title: Pronuclei_Pat_PN5_2_Dnase-seq

status: Public on Jun 03 2016

city: Brookline

first: Yuting

last: Liu

address: 82 Walnut St. Unit 3

department: Department of Genetics

institute: Harvard Medical School

data_processing:
Dnase-seq data analysis: Reads of DNase-seq were mapped to the mouse genome (mm9) using Bowtie v0.12.9. We use ‘-m 1’ parameter for unique hits mapping. The reads with mapping quality (MAPQ) ≤ 10 or redundant reads that mapped to the same location with the same orientation were removed from further analysis in each library. The DHS peaks in liDNase-seq data were identified by Hotspot program with FDR <= 0.01 (John et al., 2011). The tag density at each DHS was calculated by normalizing the number of reads in the DHS to the total number of reads in the library (possibility of a tag located on a base- pair per million reads). For identifying DHS peaks in preimplantation embryos, we used an additional stringent cutoff requirement of RPM > 1 for the peak in both replicates.
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited text files include DHS peaks for each Sample

source_name: Paternal pronuclei from PN5 (12 hrs after fertlization) zygote

growth_protocol: All animal studies were performed in accordance with guidelines of the Institutional Animal Care and Use Committee at Harvard Medical School. MII-stage oocytes were collected from 8 week-old superovulated BDF1 females by injecting 7.5 I.U. of PMSG (Millipore) and hCG (Millipore). For in vitro fertilization (IVF), MII oocytes were transferred into HTF medium supplemented with 10 mg/ml bovine serum albumin (BSA; Sigma-Aldrich) and inseminated with activated spermatozoa obtained from the caudal epididymides of adult JF1 male mice. Spermatozoa capacitation was attained by 1 h incubation in HTF medium. Six hours post-insemination (hpi), fertilized oocytes were transferred and cultured in KSOM in a humidified atmosphere of 5% CO2/95% air at 37.8°C. The 1-cell, 2-cell, 4-cell, 8-cell, and morula stage embryos were harvested at 12, 30, 46, 60, 78 hpi, respectively. The numbers of the embryos collected for DNase-seq are around 100, 100, 50, 25, and 10, respectively. For the 1-cell stage zygotes, BDF1 sperm was used for IVF.

extract_protocol:
Just before sampling the embryos for DNase-seq, we removed the polar bodies to avoid genomic contamination from polar body. First, the embryos were briefly exposed to the Acid Tyrode's solution (Sigma-Aldrich) supplemented with 0.5% polyvinylpyrrolidone (PVP, Irvine Scientific, 90123) and 50 mM NaCl to gently remove zona pellucida. After washing with M2 media (Millipore), the embryos were incubated with Trypsin/EDTA (Life technologies) supplemented with 0.5% PVP and 50 mM NaCl for 2-3 min on multitest glass slide (MP Biomedicals, 6040805). The embryos were then washed with M2 media followed by washing with PBS with 0.2% BSA and transferred into an Eppendorf LoBind 1.5 ml tube (Eppendorf) on ice.
Sequencing library was prepared using NEBNext Ultra DNA Library Prep Kit for Illumina (New England Biolabs) with the exception that PCR amplification was done using Kapa hifi hotstart readymix (Kapa Biosystems) for 8-cycles. The PCR product was purified with 1.3 volume SPRIselect beads (Beckman Coulter) and then size selected with 0.7 volume plus 0.7 volume SPRIselect beads. Then the sample was amplified again with Kapa hifi hotstart readymix for 6 to 11 cycles. Repeat the previous purification and size selection step. The libraries were sequenced on a Hiseq2500 with single-end 100 bp reads (Illumina).

treatment_protocol: Fresh cells were collected and resuspended in 36 μl lysis buffer (10mM Tris-HCl, pH 7.5, 10mM NaCl, 3mM MgCl2, 0.1% Triton X-100). Put on ice for 5 to 30 minutes. Add 4 μl 1:12.5 (for 1-cell embryos) or 1:25 (for all the other samples in this study) diluted DNase I (10 U/μl, Roche). Incubate at 37°C for 5 minutes. The reaction was stopped by adding 80 μl Stop Buffer (10 mM Tris-HCl, pH 7.5, 10 mM NaCl, 0.15% SDS, 10 mM EDTA) containing 2 μl Proteinase K (20 mg/ml, Life technologies). Then 20 ng circular carrier DNA (pure plasmid DNA without mammalian sequence) was added. The mixture was incubated at 50 °C for 1 hour. DNA was purified by extraction with phenol-chloroform and precipitated by ethanol in the presence of linear acrylamide (Life technologies) overnight. Precipitated DNA was resuspended in 55.5 μl TE (2.5 mM Tris, pH 7.6, 0.05 mM EDTA).

value: PN5 (12 hrs after fertilization) zygote

tag: cell/embruos number

value: 170 pronuclei


title: Pronuclei_Mat_PN5_2_Dnase-seq

type: SRA

state: MA

country: USA

library_source: genomic

molecule: genomic DNA

organism: Mus musculus

source_name: Maternal pronuclei from PN5 (12 hrs after fertlization) zygote

tag: strain

tag: developmental stage


title: Pronuclei_Pat_PN5_1_Dnase-seq


title: A3 Zfp57-/- BS-seq

status: Public on Jun 04 2016

city: Naples

first: Marco

last: Cammisa

state: Naples

address: Via Castellino Pietro, 111

country: Italy

institute: Institute of Genetics and Biophysics "A. Buzzati Traverso" (IGB-ABT)

data_processing:
Basecalls and adaptor trimming was performed using Illumina Pipeline Casava (v1.8.2).
Data was filtered for low quality reads Trim Galore (v0.2.8).
Reads were aligned to the NCBI37/mm9 genome assembly using Bismark (v0.7.9).
CpG methylation calls were extracted using the Bismark methylation extractor (v0.7.9).
Genome_build: NCBI37/mm9
Supplementary_files_format_and_content: Bedgraph file report methylation level of symmetric CpGs at least 5x covered.

growth_protocol: Embryonic stem cells were cultured under standard feeder-free conditions on gelatinized tissue culture dishes with media containing DMEM (EuroClone ECM0101L) supplemented with 100 µM 2-mercaptoethanol (Sigma), 1 x non-essential amino acids (only for A3), 1 mM sodium pyruvate, 2 mM L-glutamine, 1 x penicillin-streptomycin, 10% (15% for E14) foetal calf serum (HyClone) and 103 U/ml  leukemia inhibitory factor (LIF, Millipore) at 37 °C under an atmosphere of 5% CO2.

extract_protocol: For library preparation 100-200 ng of genomic DNA was used with the NuGEN Ovation Ultralow Methyl-Seq Library System.

tag: cell type

value: Embryonic stem cells

tag: genotype/variation


title: Pronuclei_Mat_PN5_1_Dnase-seq


title: Pronuclei_Pat_PN3_2_Dnase-seq

source_name: Paternal pronuclei from PN3 (7.5 hrs after fertlization) zygote

value: PN3 (7.5 hrs after fertilization) zygote

value: 140 pronuclei


title: Pronuclei_Pat_PN3_1_Dnase-seq

value: 150 pronuclei


title: Pronuclei_Mat_PN3_2_Dnase-seq

source_name: Maternal pronuclei from PN3 (7.5 hrs after fertlization) zygote


title: Pronuclei_Mat_PN3_1_Dnase-seq


title: Sperm_2_Dnase-seq

source_name: Sperm

value: 5000 cells


title: Sperm_1_Dnase-seq


title: GV Oocyte_14d_2_Dnase-seq

source_name: GV Oocyte from 14 day-old females

value: 14 day-old

value: 150 cells


title: WT.Epid.NoUV_2

city: Lausanne

first: Mark

last: Ibberson

address: Genopode building

department: Vital-IT

country: Switzerland

institute: SIB Swiss Institute of Bioinformatics

library_source: transcriptomic

data_processing:
Base calling was done with Illumina GAP Pipeline Software v1.82
Sequence reads were processed to remove the adaptor sequences and reformatted to FASTA files using the FASTX-Toolkit
Sequences were aligned to mouse mature microRNA sequences (from miRBase Version 19) and non-coding RNA sequences (Rfam Version 11) using MEGABLAST with a word size of 8 nucleotides. The criteria for counting a sequence match were if the % query was >=90% of the target sequence and if there were <= 2 mismatches over the alignment. The % query was calculated as (a/q) x p where a= alignment length, q= query length and p= percent identity over aligned region. The matches against miRBase were parsed and the top matches (based on % query) were selected. If a sequence had more than one top match against different database sequences, it was excluded from the subsequent analysis. Matches to Rfam were only taken into account for sequences not matching miRBase.
Supplementary_files_format_and_content: Raw count data for microRNAs were normalized to the relative size of each library using R/Bioconductor package DESeq, estimateSizeFactors function. Count data are provided in tab-delimited format

molecule: total RNA

source_name: Skin, Epidermis_WT_NoUV

extract_protocol:
Total RNA was isolated using Trizol (Invitrogen). RNA quantities and quality were assessed using a NanoDrop ND-1000 spectrophotometer or an Agilent 2100 BioanalyzerRNA quantities and quality were assessed using a NanoDrop ND-1000 spectrophotometer or an Agilent 2100 Bioanalyzer.
Small RNA libraries were prepared using 1 μg of total RNA according to the TruSeq Small RNA Sample Preparation Guide (Illumina, San Diego, CA). Libraries were sequenced HiSeq 2500 using v3 chemistry.

tag: tissue

value: skin

tag: tissue compartment

value: epidermis

tag: genotype

value: wildtype


title: WT.Derm.UV_3

source_name: Skin, Dermis_WT_UV

value: dermis


title: WT.Epid.UV_4

source_name: Skin, Epidermis_WT_UV


title: WT.Epid.UV_2


title: WT.Epid.NoUV_1


title: WT.Derm.UV_1


title: WT.Derm.UV_2


title: KO.Epid.UV_2

source_name: Skin, Epidermis_KO_UV

value: PPARB KO


title: KO.Epid.NoUV_4

source_name: Skin, Epidermis_KO_noUV


title: KO.Epid.UV_1


title: WT.Epid.NoUV_3


title: KO.Epid.UV_4


title: WT.Epid.UV_1


title: WT.Epid.UV_3


title: WT.Derm.NoUV_3

source_name: Skin, Dermis_WT_noUV


title: WT.Derm.NoUV_1


title: KO.Epid.NoUV_2


title: KO.Epid.UV_3


title: KO.Epid.NoUV_3


title: KO.Epid.NoUV_1


title: GV Oocyte_14d_1_Dnase-seq


title: WT.Derm.NoUV_2


title: KO.Derm.UV_1

source_name: Skin, Dermis_KO_UV


title: KO.Derm.UV_2


title: KO.Derm.UV_3


title: KO.Derm.NoUV_2

source_name: Skin, Dermis_KO_noUV


title: KO.Derm.NoUV_3


title: KO.Derm.NoUV_1


title: RNA-seq 2-Cell Nfya-KD_1

data_processing:
Dnase-seq data analysis: Reads of DNase-seq were mapped to the mouse genome (mm9) using Bowtie v0.12.9. We use ‘-m 1’ parameter for unique hits mapping. The reads with mapping quality (MAPQ) ≤ 10 or redundant reads that mapped to the same location with the same orientation were removed from further analysis in each library. The DHS peaks in liDNase-seq data were identified by Hotspot program with FDR <= 0.01 (John et al., 2011). The tag density at each DHS was calculated by normalizing the number of reads in the DHS to the total number of reads in the library (possibility of a tag located on a base- pair per million reads). For identifying DHS peaks in preimplantation embryos, we used an additional stringent cutoff requirement of RPM > 1 for the peak in both replicates.
RNA-seq data analysis: Reads of RNA-seq were mapped to the mouse genome (mm9) with TopHat v2.0.6. All programs were run with default parameters unless otherwise specified. Uniquely mapped reads were subsequently assembled into transcripts guided by the reference annotation (UCSC gene models) with Cufflinks v2.2.1. Expression level of each gene was quantified with normalized FPKM (fragments per kilobase of exon per million mapped fragments). The 2-cell activated genes were identified by comparing the 2-cell control RNA-seq with the 1-cell RNA-seq data using five-fold change as a cut-off.
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited text files include DHS peaks for each Sample
Supplementary_files_format_and_content: RNA abundance

source_name: 2-cell IVF embryos siNfya-1 (30 hrs after fertilization)

value: 10 embryos

value: BDF1 oocyte x BDF1 sperm


title: RNA-seq 2-Cell Nfya-KD_2

source_name: 2-cell IVF embryos siNfya-2 (30 hrs after fertilization)

value: 13 embryos


title: RNA-seq 2-Cell siControl_2

source_name: 2-cell IVF embryos siControl (30 hrs after fertilization)

value: 16 embryos


title: 8-Cell siControl_1_Dnase-seq

description: Decreased 8-Cell-gained DHS upon Oct4 knockdown.xlsx

source_name: 8-cell IVF embryos siControl (60 hrs after fertilization)

value: 25 embryos


title: 8-Cell Oct4-KD_2_Dnase-seq

source_name: 8-cell IVF embryos siOct4-2 (60 hrs after fertilization)


title: RNA-seq 1-Cell

source_name: 1-cell IVF embryos (12 hrs after fertilization)

value: 20 embryos


title: 2-Cell Nfya-KD_2_Dnase-seq

description: Decreased 2-Cell DHS upon Nfya knockdown.xlsx

value: 90 embryos


title: Morula_2_Dnase-seq

description: All DHSs for each stages.xlsx

source_name: Morula IVF embryos (78 hrs after fertilization)

