title: mESC-4sU-TFEA-SSC

description: Remove the medium from plates and add mESC medium supplemented with 200 μM 4sU. The fresh medium with 4sU was changed every 4 h to enhance 4sU incorporation and mESCs were labeled for 24 hrs. After 4sU-labeling, the 4sU-containing medium was removed. Cells were harvested and fixed with methanol and stored at -80°C for future use. For sample rehydration, cells were taken out of -80°C freezer and kept on ice throughout the procedure. After cells were spun down at 1,000 g for 5 min at 4°C, methanol-PBS solution was removed, and cells were resuspended in 1 mL of rehydration buffer (SSC).  After cell counting, the single cell suspension was diluted to 100 cells/μL and immediately used for scNT-seq analysis.

value: Methanol fixation followed by rehydration (SSC); 200 μM 4sU, 24 h; TFEA/NaIO4


title: mESC-TFEA

status: Public on Jul 21 2020

type: SRA

city: Philadelphia

first: Peng

last: Hu

state: PA

address: 415 Curie Blvd, 545 CRB

department: Genetics

country: USA

institute: University of Pennsylvanian

description: Remove the medium from plates and add mESC medium supplemented without 4sU. Cells were harvested and used for scNT-seq analysis. For data analysis, down sampling was performed using “samtools view -s 0.73” to subsample the reads from bam files to match the sequencing depth with other conditions

library_source: transcriptomic

data_processing:
Library strategy: scNT-seq
Paired-end sequencing reads of scNT-Seq were processed as previously described (Hu et al., Molecular Cell, 2017) with some modifications. Briefly, each mRNA read (read2) was tagged with the cell barcode (bases 1 to 12 of read 1) and unique molecular identifier (UMI, bases 13 to 20 of read 1), trimmed of sequencing adaptors and poly-A sequences, and aligned using STAR v2.5.2a to the mouse (mm10, Gencode release vM13),  human genome (GRCh38, Gencode release v23), or a concatenation of the mouse and human (for the species mixing experiment) reference genome assembly. Both exonic and intronic reads mapped to predicted strands of annotated genes were retained for the downstream analysis. To qualify the labeled and unlabeled transcripts, uniquely mapped reads with mapping score > 10 were grouped by UMI barcodes in every cell and were used to determine the T > C substitution using sam2tsv (https://github.com/lindenb/jvarkit) , T > C substitutions with a base quality of Phred score > 27 were retained. For each experiment, locus with background T > C substitutions (detected in the sample without TFEA/NaIO4 treatment) was determined and was excluded for T > C substitution identification. After background correction of T > C substitution, a UMI was defined as labeled if there is a T > C substitution in any one of the reads belonging to that particular UMI. To this end, every UMI will be assigned to labeled or unlabeled based on the existence of T > C substitution. For each transcript, the total number of uniquely labeled (with T->C mutations) and unlabeled UMI sequences were counted and finally were assembled into 2 matrics using gene name as rows and cell barcode as columns.
Genome_build: Mouse (mm10) and human reference genome (GRCh38)
Supplementary_files_format_and_content: Two gene expression matrics containing labeled and unlabeled UMI count or total UMI count for each gene (rows) in each cell (columns), respectively

molecule: total RNA

organism: Mus musculus

source_name: Mouse embryonic stem cell

growth_protocol: Wild-type mESCs were initially cultured in presence of Mitomycin C inactivated mouse embryonic fibroblasts on 0.1% gelatin-coated (Millipore) 6-well-plates in Dulbecco’s Modified Eagle’s Medium (DMEM) (Gibco) supplemented with 15% fetal bovine serum (Gibco), 0.1 mM nonessential amino acid (Gibco), 1 mM sodium pyruvate (Gibco), 2 mM L-glutamine (Gibco), 50 μM 2-mercaptoethanol (Gibco), 1 μM MEK inhibitor PD0325901 and 3 μM GSK3 inhibitor CHIR99021 (Axon MedChem), and 1,000 U/mL LIF (Gemini Bio-Products). Human K562 cells (ATCC, CCL-243) were cultured in RPMI media supplemented with 10% FBS (Sigma, F6178).

extract_protocol:
After 4sU-labeling, K562 cells were pelleted and washed once with DPBS. mESCs were rinsed once with DPBS and dissociated with TrypLE-Express (Gibco, 12605010) for 5 min at 37°C. After neutralizing with culture medium, cells were pelleted at 1,000 rpm for 3 min. The cells were resuspended with DPBS-0.01% BSA. After cell counting with Countess II system, the single cell suspension was diluted to 100 cells/uL with DPBS containing 0.01% BSA and mixed at 1:1 ratio (mESCs:K562) for Drop-seq or scNT-seq analysis.
Single cells, polyA-containing barcoded beads (ChemGenes), and droplet generation oil (Bio-Rad) were each flowed into individual input ports on a microfluidic device (uFluidix). Cells were lysed and hybridized to beads in droplets. Beads were isolated and subjected to reverse transcription (Thermo Fisher) and exonuclease I (NEB) treatment, followed by PCR of full length transcripts (Kapa). Final libraries were created by tagmentation of cDNA, amplified by PCR (Illumina, Nextera xT DNA Library Preparation kit) and purified by SPRI beads. For scNT-seq, the beads were treated with either TimeLapse chemistry (TFEA) or SLAM-seq chemistry (IAA) to convert 4sU to cytidine-analog before reverse transcription. For second-strand synthesis, after exonuclease I treatment, the beads were washed once with TE-SDS buffer and twice with TE-TW buffer. After aspiration of TE-TW wash, the beads were resuspended in 500 μL 0.1 M NaOH and incubated for 5 min at room temperature with rotation. Then 500 μL 0.2 M Tris-HCl (pH 7.5) was added to neutralize the solution. The beads were washed once with TE-TW buffer and once with 10 mM Tris-HCl (pH 8.0), then 200 μL 2nd-strand synthesis mix (1x Blue buffer (Enzymatics), 4% Ficoll PM-400, 1 mM dNTPs (Clontech), 2.5 μM Template Switch Oligo-GAATG (TSO-GAATG: /5SpC3/AAGCAGTGGTATCAACGCAGAGTGAATG), 5 μM TSO-N9 (TSO-N9: /5SpC3/AAGCAGTGGTATCAACGCAGAGTGAAT(N1:25252525)(N1) (N1)(N1)(N1)(N1)(N1)(N1)(N1)), and 1.25 U/ μL Klenow exo- (Enzymatics)) were added [N1 stands for mixture of A,T,C,G at 25:25:25:25 ratio]. The reaction was incubated at room temperature for 10 min, followed by incubation at 37°C for 1 hour with end-over-end rotation.  The reaction was stopped by washing the beads once with TE-SDS buffer and twice with TE-TW buffer.

