city: Auckland

first: Justin

middle: M.

last: O'Sullivan

address: 85 Park Road, Grafton

department: Liggins Institute

country: New Zealand

institute: University of Auckland

description:
Genome Conformation Capture (GCC) performed on Glucose grown S. cerevisiae
No experimental enrichment

data_processing: Sequences were mapped onto the S. cerevisiae S288c genome (The reference genome used in this study consists of the sixteen S. cerevisiae nuclear chromosomes [NC_001133.7, NC_001134.7, NC_001135.4, NC_001136.8, NC_001137.2, NC_001138.4, NC_001139.8, NC_001140.5, NC_001141.1, NC_001142.7, NC_001143.7, NC_001144.4, NC_001145.2, NC_001146.6, NC_001147.5, NC_001148.3] as well as the mitochondrial genome [NC_001224.1] and the 2-micron plasmid [YSCPLASM]) using Topography v1.19 which is coupled to the SOAP algorithm (Li. R., et al 2008). The GCC protocol incorporates restriction digestion (MspI) and a ligation steps to capture inter- and intra- chromosomal interactions. The Topography software aligns each "paired" sequence tag end against the genome using the SOAP algorithm with no mismatches allowed. If each sequence tag end aligns to a different MspI fragment then the Topography software records an interaction between these two MspI restriction fragments. If the sequence tag aligns uniquely to the genome it is placed into the .dat file (this means that the .dat processed data file contains only interactions which are unique), a copy of the unique interactions are also placed in the .datcomb file. If the sequence tag aligns against the genome more than once it is considered repetitive and is placed in the .datcomb file only. Interactions derived from sequence tags which aligned to multiple sites in the genome can be isolated by subtracting all the interactions in the .dat file from the .datcomb files. For further information see Rodley et al. (2009) supplemental program information. Topography is avaliable upon request from the corresponding author.

source_name: Glucose grown S. cerevisiae

growth_protocol: S. cerevisiae strain (BY4741) was grown (30oC, 160 rpm) to an OD600 = 0.6 in synthetic complete media containing amino acid supplements and glucose (2% w/v).

extract_protocol: Cells were cross-linked (2% v/v formaldehyde, 10 mins, room temperature, with stirring) and quenched (0.125M glycine). Samples (residuals of 108 cells in 52.6μl) were digested (37°C, 2hr) with MspI (100U, Fermentas) in a final volume of 100 ml.  MspI was inactivated by the addition of 1% SDS (final v/v) and incubation (65°C, 20mins). Reactions were diluted in 2ml T4 ligation buffer (NEB) containing 1% Triton X-100. T4 DNA Ligase (20U, Invitrogen) was added and the reaction incubated (16°C, 2hrs). Samples were reverse cross-linked (65°C, O/N) in the presence of Proteinase K (7-11ug, Roche), 20μl EDTA (0.5M), 12μl NaCl (5M), 1.2μl Tris-HCl (1M).  RNase (20μg) was added and incubated (37°C, 15mins) prior to three Phenol:Chloroform (1:1) extractions.  Samples were purified using Zymo Clean and Concentrator columns. The sequencing centre nebulized the samples to 200bp+/-20bp. Sequencing was performed (38 bp paired end) on an Illumina Genome Analyser (FMI, Basel, Switzerland).

tag: growth media

value: glucose


title: Saccharomyces_cerevisiae_GycerolLactate_GCC_Reps1and2

description:
Genome Conformation Capture (GCC) performed on Glycerol Lactate grown S. cerevisiae
No experimental enrichment

data_processing: Sequences were mapped onto the S. cerevisiae S288c genome  (The reference genome used in this study consists of the sixteen S. cerevisiae nuclear chromosomes [NC_001133.7, NC_001134.7, NC_001135.4, NC_001136.8, NC_001137.2, NC_001138.4, NC_001139.8, NC_001140.5, NC_001141.1, NC_001142.7, NC_001143.7, NC_001144.4, NC_001145.2, NC_001146.6, NC_001147.5, NC_001148.3] as well as the mitochondrial genome [NC_001224.1] and the 2-micron plasmid [YSCPLASM])  using Topography v1.19 which is coupled to the SOAP algorithm (Li. R., et al 2008). The GCC protocol incorporates restriction digestion (MspI) and a ligation steps to capture inter- and intra- chromosomal interactions. The Topography software aligns each "paired" sequence tag end against the genome using the SOAP algorithm with no mismatches allowed. If each sequence tag end aligns to a different MspI fragment then the Topography software records an interaction between these two MspI restriction fragments. If the sequence tag aligns uniquely to the genome it is placed into the .dat file (this means that the .dat processed data file contains only interactions which are unique), a copy of the unique interactions are also placed in the .datcomb file. If the sequence tag aligns against the genome more than once it is considered repetitive and is placed in the .datcomb file only. Interactions derived from sequence tags which aligned to multiple sites in the genome can be isolated by subtracting all the interactions in the .dat file from the .datcomb files. For further information see Rodley et al. (2009) supplemental program information. Topography is avaliable upon request from the corresponding author.

source_name: Glycerol lactate grown S. cerevisiae

growth_protocol: S. cerevisiae strain (BY4741) was grown (30oC, 160 rpm) to an OD600 = 0.6 in synthetic complete media containing amino acid supplements and glycerol lactate or galactose (2% w/v).

value: Glycerol Lactate


title: Saccharomyces_cerevisiae_Galactose_GCC_Reps1and2

description:
Genome Conformation Capture (GCC) performed on Galactose grown S. cerevisiae
No experimental enrichment

source_name: Galactose grown S. cerevisiae

value: Galactose


title: IgG_REP1

description: Chromatin IP against non specific IgG

data_processing:
Single end 36-bp ChIP-seq data were generated by the Illumina analysis pipeline version 1.6.1. For samples JC830, JC832, JC888, JC907 and JC945, version 1.7.0 was used.
Reads were aligned to the Human Reference Genome (assembly hg18, NCBI Build 36.6, March 2008 using BWA version 0.5.5; Li and Durbin, 2010). For samples JC830, JC832, JC888, JC907 and JC945, reads were aligned to the Human Reference Genome (assembly hg18, NCBI Build 36.6, March 2008 using BWA version 0.6.1-r104; Li and Durbin, 2010).
Reads were filtered by an alignment quality score removing all reads with a score less than 15. For samples JC830, JC832, JC888, JC907 and JC945, reads were filtered by an mapping quality score removing all reads with a score less than 5.
Peaks were called using MACS, version 1.3.7.1. For samples JC830, JC832, JC888, JC907 and JC945, peaks were called using MACS, version 1.4.1.
Genome_build: hg18
Supplementary_files_format_and_content: MACS peak files in .bed format

tag: cell line

tag: library id

tag: antibody

value: IgG;  sc-2027 (santa cruz)-Lot(c2712)

tag: treatment

value: MCF7 cells in Complete DMEM media


title: FAIRE_UT

first: Iros

last: Barozzi

address: Borschkegasse 8a

institute: Medical University Vienna

description: FAIRE sample.

data_processing:
Library strategy: FAIRE-Seq
Reads were quality filtered according to the Illumina pipeline.
Reads were mapped to the mouse mm9 genome using Bowtie 0.12.7 (PMID 19261174). All reads with a unique match to the genome with two or fewer mismatches were retained.
Accessible regions were called using MACS 1.4 (PMID 18798982) with parameters bw=100, p<=1e-10, no model and no local lambda.
Genome_build: mm9 (NCBI Build 37)
Supplementary_files_format_and_content: *.bed file of accessible regions.

source_name: Primary bone marrow-derived macrophages, no treatment

growth_protocol: Bone marrow cells isolated from mice were plated in 10 cm plates in 5ml of BM-medium (high glucose DMEM supplemented with 20% low endotoxin fetal bovine serum, 30% L929-conditioned medium, 1% glutamine, 1%, Pen/Strep, 0.5% Sodium Pyruvate, 0.1% β-mercaptoethanol). Cultures were fed with 2.5 ml of fresh medium every two days.