value: BDF1 oocyte x JF1 sperm


title: 2-Cell siControl_1_Dnase-seq

value: 70 embryos


title: 8-Cell Oct4-KD_1_Dnase-seq

source_name: 8-cell IVF embryos siOct4-1 (60 hrs after fertilization)


title: 2-Cell siControl_2_Dnase-seq


title: 2-Cell Nfya-KD_1_Dnase-seq


title: 8Cell_2_Dnase-seq

description: All DHSs for each stages.xlsx

source_name: 8-cell IVF embryos (60 hrs after fertilization)


title: 4Cell_2_Dnase-seq

description: All DHSs for each stages.xlsx

source_name: 4-cell IVF embryos (46 hrs after fertilization)

value: 50 embryos


title: Morula_1_Dnase-seq

description: All DHSs for each stages.xlsx


title: 100-cell ESC_Dnase-seq

description: All DHSs for each stages.xlsx

source_name: Mouse ESCs

value: 100 cells


title: 8Cell_1_Dnase-seq

description: All DHSs for each stages.xlsx


title: 2Cell_2_Dnase-seq

description: All DHSs for each stages.xlsx

source_name: 2-cell IVF embryos (30 hrs after fertilization)

value: 100 embryos


title: 4Cell_1_Dnase-seq

description: All DHSs for each stages.xlsx


title: 2Cell_1_Dnase-seq

description: All DHSs for each stages.xlsx


title: 1Cell_2_Dnase-seq

description: All DHSs for each stages.xlsx


title: CFCF-WT

city: Seoul

first: Tae-Young

last: Roh

state: --- Select One ---

address: 52 Ewhayeodae-gil, Seodaemun-gu

department: Life Sciences

country: South Korea

institute: Ewha Womans University

description: anti-CTCF

data_processing:
Illumina Casava1.6 software used for basecalling.
Sequenced reads were mapped to mm9 genome using CASAVA 1.6
The sequence reads were normalized as reads per million per nucleotide (RPM).
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited text files with BED graph format

source_name: MEF Cell Line

growth_protocol: Cells were maintained in Dulbecco’s Modified Eagles Medium (DMEM, WelGENE Inc., Daegu, Korea) supplemented with 10% FBS (Fetal bovine serum; WelGENE Inc.) and 100 µg/ml penicillin-streptomycin (WelGENE Inc.), at 37°C, 5% CO2.

extract_protocol:
Cells harvested from 100 mm dishes were cross-linked with 1% formaldehyde for 15 minutes at room temperature and cross-linking was quenched with 125 mM glycine. After lysing cells in buffer containing detergents (1% Triton X-100, 0.5% sodium deoxycholate, 1% SDS, 10 mM EDTA, 50 mM Tris-Cl [pH 8.0]), the chromatin was sonicated to an average length of 300 bp by 30% pulses at a 10 seconds interval, repeated 30 times, and then  immunoprecipitated overnight using an anti-CTCF antibody (CTCF (D31H2) XP Rabbit mAb #3418; Cell Signaling Technology).
Sequencing libraries were manually generated for high-throughput sequencing. Either end of DNA strands was filled using the Epicentre DNA END-Repair kit (Epicentre Biotechnologies, Madison, WI). Adenine was added at 3’ ends by  Taq DNA polymerase (New England Biolabs, Beverly, MA) under dATP conditions. Illumina’s adaptor oligomers was ligated at both ends using Quick Ligation kit (Qiagen, Germantown, MD). After purified twice, DNA libraries are amplified using Solexa primers (Illumina, San Diego, CA) and Phusion PCR master mix (Thermo Scientific, Hudson, NH).

value: MEF Cell Line

tag: background strain


title: CFCF-KO

description: anti-CTCF


title: 1Cell_1_Dnase-seq

description: All DHSs for each stages.xlsx


title: E14_serum_To_2i_8h_H2A_TAB_seq

city: Nijmegen

first: Hendrik

middle: G

last: Stunnenberg

address: Geert Grooteplein 28

department: Molecular Biology

country: Netherlands

institute: Radboud University

data_processing:
FASTQ sequence files generate for both BS-seq and TAB-seq were aligned to Mus musculus NCBIM37 genome assembly (mm9) including lambda and pUC19 sequences using rmapbs-pe (Smith et al., 2009). 5mC and 5hmC levels were directly determined using MethPipe (Song et al., 2013). For TAB-seq, hmC protection/non-protection and mC non-conversion rates were quantified and estimated levels were corrected as described in details previously (Lister et al., 2013; Yu et al., 2012). A maximum likelihood method (Qu et al., 2013) on the combined data from BS-seq and TAB-seq were used to estimate absolute levels for 5mC and 5hmC.
Genome_build: mm9
Supplementary_files_format_and_content: bedGraph including 5hmC level

source_name: Embryonic Stem cells

growth_protocol: Mouse embryonic stem were cultured without feeders in the presence of  LIF in DMEM containing 15% fetal calf serum. For serum-to-2i transition, cells were washed two times with PBS and serum medium was replaced by serum-free NDiff 227 (StemCells, Inc.) supplemented with LIF, MEK inhibitor PD0325901 (1 mM) and GSK3 inhibitor CHIR99021 (3 mM), together known as 2i (Ying et al., 2008). For serum-to-2i switching in the presence of vitamin C (L-ascorbic acid, Sigma), 17 μM of vitamin C (dissolved in water) was added to 2i. The media were refreshed every day and cells were passaged every 2–3 days.

extract_protocol:
DNA was isolated using the Cell Culture DNA Midi Kit from QIAGEN (Cat No 13343) or GenElute™ Mammalian Genomic DNA Miniprep Kit (Cat No G1N350, Sigma).
TAB-seq was performed as described previously (Yu et al., 2012).

value: ES-cells

tag: measurement

value: DNA hydroxymethylation

tag: time point

tag: treatment


title: E14_serum_H9B_TAB_seq

value: control (without treatment)


title: E14_serum_To_2i_VitC_8h_H6A_TAB_seq


title: RNA-seq 2-Cell siControl_1


title: E14_serum_To_2i_VitC_32h_H8B_TAB_seq


title: E14_serum_To_2i_4h_H1A_TAB_seq


title: E14_serum_To_2i_VitC_4h_H5A_TAB_seq


title: 72H_M2

city: Chapel Hill

first: Bailey

last: Peck

state: NC

address: 120 Mason Farm Rd, 5100H Genetic Medicine Bldg

department: Genetics

institute: University of North Carolina at Chapel Hill

data_processing:
Following sequencing and demultiplexing by the UNC HTSF using bcl2fastq 1.8.4
Reads were aligned to the hg19 genome using Mapsplice by the UNC Bioinformatics Core Facility
Transcripts quantified using RSEM by the UNC Bioinformatics Core Facility
Differential gene expression was evaluated using edgeR. Genes with low expression (CPM < 10 in more than half the samples) were filtered out of our analysis. Gene counts were then TMM normalized and evaluated for differential gene expression using GLM.
Genome_build: hg19
Supplementary_files_format_and_content: Full counts table from RSEM (TXT); normalized and filtered gene counts in counts per million (txt)

organism: Homo sapiens

source_name: Human Intestinal Epithelial Cells (HIECs)

growth_protocol: HIECs were acquired from the Beaulieu laboratory and were cultured in OptiMEM 1 (Life Technologies) supplemented with 10% FBS (Life Technologies), 0.01 M HEPES (Life Technologies), and 5 ng/mL hEGF (Invitrogen, Grand Island, NY).

extract_protocol:
HIECs were trypsinized and spun down, then lysed and RNA was extracted using Norgen Total RNA isolation kits as per the manufacturer's instructions. RNA quality was assessed using Agilent RNA Nano 6000 kit (Agilent Technologies, Inc, Santa Clara, CA) run on a Bioanalyzer 2100 (Agilent). All samples had high integrity with RINs above 9.2 (with an average of 9.7).
Samples were submitted to the UNC High Throughput Sequencing Facility (HTSF) for TruSeq Stranded Total RNA library preparation (Illumina, San Diego, CA) and paired-end 50 bp sequencing on a HiSeq 2000 (Illumina) multiplexing 6 samples per lane.

treatment_protocol: HIECs at passage 25 were seeded onto tissue-culture treated plates and transfected at 70% confluency with 3.25 uL/mL Lipofectamine 2000 (Life Technologies). Each replicate represents a single 10cm plate that was either Mock transfected or transfected with 100 nM LNA30bcd

value: Mock

tag: replicate


title: E14_serum_To_2i_VitC_32h_RNAseq

data_processing:
Fastq files were aligned to mouse genome assembly NCBIM37 (mm9) using GSNAP version 2012-07-20 with following setting: -B 4 -N 1 -n 1 -m 1.
Genome_build: mm9
Supplementary_files_format_and_content: raw counts of sequencing reads against Mus_musculus.NCBIM37.67 gtf file using the htseq-count python library

extract_protocol:
Total RNA for each sample was extracted using RNeasy mini kit (QIAGEN) following the manufacture’s instructions.
5ug of extracted RNA was depleted from ribosomal RNA using Ribo-Zero Gold Kit (Epicentre Madison, Winsconsin, USA). Then, rRNA-depleted RNA was used for library preparation using the TruSeq RNA Sample Prep Kit (Illumina) following the manufacturer’s instructions. Libraries were indexed using NEXTflex adapters (Bioo-Scientific Corporation, Austin, TX, USA) and 50 bp single-end sequencing was performed on Illumina HiSeq 2000 instruments using TruSeq reagents (Illumina, San Diego, CA, USA), according to manufacturer’s instructions.

value: RNA-seq


title: E14_serum_To_2i_VitC_24h_RNAseq


title: 72H_L3

value: 100 nM LNA30bcd


title: E14_serum_To_2i_VitC_4h_RNAseq


title: 72H_L2


title: E14_serum_To_2i_32h_RNAseq


title: 8-Cell siControl_2_Dnase-seq

value: BDF1 oocyte x BDF1 sperm


title: 72H_L1


title: E14_serum_To_2i_VitC_16h_RNAseq


title: 48H_M2


title: E14_serum_To_2i_24h_RNAseq


title: 48H_M1


title: 48H_L3


title: 48H_L2


title: E14_serum_To_2i_32h_H4A_TAB_seq


title: 72H_M3


title: E14_serum_To_2i_16h_RNAseq


title: 48H_L1


title: E14_serum_To_2i_4h_RNAseq


title: 24H_M3


title: E14_serum_To_2i_8h_H2A_WGBS

data_processing:
FASTQ sequence files generate for both BS-seq and TAB-seq were aligned to Mus musculus NCBIM37 genome assembly (mm9) including lambda and pUC19 sequences using rmapbs-pe (Smith et al., 2009). 5mC and 5hmC levels were directly determined using MethPipe (Song et al., 2013). For TAB-seq, hmC protection/non-protection and mC non-conversion rates were quantified and estimated levels were corrected as described in details previously (Lister et al., 2013; Yu et al., 2012). A maximum likelihood method (Qu et al., 2013) on the combined data from BS-seq and TAB-seq were used to estimate absolute levels for 5mC and 5hmC.
Genome_build: mm9
Supplementary_files_format_and_content: bedGraph including 5mC level

extract_protocol:
DNA was isolated using the Cell Culture DNA Midi Kit from QIAGEN (Cat No 13343) or GenElute™ Mammalian Genomic DNA Miniprep Kit (Cat No G1N350, Sigma).
BS-seq was performed as described previously (Lister et al., 2008).