treatment_protocol: Before 4sU labeling experiments, J1 WT mESCs were passaged and cultured in feeder-free conditions for 48 hrs. For 4sU labeling, the mESC or K562 media was replaced with media supplemented with 4sU (100 μM) and cultured for 4 hours.

tag: cell line

value: J1 mESCs (ATCC, SCRC-1010)

tag: cell type

value: Mouse embryonic stem cell

tag: genotype

value: Wild-type

tag: Sex

value: male

tag: treatment

tag: datatype

value: UMI count matrix of total RNAs


title: mESC-4sU-TFEA-fresh

description: Remove the medium from plates and add mESC medium supplemented with 200 μM 4sU. The fresh medium with 4sU was changed every 4 h to enhance 4sU incorporation and mESCs were labeled for 24 hrs. After 4sU-labeling, the 4sU-containing medium was removed. Cells were harvested and used for scNT-seq analysis.

value: 200 μM 4sU, 24 h; TFEA/NaIO4

value: UMI count matrix of new (C) and old (T) RNAs


title: OT-rich bone CrtapKO rep3

city: Little Rock

first: Roy

last: Morello

state: AR

address: 4301 W. Markham St #505

department: Physiology and Biophysics

institute: University of Arkansas for Medical Sciences

data_processing:
Initial RNA sequencing data analysis was done using Illumina’s software portfolio including the BaseSpace (Illumina) analysis pipeline and the Tuxedo suite consisting of Bowtie, TopHat and Cufflinks to align sequence reads to the murine reference genome GRCm38, identify splice isoforms, and estimate transcript abundance.
Quantification, quality control, and differential expression analysis of the RNA sequencing data was performed using SeqMonk software (version 0.31.1) which runs R-based statistical analyses and uses Bioconductor packages (DESeq2).
Genome_build: GRCm38
Supplementary_files_format_and_content: TXT file with table of FPKM values for all genes and all samples

source_name: osteocyte-enriched cortical bone

growth_protocol: Mice were housed in a pathogen free facility with 12-hour light/dark cycle and unlimited access to water and standard chow diet. All animal work was conducted in accordance to local, state and U.S. federal regulations. Mice were euthanized at 3 months of age and relevant tissues were harvested according to the recommendations of the Guide for Care and Use of Laboratory Animals (8th Edition).

extract_protocol:
Femora and tibiae were harvested from WT, oim/oim, and CrtapKO mice at 3 months of age, N=4/genotype. Osteocyte-enriched bone was obtained by removing the epiphyses, thoroughly flushing the bone marrow, and scraping off the periosteal membrane; all these procedures were performed on ice in less than 10 minutes after the initial harvest. The osteocyte-enriched bone was immediately homogenized in 1 mL of TriPure Isolation Reagent (Roche REF 11667157001) and processed for RNA extraction according to the manufacturer’s instructions until the chloroform separation step. An equal volume of freshly-made, RNase-free 75% ethanol was added to the clear supernatant obtained from the chloroform separation, mixed, and then transferred to an RNeasy Micro column (Qiagen cat# 74004). Digestion with DNase I and elution of RNA was performed according to the manufacturer’s instructions. RNA samples were tested for quality using an Agilent 2100 Bioanalyzer. Only RNA samples with an RNA integrity number (RIN) of ≥7.0 were accepted for sequencing.
Sequencing libraries were prepared from the RNA samples by use of the TruSeq mRNA Sample Prep Kit (Illumina). Briefly, total RNA (1μg) was subjected to polyA selection, then chemically fragmented and converted to single-stranded cDNA using random hexamer-primed reverse transcription. The second strand was then generated to create double-stranded cDNA, followed by fragment end repair and addition of a single A base on each end of the cDNA. Adapters were ligated to each fragment end to enable attachment to the sequencing flow cell. The adapters also contained unique index sequences that allowed the libraries from different samples to be pooled and individually identified during downstream analysis. Library cDNA was PCR amplified and enriched for fragments containing adapters at each end to create the final cDNA sequencing library. Libraries were validated on the Fragment Analyzer for fragment size and quantified by use of a Qubit fluorometer (Life Technologies). Equal amounts of each library were pooled for sequencing on the NextSeq 500 platform using a high output flow cell to generate approximately 25 million 75-base reads per sample.