extract_protocol: Lysates were generated from 5x10^7 cells following a ChIP protocol. Cells were fixed in 1% formaldehyde for 10min. Three rounds of phenol/chloroform (Invitrogen) extraction were performed in order to recover the protein-free aqueous phase (PMID 17179217). DNA was then purified by QIAquick columns (Qiagen) and quantified using PicoGreen (Invitrogen). DNA was prepared for Illumina HiSeq 2000 sequencing using a standard protocol consisting of blunting, addition of dA overhangs, ligation of Illumina adapters, selection on gel and PCR with index primers. A mixture of T4 DNA polymerase, DNA polymerase I and T4 kinase was used according to the manufacturer's instruction. Library preparation was carried out on SPRIworks Fragment Library System.

treatment_protocol: No treatments.

tag: strain/background

tag: cell type

value: bone marrow-derived macrophages (7th day of differentiation)


title: OSC RNA-seq biological replicate 2

value: OSC cells


title: JunB_UT

description: Chromatin IP against JunB.

data_processing:
Reads were quality filtered according to the Illumina pipeline.
Reads were mapped to the mouse mm9 genome using Bowtie 0.12.7 (PMID 19261174). All reads with a unique match to the genome with two or fewer mismatches were retained.
Peak calling was performed using MACS 1.4 (PMID 18798982) with default parameters and bw=100. Each IP was compared to input DNA derived from BMDM (GEO Sample GSM487453).
Genome_build: mm9 (NCBI Build 37)
Supplementary_files_format_and_content: *_peaks.bed: List of the peaks called by MACS.

source_name: Primary bone marrow-derived macrophages, no treatment, JunB ChIP

extract_protocol: ChIP lysates were generated from 2x10^8 cells. Cells were fixed in 1% formaldehyde for 10min. Lysate was immunoprecipitated with 10ug of antibody. Antibodies were pre-bound overnight to 100ul of G protein-coupled paramagnetic beads in PBS/BSA 0.5%. Beads were then added to lysates (the preclearing step was omitted) and incubation was let to proceed overnight. Beads were washed 6 times in a modified RIPA buffer (50mM Hepes pH 7.6, 500mM LiCl, 1mM EDTA, 1% NP-40, 0.7% Na-deoxycholate) and once in TE containing 50mM NaCl. DNA was eluted in TE containing 2% SDS and crosslinks were reversed by incubation overnight at 65ºC. DNA was then purified by QIAquick columns (Qiagen) and quantified using PicoGreen (Invitrogen). ChIP DNA was prepared for Illumina HiSeq 2000 or Genome Analyzer II sequencing using a standard protocol consisting of blunting, addition of dA overhangs, ligation of Illumina adapters, selection on gel and PCR with index primers. A mixture of T4 DNA polymerase, DNA polymerase I and T4 kinase was used according to the manufacturer's instructions. Library preparation was carried out on SPRIworks Fragment Library System.

treatment_protocol: Macrophages were subjected to different treatments (see individual samples for details).

tag: chip antibody

value: anti-JunB

tag: antibody vendor, catalog #

value: Santa Cruz Biotechnology, sc-46


title: JunB_LPS_4h

description: Chromatin IP against JunB.

source_name: Primary bone marrow-derived macrophages, LPS for 4hrs, JunB ChIP

value: LPS (10 ng/ml) for 4hrs


title: Stat1_UT

description: Chromatin IP against Stat1.

source_name: Primary bone marrow-derived macrophages, no treatment, Stat1 ChIP

value: Santa Cruz Biotechnology, sc-592


title: Stat1_tc_IFNg_2h

description: Chromatin IP against Stat1.

source_name: Primary bone marrow-derived macrophages, IFNg for 2hrs, Stat1 ChIP

value: IFNg (100 ng/ml) for 2hrs


title: Hela_PTB_3end_RNAseq_control_0h and 4h

description: 0h and 4h

value: Hela

value: treated with control shRNA and further treated with actinomycin D (2 ug/ml) for 4 hours


title: Hela_PTB_RNAseq_knockdown

value: treated with PTB shRNA


title: Hela_PTB_RNAseq_control

value: treated with control shRNA


title: Stat1_tc_IFNg_1h

description: Chromatin IP against Stat1.

source_name: Primary bone marrow-derived macrophages, IFNg for 1hrs, Stat1 ChIP

value: IFNg (100 ng/ml) for 1hrs


title: Stat1_tc_IFNg_30min

description: Chromatin IP against Stat1.

source_name: Primary bone marrow-derived macrophages, IFNg for 30min, Stat1 ChIP

value: IFNg (100 ng/ml) for 30min


title: Stat1_tc_IFNg_UT

description: Chromatin IP against Stat1.


title: PU.1_tc_IFNg_2h

description: Chromatin IP against PU.1.

source_name: Primary bone marrow-derived macrophages, IFNg for 2hrs, PU.1 ChIP

value: anti-PU.1 (rabbit polyclonal against the N-terminus (aa. 1-100))

value: in-house


title: Hela_AGO2_CLIP_ptb_knockdown

value: Anti-AGO2 mAb, clone 2E12-1C9(Abnova)


title: H3K4me1_IL4_24h

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IL4 for 24hrs, H3K4me1 ChIP

value: IL4 (10 ng/ml) for 24hrs

value: Abcam, ab8895


title: Stat1_IFNg_4h

description: Chromatin IP against Stat1.

source_name: Primary bone marrow-derived macrophages, IFNg for 4hrs, Stat1 ChIP

value: bone marrow-derived macrophages (7th day of differentiation)

value: IFNg (100 ng/ml) for 4hrs


title: Hela_AGO2_CLIP_control

source_name: Hela cells


title: H3K4me1_tc_IFNg_2h

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IFNg for 2hrs, H3K4me1 ChIP


title: PU.1_tc_IFNg_30min

description: Chromatin IP against PU.1.

source_name: Primary bone marrow-derived macrophages, IFNg for 30min, PU.1 ChIP


title: H3K4me1_tc_IFNg_UT

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, no treatment, H3K4me1 ChIP


title: pol_II_LPS_4h

description: Chromatin IP against total RNA polymerase II

data_processing:
Reads quality filtered according to the Illumina pipeline
Reads were mapped to the mouse mm9 genome using Bowtie 0.12.7 (PMID: 19261174). All reads with a unique match to the genome with two or fewer mismatches were retained.
Peak calling was performed using MACS 1.4 (PMID 18798982) with default parameters and bw=100 (except for the pol_II for which a bw=300 was used). Each IP was compared to input DNA derived from BMDM (GSM487453).
Genome_build: mm9 (NCBI Build 37)
Supplementary_files_format_and_content: *_peaks.bed, list of the peaks called by MACS

source_name: Primary bone marrow-derived macrophages (BMDM)

growth_protocol: Bone marrow cells isolated from C57BL/6 mice were plated in 10 cm plates in 5ml of BM-medium (high glucose DMEM supplemented with 20% low endotoxin fetal bovine serum, 30% L929-conditioned medium, 1% glutamine, 1%, Pen/Strep, 0.5% Sodium Pyruvate, 0.1% β-mercaptoethanol). Cultures were fed with 2.5 ml of fresh medium every two days.

extract_protocol: ChIP lysates were generated from 2x10^8 cells. Cells were fixed in 1% formaldehyde for 10min.  Lysate was immunoprecipitated with 10ug of antibody. Antibodies were pre-bound overnight to 100ul of G protein-coupled paramagnetic beads in PBS/BSA 0.5%. Beads were then added to lysates (the preclearing step was omitted) and incubation was let to proceed overnight. Beads were washed 6 times in a modified RIPA buffer (50mM Hepes pH 7.6, 500mM LiCl, 1mM EDTA, 1% NP-40, 0.7% Na-deoxycholate) and once in TE containing 50mM NaCl. DNA was eluted in TE containing 2% SDS and crosslinks reversed by incubation overnight at 65ºC. DNA was then purified by Qiaquick columns (Qiagen) and quantified using PicoGreen (Invitrogen). ChIP DNA was prepared for HiSeq 2000 Illumina sequencing using a standard protocol consisting in blunting, addition of dA overhangs, ligation of Illumina adapters, selection on gel and PCR with index primers. A mixture of T4 DNA polymerase, DNA polymerase I and T4 kinase was used according to manufacturer’s instruction. Library preparation is carried out on SPRIworks Fragment Library System.