value: DNA methylation


title: 24H_M2


title: 24H_M1


title: 72H_M1


title: 24H_L2


title: E14_serum_To_2i_4h_H1A_WGBS


title: E14_serum_To_2i_32h_H4A_WGBS


title: 48H_M3


title: 24H_L1


title: bCtr.8

city: London

first: Kathy

last: Niakan

address: Mill Hill

country: United Kingdom

institute: The Francis Crick Institute

data_processing:
Paired-end reads were aligned to the human genome (hg19/GRCh37 ) using TopHat2 (Kim et al., 2013).
BAM files were sorted and converted into SAM files using SAMtools (Li et al., 2009).
The python package HTSeq (Anders et al., 2015) was then used to calculate read counts for each ensemble gene model (release 74), and these values were normalized by both gene length and sequencing depth to generate RPKM expression values.
Genome_build: GRCh37.74
Supplementary_files_format_and_content: RPKM table

molecule: polyA RNA

source_name: Human preimplantation embryo

extract_protocol:
cDNA was generated from single cells using the SMARTer Ultra Low Input RNA kit for Illumina Sequencing–HV (Clontech Laboratories, Inc.) according to maunfacturers’ guidelines. Single cells were picked using 100 micromteer inner diameter Stripper pipette (Origio) and transferred to individual low bind RNAse-free tube containing 0.25 μl RNase inhibitor, 4.75 μl Dilution buffer and 5 microliter nuclease-free water on a -80°C pre-chilled CoolRack (Biocision, CA). Samples were stored at -80°C until ready to be processed. 1 microliter of 3’ SMART CDS Primer II A was added to the sample, mixed well and incubated at 72°C for 3 min. First strand cDNA was synthesised by adding 4microliter 5X First-Strand Buffer, 0.5 microliter 100 mM DTT, 1 microliter 20 mM dNTP mix, 1 microliter  SMARTer IIA Oligonucleotide, 0.5 microliter RNase Inhibitor and 2 microliter SMARTScribe Reverse Transcriptase (100 U/microliter) directly to a tube containing the sample and incubating at 42°C for 2 hours followed by 10 min at 70°C.
First strand cDNA was purified by adding 36 microliter of room temperature SPRI Ampure XP beads (Beckman Coulter Genomics), mixing well and incubating at room temperature for 8 min. Tubes were placed on a MagnaBot II Magnetic Separation device (Promega) and allowed to stand until all beads were immobilised into a pellet. The supernatant was removed and discarded. Tubes were briefly spun and any residual liquid was removed.
Double stranded cDNA was amplified from the template bound to the beads using Advantage 2 PCR kit (Clontech Laboratories, Inc.). 5 microlite r10X Advantage 2 PCR Buffer, 2 microliter 10 mM dNTP Mix, 2 microliter IS PCR Primer, 2 microliter 50X Advantage 2 Polymerase Mix and 39 microliter Nuclease-Free water were added to the tube containing the sample to give a total volume of 50 microliter. PCR amplification was performed at 95°C for 1 min, followed by 18 cycles of 15 sec at 95°C, 30 sec at 65°C and 6 min at 68°C followed by final extension step of 10 min at 72°C. Amplified cDNA was purified by adding 90 microliter SPRI Ampure XP beads, mixing well and incubating at room temperature for 8 min to allow amplified cDNA bind to the beads. Sample tubes were placed on the magnet and allowed to stand until all beads had been immobilised. Supernatant was removed and discarded and beads were washed twice by adding 200 microliterl freshly prepared 80% ethanol and leaving for 30 sec before discarding the supernatant. Tubes were spun briefly to collect residual liquid. The bead pellet was allowed to air dry. 12 microliter of purification buffer was added to rehydrate the pellet and incubated for 2 min at room temperature. cDNA was eluted by pipetting up and down 10 times before returning the tube to the magnet. The clear supernatant containing the cDNA was removed from the immobilised beads and transferred to a new low-bind tube. cDNA was stored at -80°C until library preparation.
cDNA quality was assessed by High Sensitivity DNA assay on an Agilent 2100 Bioanalyser with good quality cDNA showing a broad peak from 300 to 9000 bp. cDNA concentration was measured using QuBit dsDNA HS kit (Life Technologies UK Ltd.) Typical yields from a single cell ranged from 1 ng to 9 ng.
In preparation for library generation cDNA was sheared using Covaris S2 to achieve cDNA in 200-500 bp range. 10 μl of cDNA sample and 65 microliter purification buffer was added to Covaris AFA Fiber Pre-Slit Snap Cap microTUBE. cDNA was sheared using the settings 10% Duty, Intensity 5, Burst Cycle 200 for 2 min. Sheared cDNA was transferred to a new 0.2 ml low-bind tube.Libraries were prepared using Low Input Library Prep Kit (Clontech Laboratories, Inc.) according to manufacturer’s instructions. The amount of input cDNA was calculated from the concentration measured by the Bioanalyser assay prior to shearing, taking into account the dilution involved in the shearing step. The appropriate amplification cycle number was selected according to manufacturer’s guidelines. Library quality was assessed by Bioanalyser and the concentration was measured by QuBit assay.  The molar concentration of library was calculated thus:
Library molecular weight = average size in bp (from Bioanalyser) x 650 g/mol per bp
Molar concentration = library concentration from QuBit/library molecular weight
Libraries with a molar concentration greater than 2nM were submitted for 50-bp paired-end sequencing on Illumina HiSeq 2000.

treatment_protocol: Single cell isolation protocol: Single cells were isolated from blastocyst stage embryos (6-7 days post fertilisation) for subsequent analysis by micromanipulation, with a duration of less than 20 minutes. Embryos were placed in drops of G-MOPS solution (Vitrolife) on a petri dish overlaid with mineral oil. The plate was placed on a microscope stage (Olympus IX70) and the embryos were held with an opposing holding pipette and 30 micrometer inner diameter blastomere biopsy pipette (Research Instruments) using Narishige micromanipulators (Narishige, Japan). The biopsy mode of a Saturn 5 laser (Research Instruments) was used to separate the majority of the mural TE from the ICM and polar TE. The ICM and polar TE were washed quickly in PBS without Ca2+ and Mg2+ (Invitrogen) then placed in 0.05% trypsin/EDTA (Invitrogen) for 5 minutes at room temperature. Trypsin was quenched using Global Media supplemented with 5 mg/mL LifeGlobal Protein Supplement. After quenching, the cell clump was placed back on the stage in a drop of G-MOPS solution and pipetted up and down several times using the blastomere biopsy pipette.

value: Blastocyst single cell

tag: sample type

value: Control embryo b


title: E14_serum_To_2i_VitC_8h_H6A_WGBS


title: bCtr.10


title: 24H_L3


title: E14_serum_To_2i_VitC_32h_H8B_WGBS


title: bCtr.6


title: bCtr.9


title: bCtr.7


title: E14_serum_To_2i_VitC_4h_H5B_WGBS


title: bCtr.5


title: H2217-WT-XYL.5-12-3-12.16.09-239-8-18

city: Saint Louis

first: Ezekiel

middle: John

last: Maier

state: MO

address: 4444 Forest Park Ave

department: Computer Science

institute: Washington University

description: wildtype in 5% Xylose for 12 hours

data_processing:
Sequenced reads were aligned to the S. cerevisiae S288C reference sequence genome release R63 (Engel et al. 2013) using TopHat version 2.0.4 (Trapnell et al. 2009) and Bowtie version 0.12.8 (Langmead et al. 2009).
Reads that aligned uniquely to the reference sequence were considered for gene expression quantification with Cufflinks version 2.0.2 (Trapnell et al. 2010). Gene expression was quantitated against known gene boundaries from the gene annotations provided by the SGD S288C reference genome release R63. Gene expression was normalized using the Cufflinks upper-quartile normalization option.
After gene expression quantification, samples not passing the following quality control filters were removed from consideration. Each sample was required to be sequenced at a depth of at least 750,000 reads. The expression of the deleted gene in mutant expression profiles was required to be less than 10% of its expression in WT. The mean and median expression of the deleted genes were 1.3% and 0% of the WT levels respectively. The low levels of expression that remained for some samples were likely the result of errors in the sequencing process itself, either due to contamination between samples or a low level of error in matching barcodes for multiplexed samples to reads. In addition, within each replicate set containing 3 biological replicates outlier gene expression profiles were detected and removed if the outlier caused the median of all genes’ coefficients of variation (CoV -- the standard deviation divided by the mean) to be greater than 0.5 for the replicate set or if the outlier caused the Pearson Correlation Coefficient between replicate gene expression profiles to be less than 0.8.
Counts of reads mapping to each gene were quantitated using the htseq-count tool in version 0.5.3 of the HTSeq Python package (Anders et al. 2014).
Batch effects were removed from RNA-seq read counts using Remove Unwanted Variation (RUV) (Risso et al. 2014).  Unwanted variation due to technical batch effects was identified and removed, using RUVg, through factor analysis on a set of negative control genes that were not expected to respond to the biological treatments. The set of 192 negative control genes used to identify batch effects are not predicted to be regulated by any of the deleted regulators in the top 100,000 network predicted interactions, and their expression did not change significantly (log2 fold change < 1.3 and adjusted p-value > 0.1) between any pair of expression profiled single carbon growth conditions. Dropping the first factor of unwanted variation, by using RUVg with the parameter k set to 1, was sufficient to remove the majority of the batch effect.
Differential expression analysis comparing expression profiles sets was performed by using LIMMA(Smyth 2004) with the voom transformation applied to read counts (Law et al. 2014).
Genome_build: S. cerevisiae S288C reference sequence genome release R63 (Engel et al. 2013)
Supplementary_files_format_and_content: cuff: Gene expression was quantified by processing tophat produced bam alignments with Cufflinks v2.0.2 with S. cerevisiae S288c reference sequence genome release R63 (Engel et al. 2013). Expression was normalized by applying the Cufflinks supplied upper-quartile normalization routine. counts: Counts of reads mapping to each gene were quantitated using the htseq-count tool in version 0.5.3 of the HTSeq Python package (Anders et al. 2014).The expression.mtr file is a matrix of fully processed gene expression values (upper-quartile normalized Cufflinks reported FPKM values). The matrix is of size: number of S. cerevisiae genes (rows) BY number of expression profiles (columns). Each column in the expression.mtr file lists gene FPKM values from the matching .cuff file.

organism: Saccharomyces cerevisiae

source_name: S. cerevisiae cells grown in a single carbon source

growth_protocol: All strains were constructed in the S. cerevisiae H2217-7 background. Cells were cultured overnight in synthetic complete medium with 2% glucose to a density of 1-2 OD600.

extract_protocol:
Total mRNA was isolated using the yeast RiboPure kit (Life Technologies, Carlsbad CA).
Libraries for RNA-Seq were prepared as in (Haynes, et al., 2011) except that the selection of poly(A) RNA from the total RNA isolated was accomplished using an mRNA Catcher Plus Kit (Life Technologies) and the resulting poly(A) RNA was subsequently sheared by incubating in TURBO DNA-free buffer at 75°C for 10 minutes and purified with a QIAquick PCR Purification Kit (Qiagen).

treatment_protocol: All S. cerevisiae strains were grown aerobically in 60 mL of synthetic complete at 30°C in 250 mL baffled Erlenmeyer flasks shaken at 225 RPM. Cultures of cells for evaluating the impact of transcriptional interventions were performed in triplicate and initiated by inoculating overnight cultures into 60 mL of synthetic complete media supplemented with 20 g/L glucose and 50 g/L xylose resulting in a density of OD600=1.0+/- 0.2. Samples taken for RNA-seq were aliquoted from the primary culture, spun down at 3000 x g and frozen in liquid nitrogen. Cultures for identification of differential expression associated with carbon sources were grown in triplicate in either 50 g/L glucose, 50 g/L xylose or 1.3 g/L ethanol for 8 hours prior to collection of biomass for RNA sequencing.

tag: condition description

value: single carbon source for 12 hours


title: H2217-WT-XYL.5-12-2-12.16.09-239-5-17

description: wildtype in 5% Xylose for 12 hours


title: bCtr.4


title: H2217-WT-GLU.5-12-3-12.16.09-239-7-15

description: wildtype in 5% Glucose for 12 hours


title: H2217-WT-GLU.5-12-2-12.16.09-239-6-14

description: wildtype in 5% Glucose for 12 hours


title: H2217-WT-XYL.5-12-1-12.16.09-222-1-16

description: wildtype in 5% Xylose for 12 hours


title: bCtr.3


title: H2217-WT-GLU.5-12-1-12.16.09-222-1-13

description: wildtype in 5% Glucose for 12 hours


title: H2217-WT-GLU.0p2-12-2-12.16.09-239-5-11

description: wildtype in 0.2% Glucose for 12 hours


title: aCtr.5

value: Control embryo a


title: H2217-WT-GLU.0p2-12-3-12.16.09-239-8-12

description: wildtype in 0.2% Glucose for 12 hours


title: H2217-WT-GLY.5-12-3-12.16.09-239-7-9

description: wildtype in 5% Glycerol for 12 hours


title: aCtr.7


title: H2217-WT-GLY.5-12-2-12.16.09-239-6-8

description: wildtype in 5% Glycerol for 12 hours


title: aCtr.3


title: H2217-WT-GAL.5-12-3-12.16.09-239-8-6

description: wildtype in 5% Galactose for 12 hours


title: aCtr.4


title: bCtr.1


title: aCtr.1


title: H2217-WT-GAL.5-12-2-12.16.09-239-6-5

description: wildtype in 5% Galactose for 12 hours


title: H2217-WT-GLY.5-12-1-12.16.09-239-5-7

description: wildtype in 5% Glycerol for 12 hours


title: H2217-WT-GAL.5-12-1-12.16.09-222-1-4

description: wildtype in 5% Galactose for 12 hours


title: bCtr.2


title: H2217-WT-ETH.1p3-12-1-12.16.09-239-5-1

description: wildtype in 1.3% Ethanol for 12 hours


title: eCtr.4

value: Control embryo e


title: aCtr.6


title: H2217-WT-ETH.1p3-12-2-12.16.09-239-6-2

description: wildtype in 1.3% Ethanol for 12 hours


title: aCtr.2


title: bCtr.11


title: H2217-WT-ETH.1p3-12-3-12.16.09-239-7-3

description: wildtype in 1.3% Ethanol for 12 hours


title: 27PNT.4

value: Heterologous pronuclear transfer embryo 27


title: 27PNT.2


title: 27PNT.1


title: 28PNT.2

value: Heterologous pronuclear transfer embryo 28


title: 28PNT.7


title: 27PNT.10


title: hCtr.2

value: Control embryo h


title: hCtr.3


title: hCtr.1


title: 20PNT.8

value: Heterologous pronuclear transfer embryo 20


title: dCtr.3

value: Control embryo d


title: eCtr.2


title: 27PNT.7


title: cCtr.1

value: Control embryo c


title: cCtr.2


title: fCtr.2

value: Control embryo f


title: 20PNT.6


title: eCtr.1


title: S2P ChIP rep1

city: Göttingen

first: Björn

last: Schwalb

address: Fassberg 11

country: Germany

institute: MPI

data_processing:
Sequencing data processing. Paired-end 50 base reads with additional 6 base reads of barcodes were obtained for each of the samples, i.e. total RNA (RNA-Seq), total fragmented RNA (RNA-Frag-Seq), labeled RNA (4sU-Seq), labeled fragmented RNA (TT-Seq) and ChIP-Seq samples (N20 and S2P). Reads were demultiplexed and mapped with STAR 2.3.0 (for RNA-Seq) (Dobin and Gingeras, Current protocols in bioinformatics, 2015) and Bowtie 2.1.0. (Langmead and Salzberg, Nat Methods, 2012) (for ChIP-Seq) to the hg20/hg38 (GRCh38) genome assembly (Human Genome Reference Consortium). Samtools (Li et al., Bioinformatics, 2009) was used to quality filter SAM files, whereby alignments with MAPQ smaller than 7 (-q 7) were skipped and only proper pairs (-f99, -f147, -f83, -f 163) were selected. Further processing of the RNA-Seq data was carried out using the R/Bioconductor environment. For visualization purposes piled-up read counts for every genomic position were summed up over replicates, size factors for each condition were calculated as described in (Anders and Huber, Genome Biol, 2010) and used to correct for library size and sequencing depth variations.
Genome_build: hg20/hg38 (GRCh38)
Supplementary_files_format_and_content: The file binned_count_table.txt gives the fragment counts for all experiments using the binning_anno.txt annotation file
Supplementary_files_format_and_content: The Series file README.txt includes information about the contents of the processed data files.