tag: strain

value: Crtap -/-

tag: tissue

value: femur and tibia

tag: mouse id

tag: age

value: 3 months

value: female


title: 1_01XZ_38693Tokyo_ISC-rep1_IRF2_mm10_i68

city: Tokyo

first: Taku

last: Sato

address: 1-5-45 Yushima Bunkyo-ku

department: Department of Biodefense Research

country: Japan

institute: Medical Research Institute, Tokyo Medical and Dental University (TMDU)

library_source: genomic

data_processing:
Sequences: Standard Illumina software base-calling and quality-control filtering was applied
Sequences (75 nt reads, single end) were aligned to the mouse genome (mm10) using the BWA algorithm (v0.7.12, default parameters).
Aligns were extended in silico at their 3’-ends to a length of 200 bp, which is the average genomic fragment length in the size-selected library, and assigned to 32-nt bins along the genome. The resulting histograms (genomic “signal maps”) were stored in bigWIG files.
Peak locations were determined using the MACS algorithm (v2.1.0) with a cutoff of pvalue = 1E-7.
Signal maps and peak locations were used as input data to Active Motifs proprietary analysis program, which creates excel tables containing detailed information on sample comparison, peak metrics, peak locations and gene annotations
Genome_build: mm10
Supplementary_files_format_and_content: BAM = standard BAM alignment format
Supplementary_files_format_and_content: BW = signal map in bigWIG format (UCSC)
Supplementary_files_format_and_content: BED = standard BED file containing MACS peaks

molecule: genomic DNA

source_name: Intestinal stem cells

extract_protocol:
Cells were fixed with 1% formaldehyde for 15 min and quenched with 0.125 M glycine.  Lysates were sonicated with a microtip sonicator to shear the DNA. Genomic DNA (Input) was prepared by treating aliquots of chromatin with RNase, proteinase K and heat for de-crosslinking, followed by ethanol precipitation. For each ChIP reaction, 30 ug of chromatin was precleared with protein A agarose beads (Invitrogen). ChIP reaction was set up using precleared chromatin and antibody and incubated overnight at 4 C. Protein A agarose beads were added and incubation at 4 C was continued for another 3 hr. Immune complexes were washed, eluted from the beads with SDS buffer, and subjected to RNase and proteinase K treatment. Crosslinks were reversed by incubation overnight at 65 C, and ChIP DNA was purified by phenol-chloroform extraction and ethanol precipitation.
Illumina sequencing libraries were prepared from the ChIP and Input DNAs by the standard consecutive enzymatic steps of end-polishing, dA-addition, and adaptor ligation. After a final PCR amplification step, the resulting DNA libraries were quantified and sequenced on NextSeq 500.

value: small intestine

tag: chip antibody

value: Anti-IRF2 (Abcam, ab124744)


title: mESC-TKO-rep1

data_processing:
Paired-end sequencing reads of scNT-Seq were processed as previously described (Hu et al., Molecular Cell, 2017) with some modifications. Briefly, each mRNA read (read2) was tagged with the cell barcode (bases 1 to 12 of read 1) and unique molecular identifier (UMI, bases 13 to 20 of read 1), trimmed of sequencing adaptors and poly-A sequences, and aligned using STAR v2.5.2a to the mouse (mm10, Gencode release vM13), human genome (GRCh38, Gencode release v23), or a concatenation of the mouse and human (for the species mixing experiment) reference genome assembly. Both exonic and intronic reads mapped to predicted strands of annotated genes were retained for the downstream analysis. To qualify the labeled and unlabeled transcripts, uniquely mapped reads with mapping score > 10 were grouped by UMI barcodes in every cell and were used to determine the T > C substitution using sam2tsv (https://github.com/lindenb/jvarkit) , T > C substitutions with a base quality of Phred score > 27 were retained. For each experiment, locus with background T > C substitutions (detected in the sample without TFEA/NaIO4 treatment) was determined and was excluded for T > C substitution identification. After background correction of T > C substitution, a UMI was defined as labeled if there is a T > C substitution in any one of the reads belonging to that particular UMI. To this end, every UMI will be assigned to labeled or unlabeled based on the existence of T > C substitution. For each transcript, the total number of uniquely labeled (with T->C mutations) and unlabeled UMI sequences were counted and finally were assembled into 2 matrics using gene name as rows and cell barcode as columns.
Genome_build: Mouse (mm10) and human reference genome (GRCh38)
Supplementary_files_format_and_content: Two gene expression matrics containing labeled and unlabeled UMI count for each gene (rows) in each cell (columns), respectively

growth_protocol:
4-thiouridine (4sU) (Alfa Aesar, J60679) were dissolved in DMSO to make 1 M stock. Before 4sU labeling experiments, WT and Tet-TKO mESCs were passaged and cultured in feeder-free conditions (0.1% gelatin-coated plates) for 48 hrs. For 4sU labeling in WT and Tet TKO mESC cultures, the medium was replaced with fresh mESC medium supplemented with 100 μM 4sU. After 4 h, mESCs were rinsed once with PBS and dissociated with TrypLE-Express (Gibco, 12605010) for 5 min at 37°C. After neutralizing with culture medium, cells were pelleted at 1,000 rpm for 3 min. After cell counting with the Countess II system, the single cell suspension was diluted to 100 cells/uL with DPBS containing 0.01% BSA for scNT-Seq analysis.
Wild-type mESCs were initially cultured in presence of Mitomycin C inactivated mouse embryonic fibroblasts on 0.1% gelatin-coated (Millipore) 6-well-plates in Dulbecco’s Modified Eagle’s Medium (DMEM) (Gibco) supplemented with 15% fetal bovine serum (Gibco), 0.1 mM nonessential amino acid (Gibco), 1 mM sodium pyruvate (Gibco), 2 mM L-glutamine (Gibco), 50 μM 2-mercaptoethanol (Gibco), 1 μM MEK inhibitor PD0325901 and 3 μM GSK3 inhibitor CHIR99021 (Axon MedChem), and 1,000 U/mL LIF (Gemini Bio-Products). Human K562 cells (ATCC, CCL-243) were cultured in RPMI media supplemented with 10% FBS (Sigma, F6178).