treatment_protocol: Macrophages were subjected to different treatment (see individual samples for details)

value: BMDM cells (7th day of differentiation)

value: Anti-polII (Santa Cruz # sc-899)


title: pol_II_LPS_24h

value: LPS (10 ng/ml) for 24hrs


title: H3K4me1_tc_IFNg_30min

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IFNg for 30min, H3K4me1 ChIP


title: pol_II_UT

value: No treatment


title: Hela_PTB_3end_RNAseq_knockdown_0h and 4h

description: 0h and 4h

value: Hela

value: treated with PTB shRNA and further treated with actinomycin D (2 ug/ml) for 4 hours


title: PU.1_MALP2

description: Chromatin IP against PU.1

value: Anti-PU.1 (in-house rabbit polyclonal against the N-Terminal (aa. 1-100))

value: MALP2 (1 ng/ml) for 4hrs


title: PU.1_CpG

description: Chromatin IP against PU.1

value: CpG (100 nM) for 4hrs


title: H3K4me1_tc_IFNg_1h

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IFNg for 1hrs, H3K4me1 ChIP

value: IFNg (100 ng/ml) for 1hrs


title: PU.1_IL1b_4h

description: Chromatin IP against PU.1

value: IL1b (10 ng/ml) for 4hrs


title: PU.1_TGFb1_4h

description: Chromatin IP against PU.1

value: TGFb1 (1 ng/ml) for 4hrs


title: Stat6_tc_IL4_1h

description: Chromatin IP against Stat6.

source_name: Primary bone marrow-derived macrophages, IL4 for 1hrs, Stat6 ChIP

value: IL4 (10 ng/ml) for 1hrs

value: Santa Cruz Biotechnology, sc-981


title: Stat6_tc_IL4_2h

description: Chromatin IP against Stat6.

source_name: Primary bone marrow-derived macrophages, IL4 for 2hrs, Stat6 ChIP

value: IL4 (10 ng/ml) for 2hrs


title: Stat6_tc_IL4_30min

description: Chromatin IP against Stat6.

source_name: Primary bone marrow-derived macrophages, IL4 for 30min, Stat6 ChIP

value: bone marrow-derived macrophages (7th day of differentiation)

value: IL4 (10 ng/ml) for 30min


title: H3K4me1_IFNg_24h

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IFNg for 24hrs, H3K4me1 ChIP

value: IFNg (100 ng/ml) for 24hrs

value: Abcam, ab8895


title: Stat6_tc_IL4_15min

description: Chromatin IP against Stat6.

source_name: Primary bone marrow-derived macrophages, IL4 for 15min, Stat6 ChIP

value: IL4 (10 ng/ml) for 15min


title: PU.1_tc_IFNg_UT

description: Chromatin IP against PU.1.

source_name: Primary bone marrow-derived macrophages, no treatment, PU.1 ChIP

value: anti-PU.1 (rabbit polyclonal against the N-terminus (aa. 1-100))

value: in-house


title: PU.1_IL4_4h

description: Chromatin IP against PU.1

value: BMDM cells (7th day of differentiation)

value: IL4 (10 ng/ml) for 4hrs


title: Stat6_tc_IL4_UT

description: Chromatin IP against Stat6.

source_name: Primary bone marrow-derived macrophages, no treatment, Stat6 ChIP


title: PU.1_TNFa_4h

description: Chromatin IP against PU.1

value: Anti-PU.1 (in-house rabbit polyclonal against the N-Terminal (aa. 1-100))

value: TNFa (10 ng/ml) for 4hrs


title: Stat6_tc_IL4_4h

description: Chromatin IP against Stat6.

source_name: Primary bone marrow-derived macrophages, IL4 for 4hrs, Stat6 ChIP


title: H3K27ac_tc_IL4_1h

description: Chromatin IP against H3K27ac.

source_name: Primary bone marrow-derived macrophages, IL4 for 1hrs, H3K27ac ChIP

value: Abcam, ab4729


title: PU.1_IFNg_4h

description: Chromatin IP against PU.1

value: IFNg (100 ng/ml) for 4hrs


title: H3K27ac_tc_IL4_30min

description: Chromatin IP against H3K27ac.

source_name: Primary bone marrow-derived macrophages, IL4 for 30min, H3K27ac ChIP


title: PU.1_LPS_24h

description: Chromatin IP against PU.1

value: LPS (10 ng/ml) for 24hrs


title: PU.1_LPS_4h

description: Chromatin IP against PU.1

value: LPS (10 ng/ml) for 4hrs


title: H3K27ac_tc_IL4_15min

description: Chromatin IP against H3K27ac.

source_name: Primary bone marrow-derived macrophages, IL4 for 15min, H3K27ac ChIP


title: H3K27ac_tc_IL4_4h

description: Chromatin IP against H3K27ac.

source_name: Primary bone marrow-derived macrophages, IL4 for 4hrs, H3K27ac ChIP


title: H3K4me3_MALP2

description: Chromatin IP against H3K4me3

value: Anti-H3K4me3 (Active Motif #39159)

value: MALP2 (1 ng/ml) for 4hrs


title: PU.1_UT (2)

description: Chromatin IP against PU.1

value: No treatment


title: H3K4me1_tc_IL4_4h

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IL4 for 4hrs, H3K4me1 ChIP

value: bone marrow-derived macrophages (7th day of differentiation)


title: H3K4me3_TNFa_4h

description: Chromatin IP against H3K4me3


title: H3K4me3_TGFb1_4h

description: Chromatin IP against H3K4me3

value: TGFb1 (1 ng/ml) for 4hrs


title: H3K4me3_CpG

description: Chromatin IP against H3K4me3

value: CpG (100 nM) for 4hrs


title: H3K4me1_tc_IL4_2h

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IL4 for 2hrs, H3K4me1 ChIP

value: IL4 (10 ng/ml) for 2hrs

value: Abcam, ab8895


title: H3K4me3_IL1b_4h

description: Chromatin IP against H3K4me3

source_name: Primary bone marrow-derived macrophages (BMDM)

value: IL1b (10 ng/ml) for 4hrs


title: H3K27ac_tc_IL4_2h

description: Chromatin IP against H3K27ac.

source_name: Primary bone marrow-derived macrophages, IL4 for 2hrs, H3K27ac ChIP


title: H3K27ac_tc_IL4_UT

description: Chromatin IP against H3K27ac.

source_name: Primary bone marrow-derived macrophages, no treatment, H3K27ac ChIP


title: H3K4me1_tc_IL4_1h

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IL4 for 1hrs, H3K4me1 ChIP

value: IL4 (10 ng/ml) for 1hrs


title: H3K4me1_tc_IL4_30min

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IL4 for 30min, H3K4me1 ChIP

value: IL4 (10 ng/ml) for 30min


title: H3K4me1_tc_IL4_15min

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IL4 for 15min, H3K4me1 ChIP

value: IL4 (10 ng/ml) for 15min


title: H3K4me1_tc_IL4_UT

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, no treatment, H3K4me1 ChIP


title: H3K4me3_LPS_4h

description: Chromatin IP against H3K4me3

value: BMDM cells (7th day of differentiation)


title: H3K4me3_IFNg_4h

description: Chromatin IP against H3K4me3


title: H3K4me3_IL4_4h

description: Chromatin IP against H3K4me3

value: IL4 (10 ng/ml) for 4hrs


title: H3K4me3_LPS_24h

description: Chromatin IP against H3K4me3


title: H3K27ac_stat1ko_IFNg_4h

description: Chromatin IP against H3K27ac.