source_name: K562 cells

growth_protocol: K562 cells were acquired from DSMZ (Braunschweig, Germany). Cells were grown in RPMI 1640 medium (Gibco) supplemented with 10% heat-inactivated FBS (Gibco) and 1% Penicillin/Streptomycin (100x, PAA) at 37°C under 5% CO2

extract_protocol: NEBNext Ultra DNA Library Prep Kit for Illumina

treatment_protocol: This protocol was adapted from the laboratory of Richard Myers (Johnson et al., Science, 2007). The protocol was followed except for some exceptions. 3x107 K562 cells were crosslinked for 7.5 min with 1% methanol-free formaldehyde (Pierce™ 16% Formaldehyde (w/v), Methanol-free, Thermo Scientific). The chromatin was sonicated on a BioRuptor Next Gen (Diagenode) at high power for 40 cycle of 30’’/30’’ ON/OFF. Antibodies were bound to Dynabeads Protein G (Thermo Scientific) following the manufacturer’s instructions. 75 μl of the antibody 3E10 (Chapman et al., Science, 2007) was bound to 1.5 mg Dynabeads. Samples were immunoprecipitated overnight. Illumina libraries were prepared with NEBNext Ultra DNA Library Prep kit. Samples were sequenced on an Illumina HiSeq 1500 machine.

tag: passage

tag: chip-antibody

value: Serine-2-P (3E10)

tag: cell line


title: N20 ChIP rep2

treatment_protocol: This protocol was adapted from the laboratory of Richard Myers (Johnson et al., Science, 2007). The protocol was followed except for some exceptions. 3x107 K562 cells were crosslinked for 7.5 min with 1% methanol-free formaldehyde (Pierce™ 16% Formaldehyde (w/v), Methanol-free, Thermo Scientific). The chromatin was sonicated on a BioRuptor Next Gen (Diagenode) at high power for 40 cycle of 30’’/30’’ ON/OFF. Antibodies were bound to Dynabeads Protein G (Thermo Scientific) following the manufacturer’s instructions. 1.5 μg of the N-20 antibody (Santa Cruz, sc-899) was bound to 1.5 mg Dynabeads. Samples were immunoprecipitated overnight. Illumina libraries were prepared with NEBNext Ultra DNA Library Prep kit. Samples were sequenced on an Illumina HiSeq 1500 machine.

value: N-20 (Santa Cruz)


title: S2P ChIP rep2


title: Total RNA-Seq rep2

extract_protocol:
RNA was extracted using TRIzol (Life Technologies), 4sU-labeled RNA was exctracted according to Dolken et al., RNA, 2008.
fragmentation: no
NuGEN Human Whole Blood RNA-Seq System

treatment_protocol: Cells  were labeled in media for 5 min with 500 μM 4-thiouridine (4sU, Sigma-Aldrich) and harvested through centrifugation for 2 min at 3,000 rpm.


title: kCtr.2

value: Control embryo k


title: N20 ChIP rep1


title: Total RNA-Seq rep1


title: 20PNT.5


title: eCtr.3


title: gCtr.1

value: Control embryo g


title: 4sU-Seq rep2


title: 20PNT.3


title: 1PNT.7

value: Heterologous pronuclear transfer embryo 1


title: 20PNT.1

value: Blastocyst single cell


title: 8PNT.14

value: Heterologous pronuclear transfer embryo 8


title: 8PNT.15


title: jCtr.3

value: Control embryo j


title: H2217-WT-GLU.2.XYL.5-24-2-03.20.14-1169_redo-4-98

description: wildtype in 2% Glucose and 5% Xylose for 24 hours

source_name: S. cerevisiae cells put into aerobic fermentations of SC medium with 2% glucose and 5% xylose