extract_protocol:
After 4sU-labeling, K562 cells were pelleted and washed once with DPBS. mESCs were rinsed once with DPBS and dissociated with TrypLE-Express (Gibco, 12605010) for 5 min at 37°C. After neutralizing with culture medium, cells were pelleted at 1,000 rpm for 3 min. The cells were resuspended with DPBS-0.01% BSA. After cell counting with Countess II system, the single cell suspension was diluted to 100 cells/uL with DPBS containing 0.01% BSA and mixed at 1:1 ratio (mESCs:K562) for Drop-seq or scNT-seq analysis.
Single cells, polyA-containing barcoded beads (ChemGenes), and droplet generation oil (Bio-Rad) were each flowed into individual input ports on a microfluidic device (uFluidix). Cells were lysed and hybridized to beads in droplets. Beads were isolated and subjected to reverse transcription (Thermo Fisher) and exonuclease I (NEB) treatment, followed by PCR of full length transcripts (Kapa). Final libraries were created by tagmentation of cDNA, amplified by PCR (Illumina, Nextera xT DNA Library Preparation kit) and purified by SPRI beads. For scNT-seq, the beads were treated with either TimeLapse chemistry (TFEA) or SLAM-seq chemistry (IAA) to convert 4sU to cytidine-analog before reverse transcription. For second-strand synthesis, after exonuclease I treatment, the beads were washed once with TE-SDS buffer and twice with TE-TW buffer. After aspiration of TE-TW wash, the beads were resuspended in 500 μL 0.1 M NaOH and incubated for 5 min at room temperature with rotation. Then 500 μL 0.2 M Tris-HCl (pH 7.5) was added to neutralize the solution. The beads were washed once with TE-TW buffer and once with 10 mM Tris-HCl (pH 8.0), then 200 μL 2nd-strand synthesis mix (1x Blue buffer (Enzymatics), 4% Ficoll PM-400, 1 mM dNTPs (Clontech), 2.5 μM Template Switch Oligo-GAATG (TSO-GAATG: /5SpC3/AAGCAGTGGTATCAACGCAGAGTGAATG), 5 μM TSO-N9 (TSO-N9: /5SpC3/AAGCAGTGGTATCAACGCAGAGTGAAT(N1:25252525)(N1) (N1)(N1)(N1)(N1)(N1)(N1)(N1)), and 1.25 U/ μL Klenow exo- (Enzymatics)) were added [N1 stands for mixture of A,T,C,G at 25:25:25:25 ratio]. The reaction was incubated at room temperature for 10 min, followed by incubation at 37°C for 1 hour with end-over-end rotation. The reaction was stopped by washing the beads once with TE-SDS buffer and twice with TE-TW buffer.

value: Tet1/2/3 triple knockout

value: 4sU labeling (100 μM 4hr) and TFEA treatment


title: mESC-chase-24h-rep2

growth_protocol:
Remove the medium from plates and add mESC medium supplemented with 200 μM 4sU. The fresh medium with 4sU was changed every 4 h to enhance 4sU incorporation and mESCs were labeled for 24 hrs. After 4sU-labeling, the 4sU-containing medium was removed and cells were washed once with DPBS. Then mESC medium containing 10 mM Uridine (Sigma, U6381) was added to the culture before cells were harvested at different time-points ranging from 0.5 hour to 24 hours. Cells were fixed with methanol as aforementioned and stored at -80°C for future use.
Wild-type mESCs were initially cultured in presence of Mitomycin C inactivated mouse embryonic fibroblasts on 0.1% gelatin-coated (Millipore) 6-well-plates in Dulbecco’s Modified Eagle’s Medium (DMEM) (Gibco) supplemented with 15% fetal bovine serum (Gibco), 0.1 mM nonessential amino acid (Gibco), 1 mM sodium pyruvate (Gibco), 2 mM L-glutamine (Gibco), 50 μM 2-mercaptoethanol (Gibco), 1 μM MEK inhibitor PD0325901 and 3 μM GSK3 inhibitor CHIR99021 (Axon MedChem), and 1,000 U/mL LIF (Gemini Bio-Products). Human K562 cells (ATCC, CCL-243) were cultured in RPMI media supplemented with 10% FBS (Sigma, F6178).

value: 4sU labeling (200 μM 24hr), Chase 24h, and TFEA treatment


title: mESC-WT-rep2


title: mESC-TKO-rep2


title: mESC-WT-rep1


title: OT-rich bone WT rep1

value: wildtype


title: mESC-chase-12h-rep2

value: J1 mESCs (ATCC, SCRC-1010)

value: 4sU labeling (200 μM 24hr), Chase 12h, and TFEA treatment


title: mESC-chase-6h-rep2

value: Mouse embryonic stem cell

value: Wild-type

value: 4sU labeling (200 μM 24hr), Chase 6h, and TFEA treatment


title: mESC-chase-12h-rep1

value: male


title: mESC-chase-24h-rep1

value: UMI count matrix of new (C) and old (T) RNAs


title: mESC-chase-0h-rep2

growth_protocol:
Remove the medium from plates and add mESC medium supplemented with 200 μM 4sU. The fresh medium with 4sU was changed every 4 h to enhance 4sU incorporation and mESCs were labeled for 24 hrs. After 4sU-labeling, the 4sU-containing medium was removed and cells were washed once with DPBS. Then mESC medium containing 10 mM Uridine (Sigma, U6381) was added to the culture before cells were harvested at different time-points ranging from 0.5 hour to 24 hours. Cells were fixed with methanol as aforementioned and stored at -80°C for future use. Two batches of experiment were performed.
Wild-type mESCs were initially cultured in presence of Mitomycin C inactivated mouse embryonic fibroblasts on 0.1% gelatin-coated (Millipore) 6-well-plates in Dulbecco’s Modified Eagle’s Medium (DMEM) (Gibco) supplemented with 15% fetal bovine serum (Gibco), 0.1 mM nonessential amino acid (Gibco), 1 mM sodium pyruvate (Gibco), 2 mM L-glutamine (Gibco), 50 μM 2-mercaptoethanol (Gibco), 1 μM MEK inhibitor PD0325901 and 3 μM GSK3 inhibitor CHIR99021 (Axon MedChem), and 1,000 U/mL LIF (Gemini Bio-Products). Human K562 cells (ATCC, CCL-243) were cultured in RPMI media supplemented with 10% FBS (Sigma, F6178).