source_name: Primary bone marrow-derived macrophages, IFNg for 4hrs, H3K27ac ChIP

tag: genotype/variation

value: Stat1 KO

value: IFNg (100 ng/ml) for 4hrs

value: Abcam, ab4729


title: H3K27ac_stat1ko_UT

description: Chromatin IP against H3K27ac.


title: H3K4me3_UT (2)

description: Chromatin IP against H3K4me3

value: Anti-H3K4me3 (Active Motif #39159)


title: H3K27ac_stat1ko_ctrl_IFNg_4h

description: Chromatin IP against H3K27ac.


title: H3K27ac_MALP2

description: Chromatin IP against H3K27ac

value: Anti-H3K27ac (Abcam # ab4729)

value: MALP2 (1 ng/ml) for 4hrs


title: H3K27ac_CpG

description: Chromatin IP against H3K27ac


title: H3K27ac_stat1ko_ctrl_UT

description: Chromatin IP against H3K27ac.

value: bone marrow-derived macrophages (7th day of differentiation)


title: PU.1_stat1ko_ctrl_IFNg_4h

description: Chromatin IP against PU.1.

source_name: Primary bone marrow-derived macrophages, IFNg for 4hrs, PU.1 ChIP

value: anti-PU.1 (rabbit polyclonal against the N-terminus (aa. 1-100))

value: in-house


title: PU.1_stat1ko_IFNg_4h

description: Chromatin IP against PU.1.


title: PU.1_stat1ko_UT

description: Chromatin IP against PU.1.

source_name: Primary bone marrow-derived macrophages, no treatment, PU.1 ChIP


title: H3K27ac_TGFb1_4h

description: Chromatin IP against H3K27ac

value: TGFb1 (1 ng/ml) for 4hrs


title: H3K4me3_UT

description: Chromatin IP against H3K4me3

value: No treatment


title: H3K27ac_IL1b_4h

description: Chromatin IP against H3K27ac

value: IL1b (10 ng/ml) for 4hrs


title: H3K27ac_TNFa_4h

description: Chromatin IP against H3K27ac

value: TNFa (10 ng/ml) for 4hrs


title: H3K27ac_IFNg_4h

description: Chromatin IP against H3K27ac


title: H3K4me1_stat1ko_UT

description: Chromatin IP against H3K4me1.

value: Abcam, ab8895


title: H3K27ac_IL4_4h

description: Chromatin IP against H3K27ac


title: H3K4me1_stat1ko_IFNg_4h

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IFNg for 4hrs, H3K4me1 ChIP


title: H3K27ac_LPS_24h

description: Chromatin IP against H3K27ac

value: LPS (10 ng/ml) for 24hrs


title: H3K27ac_LPS_4h

description: Chromatin IP against H3K27ac

value: BMDM cells (7th day of differentiation)

value: LPS (10 ng/ml) for 4hrs


title: H3K27ac_UT (2)

description: Chromatin IP against H3K27ac


title: H3K4me1_stat1ko_ctrl_UT

description: Chromatin IP against H3K4me1.


title: H3K4me1_stat1ko_ctrl_IFNg_4h

description: Chromatin IP against H3K4me1.


title: H3K4me1_washout_IFNg_24h+48h

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IFNg for 24hrs, then washed, H3K4me1 ChIP

value: IFNg (100 ng/ml) for 24hrs, then washed for 48hrs


title: H3K4me1_washout_IFNg_24h

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IFNg for 24hrs, H3K4me1 ChIP

value: IFNg (100 ng/ml) for 24hrs


title: H3K4me1_washout_IFNg_72h

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IFNg for 72hrs, H3K4me1 ChIP

value: IFNg (100 ng/ml) for 72hrs


title: H3K4me1_CpG

description: Chromatin IP against H3K4me1

value: Anti-H3K4me1 (Abcam # ab8895)

value: CpG (100 nM) for 4hrs


title: H3K4me1_MALP2

description: Chromatin IP against H3K4me1

value: MALP2 (1 ng/ml) for 4hrs


title: H3K4me1_washout_IFNg_UT

description: Chromatin IP against H3K4me1.

value: bone marrow-derived macrophages (7th day of differentiation)


title: H3K4me1_washout_IL4_24h+48h

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IL4 for 24hrs, then washed, H3K4me1 ChIP

value: IL4 (10 ng/ml) for 24hrs, then washed for 48hrs


title: H3K4me1_IL1b_4h

description: Chromatin IP against H3K4me1


title: H3K4me1_TGFb1_4h

description: Chromatin IP against H3K4me1


title: H3K4me1_washout_IL4_72h

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IL4 for 72hrs, H3K4me1 ChIP

value: IL4 (10 ng/ml) for 72hrs


title: H3K4me1_IFNg_4h

description: Chromatin IP against H3K4me1

value: IFNg (100 ng/ml) for 4hrs


title: H3K4me1_TNFa_4h

description: Chromatin IP against H3K4me1


title: PU.1_stat1ko_ctrl_UT

description: Chromatin IP against PU.1.

value: anti-PU.1 (rabbit polyclonal against the N-terminus (aa. 1-100))

value: in-house


title: H3K4me1_washout_IL4_UT

description: Chromatin IP against H3K4me1.


title: H3K4me1_IL4_4h

description: Chromatin IP against H3K4me1

value: IL4 (10 ng/ml) for 4hrs


title: H3K4me1_LPS_4h

description: Chromatin IP against H3K4me1


title: H3K4me1_LPS_24h

description: Chromatin IP against H3K4me1


title: H3K27ac_stat6ko_ctrl_IL4_4h

description: Chromatin IP against H3K27ac.

source_name: Primary bone marrow-derived macrophages, IL4 for 4hrs, H3K27ac ChIP

value: Balb/c

value: Abcam, ab4729


title: H3K27ac_stat6ko_IL4_4h

description: Chromatin IP against H3K27ac.

value: Stat6 KO


title: H3K4me1_UT (2)

description: Chromatin IP against H3K4me1

source_name: Primary bone marrow-derived macrophages (BMDM)

value: BMDM cells (7th day of differentiation)

value: No treatment


title: H3K4me1_UT (1)

description: Chromatin IP against H3K4me1


title: PU.1_stat6ko_IL4_4h

description: Chromatin IP against PU.1.

source_name: Primary bone marrow-derived macrophages, IL4 for 4hrs, PU.1 ChIP


title: PU.1_stat6ko_UT

description: Chromatin IP against PU.1.


title: PU.1_stat6ko_ctrl_IL4_4h

description: Chromatin IP against PU.1.


title: BMDM cells IL1b

data_processing:
Reads were quality filtered according to the Illumina pipeline
Reads were mapped to the mouse mm9 genome and transcripts using TopHat 1.3.1 (PMID: 19289445), allowing up to two mismatches and with a mean distance between pairs (-r) of 250bp
Transcripts abundance were quantified using Cufflinks 1.2.1 (PMID: 20436464)
Differentially expressed genes were called using Cuffdiff 1.2.1 (PMID: 20436464)
Genome_build: mm9 (NCBI Build 37), transcripts from Mus Musculus Ensembl Build 63
Supplementary_files_format_and_content: *.fpkm_tracking list the absolute value of expression (FPKM), *.diff list the statistics for differential expression compared to the untreated (refer to http://cufflinks.cbcb.umd.edu/manual.html for details)

extract_protocol: The poly-A containing mRNA molecules were purified using poly-T oligo-attached magnetic beads. Following purification, the mRNA is fragmented into small pieces using divalent cations under elevated temperature. The cleaved RNA fragments are copied into first strand cDNA using reverse transcriptase and random primers. This is followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. These cDNA fragments then go through an end repair process, the addition of a single A base, and then ligation of the adapters. The products are then purified and enriched with PCR to create the final cDNA library.

value: IL1b (10 ng/ml) for 4hrs


title: BMDM cells CpG


title: BMDM cells MALP2


title: BMDM cells TGFb1

value: TGFb1 (1 ng/ml) for 4hrs


title: H3K4me1_washout_IL4_24h

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IL4 for 24hrs, H3K4me1 ChIP

value: IL4 (10 ng/ml) for 24hrs

value: Abcam, ab8895


title: H3K4me1_stat6ko_IL4_4h

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, IL4 for 4hrs, H3K4me1 ChIP


title: PU.1_stat6ko_ctrl_UT

description: Chromatin IP against PU.1.