value: aerobic batch fermentation of SC + 2% gluocse and 5% xylose for 24 hours


title: H2217-USV1-GLU.2.XYL.5-24-2-03.20.14-1169_redo-5-94

description: USV1(YPL230W) in 2% Glucose and 5% Xylose for 24 hours


title: H2217-USV1-GLU.2.XYL.5-24-1-03.20.14-1169_redo-4-93


title: H2217-WT-GLU.2.XYL.5-4-2-03.20.14-1169_redo-4-96

description: wildtype in 2% Glucose and 5% Xylose for 4 hours

value: aerobic batch fermentation of SC + 2% gluocse and 5% xylose for 4 hours


title: H2217-MSN4-GLU.2.XYL.5-24-2-03.20.14-1169_redo-5-90

description: MSN4(YKL062W) in 2% Glucose and 5% Xylose for 24 hours


title: 7PNT.3

value: Heterologous pronuclear transfer embryo 7


title: 10PNT.10

value: Autologous pronuclear transfer embryo 10


title: H2217-WT-GLU.2.XYL.5-24-3-03.20.14-1169_redo-5-99


title: 8PNT.10


title: H2217-USV1-GLU.2.XYL.5-4-2-03.20.14-1169_redo-5-92

description: USV1(YPL230W) in 2% Glucose and 5% Xylose for 4 hours


title: H2217-WT-GLU.2.XYL.5-4-1-03.20.14-1169_redo-4-95


title: 8PNT.16


title: 1PNT.4


title: 10PNT.8


title: jCtr.5


title: H2217-MSN4-GLU.2.XYL.5-24-1-03.20.14-1169_redo-4-89


title: 4sU-Seq rep1


title: 8PNT.6


title: H2217-MSN2-GLU.2.XYL.5-24-1-03.20.14-1169_redo-4-85

description: MSN2(YMR037C) in 2% Glucose and 5% Xylose for 24 hours


title: H2217-MSN4-GLU.2.XYL.5-4-1-03.20.14-1169_redo-4-87

description: MSN4(YKL062W) in 2% Glucose and 5% Xylose for 4 hours


title: H2217-MSN2-GLU.2.XYL.5-24-2-03.20.14-1169_redo-5-86


title: 8PNT.9


title: 7PNT.1


title: H2217-USV1-GLU.2.XYL.5-4-1-03.20.14-1169_redo-4-91


title: H2217-MSN4-GLU.2.XYL.5-4-2-03.20.14-1169_redo-5-88


title: 8PNT.12


title: H2217-HAP4-GLU.2.XYL.5-24-2-03.20.14-1169_redo-4-81

description: HAP4(YKL109W) in 2% Glucose and 5% Xylose for 24 hours


title: H2217-HAP4-GLU.2.XYL.5-4-2-03.20.14-1169_redo-4-78

description: HAP4(YKL109W) in 2% Glucose and 5% Xylose for 4 hours


title: H2217-HAP4-GLU.2.XYL.5-4-1-03.20.14-1169_redo-4-77


title: 8PNT.2


title: H2217-HAP4-GLU.2.XYL.5-24-3-03.20.14-1169_redo-5-82


title: H2217-GIS1-GLU.2.XYL.5-24-1-03.20.14-1169_redo-4-75

description: GIS1(YDR096W) in 2% Glucose and 5% Xylose for 24 hours


title: 2PNT.11

value: Heterologous pronuclear transfer embryo 2


title: iCtr.2

value: Control embryo i


title: H2217-GIS1-GLU.2.XYL.5-24-2-03.20.14-1169_redo-5-76


title: 2PNT.5


title: iCtr.4


title: 1PNT.5


title: iCtr.1


title: H2217-GIS1-GLU.2.XYL.5-4-2-03.20.14-1169_redo-5-74

description: GIS1(YDR096W) in 2% Glucose and 5% Xylose for 4 hours


title: H2217-GIS1-GLU.2.XYL.5-4-1-03.20.14-1169_redo-4-73


title: 1PNT.1


title: H2217-CAT8-GLU.2.XYL.5-24-3-03.20.14-1169_redo-5-72

description: CAT8(YMR280C) in 2% Glucose and 5% Xylose for 24 hours


title: H2217-HAP4-GLU.2.XYL.5-4-3-03.20.14-1169_redo-5-79


title: 9PNT.12

value: Heterologous pronuclear transfer embryo 9


title: H2217-HAP4-GLU.2.XYL.5-24-1-03.20.14-1169_redo-4-80


title: 9PNT.9


title: iCtr.6


title: 9PNT.8


title: H2217-CAT8-GLU.2.XYL.5-24-1-03.20.14-1169_redo-4-70


title: H2217-CAT8-GLU.2.XYL.5-24-2-03.20.14-1169_redo-5-71


title: H2217-CAT8-GLU.2.XYL.5-4-2-03.20.14-1169_redo-5-68

description: CAT8(YMR280C) in 2% Glucose and 5% Xylose for 4 hours


title: H2217-MSN2-GLU.2.XYL.5-4-1-03.20.14-1169_redo-4-83

description: MSN2(YMR037C) in 2% Glucose and 5% Xylose for 4 hours


title: 9PNT.5


title: H2217-CAT8-GLU.2.XYL.5-4-1-03.20.14-1169_redo-4-67


title: iCtr.3


title: 9PNT.4


title: H2217-SNF1-GLU.2.XYL.5-24-2-02.10.11-392-3-41

description: SNF1(YDR477W) in 2% Glucose and 5% Xylose for 24 hours


title: 9PNT.2


title: H2217-SNF1-GLU.2.XYL.5-24-1-02.10.11-392-4-40


title: H2217-CAT8.HAP4-GLU.2.XYL.5-24-3-03.20.14-1169_redo-5-66

description: CAT8(YMR280C)HAP4(YKL109W) in 2% Glucose and 5% Xylose for 24 hours


title: H2217-CAT8.HAP4-GLU.2.XYL.5-24-1-03.20.14-1169_redo-4-64


title: 9PNT.3


title: H2217-SNF1-GLU.2.XYL.5-4-3-02.10.11-392-2-39

description: SNF1(YDR477W) in 2% Glucose and 5% Xylose for 4 hours


title: H2217-SNF1-GLU.2.XYL.5-24-3-02.10.11-392-2-42


title: 4PNT.12

value: Heterologous pronuclear transfer embryo 4


title: 9PNT.7


title: 1PNT.2


title: H2217-CAT8.HAP4-GLU.2.XYL.5-4-1-03.20.14-1169_redo-4-61

description: CAT8(YMR280C)HAP4(YKL109W) in 2% Glucose and 5% Xylose for 4 hours


title: 9PNT.1


title: H2217-SNF1-GLU.2.XYL.5-4-1-02.10.11-392-4-37


title: H2217-ADR1-GLU.2.XYL.5-24-3-02.10.11-392-3-36

description: ADR1(YDR216W) in 2% Glucose and 5% Xylose for 24 hours


title: H2217-SNF1-GLU.2.XYL.5-4-2-02.10.11-392-3-38


title: H2217-CAT8.AFT2-GLU.2.XYL.5-4-3-03.20.14-1169_redo-5-59

description: CAT8(YMR280C)AFT2(YPL202C) in 2% Glucose and 5% Xylose for 4 hours


title: 4PNT.10


title: 4PNT.11


title: H2217-ADR1-GLU.2.XYL.5-24-1-02.10.11-392-5-34


title: H2217-ADR1-GLU.2.XYL.5-4-3-02.10.11-392-3-33

description: ADR1(YDR216W) in 2% Glucose and 5% Xylose for 4 hours


title: H2217-CAT8.AFT2-GLU.2.XYL.5-4-2-03.20.14-1169_redo-4-58


title: 4PNT.9


title: H2217-CAT8.AFT2-GLU.2.XYL.5-24-3-03.20.14-1169_redo-5-60

description: CAT8(YMR280C)AFT2(YPL202C) in 2% Glucose and 5% Xylose for 24 hours


title: H2217-AFT2-GLU.2.XYL.5-24-2-03.20.14-1169_redo-5-57

description: AFT2(YPL202C) in 2% Glucose and 5% Xylose for 24 hours


title: H2217-CAT8.HAP4-GLU.2.XYL.5-4-3-03.20.14-1169_redo-5-63


title: 4PNT.8


title: H2217-AFT2-GLU.2.XYL.5-4-2-03.20.14-1169_redo-5-56

description: AFT2(YPL202C) in 2% Glucose and 5% Xylose for 4 hours


title: 4PNT.7


title: 4PNT.6


title: H2217-ADR1-GLU.2.XYL.5-4-1-02.10.11-392-5-31


title: H2217-AFT2-GLU.2.XYL.5-4-1-03.20.14-1169_redo-4-55


title: H2217-ADR1-GLU.2.XYL.5-4-2-02.10.11-392-4-32


title: 4PNT.4


title: H2217-CAT8.ADR1-GLU.2.XYL.5-24-2-04.30.13-881-5-53

description: CAT8(YMR280C)ADR1(YDR216W) in 2% Glucose and 5% Xylose for 24 hours


title: H2217-WT-GLU.2.XYL.5-24-1-02.10.11-392-3-28

description: wildtype in 2% Glucose and 5% Xylose for 24 hours


title: H2217-WT-GLU.2.XYL.5-24-2-02.10.11-392-2-29


title: 4PNT.3


title: 4PNT.2


title: 4PNT.1


title: H2217-CAT8.ADR1-GLU.2.XYL.5-4-3-04.30.13-881-5-51

description: CAT8(YMR280C)ADR1(YDR216W) in 2% Glucose and 5% Xylose for 4 hours


title: BY4741-WT-GLU.2.XYL.5-24-3-02.10.11-392-4-24


title: 3PNT.11

value: Heterologous pronuclear transfer embryo 3


title: H2217-CAT8.ADR1-GLU.2.XYL.5-4-1-04.30.13-881-5-49


title: H2217-WT-GLU.2.XYL.5-4-2-02.10.11-392-2-26

description: wildtype in 2% Glucose and 5% Xylose for 4 hours


title: H2217-WT-GLU.2.XYL.5-4-3-02.10.11-392-5-27


title: H2217-CAT8-GLU.2.XYL.5-24-3-04.30.13-881-5-48

description: CAT8(YMR280C) in 2% Glucose and 5% Xylose for 24 hours


title: H2217-CAT8-GLU.2.XYL.5-24-2-04.30.13-881-5-47


title: H2217-CAT8.ADR1-GLU.2.XYL.5-4-2-04.30.13-881-5-50


title: BY4741-WT-GLU.2.XYL.5-24-2-02.10.11-392-5-23


title: H2217-WT-GLU.2.XYL.5-24-3-02.10.11-392-5-30


title: 3PNT.9


title: H2217-CAT8.ADR1-GLU.2.XYL.5-24-3-04.30.13-881-5-54


title: BY4741-WT-GLU.2.XYL.5-24-1-02.10.11-392-2-22


title: BY4741-WT-GLU.2.XYL.5-4-2-02.10.11-392-5-20


title: BY4741-WT-GLU.2.XYL.5-4-3-02.10.11-392-4-21


title: 30-cell ESC_Dnase-seq

description: All DHSs for each stages.xlsx

tag: cell/embruos number

value: 30 cells

tag: strain


title: H2217-CAT8-GLU.2.XYL.5-4-3-04.30.13-881-5-45

description: CAT8(YMR280C) in 2% Glucose and 5% Xylose for 4 hours


title: 3PNT.6

tag: cell type


title: H2217-CAT8-GLU.2.XYL.5-4-2-04.30.13-881-5-44


title: 3PNT.4


title: H2217-CAT8-GLU.2.XYL.5-4-1-04.30.13-881-5-43


title: 3PNT.8


title: MCF-10A wildtype BRG1 ChIPseq Replicate1 Pulldown

status: Public on Jun 02 2016

city: Durham

first: Rasim

last: Barutcu

address: 6 Davis Drive

institute: ScitoVation

description:
processed data file: MCF10A_BRG1_CHIP.ucsc.bedGraph.gz
processed data file: MCF10A_BRG1_peaks.bed

data_processing:
The adapters were trimmed from the sequencing reads, and the reads were aligned to the hg19 human genome using the Bowtie2 tool.  Quality controls, peak calling, motif analysis and peak annotation was performed using the HOMER suite. As the ChIP signal across the biological replicates showed high correlation, we performed ChIP-seq peak calling on the pooled replicates.
genome build: hg19
processed data files format and content: ChIP-seq: BedGraph UCSC visualization files and the MCF10A BRG1 peak bed file

source_name: Mammary epithelial cell line

growth_protocol: MCF-10A cells were obtained from the Barbara Ann Karmanos Cancer Institute (Detroit, MI). The cells were maintained in monolayer in Dulbecco's modified Eagle's medium-F12 (DMEM/F12) (Invitrogen, 21041025) supplemented with 5% horse serum (Invitrogen, 16050122), 1% penicillin/streptomycin (Invitrogen, 15140122), 0.5 μg/ml hydrocortisone (Sigma, H-0888), 100 ng/ml cholera toxin (Sigma, C-8052), 10 μg/ml insulin (Sigma, I-1882), and 20 ng/ml recombinant human EGF (Peprotech, 100-15) as previously described (Debnath et al. 2003).  MCF-7 cells were obtained from ATCC and were cultured in DMEM supplemented with 10% FBS and pen-strep.

extract_protocol:
~1x107 parental MCF-10A cells were crosslinked with formaldehyde at room temperature for 10 minutes.  Then, the cells were lysed using lysis buffer A (50mM HEPES, 140mM NaCl, 1mM EDTA pH=8, 10% Glycerol, 0.5% NP-40, 0.25% Triton X-100), and the residual cytoplasmic protein was removed using lysis buffer B (10mM Tris-HCl pH=8, 200mM NaCl, 1mM EDTA, 1mM EGTA).  The nuclear fraction was released using lysis buffer C (10mM Tris-Hcl pH=8, 100mM NaCl, 1mM EDTA, 1mM EGTA, 0.1% Sodium Deoxycholate, 0.5% N-lauroylsarcosine).  The chromatin was then sheared by using a Bioruptor instrument in high setting, 30' on , 30' off for 5 minutes for 5 cycles. The pull-down was performed by using the BRG1 antibody (Santa Cruz #G-7).  Samples were washed three times with RIPA buffer (Tris-HCl pH=8, 150mM NaCl, 1mM EDTA, 1% NP-40, 0.25% Sodium deoxycholate, 0.1% SDS) and were eluted.
The pull-down and input control sequencing libraries were generated by using the NEXTflex Rapid DNA Sequencing Kit (Bioo Scientific #5144-02) and were sequenced by using SE100 reads with a HiSeq 2000 instrument.

treatment_protocol: The doxocyline induction was performed by the addition of 0.05 microgram per mililiter DOX to the cells and incubating them for 3 to 4 days.

value: Mammary epithelial cell line


title: MCF-10A wildtype BRG1 ChIPseq Replicate2 Pulldown


title: BY4741-WT-GLU.2.XYL.5-4-1-02.10.11-392-2-19


title: MCF-10A wildtype BRG1 ChIPseq Replicate2 Input

description:
processed data file: MCF10A_INPUT.ucsc.bedGraph.gz
processed data file: MCF10A_BRG1_peaks.bed


title: MCF-10A shSCRAM polyA RNA-seq Replicate2

description:
processed data file: genes_expression_expected_count.tsv
processed data file: shBRG1_over_shSCRAM_SignificantlyExpressedGenes.csv

data_processing:
For  RNA-Seq analysis, first, the adapter sequences were removed from the RNA-seq reads.  Ribosomal RNA reads were then filtered out using Bowtie (Song et al. 2014).  After these filtration and adapter removal steps, the reads were aligned to a transciptome and quantified using RSEM v1.2.7 (Li and Dewey 2011). Differential gene expression was calculated by using the Deseq2 package in R 3.1.0 by using the mean value of gene-wise dispersion estimates (Love et al. 2014).
genome build: hg19
processed data files format and content:  Expected count file for all genes and a txt file showing the shBRG1/shSCRAM log2 Fold Change and pvalues for all genes

extract_protocol:
For the RNA-seq, total RNA was isolated from ~%75 confluent MCF-10A cells by using the TRizol Reagent (Life Technologies #15596-026) treated with DNase1.
The poly(A)-selected RNA-seq libraries were generated with TruSeq RNA Sample Preparation Kit v2 and SE100 sequencing was performed using a Hi-Seq 2000 instrument.


title: MCF-10A shBRG1 polyA RNA-seq Replicate1


title: MCF-10A shSCRAM polyA RNA-seq Replicate1


title: MCF-10A shSCRAM Hi-C Replicate2

description: processed data file: shSCRAM_shBRG1_processed_HiC_files.tar.gz

data_processing:
Library strategy: HiC
The Hi-C data were mapped with Bowtie and  binned at 6.5Mb, 1Mb, 250kb, 100kb and 40kb non-overlapping genomic intervals.  Iterative mapping and correction of Hi-C data were performed as previously described (Imakaev et al. 2012).  Biological replicates showed high reproducibility (Pearson's correlation coefficient > 0.9 for 1Mb resolution data).  Similarly, the first eigenvector comparison of the replicates showed high reproducibility.  For the downstream analyses, sequences obtained from both biological replicates were pooled and ICE-corrected to serve as a combined dataset.
First, the z-scores of the interaction matrices at 250kb resolution were generated as described previously (Lieberman, 2009).  Then, Pearson Correlation on the Z-score matrices was calculated.  By performing principal component analysis (Lieberman-Aiden et al. 2009; Zhang et al. 2012), the first eigenvector typically represents the compartment profile(Lieberman-Aiden et al. 2009), where the positive and negative 1st eigenvalues represent different compartments.   Gene density for each compartment was calculated to call the “A” and “B” compartmentalization.
TAD calling was performed as calculating the “insulation” score of each bin using the 40kb resolution combined Hi-C data (as previously described.  The mean of the interactions across each bin were calculated.  By sliding a 1Mb x 1Mb (25bins x 25bins) square along the diagonal of the interaction matrix for every chromosome, we obtained the “insulation score” of the interaction matrix.  Valleys in the insulation score indicated the depletion of Hi-C interactions occurring across a bin.  These 40kb valleys represent the TAD boundaries.  Based on the variation of boundaries between replicates, we chose to add a total of 160kb (80kb to each side) to the boundary to account for replicate variation.  The final boundaries span a 200kb region. All boundaries with a boundary strength < 0.15 were excluded as they were considered weak and non-reproducible.  The insulation plots for the biological replicates showed high reproducibility, suggesting the robustness of the method.  Similarly, the overlap of detected boundaries also showed high reproducibility between the biological replicates.  Therefore, we used the combined Hi-C replicates for the TAD analyses.
We modeled the overall Hi-C decay with distance using a modified LOWESS method (alpha = 1%, IQR filter), as described previously (Sanyal et al. 2012).  LOWESS calculates the weighted-average and weighted-standard deviation for every genomic distance and therefore normalizes for genomic distance signal bias.
genome build: hg19
processed data files format and content: HiC: Interaction matrices at 2.5Mb, 1Mb, 250kb and 40kb resolution

extract_protocol:
not provided
The Hi-C libraries were prepared as previously described (PMID:22652625).  Hi-C was sequenced using PE150 reads


title: MCF-10A shBRG1 polyA RNA-seq Replicate2


title: MCF-10A shSCRAM Hi-C Replicate1


title: RRHP_hESC_22

city: Oslo

first: Xavier

last: Tekpli

address: Montebello

country: Norway

institute: Institute for Cancer Research

data_processing:
FASTQ files were aligned to the human reference genome (hg19) with Bowtie-0.12.8 using its default parameters and --best. Each sequencing/aligned read with a CCGG tag at the beginning of the read represented one 5hmC site. The number of sequencing reads at a CpG-site in a CCGG context was considered to be proportional to the level of hydroxymethylation
Genome_build: hg19
Supplementary_files_format_and_content: Bed files of the number of sequencing reads representing the level of hydroxymethylation at CpG sites in a CCGG context
For more details on RRHP: see RRHP: a tag-based approach for 5-hydroxymethylcytosine mapping at single-site resolution (Petterson et al; 2014).

source_name: Embryonic stem cells

extract_protocol: DNA was isolated from human cells using proteinase K (Sigma, St Louis, MO) and RnaseA digestion (Qiagen, Hilden, Germany) followed by phenol–chloroform extraction and ethanol precipitation.

value: Embryonic stem cells

tag: status

value: WIBR #22


title: MCF-10A shBRG1 Hi-C Replicate2


title: RRHP_hESC_38

value: WIBR #38


title: MCF-10A shBRG1 Hi-C Replicate1


title: MCF-10A wildtype BRG1 ChIPseq Replicate1 Input


title: NCCIT.CBFA2T2

city: New York City

first: Danny

last: Reinberg

state: New York

address: 522 First Avenue, 2nd floor Rm 211

institute: NYU School of Medicine / HHMI

data_processing:
Reads were mapped to mm9 or hg19 with BOWTIE -m4 -v2
1nt resolution bedgraphs were generated with the BEDtools bamToBed and genomeCoverageBed utilities after extending each read to 200
coverage was normalized to FP10M (fragments per 10 million reads sequenced)
bigWig files were generated with bedGraphToBigWig
Genome_build: mm9 and hg19
Supplementary_files_format_and_content: FP10M-normalized bigWigs

source_name: NCCIT cells

growth_protocol: mESCs were grown in standard ESC medium containing Lif, 1 μM MEK1/2 inhibitor (PD0325901) and 3 μM GSK3 inhibitor (CHIR99021). NCCIT cells were grown in RPMI1640 with 10% FBS.