value: 4sU labeling (200 μM 24hr), Chase 0min, and TFEA treatment


title: mESC-chase-1h-rep1

value: 4sU labeling (200 μM 24hr), Chase 60min, and TFEA treatment


title: mESC-chase-6h-rep1


title: mESC-chase-3h-rep1

value: 4sU labeling (200 μM 24hr), Chase 3h, and TFEA treatment


title: OT-rich bone WT rep4

value: femur and tibia

value: 3 months


title: mESC-chase-0.5h-rep2

value: 4sU labeling (200 μM 24hr), Chase 30min, and TFEA treatment


title: mESC-chase-1h-rep2


title: mESC-chase-0h-rep1


title: mESC-chase-0.5h-rep1


title: 2_026V_0082Takara_IntestinalStem_Input_mm10_i80

data_processing:
Sequences: Standard Illumina software base-calling and quality-control filtering was applied
Sequences (75 nt reads, single end) were aligned to the mouse genome (mm10) using the BWA algorithm (v0.7.12, default parameters).
Aligns were extended in silico at their 3’-ends to a length of 200 bp, which is the average genomic fragment length in the size-selected library, and assigned to 32-nt bins along the genome. The resulting histograms (genomic “signal maps”) were stored in bigWIG files.
Peak locations were determined using the MACS algorithm (v2.1.0) with a cutoff of pvalue = 1E-7.
Signal maps and peak locations were used as input data to Active Motifs proprietary analysis program, which creates excel tables containing detailed information on sample comparison, peak metrics, peak locations and gene annotations
Genome_build: mm10
Supplementary_files_format_and_content: BW = signal map in bigWIG format (UCSC)
Supplementary_files_format_and_content: BED = standard BED file containing MACS peaks

value: small intestine


title: 1_026I_0082Takara_IntestinalStem_IRF2_mm10_i81

value: Anti-IRF2 (Abcam, ab124744)


title: mESC-chase-3h-rep2


title: Cortical-KCl-0min

source_name: Primary mouse cortical cultures

growth_protocol:
Mouse cortices were dissected from embryonic day 16 (E16) C57BL/6 embryos of mixed sex (Charles River). Cortical neurons were dissociated with papain (Worthington) and plated on 6-well plates (at a density of ~600,000 cells/well) coated with poly-ornithine (30mg/mL, Sigma, P2533). Mouse cortical neuronal cultures were maintained in neurobasal media (Gibco, 21103049) supplemented with B27 (Gibco, 17504044), 2 mM GlutaMAX (Gibco, 35050061), and 1X Penicillin/streptomycin (Gibco, 15140122). After 4 days in vitro culture, primary cortical cultures were stimulated with a final concentration of 55mM potassium chloride (KCl) for various periods (0, 15, 30, 60, and 120 minutes). Before neuronal activation, the fresh media supplemented with 4sU (200 μM) was added. After 4sU labeling (2 h), the cells were washed with PBS once and were digested with 0.05% Trypsin-EDTA (Gibco, 25300054) for 20 min at 37°C, then replaced the buffer with 1 mL of DPBS and dissociated cells with a cell-scraper. After cell counting with Countess II system, the single cell suspension was diluted to 100 cells/μL with DPBS containing 0.01% BSA for scNT-Seq analysis.
Wild-type mESCs were initially cultured in presence of Mitomycin C inactivated mouse embryonic fibroblasts on 0.1% gelatin-coated (Millipore) 6-well-plates in Dulbecco’s Modified Eagle’s Medium (DMEM) (Gibco) supplemented with 15% fetal bovine serum (Gibco), 0.1 mM nonessential amino acid (Gibco), 1 mM sodium pyruvate (Gibco), 2 mM L-glutamine (Gibco), 50 μM 2-mercaptoethanol (Gibco), 1 μM MEK inhibitor PD0325901 and 3 μM GSK3 inhibitor CHIR99021 (Axon MedChem), and 1,000 U/mL LIF (Gemini Bio-Products). Human K562 cells (ATCC, CCL-243) were cultured in RPMI media supplemented with 10% FBS (Sigma, F6178).

tag: developmental stage

value: Primary mouse cortical cultures

value: mixed

value: 4sU labeling (200μM 2hr), No KCl induction (0 min), and TFEA treatment


title: Cortical-KCl-30min

value: 4sU labeling (200μM 2hr), 30min KCl induction, and TFEA treatment


title: Cortical-KCl-120min

value: 4sU labeling (200μM 2hr), 120min KCl induction, and TFEA treatment


title: Cortical-KCl-60min

value: Wild-type

value: 4sU labeling (200μM 2hr), 60min KCl induction, and TFEA treatment


title: HeLa_shINTS11_ctrl_r1_H3k36me3_ChIP-seq

city: Miami

first: Felipe

last: Beckedorff

state: FL

address: 1501 NW 10th Avenue room 731.G , Miami, FL

institute: University of Miami

data_processing:
Total RNA-seq and ChIP-seq: FASTQ data were processed with Trimmomatic v0.32 to remove adapters and low-quality reads. Small_RNA:  FASTQ data were processed with Cutadapt v1.14 adapter trimmed (AAAAAAA). Proseq: FASTQ data were trimmed by Cutadapt 1.14 and Trimmomatic v0.32.
ChIP-seq: reads were aligned to the hg19 human genome using STAR aligner v2.5.3a with default parameters. RNA-seq: reads were aligned to the hg19 human genome using STAR aligner v 2.5.3a with default parameters and RSEM v1.2.31 to obtain expected gene counts against the human Ensembl (release 87).
Genome_build: hg19
Supplementary_files_format_and_content: The processed data files are in bigWig format : ChIP-seq/Total_RNA-seq bigwigs were normalized by Count Per Million produced from DeepTools2.