title: H3K4me1_stat6ko_UT

description: Chromatin IP against H3K4me1.


title: BMDM cells TNFa

value: TNFa (10 ng/ml) for 4hrs


title: BMDM cells IL4


title: H3K4me1_stat6ko_ctrl_IL4_4h

description: Chromatin IP against H3K4me1.

value: bone marrow-derived macrophages (7th day of differentiation)


title: E13.5_m_1

status: Public on Jan 16 2013

city: La Jolla

first: Kun

last: Zhang

address: 9500 Gilman Dr Mailcode: 0412

department: Bioengineering

institute: University of California, San Diego

description: sequencing barcode id: Indx5

data_processing:
mRNA-seq reads generated from each sample were aligned to the mouse genome (mm9, NCBI build 37) with Bowtie/Tophat v1.3.1 (http://tophat.cbcb.umd.edu) which allows mapping across splice sites by reads segmentation. All programs were used with default setting unless otherwise specified.
Mapped reads (94-95% of total reads) were subsequently assembled into transcripts guided by reference annotation (mm9, USCS gene annotation) with Cufflinks v1.2.1 (http://cufflinks.cbcb.umd.edu).
Expression level of each transcript was quantified with normalized FPKM (fragments per kilobase of exon per million mapped fragments).  Two or three biological replicates were used in each genotype to identify transcripts that showed significant differences at a FDR cutoff <0.05 between wild type and Tet1Gt/Gt by Cuffdiff v1.2.1.
Functional annotation of significantly different transcripts and enrichment analysis was performed with DAVID (http://david.abcc.ncifcrf.gov).
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited text files include average RPKM for different developmental stages and statistics generated by CuffLinks (cuffdiff)

source_name: Primordial germ cells

extract_protocol: Total RNA was purified from 400-5,000 PGCs using ZR RNA microprep kit (Zymo Research, UAS). The cDNA synthesis and amplification was performed with the SMARTer ultra low input RNA kit (Clontech, US). The amplified cDNA (2-70 ng) was then fragmented by Covaris S2 sonicator (Covaris, US) and converted to sequencing libraries following the Illumina’s construction protocol for low input DNA (Illumina, US). Barcoded libraries were pooled and sequenced in three lanes of Illumina Hiseq 2000 instrument.

tag: tissue

value: Primordial germ cells

tag: age

value: embryonic 13.5 day

tag: genotype

value: wild type


title: BMDM cells IFNg

value: IFNg (100 ng/ml) for 4hrs


title: E13.5_m_2

description: sequencing barcode id: Indx9


title: H3K27ac_stat6ko_UT

description: Chromatin IP against H3K27ac.

source_name: Primary bone marrow-derived macrophages, no treatment, H3K27ac ChIP


title: Bcd6x Rep 3 Slice 4

city: Stanford

first: Peter

middle: Acuña

last: Combs

address: Gilbert Rm 309, 371 Serra Mall

department: Biology

institute: Stanford University

description: 60um slice of D. melanogaster pooled with D. persimilis Index Barcode 16

data_processing:
Bases were called using HiSeq Control Software v1.8 and Real Time Analysis v2.8.
Reads were aligned to concatenated D.melanogaster and carrier species reference genomes using Tophat v.2.0.6, tophat -p8 --no-novel-juncs --read-edit-dist 6 --report-secondary-alignments
Aligned reads were filtered using custom software (http://github.com/petercombs/EisenLab-Code/)
Only unambiguous D. melanogaster reads were used for FPKM estimation and differential expression calling using Cufflinks and Cuffdiff v2.0.2
Genome_build: FlyBase FB2012_05
Supplementary_files_format_and_content: genes.cuff: cuffdiff generated FPKM_tracking files for the combined replicates

source_name: Cryo-sliced embryo

growth_protocol: embryos were collected, fixed in methanol, imaged under a light microscope, and selected at 40% cellularization.

extract_protocol:
60 um transverse sections of embryos were taken, then RNA was extracted in TRIzol.
Total RNA was pooled with total RNA from a single embryo of a carrier species (D. persimilis, D. willistoni, or D. mojavensis). RNA libraries were prepared for sequencing using standard barcoded Illumina TruSeq protocols, scaling all reactions by half

treatment_protocol: Selected embryos were dyed with bromophenol blue, aligned, submerged in TFM, and flash-frozen in LN2.

tag: genetic background

value: Canton S

value: embryo

tag: developmental stage

value: blastoderm


title: H3K4me1_stat6ko_ctrl_UT

description: Chromatin IP against H3K4me1.

source_name: Primary bone marrow-derived macrophages, no treatment, H3K4me1 ChIP

value: Balb/c


title: Bcd6x Rep 2 Slice 7

description: 60um slice of D. melanogaster pooled with D. willistoni Index Barcode 12

tag: sample type


title: Bcd6x Rep 3 Slice 3

description: 60um slice of D. melanogaster pooled with D. persimilis Index Barcode 15


title: H3K27ac_stat6ko_ctrl_UT

description: Chromatin IP against H3K27ac.

value: Abcam, ab4729


title: Bcd6x Rep 3 Slice 5

description: 60um slice of D. melanogaster pooled with D. persimilis Index Barcode 18


title: Bcd6x Rep 3 Slice 2

description: 60um slice of D. melanogaster pooled with D. persimilis Index Barcode 14


title: Bcd6x Rep 2 Slice 6

description: 60um slice of D. melanogaster pooled with D. willistoni Index Barcode 11


title: Bcd6x Rep 2 Slice 5

description: 60um slice of D. melanogaster pooled with D. willistoni Index Barcode 10


title: Bcd6x Rep 2 Slice 3

description: 60um slice of D. melanogaster pooled with D. willistoni Index Barcode 8


title: Bcd6x Rep 2 Slice 1

description: 60um slice of D. melanogaster pooled with D. willistoni Index Barcode 6


title: Bcd6x Rep 2 Slice 4

description: 60um slice of D. melanogaster pooled with D. willistoni Index Barcode 9


title: Bcd6x Rep 1 Slice 4

description: 60um slice of D. melanogaster pooled with D. mojavensis Index Barcode 4


title: Bcd6x Rep 1 Slice 5

description: 60um slice of D. melanogaster pooled with D. mojavensis Index Barcode 5


title: Bcd6x Rep 1 Slice 2

description: 60um slice of D. melanogaster pooled with D. mojavensis Index Barcode 2


title: Bcd6x Rep 1 Slice 3

description: 60um slice of D. melanogaster pooled with D. mojavensis Index Barcode 3


title: WT Rep 3 Slice 5

description: 60um slice of D. melanogaster pooled with D. mojavensis Index Barcode 18


title: Bcd6x Rep 1 Slice 1

description: 60um slice of D. melanogaster pooled with D. mojavensis Index Barcode 1


title: WT Rep 3 Slice 4

description: 60um slice of D. melanogaster pooled with D. mojavensis Index Barcode 16


title: WT Rep 3 Slice 3

description: 60um slice of D. melanogaster pooled with D. mojavensis Index Barcode 15


title: WT Rep 3 Slice 2

description: 60um slice of D. melanogaster pooled with D. mojavensis Index Barcode 14


title: WT Rep 2 Slice 6

description: 60um slice of D. melanogaster pooled with D. willistoni Index Barcode 12


title: WT Rep 2 Slice 5

value: 60um slice of D. melanogaster pooled with D. willistoni Index Barcode 11


title: WT Rep 2 Slice 4

value: 60um slice of D. melanogaster pooled with D. willistoni Index Barcode 10


title: WT Rep 2 Slice 3

value: 60um slice of D. melanogaster pooled with D. willistoni Index Barcode 9


title: E13.5_m_3

description: sequencing barcode id: Indx10

tag: strain


title: WT Rep 1 Slice 6

description: 60um slice of D. melanogaster pooled with D. persimilis Index Barcode 6