extract_protocol:
Cells were lysed in ChIP buffer (50 mM Tris-HCl pH 7.9, 150 mM NaCl, 1% Triron X-100, 0.5% NP-40, 5 mM EDTA pH 8.0, 1 mM PMSF and protease inhibitor) and chromatin was sonicated to ~200 bp with a Diagenode Bioruptor
Libraries were prepared according to manufacturer’s instructions (Illumina). Immunoprecipitated DNA was first end-repaired using End-It Repair Kit (Epicenter), tailed with an A using Klenow exo minus (NEB M0212), and ligated to custom adapters. Fragments of 300±100 bp were size-selected and subjected to ligation-mediated PCR amplification (LM-PCR), using Q5 polymerase (NEB M0530).

value: NCCIT Embryonal Carcinoma cell line

tag: chip antibody

value: CBFA2T2 (Bethyl, catalog# A303-593A)


title: NCCIT.input


title: NCCIT.PRDM14

value: PRDM14 (Homemade)


title: RNA-seq_PB_CD19+

data_processing: RNA was purified using the RNeasy Mini Kit (Qiagen), and RNA samples with RNA Integrity number values > 8 were used for sequencing. Libraries were prepared using TruSeq RNA Sample Preparation Kit v2 (Illumina) following the manufacturer’s instructions and sequenced with 100-bp paired-end reads on a HiSeq2000 Illumina. Raw reads (FASTQ) were aligned to the human reference genome (hg19) by Tophat v2.0.13 and then assembled by Cufflinks v2.1.1 with default parameters . Quantification of gene expression was performed using cufflink with default parameters to obtain FPKM value

source_name: Venous peripheral blood

extract_protocol: RNA was purified using the RNeasy Mini Kit (Qiagen, Hilden, Germany) and only RNA samples with RIN values > 8 were processed

tag: disease state

value: healthy

tag: tissue

value: Venous peripheral blood

value: B lymphocytes


title: RNA-seq_PB_CD14+

value: Monocytes


title: NCCIT.RING1B

value: RING1B (Bethyl, catalog# A302-869A)


title: RRHP_CD14+

description: library strategy: RRHP


title: RRHP_Granulocytes_2

description: library strategy: RRHP

extract_protocol: Raw RNA-seq data were aligned to the human reference genome (hg19) by Tophat v2.0.13 using p 8 parameter. Gene expression quantification by FPKM (Fragments Per Kilobase of exon per Million mapped fragments) with default parameter by using cufflinks (Trapnell et al., 2012). FPKM are shown in the processed file.

value: Granulocytes


title: RRHP_Granulocytes_1

description: library strategy: RRHP


title: RNA-seq_PB_CD4+

value: T lymphocytes


title: RRHP_CD4+

description: library strategy: RRHP


title: RRHP_CD19+

description: library strategy: RRHP


title: NCCIT.SUZ12

value: SUZ12 (Cell Signaling, catalog# 3737S)


title: RRHP_BM_CD34+

description: library strategy: RRHP

source_name: Bone marrow

value: Bone marrow

value: Hematopoietic stem/progenitor cells


title: RRHP_CB_CD34+_348

description: library strategy: RRHP

source_name: Umbilical cord blood

value: Umbilical cord blood


title: HCT116-Day42-HCG/GCH

city: Los Angeles

first: Jessica

last: Charlet

state: California

address: 1441 Eastlake Avenue

department: Norris Cancer Center

institute: USC Keck School of Medicine

description: NOMe-seq

data_processing:
genomic alignment for paired-end reads were done using MAQ with the -c bisulfite mode
Low quality reads and improperly paired reads were filtered out
Methylation calling of CpG and GpC was done using Bis-SNP (Liu et al 2011)
Genome_build: hg19
Supplementary_files_format_and_content: Methylation value of CG and GC sites were extacted and bigwig files were generated

growth_protocol: HCT116 cells were cultured in McCoy5A media supplemented with 10% FBS and penicillin/streptomycin at 37C/5%CO2

extract_protocol:
Cells were trypsinized and washed with PBS. Nuclei were isolated and treated with 300 U of M.CvIPI, a GpC methyltransferase which can methylate accessible GpC sites without affecting CpG information (Kelly et al 2012)
WGBS libraries were prepared based on previously described (Berman et al 2012) and according to Illumina's recommendation.

treatment_protocol: HCT116 cells were treated for 24h with 0.3uM 5-Aza-CdR

tag: Sex

value: male

value: day 42


title: RRHP_CB_CD34+_350

description: library strategy: RRHP

data_processing:
FASTQ files were aligned to the human reference genome (hg19) with Bowtie-0.12.8 using its default parameters and --best. Each sequencing/aligned read with a CCGG tag at the beginning of the read represented one 5hmC site. The number of sequencing reads at a CpG-site in a CCGG context was considered to be proportional to the level of  hydroxymethylation
Genome_build: hg19
Supplementary_files_format_and_content: Bed files of the number of sequencing reads representing the level of hydroxymethylation at CpG sites in a CCGG context
For more details on RRHP: see RRHP: a tag-based approach for 5-hydroxymethylcytosine mapping at single-site resolution (Petterson et al; 2014).


title: HCT116-Day25-HCG/GCH

description: NOMe-seq

value: day 25


title: Fetal liver Knockout of alpha globin regulatory elements R1 and R2, biological replicate 3 [Capture C]

city: oxford

first: Jim

last: Hughes

address: Weatherall Institute Of Molecular Me

department: MHU

institute: University of Oxford

description:
molecule: Next generation Capture-C product
3C library prepartion followed by double oligonucleotide capture
HbaCombined_Enhancer_KO_30gH_Combined_PF_BF.gfc
HbbCombined_Enhancer_KO_30gH_Combined_PF_BF.gfc
Slc25A37_Enhancer_KO_30gH_Combined_PF_BF.gfc

data_processing:
Library strategy: Capture-C
Trim_galore (to remove sequencing adaptors)
FLASH (to reconstruct paired end reads into single reads where possible)
In silico restriction enzyme digestion of FASTQ file (dpnIIPE.pl)
Alignment to the genome (Bowtie) maintaining strict read order
Removal of PCR duplicates; Parsing of informative reads and mapping to restriction enzyme fragments (CCanalyser2.pl)
Combining data from multiple replicates (custom scripts)
Removal of ploidy regions and off target capture (custom scripts)
Analysis in R to normalise between tracks using the total number of informative interactions
Genome_build: mm9
Supplementary_files_format_and_content: Custom combined data format (restriction enzyme fragment \t sample 1 \t sample 2 \t etc.)

source_name: Erythroid cells

extract_protocol:
Fetal livers were dissected from E12.5 mouse embryos and the dissociated cell suspension was expanded for 5-7 days in Stempro (Invitrogen) supplemented with Epo (1U/ml), SCF (50ng/ml) and dexamethasone (1mM). Following expansion of the cultures, mature red cells were removed by negative selection for Ter119 using anti-Ter-119 magnetic beads (Miltenyi), to obtain a population of erythroid precursors. Cultures were then switched to medium containing high Epo (5U/ml) to induce differentiation to mature erythroid cells. After 18hrs differentiation, 3C libraries were prepared.
The protocol used for generation of the 3C libraries was similar to previous methods with minor modifications. Briefly, cells were crosslinked with 2% formaldehyde (10 minutes, room temperature); quenched with cold glycine; washed in phosphate buffered saline; resuspended in cold lysis buffer (tris 10mM, NaCl 10mM, NP40 0.2%, complete proteinase inhibitor (Roche)) and snap frozen to -80.  Cells were thawed on ice, washed in milliq dH2O and Dounce homogenised on ice (x 40 strokes). Cells were then resuspended with 0.25% SDS and restriction enzyme buffer and incubated at 37C for 1h at 1400rpm on a Comfort Thermomixer (Eppendorf) followed by a further incubation of 1h following the addition of triton X100 (final concentration 1.67%). An overnight digestion was performed using Dpn II (500U /ml (NEB) at 37C / 1400 rpm). The digested chromatin was ligated overnight (Fermentas HC Ligase final concentration 10U/ml) at 16 degrees at 1400 rpm on the Thermomixer.  The samples were then decrosslinked overnight at 65C with Proteinase K (Roche) followed by a 30 min incubation at 37C with RNAse (Roche). Phenol/Chloroform extraction was then performed followed by an Ethanol precipitation and a wash with 70% Ethanol. Digestion efficiencys were assessed by gel electrophoresis (1% agarose) and RT-PCR (Taqman), which showed digestion efficiencies in excess of 80%.  DNA content of the Dpn II 3C libraries were quantitated using a Qubit fluorometer (Life technologies) and 5-10ug of each library was sheared using a Covaris S2 in milliq dH2O. Covaris settings used were: duty cycle 10%, Intensity 5, Cycles/burst 200, Time 6 cycles of 60seconds, Set Mode Frequency sweeping Temperature 4 to 7 degrees. Following shearing DNA was purified using AMPureXP beads (Agencourt) and DNA quality assessed on a Bioanalyser 2100 using a DNA High Sensitivity Chip (Agilent). DNA end repair and adapter ligation was performed using the NEB Next or NEB Ultra DNA sample preparation reagent kits, depending on the amount of DNA available, using the standard protocol.  Biotinylated capture oligonucleotides were designed to the ends of the viewpoint fragments.  Where possible 1-2ug of each adapter ligated library were hybridized with the biotinylated capture oligonucleotides, using  the Nimblegen SeqCap reagents and an adapted protocol.  The quality of the resultant captured library was assessed by Agilent tapestation or bioanalyser (D1000). The resulting libraries were sequenced using either the Illumina MiSeq (150bp paired end reads) or HiSeq (100bp paired end reads).
The experimental strategy was to generate very high complexity randomly sonicated 3C libraries with ligated Illumina sequencing adaptors (ideally with several hundred thousand fold coverage).  Oligonucleotide capture technology was then used to pull down the restricion fragments containing the promoters of the genes of interest and their interacting partners, which were subsequently idendified by high throughput sequencing.  The power of the technique is that multiple samples and genes can be analysed simultaneously by using differently indexed samples.

tag: knockout of enhancer-like component

value: regulatory elements R1 and R2

value: F1 C57Bl6 / CBA

value: Fetal Liver Culture

tag: genes analysed


title: Fetal liver Knockout of alpha globin regulatory elements R2 and R3, biological replicate 1 [Capture C]

value: regulatory elements R2 and R3


title: Fetal liver,  biological replicate control replicate  1 [Capture C]


title: Fetal liver,  biological replicate control replicate  2 [Capture C]


title: Fetal liver Knockout of alpha globin regulatory elements R2 and R3, biological replicate 2 [Capture C]


title: Fetal liver Knockout of alpha globin regulatory elements R1 and R2, biological replicate 2 [Capture C]


title: Fetal liver Knockout of alpha globin regulatory elements R1 and R2, biological replicate 1 [Capture C]


title: Fetal liver Knockout of alpha globin regulatory elements R2 and R3, biological replicate 3 [Capture C]


title: Fetal liver,  biological replicate control replicate  3 [Capture C]


title: HCT116-Day5-HCG/GCH

description: NOMe-seq

tag: time point

value: day 5

tag: treatment


title: Fetal liver Knockout of alpha globin Wild type litter mate replicate 1 [ATAC-seq]

description: Genome-wide library of transposed fragments

data_processing:
Library strategy: ATAC-seq
Trim_galore (to remove sequencing adaptors)
Alignment to the genome Bowtie 1 standard parameters with -m set to 2
Removal of PCR duplicates;
plot density of alignments in a moving 300bp window with an moving increment of 30 bp
Genome_build: mm9
Supplementary_files_format_and_content: wig

extract_protocol:
A single cell suspension was made by gently dissociating the E14.5 fetal livers and passing it through a 70um filter.  Ter119 selection was performed by staining the cells with phycoerythrin conjugated anti-ter119 antibodies (BD biosciences)).  The cells were subsequently washed and then incubated with MACS anti-phycoerythrin beads.  Cells were then separated using a magnetic column, MACS (miltenyi biotec). Nuclei were isolated by lysing the cells as  previously published (Buenrostro, 2013).
Assay for transposition of active chromatin sequencing (ATAC-Seq) was performed as previously published (Buenrostro, 2013). 60000-80000 cells were used per biological replicate. Cells were lysed and nuclei were isolated prior to transposition with Tn5 transposase (Nextera, Illumina) for 30 minutes at 37°C. DNA was purified using a MinElute kit (Qiagen). Libraries were amplified and barcoded using the NEBNext 2xMastermix (NEB) and the custom primers as published in Buenrostro et al., 2013. ATAC-Seq libraries profiles were visualized using D1000 tape on the Tapestation (Agilent). Due to the broad range of DNA fragment sizes found in these libraries, quantitation with the Qubit for DNA concentration was found to be highly variable and was omitted. The libraries were quantified using the universal library quantification kit (KAPA Biosystems). Samples were sequenced using: 75bp paired end reads (MiSeq platform) or 40 bp paired end reads (NextSeq platform).
The strategy was to isolate open chromatin fragments specifically as they underly regulatory genomic regions. These are short stretches of DNA, with low abundance in the genome and thus require to be processed efficiently. The use of the transposase is ideal as the modified Tn5 can in one step identify the open chromatin fragments, cut them from the chromatin environment and ligate adaptors. The fragments can then be multiplexed and sequenced

value: wild type litter mate

value: Fetal Liver

tag: age

value: Genome-wide


title: Celf2 nuc CLIPseq

city: San Diego

first: Bing

last: Zhou

state: CALIFORNIA

address: 9500 Gilman Drive

department: Cellular and Molecular Medicine

institute: University of California San Diego

data_processing:
The quality of sequencing reads from fastq files were evaluated by FastQC. The sequencing adapters and over-represented short sequences in the reads found by fastqc were then trimmed using cutadapt.
These trimmed reads from unc-75 CLIP-seq or Celf2 CLIP-seq were mapped to Caenorhabditis elegans genome (WS220/ce10) or Mus musculus genome (GRCm38/mm10) respectively by GSNAP (version 2015-09-29) with parameters of “-t 4 -k 10 -N 1”. The coordinates of mapped reads were evaluated and filtered by custom scripts. Specifically, reads with multiple coordinates and coordinates with duplicated reads were removed by using samtools and bedtools
The peaks calling procedure for the CLIP-seq data was mainly followed the protocols published previously (Moore MJ. et al., Nature Protocols 2014) with some modifications. With samtools and bedtools, the mutation information in the read mapping were extracted, and separated into substitution, deletion and insertion. With mutation information, CIMS pipeline (Zhang C. et al., Nature Biotech. 2011) was then applied to the read coordinates to detect significant peaks at FDR < 0.01 and m > 5.
The de novo motifs were discovered by MEME-ChIP in the DNA sequences underlying the CLIP-seq peaks with 20 bp extensions with both directions, with nearby flanking sequences (100 bp length with 100 bp gap) extracted with bedtools as background control.
Genome_build: Caenorhabditis elegans genome (WS220/ce10), Mus musculus genome (GRCm38/mm10)
Supplementary_files_format_and_content: CLIPseq peaks found by CIMS in BedGraph format

source_name: N2A cell line

growth_protocol: N2A cells were cultured in DMEM media supplemented with 10% FBS in a 5% CO2 humidified incubator at 37°C.