organism: Homo sapiens

growth_protocol: HeLa stable containing INTS11 shRNA cell lines were maintained at 37°C with 5% CO2 and passaged every 2-3 days at a 1:3 split ratio, according to ATCC recommendations. DMEM medium was supplemented with 10% FBS and puromycin (2 μg/ml). Knockdowns were induced by adding of 1 μg/ml oxycycline (Dox) into the culture medium daily for 3 days.

extract_protocol:
ChIP-seq was performed as previously described (Gardini, A. et al. 2014, Lai, F. et a.l 2015, Yue, J.  et. al 2017) with slight modifications. 2 x 107 cells were crosslinked in 1% formaldehyde for 10 minutes at room temperature and quenched with 125 mM glycine. Samples were resuspended in ChIP lysis buffer (20 mM Tris-HCl, 50 mM NaCl, 0.1% SDS, 0.5% Triton-X-100, 1 mM EDTA) and sonicated in a Covaris M220 focused sonicator until chromatin fragments of 150-300 bp were produced. Protein A/G magnetic beads were bound with antibody and incubated with 1 mg of sonicated chromatin overnight. The following day, beads were washed twice with Mixed Micelle Buffer (150 mM NaCl, 1% Triton X-100, 0.2% SDS, 20 mM Tris-HCl, 5 mM EDTA, 65% sucrose), Buffer 500 (500 mM NaCl, 1% Triton X-100, 0.1% Na deoxycholate, 25 mM HEPES, 10 mM Tris-HCl, 1 mM EDTA), LiCl/detergent wash (250 mM LiCl, 0.5% Na deoxycholate, 0.5% NP-40, 10 mM Tris-HCl, 1 mM EDTA), and eluted in Decrosslinking Buffer (20 mM Tris-HCl, 300 mM NaCl, 0.5% SDS, 1 mM EDTA) at 65 °C. Eluates were treated with RNAse A for 1 hour at 37 °C and then Proteinase K for 3 hours at 56 °C. The DNA was finally purified by phenol/chloroform and eluted in 1x TE.
ChIP-seq libraries were generated using the NEBNext Ultra II DNA library prep kit (New England Biolabs, #E7645S) with at least 10 ng of input DNA.

value: HeLa

value: cervical adenocarcinoma

tag: modification

value: Tet-pLKO_shINTS11 stable cells

value: Histone H3k36me3 [Abcam#9050]


title: HeLa_shINTS11_ctrl_r2_H3k36me3_ChIP-seq


title: HeLa_shINTS11_ctrl_r1_H3k4me1_ChIP-seq

value: Histone H3K4me1 [Abcam#8895]


title: HeLa_shINTS11_dox_r1_H3k36me3_ChIP-seq


title: HeLa_shINTS11_dox_r1_H3k4me1_ChIP-seq


title: HeLa_shINTS11_dox_r1_H3k4me3_ChIP-seq

value: Histone H3k4me3 [Diagenode#pAb-003-050]


title: HeLa_shINTS11_dox_r2_H3k36me3_ChIP-seq


title: HeLa_shINTS11_ctrl_r1_H3k4me3_ChIP-seq


title: HeLa_shINTS11_dox_r2_RNAPII-Ser2_ChIP-seq

value: RPB1 Ser-2 [Abcam#5095]


title: HeLa_shINTS11_dox_r1_RNAPII-Ser2_ChIP-seq


title: HeLa_shINTS11_ctrl_r1_RNAPII-Ser2_ChIP-seq


title: HeLa_shINTS11_ctrl_r2_RNAPII-Ser2_ChIP-seq


title: HeLa_shINTS4_total_RNA_dox_r2

extract_protocol:
Total RNA was extracted using Trizol reagent (Thermo Fisher Scientific, #15596026) according to the manufacturer’s instructions. Genomic DNA was removed by Turbo DNAse treatment (Invitrogen, #AM1907).
Total RNA-seq libraries were produced using Stranded Total RNA library prep kit (Illumina, #20020596) with 500 ng of DNAse-treated Input RNA.

value: Tet-pLKO_shINTS4 stable cells


title: GBM14 control 2

status: Public on Jul 22 2020

type: RNA

city: Boston

first: Boston University

last: Microarray and Sequencing Resource

state: MA

address: 72 East Concord Street, E631

department: Microarray and Sequencing Resource

institute: Boston University

description: Gene expression data from GBM14 glioblastoma cell line treated with control.

hyb_protocol: The labeled, fragmented DNA was hybridized to the Affymetrix Human Gene 2.0 ST Array for 18 hours in a GeneChip Hybridization oven 640 at 45oC with rotation (60 rpm). The hybridized samples were washed and stained using an Affymetrix fluidics station 450.

scan_protocol: After staining, microarrays were immediately scanned using an Affymetrix GeneArray Scanner 3000 7G Plus.

data_processing: Raw Affymetrix CEL files were normalized to produce Entrez Gene-identifier-specific expression values using the implementation of the Robust Multiarray Average (RMA) in the affy Bioconductor package (version 1.36.1), using R version 2.15.1 and the Brainarray hugene20sthsentrezgcdf R package (version 23.0.0).

label: biotin

source_name: GBM14 glioblastoma cell line

label_protocol: Biotin labeling was performed using the WT Plus reagent kit (Affymetrix, Santa Clara, CA) according to the manufacturer's protocol.

growth_protocol: DMEM + 10%FBS + primocin

extract_protocol: RNA was isolated according to the manufacturer's instructions. RNA extraction kit was purchased from QIAGEN (miRNeasy Mini Kit: Cat #217004).