title: WT Rep 3 Slice 1

description: 60um slice of D. melanogaster pooled with D. mojavensis Index Barcode 13


title: WT Rep 2 Slice 2

value: 60um slice of D. melanogaster pooled with D. willistoni Index Barcode 8


title: WT Rep 1 Slice 4

description: 60um slice of D. melanogaster pooled with D. persimilis Index Barcode 4


title: WT Rep 1 Slice 3

description: 60um slice of D. melanogaster pooled with D. persimilis Index Barcode 3


title: WT Rep 1 Slice 5

description: 60um slice of D. melanogaster pooled with D. persimilis Index Barcode 5


title: WT Rep 1 Slice 2

description: 60um slice of D. melanogaster pooled with D. persimilis Index Barcode 2


title: Bcd6x Rep 3 Slice 1

description: 60um slice of D. melanogaster pooled with D. persimilis Index Barcode 13


title: WT Rep 1 Slice 1

description: 60um slice of D. melanogaster pooled with D. persimilis, Index Barcode 1


title: 293T_Hspa1a_2hrs_severe_Heat_Shock

status: Public on Jan 15 2013

city: Haifa

first: Reut

last: Shalgi

address: 1 Efron st.

department: Faculty of Medicine

country: Israel

institute: Technion

data_processing:
Riboseq: first, reads were trimmed to a length of 32, then polyAs were further trimmed off. reads with lengthes of 22-32 (for the Hsp70 inhibitor sample, 28-32 were used) were seperately uniquely mapped to the same genome file using bowtie, and the output sam files were unified to create a single sam file for each sample.
Genome_build: mm9 and hg18
Supplementary_files_format_and_content: BED files

source_name: 293T cells

extract_protocol: Riboseq - described in Shalgi et al. 2013. similar to Ingolia et al. Science, 2009

treatment_protocol:
293T cells were transfected with GFP or Hspa1a plasmid. 48 hours later, cells were either treated with 2 hours of severe heat shock at 44C (HS) or not.
Hsp70 inhibitor: VER-155008 (Massey et al. 2010) at 20microM concentration for 3 hours.

value: 293T cells

tag: transfected with

value: severe heat shock (44C 2 hours)

tag: assay type

value: Riboseq


title: 293T_GFP_2hrs_severe_Heat_Shock

tag: cell line


title: WT Rep 3 Slice 6

description: 60um slice of D. melanogaster pooled with D. mojavensis Index Barcode 19


title: input DNA_rep2 [primary pre-B cells]

city: London

first: Gopuraja

last: Dharmalingam

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

department: Epigenetics section

institute: MRC London Institute of Medical Sciences

description:
Input data from primary pre-B Cells
input-2
input DNA, rep2

data_processing:
Base-calling was performed using CASAVA version 1.7
Reads of 36 bases were aligned to the Mus musculus mm9 genome assembly using ELAND (Illumina) with default parameter. Multiple sequencing runs of the same samples were combined for further processing.
Duplicate alignments were filtered using a Shell script
BED files were generated from ELAND aligned files using a PERL script
Peaks were identified using Swembl (http://www.ebi.ac.uk/~swilder/SWEMBL/)
Genome_build: mm9 build 37
Supplementary_files_format_and_content: BED files

source_name: primary pre-B cell

growth_protocol: Primary pre-B cells were established from fetal liver cells plated on irradiated S17 stromal cells in IMDM, 10% FCS, 1% penicillin-streptomycin, 2 mM 2-mercaptoethanol, and 5 ng/ml IL-7.

extract_protocol: Chromatin isolated from fixed nuclei was sonicated and incubated at 4ºC overnight with either Protein A Dynabeads (Invitrogen) coated with rabbit anti-C-terminal Ikaros antibody or Protein G Dynabeads (Invitrogen) coated with anti-haemagglutinin (HA) (Covance). Single read high throughput sequencing libraries were prepared from 10ng of ChIP DNA using Illumina Chip-Seq Sample Prep. Kit and sequenced according to the manufacturer's protocols using Illumina Genome Analyzer II.

value: a mix of C57BL/6 and 129

value: Fetal liver

value: primary pre-B cell


title: 3173-67B6-ACT

first: Luba

last: Pardo

address: de Boelelaan

department: Clinical Genetics

institute: VU Medical Center

description:
Sample 25
barcode: 3173-67B6-ACT

data_processing:
After sequencing fastq files were generated.  To extract linkers and adapters from the CAGE tags, custom perl scripts were written. Other sequencing artefacts were removed with the TAGDUST software. The scripts were developed as part of a mapping pipeline written at the RIKEN.
CAGE tags were mapped to the human genome (Hg18 build) with BWA.
Mapped CAGE raw tags were hierarchically  clustered into Level1 , level 2 and Level3 Tag clusters using a clustering pipeline developed by the ibioinformatics department at the RIKEN, Japan.
Genome_build: Hg 18
Supplementary_files_format_and_content: tab-delimited text file contains all Tag clusters after mapping and clustering procedures for 25 CAGE libraries (contains both raw tag counts and a normalized tags counts)

extract_protocol:
Total RNA was extracted and purified from tissues using the Trizol tissue kit according to instructions provided by the manufacturer (Invitrogen).
CAGE and massive parallel sequencing with Illumina;
The standard CAGE protocol (Kodzius et al., 2006) was adapted for sequencing on an Illumina platform. A thorough description of the protocol to prepare CAGE libraries and to sequence CAGE tags is presented in Takahashi et al.(Takahashi et al., 2012)

tag: subject id

tag: age at death

value: 70 years

value: Post morten brain

tag: tissue region

value: Putamen


title: input DNA_rep1 [primary pre-B cells]

description:
Input data from primary pre-B Cells
input-1
input DNA, rep1

tag: cell type


title: 3172-67A6-ACA

description:
Sample 24
barcode: 3172-67A6-ACA

value: Caudate


title: endogenous Ikaros_rep2, lanes 1 and 2 [primary pre-B cells]

description:
ChIP-Seq data from primary pre-B  Cells using Rabbit anti-C-terminal Ikaros antibody
Ikaros-2.1
Ikaros-2.2
endogenous Ikaros, rep2, lane 1
endogenous Ikaros, rep2, lane 2

tag: chip antibody

value: Rabbit anti-C-terminal Ikaros [from Stephen Smale Lab]


title: 3171-67I5-GAT

description:
Sample 23
barcode: 3171-67I5-GAT

value: 89 years


title: WT Rep 2 Slice 1

description: 60um slice of D. melanogaster pooled with D. willistoni Index Barcode 7

value: wild type

value: Canton S

value: embryo

value: blastoderm


title: endogenous Ikaros_rep1, lanes 1 and 2 [primary pre-B cells]

description:
ChIP-Seq data from primary pre-B  Cells using Rabbit anti-C-terminal Ikaros antibody
Ikaros-1.1
Ikaros-1.2
endogenous Ikaros, rep1, lane 1
endogenous Ikaros, rep1, lane 2


title: 3170-67H5-CTT

description:
Sample 22
barcode: 3170-67H5-CTT


title: PU.1_tc_IFNg_1h

description: Chromatin IP against PU.1.

source_name: Primary bone marrow-derived macrophages, IFNg for 1hrs, PU.1 ChIP

tag: strain/background

value: bone marrow-derived macrophages (7th day of differentiation)

tag: treatment

value: IFNg (100 ng/ml) for 1hrs

value: anti-PU.1 (rabbit polyclonal against the N-terminus (aa. 1-100))

tag: antibody vendor, catalog #

value: in-house


title: 2902-72I6-ATG

description:
Sample 21
barcode: 2902-72I6-ATG

value: 97 years

tag: tissue


title: 2894-72A6-ACG

description:
Sample 19
barcode: 2894-72A6-ACG

value: 91 years


title: 2895-72B6-ATC

description:
Sample 20
barcode: 2895-72B6-ATC

value: 87 years

value: Hippocampus


title: input DNA, rep2

description: Input data from Wildtype B3 Cells

data_processing:
Base-calling was performed using CASAVA version 1.7
Reads of 36 bases were aligned to the Mus musculus mm9 genome assembly using ELAND (Illumina) with default parameters
Duplicate alignments were filtered using a Shell script
BED files were generated from ELAND aligned files using a PERL script
Peaks were identified using Swembl (http://www.ebi.ac.uk/~swilder/SWEMBL/)
Genome_build: mm9 build 37
Supplementary_files_format_and_content: BED files

source_name: Input data from Wildtype B3 Cells

growth_protocol: Mouse pre-B cell line B3 was grown in IMDM, 10% FCS and  1% penicillin-streptomycin.