extract_protocol:
Nuclear fraction was incubated with NanoLink streptavidin magnetic beads for overnight at 4 °C followed by washes
Samples were treated with micrococcal nuclease followed by phosphatase treatment, linker addition and 5’-labeling with 32P-γ-ATP. Complexes were eluted and separated with gel electrophoresis. A band corresponding to UNC-75/RNA complexes was cut out of the membrane and proteins degraded by Proteinase K. Associated RNAs were reverse-transcribed and amplified by PCR for deep sequencing.

treatment_protocol: N2A cells were transfected with plasmids of BirA enzyme and BLRP tagged CELF2, stable cell line was obtained and expanded, followed by UV crosslinking.


title: Fetal liver Knockout of alpha globin regulatory elements R1 and R2 biological replicate 1 [ATAC-seq]

value: regulatory element R1 and R2


title: Ezh2 - Dicer KO

first: Gopuraja

last: Dharmalingam

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

department: Epigenetics section

institute: MRC London Institute of Medical Sciences

description: ChIP-Seq data from Dicer deficient ES Cells using anti-Ezh2 antibody (Diagenode)

data_processing:
Sequencing reads were aligned to Mouse genome mm9 using bowtie version 0.12.8
Reads were extended to fragment length estimated using SISSR
Coverage per base was computed using extended reads and visualised using UCSC genome browser
Genome_build: mm9
Supplementary_files_format_and_content: Bigwig

source_name: Dicer KO_ES

growth_protocol: DTCM23 Dicer lox/lox mouse ES cell line, Reference: Nesterova et al [Epigenetics and Chromatin, 1:2, 2008]. Both wildtype and Dicer-deficient ES cells were cultured without feeders in a 50:50 ratio of serum+LIF media, and N2B27+2i media (Ying et al., Nature (453) 2008)

extract_protocol:
50x10^6 cells were collected by trypsinisation, resuspended in complete media and fixed with 1% formaldehyde. Chromatin isolated from fixed nuclei was sonicated and incubated at 4C overnight with Protein A Dynabeads (Invitrogen) coated with 10ug of anti-Ezh2 antibody (Diagenode). Further steps were carried out according to Seitan et al Nature (2011).
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 Hiseq 2500

tag: strain background

value: mixed

tag: cell line background

value: DTCM23 Dicer lox/lox mouse ES cell line

tag: genotype/variation

value: Dicer KO; Dicer deficient

value: embryonic stem cells

value: anti-Ezh2 antibody

tag: chip antibody vendor

value: Diagenode


title: Input - WT

description: Input data from Wildtype ES cells

value: wild type


title: Ezh2 - WT

description: ChIP-Seq data from Wildtype ES Cells using anti-Ezh2 antibody (Diagenode)


title: Input - Dicer KO

description: Input data from Dicer deficient ES cells


title: Control for IP- chromatin only

description: DNA fragments from sonicated chromatin

data_processing:
Trim_galore (to remove sequencing adaptors)
FLASH (to reconstruct paired end reads into single reads where possible)
Alignment to the genome (Bowtie) maintaining strict read order
Removal of PCR duplicates;
Combining data from multiple replicates (custom scripts)
Removal of ploidy regions
Genome_build: mm9
Supplementary_files_format_and_content: wig

extract_protocol:
A single cell suspension was made by gently dissociating the spleen of a phenyhydrazine treated mouse and passing it through a 70um filter.  Ter119 selection was performed by staining the cells with phycoerythrin conjugated anti-ter119 antibodies (BD biosciences)).  The cells were subsequently washed and then incubated with MACS anti-phycoerythrin beads.  Cells were then separated using an auto-MACS (miltenyi biotec).
Fragments double-crosslinked were immunoprecipitated with the antibody of interest (Med1). Libraries were prepared using the NEB Ultra library preparation kit according to manufacturer’s instructions. Samples were sequenced using 40 bp paired end reads on the NextSeq platform
The strategy was to isolate at a genome-wide level the DNA regions bound by Med1. Because the binding is assumed to be indirect, stronger croslinking conditions were used (double crosslinking FA and DSG).

value: Phenylhydrazine treated spleen

tag: antibody

value: input


title: ChIP for Med1

description: DNA fragments bound by Med1

tag: strain


title: Fetal liver Knockout of alpha globin regulatory element Rm [Capture C]

description:
molecule: Next generation Capture-C product
3C library prepartion followed by double oligonucleotide capture
HbaCombined_KO_PF_BF.gfc
HbbCombined_KO_PF_BF.gfc
Slc25A37_KO_PF_BF.gfc

value: regulatory element Rm

value: F1 C57Bl6 / CBA


title: Fetal liver Knockout of alpha globin regulatory element R4 [Capture C]

value: regulatory element R4

value: Fetal Liver Culture


title: Fetal liver Knockout of alpha globin regulatory element R3 [Capture C]

value: regulatory element R3


title: Fetal liver Knockout of alpha globin regulatory element R2 [Capture C]

value: regulatory element R2


title: Fetal liver Knockout of alpha globin regulatory element R1 [Capture C]

value: regulatory element R1


title: Fetal liver Knockout of alpha globin Wild type litter mate replicate 3 [ATAC-seq]

value: wild type litter mate

value: Fetal Liver

value: Genome-wide


title: Fetal liver Knockout of alpha globin regulatory elements R2 and R3 biological replicate 3 [ATAC-seq]

value: regulatory element R2 and R3


title: Fetal liver Knockout of alpha globin regulatory elements R1 and R2 biological replicate 2 [ATAC-seq]

value: regulatory element R1 and R2


title: Fetal liver Knockout of alpha globin regulatory elements R1 and R2 biological replicate 3 [ATAC-seq]


title: Fetal liver Knockout of alpha globin regulatory element Rm, biological replicate 2 [ATAC-seq]


title: Fetal liver Knockout of alpha globin regulatory element R3, biological replicate 2 [ATAC-seq]


title: Fetal liver Knockout of alpha globin regulatory elements R2 and R3 biological replicate 2 [ATAC-seq]


title: Fetal liver Knockout of alpha globin regulatory elements R2 and R3 biological replicate 1 [ATAC-seq]


title: Mouse embryonic stem cells [Capture C]

source_name: E14 embryonic stem cells

value: Embryonic stem cell line


title: Fetal liver Knockout of alpha globin regulatory element R3, biological replicate 1 [ATAC-seq]


title: F1

city: Shenyang

first: Shengnan

last: Huang

address: 120 Dongling Road Shenhe District Shenyang P.R. China

country: China

institute: Shenyang Agricultural University

description: developing flower buds at the full-bloom stage

data_processing:
Illumina HiSeq™ 2000 used for base calling
All raw reads were filtered by removing reads with adaptors and low-quality reads. Clean reads were mapped to the reference database for the B. rapa genome (Version 1.5, http://brassicadb.org/brad/) using SOAPaligner/SOAP2 (Li et al., 2009)
Gene expression levels were calculated using the RPKM (Reads Per kb per Million reads) method (Mortazavi et al., 2008) in six libraries
The DESeq software was applied to analyze DEGs based on the RPKM-derived baseMean for each gene between samples (Anders and Huber, 2010). To identify DEGs, the False Discovery Rate (FDR ≤ 0.001) method was used to determine the threshold of P-value in multiple tests (Benjamini and Hochberg, 1995)
Genome_build: Version 1.5, http://brassicadb.org/brad/
Supplementary_files_format_and_content: tab-delimited excel files include RPKM values,length,coverage... for each gene

organism: Brassica rapa subsp. pekinensis

source_name: developing flower buds

extract_protocol:
The developing flower buds of Chinese cabbage were sampled, total RNA was extracted using Trizol reagent
cDNA libraries were prepared for sequencing using standard Illumina protocols

value: Chinese cabbage

tag: genotype

value: wild type 'FT'


title: Fetal liver Knockout of alpha globin regulatory element R2, biological replicate 2 [ATAC-seq]


title: F2

value: developing flower buds at the full-bloom stage


title: M1

value: mutant fsm


title: Fetal liver Knockout of alpha globin regulatory element R2, biological replicate 1 [ATAC-seq]


title: Fetal liver Knockout of alpha globin Wild type litter mate replicate 2 [ATAC-seq]


title: Fetal liver Knockout of alpha globin regulatory element R1, biological replicate 2 [ATAC-seq]


title: F4


title: M2


title: M3


title: FG-MQ rep1 at 1 hr repeat

city: Frederiksberg C

first: Stefan

last: Olsson

address: 40 thorvaldsensvej

department: Plant and Environmental Sciences

country: Denmark

institute: University of Copenhagen

data_processing:
Illumina casava 1.8 used for base calling. The reads are filtered on instrument using the Illumina chastity filter.
FASTQC was used to assess the quality of the read sequences
FASTQ files were processed with FASTQ Groomer for compatibility with downstream softwares
Tophat for illumina (version 1.5.0) was used to map the sequenced reads to the Fusarium graminearum genome sequence v3 (BROAD website)
Cuffdiff (version 0.0.6) was used to identify genes that were differentially expressed
Genome_build: ASM24013v3; genome information and sequences downloaded from the BROAD website.
Supplementary_files_format_and_content: For each analysis, two tab-delimited text files as produced by cuffdiff is provided. The "gene read group tracking" file provide the reading for each of the triplicate samples (test and control), and the "differential expression testing" file provide information regarding the differential expression analysis. Gene expression level reading are in FPKM.

organism: Fusarium graminearum PH-1

source_name: 4 days liquid culture

growth_protocol: Fungal mycelia were cultured in 250 ml Erlenmeyer baffled flask with 50 ml of DFM (Yoder and Christianson, 1998, Fungal Genet Biol 23: 68-80) that was inoculated with 10 fungal plugs (approximately 8 mm x 8 mm). The flasks were shaken at 150 rpm for 3 days in the dark at 21oC. The resulting  fungal biomass was filtered through Miracloth and transferred into DFM without a nitrogen source for an overnight nitrogen depletion to recreate the nitrogen limiting environment of the soil. Three milliliters of the resulting culture was filtered through Miracloth, rinsed with sterile MilliQ water prior to treatment

extract_protocol:
Freeze dried cultures of Fusarium graminearum was crushed in mortar and pestle under liquid nitrogen. The pulverised culture was processed according to manufacturer's instructions using a QIAGEN RNeasy plant mini kit with on-column Dnase digestion to remove contaminated genomic DNA.
TruSeq RNA Sample Preparation Kit v2 was used and manufacturers instruction was followed using 1ug of total RNA. Libraries were size selected to 320 bp total size (giving ~200 bp insert after subtraction of adapter length).

treatment_protocol: Fungal culture was treated with 3 ml MAMPs (100 ng/ml flagellin [containing 100µg/ml sucrose] (tlrl-pstfla; Invivogen, CA, USA); 50 µg/ml LOS that was isolated from Xanthomonas campestris pv campestris strain 8004  (LPS without the O-antigen) (Silipo et al., 2005, J Biol Chem 280: 33660-33668), 50 µg/ml PGN that was isolated from Xanthomonas campestris pv campestris strain 8004)  (Erbs et al., 2008, Chem Biol 15: 438-448) that was dissolved in sterile MilliQ water or 3 ml sterile MilliQ water as the control. The treatments were performed in 13 ml Sarstedt tubes (Germany) with shaking in the dark at 21oC for 1, 2 and 4 hours.