treatment_protocol: DMEM + 10%FBS + primocin + 1 µM crizotinib

value: glioblastoma

value: control


title: HeLa_shINTS9_total_RNA_dox_r2

value: Tet-pLKO_shINTS9 stable cells


title: HeLa_shINTS9_total_RNA_dox_r1


title: HeLa_shINTS4_total_RNA_dox_r1


title: Cortical-KCl-15min

value: Primary mouse cortical cultures

value: mixed

value: 4sU labeling (200μM 2hr), 15min KCl induction, and TFEA treatment

value: UMI count matrix of new (C) and old (T) RNAs


title: Pt_1_coll_1

city: Seattle

first: Nicole

middle: A P

last: Lieberman

state: WA

address: 850 Republican St S130

department: Laboratory Medicine

institute: University of Washington

data_processing:
Illumina Casava1.7 software used for basecalling.
Single end reads were adapter and quality trimmed by Trimmomatic v0.39, aligned to Ensembl v96 human transcriptome by Kallisto v0.46, and collapsed to gene level annotations by Biomart. Only samples with >500000 human reads were included for further analysis
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

source_name: Nasopharyngeal Swab

extract_protocol:
RNA was isolated from NP swabs in viral transport media using the Roche MagNAPure or Qiagen BioRobot
First strand cDNA was synthesized by SuperScriptIV/Random hexamers, second strand by Sequenase v2.0, and libraries constructed by Nextera XT Kit

value: male

tag: sample type

value: Nasopharyngeal Swab

tag: patient


title: Pt_2_coll_2


title: Pt_7_coll_1


title: Pt_2_coll_1


title: Pt_1_coll_2


title: TF-driven_differentiated_hiPSC 01_G09

city: Dresden

first: Volker

last: Busskamp

state: Saxony

address: Fetscherstraße 105

department: CMCB

country: Germany

institute: Technische Universität Dresden

data_processing:
Basecaller: bcl2fastq2 2.19.1
Aligner: STAR v2.5.4a was used with standard parameters.
featureCount was used for gene count quantification. Arguments: -t exon -g gene_id -a Homo_sapiens.GRCh38.94.gtf
Genome_build: hg38 release-94
Supplementary_files_format_and_content: tsv file reporting gene counts per sample, computed by featureCounts

molecule: polyA RNA

extract_protocol:
single cells  were FAC-sorted into a 96-well plate containing 2 μl of nuclease free water with 0.2% Triton-X  single cells were FAC-sorted into a 96-well plate containing 2 μl of nuclease free water with 0.2% Triton-X 100 and 4 U murine RNase Inhibitor (NEB)
Smart-seq2