treatment_protocol: B3 cells were transduced as described  (Thompson et al., 2007, Immunity 26:335-344) with mouse stem cell virus (MSCV) vectors encoding wild-type Ikaros tagged with a haemagglutinin (HA) epitope followed by an internal ribosomal entry site (IRES) and GFP (HA-Ikaros-IRES-GFP), or the DNA binding-deficient Ikaros mutant 159A (HA-159A Ikaros-IRES-GFP). Ikaros inducible cells were generated by transducing B3 cells with a construct encoding a fusion protein of HA-tagged Ikaros and the estrogen receptor hormone-binding domain (HA-Ikaros-ERt2-IRES-GFP) or an ERt2-IRES-GFP control vector.

value: pre-B cell line B3

tag: cell line reference

value: Brown et al. 1997


title: input DNA, rep3

description: Input data from HA-Ikaros transduced B3 cells


title: input DNA, rep1

description: Input data from Wildtype B3 Cells


title: 2868-72C5-ACA

description:
Sample 17
barcode: 2868-72C5-ACA

value: Post morten brain


title: 2867-72B5-GAT

description:
Sample 16
barcode: 2867-72B5-GAT

value: temporal cortex


title: 2893-72I5-ACT

description:
Sample 18
barcode: 2893-72I5-ACT

value: Caudate


title: epitope-tagged HA-Ikaros, rep2

description: ChIP-Seq data from HA-Ikaros transduced B3 cells using Anti-haemagglutinin (HA)

value: Anti-haemagglutinin (HA) (Covance).


title: 293T_Hspa1a

value: 293T cells

value: Riboseq


title: endogenous Ikaros, rep1

description: ChIP-Seq data from Wildtype B3 Cells using Rabbit anti-C-terminal Ikaros antibody

value: Rabbit anti-C-terminal Ikaros


title: endogenous Ikaros, rep2

source_name: ChIP-Seq data from Wildtype B3 Cells using Rabbit anti-C-terminal Ikaros antibody


title: 2866-72A5-CTT

description:
Sample 15
barcode: 2866-72A5-CTT

value: frontal cortex


title: 293T_GFP


title: 2906-72D7-ACG

description:
Sample 13
barcode: 2906-72D7-ACG


title: 2905-72C7-ACT

description:
Sample 12
barcode: 2905-72C7-ACT


title: 2900-72G6-ATG

description:
Sample 9
barcode: 2900-72G6-ATG


title: 2897-72D6-ACT

description:
Sample 7
barcode: 2897-72D6-ACT

value: Putamen


title: 2896-72C6-ACA

description:
Sample 6
barcode: 2896-72C6-ACA


title: 2873-72H5-ATG

description:
Sample 5
barcode: 2873-72H5-ATG

value: 82 years

value: hippocampus


title: 2901-72H6-AGC

description:
Sample 10
barcode: 2901-72H6-AGC

value: 97 years


title: 2899-72F6-ATC

description:
Sample 8
barcode: 2899-72F6-ATC


title: 2872-72G5-ATC

description:
Sample 4
barcode: 2872-72G5-ATC


title: 2871-72F5-ACG

description:
Sample 3
barcode: 2871-72F5-ACG


title: 2870-72E5-ACT

description:
Sample 2
barcode: 2870-72E5-ACT


title: 2869-72D5-ACA

description:
Sample 1
barcode: 2869-72D5-ACA


title: Bcd6x Rep 2 Slice 2

tag: genotype

tag: genetic background

value: Canton S

value: embryo

value: blastoderm

value: 60um slice of D. melanogaster pooled with D. willistoni Index Barcode 7


title: epitope-tagged HA-Ikaros, rep1

source_name: ChIP-Seq data from HA-Ikaros transduced B3 cells using Anti-haemagglutinin (HA)

value: pre-B cell line B3

value: Brown et al. 1997


title: 2908-72F7-ATC

description:
Sample 14
barcode: 2908-72F7-ATC

value: 87 years

value: Post morten brain


title: GAL-Scc1-HA, - break, Rad30del. INPUT

city: Tokyo

first: Yuki

last: KATOU

address: 1-1-1 Yayoi,Bunkyo-ku

institute: Research assistant

data_processing:
Basecalls performed using Corona
then mapped to S.c erevisiae whole genome using bowtie v0.12.2 with parameters -n3 -m10
Data were visualized using Cairo
Genome_build: Saccer1
Supplementary_files_format_and_content: wig files

source_name: GAL-Scc1-HA, - break, Rad30del. INPUT

growth_protocol: Cells grown in YEP media supplemented with 2% raffinose (YEPR)  were arrested in G2/M, by addition of benomyl.

extract_protocol:
Cells were broken by beadbeater and sonication.
The 5500 SOLiD™ Fragment Library Core Kit(#4464412)

value: input DNA

tag: cell cycle

value: G2 (benomyl)

value: rad30 deletion


title: GAL-Scc1-HA, + break, Rad30del. IP

source_name: GAL-Scc1-HA, + break, Rad30del. IP

tag: chip antibody manufacturer

value: Covance


title: GAL-Scc1-HA, - break, wt INPUT

source_name: GAL-Scc1-HA, - break, wt INPUT

value: WT RAD30


title: GAL-Scc1-HA, - break, wt IP

source_name: GAL-Scc1-HA, - break, wt IP


title: GAL-Scc1-HA, - break, Rad30del. IP

source_name: GAL-Scc1-HA, - break, Rad30del. IP


title: E11.5_1

description: sequencing barcode id: Indx4

tag: strain

value: Primordial germ cells

tag: age

value: embryonic 11.5 day

value: wild type


title: GAL-Scc1-HA, + break, Rad30del. INPUT

source_name: GAL-Scc1-HA, + break, Rad30del. INPUT


title: E9.5_2

description: sequencing barcode id: Indx7

value: embryonic 9.5 day


title: E11.5_2

description: sequencing barcode id: Indx8


title: GAL-Scc1-HA, + break, wt IP

source_name: GAL-Scc1-HA, + break, wt IP


title: GAL-Scc1-HA, + break, wt INPUT

source_name: GAL-Scc1-HA, + break, wt INPUT


title: E9.5_1 2

description: sequencing barcode id: Indx3

source_name: Primordial germ cells


title: 3T3-Hsp70_inhibitor_3hr

source_name: 3T3 cells

tag: cell line

value: 3T3 cells

value: Hsp70 inhibitor (VER-155008, Massey et al.) for 3 hours

tag: assay type

value: Riboseq


title: ovary

status: Public on Jan 14 2013

city: shenzhen

first: zhiyong

last: huang

state: Guangdong

address: F5, Main Building, Beishan Industrial Zone, Beishan Road, Yantian District

institute: BGI-Shenzhen, Shenzhen 518083, China

description: Sequenced reads were generated from ovary in Chinese tree shrew

data_processing:
use Chinese tree shrew genome as reference, sequenced reads were aligned onto the corresponding reference using TopHat v1.0.12 with default parameters except  -r 20 --mate-std-dev 10 -I 10000  --segment-mismatches 3  --segment-length 24 --solexa1.3-quals
Genome_build: This Whole Genome Shotgun project has been deposited at DDBJ/EMBL/GenBank under the accession ALAR00000000. The version described in this paper is the first version, ALAR01000000.
Supplementary_files_format_and_content: sequenced reads were aligned onto the corresponding contig

organism: Tupaia chinensis

source_name: ovary

extract_protocol: For RNA-seq, poly-A+ RNA was isolated with 30 oligo-dT-coupled beads from 20 μg total RNA of each sample. First strand cDNA synthesis was performed with random hexamers and Superscript II reverse transcriptase (Invitrogen). The second strand was synthesized with E. coli DNA PolI (Invitrogen). Double stranded cDNA was purified with Qiaquick PCR purification kit (Qiagen), and sheared with a nebulizer (Invitrogen) to 100-500 bp fragments. After end repair and addition of a 3’ dA overhang the cDNA was ligated to Illumina PE adapter oligo mix, and size selected to 200±20 bp fragments by gel purification. After 15 cycles of PCR amplification the libraries were sequenced using Illumina HiSeq 2000 and the paired-end sequencing module.