tag: mamp treatment

value: Control MQ water

tag: additional info

value: Repeated in different lab


title: Fetal liver Knockout of alpha globin regulatory element Rm, biological replicate 1 [ATAC-seq]

value: regulatory element Rm

value: F1 C57Bl6 / CBA


title: FG-MQ rep1 at 2 hr repeat


title: FG-MQ rep3 at 1 hr repeat


title: FG-MQ rep2 at 1 hr repeat


title: Fetal liver Knockout of alpha globin regulatory element R1, biological replicate 1 [ATAC-seq]


title: FG-MQ rep2 at 2 hr repeat


title: FG-MQ rep3 at 2 hr repeat


title: Fetal liver Knockout of alpha globin regulatory element R4, biological replicate 1 [ATAC-seq]

value: regulatory element R4


title: FG-MQ rep2 at 4 hr repeat


title: Fetal liver Knockout of alpha globin regulatory element R4, biological replicate 2 [ATAC-seq]


title: U2OS_PHIP3F_gal_MACS2

city: Bethesda

first: Christophe

middle: E

last: Redon

state: MD

address: 37 Convent Drive

institute: NIH

description: ChipSeq for RepID/PHIP

data_processing:
For RepID ChIP, the MACS peak calling program was used comparing reads from the ChIP –seq experiment to genomic input reads from the same cell line, with default parameters, and a p-value = 1e-9.
For nascent strand peak calling the SICER program was used. Nascent strand reads were called by comparison to HCT116 genomic reads using a window size of 200bp, gap size of 600pb, and FDR of 0.01.
Illumina single read 35bp sequencing
Genome_build: hg19
Supplementary_files_format_and_content: Bed file format of replication initiation peaks and ChipSeq

source_name: Osteosarcoma

growth_protocol: Cancer cells were grown in DMEM medium, 10% FBS

extract_protocol:
DNA from asynchronous cell cultures was isolated, size fractionated and subject to exunuclease digestion as described in Aladjem et al., Science 281, 1005, 1998. and Wang et al., Molecular and Cellular Biology 24:3373-86, 2004. For ChipSeq, DNA molecules were enriched using a FLAG antibody. Enriched DNA molecules were then sequenced by Illumina Genome Analyzer IIx.
Illumina Genomic DNA protocol (Illumina Paired-END DNA Sample Prep Kit).

tag: cell line

tag: cell type

value: Cultured cancer cell line

tag: chip antibody

value: anti-Flag antibody (F3165, Sigma, Aldrich)


title: FG-FLG rep1 at 1 hr repeat

value: FLG


title: HCT116_RepID_KO_NS

description: HCT116 RepID KO nascent strand DNA

source_name: Colon carcinoma


title: HCT116.genomic.control

description: Control sample for both HCT116_WT_NS and HCT116_RepID_KO_NS  (used to call the peaks for both HCT116_WT_NS.sam.sorted.bam and HCT116_RepID_KO_NS.bam.sorted.bam)


title: HCT116_WT_NS

description: HCT116 wt nascent strand DNA


title: FG-FLG rep3 at 1 hr repeat


title: FG-FLG rep2 at 1 hr repeat


title: H3K36me3_OHT_2

city: Copenhagen

first: Karl

last: Agger

address: Ole Maaloes Vej 5

department: BRIC

data_processing:
ChIPSeq reads were aligned to the mm10 genome assembly using bowtie (Version 0.12.7) with the following settings:  -q -p 4 -S -n 2 -e 70 -l 28 --maxbts 800 -k 1 -a -m 1 --best --strata --phred33-quals.
Mapped reads were filtered for PCR dublicates using 'samtools rmdup' (Version 0.1.19) and coverted to bed format with 'bedtools bamtobed' (Version 2.25.0)
Peaks were called in Jmjda and Jmjd2c data set for Ctrl vs OHT, using EaSeq (Version 1.0) with the following settings: for Jmjd2a fold difference above 1, for Jmjd2c fold difference above 3.
Peak annotation was based on RefSeq database annotation (UCSC refFlat, mm10 genome assembly, 19 January 2015).
Genome_build: mm10
Supplementary_files_format_and_content: text files with called peaks for anti-Jmjd2a and anti-Jmjd2c. Bigwig files of genome-wide read coverage are available for all samples.

source_name: L-GMP transformed with MLL-AF9

growth_protocol: Cells were grown in RPMI containing 20% FBS and 10ng/ml IL-3

extract_protocol:
Lysates were clarified from sonicated nuclei and histone-DNA complexes were isolated with antibody.
ChIPSeq libraries were made using NEBNext® Ultra™ II DNA Library Prep Kit for Illumina® according to the manufactures protocol

treatment_protocol: To induce knockout cells were grown in the presence of 500nM OHT 4-hydroxytamoxifen for 96hr.

value: L-GMP transformed with MLL-AF9

value: anti-H3K36me3 (Cell Signaling Technology, #4909)


title: FG-FLG rep1 at 2 hr repeat


title: H3K36me3_Ctrl_2


title: H3K9me3_Ctrl_2

value: anti-H3K9me3 (Abcam ab8898)


title: H3K36me3_OHT_1


title: H3K36me3_Ctrl_1


title: FG-FLG rep2 at 2 hr repeat


title: FG-FLG rep3 at 2 hr repeat


title: FG-FLG rep1 at 4 hr repeat


title: H3K9me3_Ctrl_1


title: Jmjd2c_Ctrl

value: anti-Jmjd2c, ktwl


title: Jmjd2c_OHT


title: H3K9me3_OHT_1


title: FG-MQ rep1 at 4 hr repeat


title: FG-MQ rep3 at 4 hr repeat


title: Jmjd2a_OHT

value: anti-Jmjd2a (Cell Signaling Technology, #5328)


title: Jmjd2a_Ctrl


title: KH2.PRDM14

source_name: KH2 mESCs

value: KH2 embryonic stem (ES) cell line

value: PRDM14 (Homemade)


title: KH2.POU5F1

value: POU5F1 (Santa Cruz, catalog# sc-5279)


title: FG-FLG rep3 at 4 hr repeat


title: H3K9me3_OHT_2


title: KH2.WT.b3

data_processing:
Reads mapped to mm9 with BOWTIE -m40 -v2
reads were assigned to the ENSEMBL annotation using DEGseq
Genome_build: mm9
Supplementary_files_format_and_content: Raw read count matching the ENSEMBL annotation

growth_protocol: mESCs were grown in standard ESC medium containing Lif on a layer of feeder cells. Feeders were removed before TRIzol extraction.

extract_protocol:
RNA was purified from cells with TRIzol (Invitrogen)
PolyA+ RNA was isolated using Dynabeads Oligo(dT)25 (Invitrogen) beads and converted into dsDNA using the dUTP method (Parkhomchuk et al., 2009). DNA was end-repaired using End-It Repair Kit (Epicenter), tailed with an A using Klenow exo minus (NEB M0212), and ligated to custom adapters. Fragments of 300±100 bp were size-selected and subjected to ligation-mediated PCR amplification (LM-PCR) using Q5 polymerase (NEB M0530) to generate strand-specific libraries.

value: Wild Type


title: KH2.WT.b1


title: KH2.CBFA2T2

value: CBFA2T2 (Bethyl, catalog# A303-593A)


title: KH2.input


title: FG-FLG rep1 at 1 hr


title: FG-FLG rep3 at 1 hr


title: KH2.CBFKO.b3

value: CBFA2T2 Knockout


title: KH2.P14KO.b2

value: PRDM14 Knockout


title: FG-FLG rep2 at 2 hr


title: KH2.P14KO.b3


title: KH2.P14KO.b1


title: KH2.CBFKO.b1


title: FG-FLG rep3 at 2 hr


title: FG-FLG rep1 at 4 hr


title: FG-FLG rep1 at 2 hr


title: FG-FLG rep2 at 4 hr repeat

value: Repeated in different lab


title: unc-75 CLIPseq

organism: Caenorhabditis elegans

source_name: worms

growth_protocol: C.elegans expressing FLAG tagged UNC-75 under Pan neuron promoter were cultured at 20°C using standard procedures

extract_protocol:
Worms were lysed by sonication in Homogenization Buffer (100 mM NaCl, 25 mM HEPES, 250 μM EDTA, 2 mM DTT, 0.1% NP-40, 25 units/ml RNasin and Protease Inhibitors). Lysates were centrifuged at 16,000g for 15 min at 4°C and supernatants collected and incubated with M2 magnetic beads (Sigma) overnight on a rotator.
Samples were treated with micrococcal nuclease followed by phosphatase treatment, linker addition and 5’-labeling with 32P-γ-ATP. Complexes were eluted and separated with gel electrophoresis. A band corresponding to UNC-75/RNA complexes was cut out of the membrane and proteins degraded by Proteinase K. Associated RNAs were reverse-transcribed and amplified by PCR for deep sequencing.

treatment_protocol: Mixed stages of worms (young adults enriched) were washed and UV cross-linked.

tag: tissue

value: whole organism


title: FG-PGN rep1 at 1 hr

value: PGN


title: FG-FLG rep3 at 4 hr


title: FG-PGN rep2 at 1 hr


title: FG-FLG rep2 at 4 hr


title: KH2.CBFKO.b2


title: FG-PGN rep3 at 1 hr


title: FG-FLG rep2 at 1 hr


title: KH2.WT.b2


title: FG-PGN rep1 at 2 hr


title: FG-PGN rep3 at 2 hr


title: FG-PGN rep2 at 2 hr


title: FG-PGN rep2 at 4 hr


title: FG-PGN rep3 at 4 hr


title: FG-LOS rep1 at 1 hr

value: LOS


title: FG-LOS rep2 at 2 hr


title: FG-LOS rep3 at 1 hr


title: FG-LOS rep2 at 1 hr


title: FG-LOS rep1 at 2 hr


title: FG-LOS rep1 at 4 hr


title: FG-LOS rep2 at 4 hr


title: FG-LOS rep3 at 4 hr


title: FG-MQ rep2 at 1 hr

description:
MH1B2
Control sample in "R2_CD_H1XXX-MQ" files at 1hr

value: Control MQ water


title: FG-MQ rep3 at 1 hr

description:
MH1B3
Control sample in "R2_CD_H1XXX-MQ" files at 1hr


title: FG-MQ rep1 at 1 hr

description:
MH1B1
Control sample in "R2_CD_H1XXX-MQ" files at 1hr


title: FG-MQ rep1 at 2 hr

description:
MH2B1
Control sample in "R2_CD_H2XXX-MQ" files at 2hr


title: FG-MQ rep3 at 2 hr

description:
MH2B3
Control sample in "R2_CD_H2XXX-MQ" files at 2hr


title: FG-MQ rep2 at 2 hr

description:
MH2B2
Control sample in "R2_CD_H2XXX-MQ" files at 2hr


title: FG-MQ rep2 at 4 hr

description:
MH4B2
Control sample in "R2_CD_H4XXX-MQ" files at 4hr


title: FG-MQ rep1 at 4 hr

description:
MH4B1
Control sample in "R2_CD_H4XXX-MQ" files at 4hr


title: FG-MQ rep3 at 4 hr

description:
MH4B3
Control sample in "R2_CD_H4XXX-MQ" files at 4hr


title: G1 phase untreated, duplicate a

city: Shanghai

first: Zefeng

last: Wang

address: 320 Yueyang Road

institute: CAS-MPG Partner Institute for Computational Biology (PICB)

data_processing:
RNA-Seq reads were mapped to the human genome (build hg19) using the MapSplice informatics tool with default parameters
The mapped reads were further analyzed with Cufflinks to calculate the level of gene expression with FPKM
The levels of alternatively spliced isoforms were quantified with MISO (Mixture-of-Isoforms) probabilistic framework (Katz et al., 2010) using the annotated AS events for human hg19version 2.
The levels of alternatively spliced isoforms were also quantified with VAST-TOOLS using the event annotation as previously described
Genome_build: hg19

growth_protocol: Cells were cultured in humidified incubators with 5% CO2. Cell cycle synchronization was adapted from the protocol of Whitfield et al. (Whitfield et al., 2002); ~ 750,000 log phase HeLa cells were plated in 15 cm dishes in complete media and allowed to attach for 16 hrs, reaching < 30% confluence.

extract_protocol:
For RNA-Seq, cells (both adherent and detached) were harvested every 1.5 hrs for 30 hrs and frozen immediately for purification of total RNA.
RNA from synchronized cells was extracted with TRIZOL (Invitrogen), treated with DNAse I (Qiagen), and purified on RNAeasy columns. (Qiagen) according to the manufacturer’s protocol. RNA-seq libraries were robotically prepared with Illumina TruSeq Total RNA Sample Prep kits according to the manufacturer’s protocol

treatment_protocol: Cells were subsequently treated with 2 mM thymidine (Sigma) for a total of 18 hrs, washed 2 times with 1xPBS, and supplemented with fresh complete media for 10 hrs. 2 mM thymidine was subsequently added for a second block of 18 hrs and washed as described previously. Mitotic block was performed by double thymidine arrest (as above) and release in fresh media for 3.5 hrs followed by addition of nocodazole 100 μM (Sigma) for 10 hrs. G1 block was performed by serum starvation for 72 hrs in DMEM containing 0.05% FBS.

value: HeLa

tag: cell cycle

value: untreated


title: G2 phase (T=6 hour), DMSO treated control sample, duplicate b

value: DMSO


title: G2 phase (T=6 hour) KHCB treated sample , duplicate b

value: KHCB


title: G2 phase (T=6 hour) KHCB treated sample , duplicate a


title: G2 phase (T=6 hour) TG003 treated sample , duplicate b


title: G2 phase (T=6 hour), DMSO treated control sample, duplicate a


title: RNA-seq_CB_CD34+

tag: disease state

value: healthy

value: Umbilical cord blood

value: Hematopoietic stem/progenitor cells


title: FG-LOS rep3 at 2 hr


title: G1 phase untreated, duplicate  b

tag: treatment