treatment_protocol: total RNA is extracted by TRIzol Reagent

value: ovary


title: 2904-72B7-ACA

description:
Sample 11
barcode: 2904-72B7-ACA

tag: subject id

tag: age at death

value: temporal cortex


title: testis

description: Sequenced reads were generated from testis in Chinese tree shrew

source_name: testis

value: testis


title: Starved

city: Oxford

first: Stephen

last: Taylor

address: Headington

institute: CBRG

data_processing:
Paired-end reads from ChIP-Seq and input samples were aligned to the mouse genome build using Bowtie (version 0.12.3) using standard parameters
Wig files of genomic read distribution were produced using a sliding 300bp window moving at 30 bp increments
Genome_build: mm9
Supplementary_files_format_and_content: Wig files of genomic distribution of aligned reads

source_name: Mouse Embryonic Fibroblasts

growth_protocol: Mouse embryo fibroblasts  were derived from E13.5 C57/BL6 embryos after removal of the head, heart  and liver and outgrowth for 15 days in DMEM supplemented with10%FCS.

extract_protocol:
Cells were fixed with 1% formaldehyde in PBS for 10 min at RT. Erf-DNA complexes were immunoprecipitated from sonicated  nuclear extracts with the S17S anti-Erf rabbit polyclonal antibody
Standard Illumina Paired End

treatment_protocol: For serum starvation, 90%-confluent cultures were grown for 4 hours in DMEM supplemented with 20mM HEPES ph7

value: Fibroblasts

tag: developmental stage

tag: antibody


title: brain

description: Sequenced reads were generated from brain in Chinese tree shrew

source_name: brain

value: brain


title: pancreas

description: Sequenced reads were generated from pancreas in Chinese tree shrew

source_name: pancreas

value: pancreas


title: heart

description: Sequenced reads were generated from heart in Chinese tree shrew

source_name: heart

value: heart


title: Exponential

value: S17S anti-Erf rabbit polyclonal antibody


title: Input SEPTIN4 replica 1

city: New York

first: Marian

last: Mellen

address: 1230 York Avenue

institute: The Rockefeller University

data_processing:
Library strategy: 5hmC-Seq
Reads were aligned to mm9 mouse genome assembly using Bowtie v0.12.7  (-m1 --best).  (-m1 --best).
Further analysis was done using Bioconductor v2.9 using packages chipseq, biomaRt,  rtracklayer, MEDIPS and custom scripts.

source_name: Bergmann glia

extract_protocol:
Isolation and sorting was performed as described in (Kriaucionis and  Heintz, 2009). Briefly, cerebella were dissected as described above and homogenized in homogenization  medium (0.25 M sucrose, 150 mM KCl, 5 mM MgCl2, 20 mM Tricine pH 7.8, 0.15 mM spermine,  0.5 mM spermidine, EDTA-free protease inhibitor cocktail (Roche) using loose (A) and tight (B) glassglass  dounce.Homogenate was supplemented with 50% iodixanol (OptiPrep, Axis-Shield,  Scotland), 150 mM KCl, 5 mM MgCl2, 20 mM Tricine pH 7.8; and laid on a 29% iodixanol cushion. Cerebella  from 12 mice (6 males and 6 females) were used for PCs, 6 (3 males and 3 females) for BGs and 4 (2 males  and 2 females) for GCs. Nuclei were pelleted by centrifugation 30 min, 10,000 g, 4 °C in swinging  bucket rotor (SW41) in a Beckman Coulter XL-70 ultracentrifuge. The nuclear pellet was resuspended in  homogenization buffer and co-stained with DyeCycle Ruby (Invirogen) to 20 μM final concentration.  Nuclei were sorted in a BD FASCAria (BD Biosiences, San Jose, CA, USA) cell sorter using 635 nm and  488 nm excitation lasers and by gating with two parameters: high GFP signal (compared to wt mice)  indicating bacTRAP positive cells and lowest signal for DC Ruby, indicating singlets. Nuclei were fast frozen in  liquid nitrogen after sorting and kept at -80 C until analysis.
5hmC was pulled down as described (Song et al., 2010). After  purification DNA was amplified as described in TruSeq DNA Sample kit. 1-0.5 μg of DNA was used for each experiment.  Sonicated DNA was end-repaired following by ligation to Illumina paired end sequencing adapters (Illumina,  PE‐ 102‐ 1003) following  by amplification with Illumina primers. Input samples were produced for  each cell types in both procedures. 5hmC enriched were then sequenced using Illumina platform obtaining more than 50 x10-6, 36-  bp single-end reads per sample. Reads were aligned to mm9 mouse genome assembly using Bowtie v0.12.7  (-m1 --best).Two biological replicas were done for each of the cell  type

tag: cell type

value: Bergmann glia

value: 8-12 weeks

value: wt


title: 5hmC-Seq SEPTIN4 replica 2


title: 5hmC-Seq SEPTIN4 replica 1


title: 5hmC-Seq ND1 replica 2

source_name: Cerebellar Granule cells

value: Cerebellar Granule cells


title: Input SEPTIN4 replica 2


title: Input ND1 replica 1


title: Input PCP2 replica 2

source_name: Purkinje cells

value: Purkinje cells


title: liver

description: Sequenced reads were generated from liver in Chinese tree shrew

source_name: liver

value: liver


title: 5hmC-Seq PCP2 replica 2


title: Input ND1 replica 2


title: Input PCP2 replica 1


title: GC WT male 1

description: 5hmC IP DNA

source_name: GRANULE CELLS

extract_protocol:
Isolation and sorting was performed as described in (Kriaucionis and  Heintz, 2009). Briefly, cerebella were dissected as described above and homogenized in homogenization  medium (0.25 M sucrose, 150 mM KCl, 5 mM MgCl2, 20 mM Tricine pH 7.8, 0.15 mM spermine,  0.5 mM spermidine, EDTA-free protease inhibitor cocktail (Roche) using loose (A) and tight (B) glassglass  dounce.Homogenate was supplemented with 50% iodixanol (OptiPrep, Axis-Shield,  Scotland), 150 mM KCl, 5 mM MgCl2, 20 mM Tricine pH 7.8; and laid on a 29% iodixanol cushion. GC nuclei from WT and Mecp2-null mice were sorted by selecting a specific FSC/SSC  population identified from GC EGFP+ mice. We observed that this FSC/SSC gated population in GC EGFP+  animals resulted >92% of the sorted nuclei being EGFP positive. Thus, the gate selected allows us  to enrich the sample from 65-70% to 92-96% on GC cells. .
5hmC was pulled down as described (Song et al., 2010). After  purification DNA was amplified as described in TruSeq DNA Sample kit. 1-0.5 μg of DNA was used for each experiment.  Sonicated DNA was end-repaired following by ligation to Illumina paired end sequencing adapters (Illumina,  PE‐ 102‐ 1003) following  by amplification with Illumina primers. Input samples were produced for  each cell types in both procedures. 5hmC enriched were then sequenced using Illumina platform obtaining more than 50 x10-6, 36-  bp single-end reads per sample. Reads were aligned to mm9 mouse genome assembly using Bowtie v0.12.7  (-m1 --best).Two biological replicas were done for each of the cell  type

value: GRANULE CELLS

value: 6 weeks