title: TF-driven_differentiated_hiPSC 01_G02


title: TF-driven_differentiated_hiPSC 01_H06


title: TF-driven_differentiated_hiPSC 01_G04


title: TF-driven_differentiated_hiPSC 01_H05


title: TF-driven_differentiated_hiPSC 01_H08


title: TF-driven_differentiated_hiPSC 01_H03


title: TF-driven_differentiated_hiPSC 01_G07


title: TF-driven_differentiated_hiPSC 01_H04


title: TF-driven_differentiated_hiPSC 01_H07


title: TF-driven_differentiated_hiPSC 01_G10


title: TF-driven_differentiated_hiPSC 01_G12


title: TF-driven_differentiated_hiPSC 01_H11


title: TF-driven_differentiated_hiPSC 01_H12


title: TF-driven_differentiated_hiPSC 01_H02


title: TF-driven_differentiated_hiPSC 01_G05


title: TF-driven_differentiated_hiPSC 01_H09


title: TF-driven_differentiated_hiPSC 01_G11


title: TF-driven_differentiated_hiPSC 01_G03


title: TF-driven_differentiated_hiPSC 01_G06


title: TF-driven_differentiated_hiPSC 01_F12


title: Pt_7_coll_2


title: TF-driven_differentiated_hiPSC 01_F09


title: TF-driven_differentiated_hiPSC 01_F10


title: TF-driven_differentiated_hiPSC 01_H10


title: TF-driven_differentiated_hiPSC 01_F07


title: TF-driven_differentiated_hiPSC 01_G01


title: TF-driven_differentiated_hiPSC 01_G08


title: TF-driven_differentiated_hiPSC 01_F03


title: TF-driven_differentiated_hiPSC 01_F05


title: TF-driven_differentiated_hiPSC 01_F06


title: TF-driven_differentiated_hiPSC 01_E09


title: TF-driven_differentiated_hiPSC 01_E07


title: TF-driven_differentiated_hiPSC 01_E02


title: TF-driven_differentiated_hiPSC 01_F04


title: TF-driven_differentiated_hiPSC 01_E10


title: TF-driven_differentiated_hiPSC 01_E12


title: TF-driven_differentiated_hiPSC 01_E11


title: TF-driven_differentiated_hiPSC 01_E04


title: TF-driven_differentiated_hiPSC 01_E06


title: TF-driven_differentiated_hiPSC 01_F01


title: TF-driven_differentiated_hiPSC 01_D10


title: TF-driven_differentiated_hiPSC 01_E05


title: TF-driven_differentiated_hiPSC 01_E08


title: TF-driven_differentiated_hiPSC 01_D12


title: TF-driven_differentiated_hiPSC 01_D11


title: TF-driven_differentiated_hiPSC 01_E01


title: TF-driven_differentiated_hiPSC 01_D09


title: TF-driven_differentiated_hiPSC 01_D07


title: TF-driven_differentiated_hiPSC 01_F08


title: TF-driven_differentiated_hiPSC 01_F11


title: TF-driven_differentiated_hiPSC 01_D06


title: TF-driven_differentiated_hiPSC 01_D05


title: TF-driven_differentiated_hiPSC 01_F02


title: TF-driven_differentiated_hiPSC 01_E03


title: TF-driven_differentiated_hiPSC 01_D04


title: TF-driven_differentiated_hiPSC 01_D02


title: TF-driven_differentiated_hiPSC 01_D03


title: TF-driven_differentiated_hiPSC 01_C07


title: TF-driven_differentiated_hiPSC 01_D08


title: TF-driven_differentiated_hiPSC 01_C02


title: TF-driven_differentiated_hiPSC 01_B11


title: TF-driven_differentiated_hiPSC 01_C05


title: TF-driven_differentiated_hiPSC 01_D01


title: TF-driven_differentiated_hiPSC 01_B08


title: TF-driven_differentiated_hiPSC 01_C11


title: TF-driven_differentiated_hiPSC 01_C09


title: TF-driven_differentiated_hiPSC 01_C08


title: TF-driven_differentiated_hiPSC 01_C03


title: TF-driven_differentiated_hiPSC 01_B04


title: TF-driven_differentiated_hiPSC 01_C04


title: TF-driven_differentiated_hiPSC 01_B06


title: TF-driven_differentiated_hiPSC 01_B03


title: TF-driven_differentiated_hiPSC 01_C06


title: TF-driven_differentiated_hiPSC 01_B10


title: TF-driven_differentiated_hiPSC 01_C01


title: TF-driven_differentiated_hiPSC 01_B12


title: TF-driven_differentiated_hiPSC 01_B01


title: TF-driven_differentiated_hiPSC 01_A10


title: TF-driven_differentiated_hiPSC 01_A11


title: TF-driven_differentiated_hiPSC 01_A09


title: TF-driven_differentiated_hiPSC 01_C10


title: TF-driven_differentiated_hiPSC 01_A12


title: TF-driven_differentiated_hiPSC 01_B05


title: TF-driven_differentiated_hiPSC 01_C12


title: TF-driven_differentiated_hiPSC 01_B09


title: TF-driven_differentiated_hiPSC 01_B07


title: TF-driven_differentiated_hiPSC 01_B02


title: TF-driven_differentiated_hiPSC 01_A07


title: TF-driven_differentiated_hiPSC 01_A06


title: TF-driven_differentiated_hiPSC 01_A05


title: TF-driven_differentiated_hiPSC 01_A02


title: TF-driven_differentiated_hiPSC 01_A01


title: TF-driven_differentiated_hiPSC 01_A04


title: TF-driven_differentiated_hiPSC 01_A03


title: K562-4sU-TFEA-2ndSS

growth_protocol:
Human K562 cells (ATCC, CCL-243) were cultured in RPMI media supplemented with 10% FBS (Sigma, F6178). The culture media was replaced with media supplemented with or without 4sU (100 μM). After 4 h, cells were rinsed with PBS and harvested for scNT-Seq analysis. Second-strand synthesis method was incorporated in the library preparation.
Wild-type mESCs were initially cultured in presence of Mitomycin C inactivated mouse embryonic fibroblasts on 0.1% gelatin-coated (Millipore) 6-well-plates in Dulbecco’s Modified Eagle’s Medium (DMEM) (Gibco) supplemented with 15% fetal bovine serum (Gibco), 0.1 mM nonessential amino acid (Gibco), 1 mM sodium pyruvate (Gibco), 2 mM L-glutamine (Gibco), 50 μM 2-mercaptoethanol (Gibco), 1 μM MEK inhibitor PD0325901 and 3 μM GSK3 inhibitor CHIR99021 (Axon MedChem), and 1,000 U/mL LIF (Gemini Bio-Products). Human K562 cells (ATCC, CCL-243) were cultured in RPMI media supplemented with 10% FBS (Sigma, F6178).

value: K562 cells (ATCC, CCL-243)

value: Wild-type

value: female

value: 4sU labeling (100 μM 4hr) and TFEA treatment , and 2nd strand systhesis


title: TF-driven_differentiated_hiPSC 01_A08


title: M2B_K5E_SELEXseq_R1

city: Los Angeles

first: Remo

last: Rohs

state: CA

address: 1050 Childs Way

department: Molecular and Computational Biology

institute: University of Southern California

data_processing:
Library strategy: SELEX-seq
Sequences were trimmed to remove adapters using cutadapt 1.12 with Python 3.5.2 with the following command: cutadapt -u -23 -M 22 -m 22
Sequencing data from selection experiments was analyzed with a selex-seq protocol available under the Bioconductor package "SELEX"

source_name: in vitro selection experiment

extract_protocol: Sequencing library was constructd by PCR using primers complementary to regions found on the DNA sequence pool used for selection experiments

tag: selex round

tag: protein


title: M2B_R15G_SELEXseq_R1


title: M2B_R15G_SELEXseq_R2


title: K562-4sU-TFEA

growth_protocol:
Human K562 cells (ATCC, CCL-243) were cultured in RPMI media supplemented with 10% FBS (Sigma, F6178). The culture media was replaced with media supplemented with or without 4sU (100 μM). After 4 h, cells were rinsed with PBS and harvested for scNT-Seq analysis.
Wild-type mESCs were initially cultured in presence of Mitomycin C inactivated mouse embryonic fibroblasts on 0.1% gelatin-coated (Millipore) 6-well-plates in Dulbecco’s Modified Eagle’s Medium (DMEM) (Gibco) supplemented with 15% fetal bovine serum (Gibco), 0.1 mM nonessential amino acid (Gibco), 1 mM sodium pyruvate (Gibco), 2 mM L-glutamine (Gibco), 50 μM 2-mercaptoethanol (Gibco), 1 μM MEK inhibitor PD0325901 and 3 μM GSK3 inhibitor CHIR99021 (Axon MedChem), and 1,000 U/mL LIF (Gemini Bio-Products). Human K562 cells (ATCC, CCL-243) were cultured in RPMI media supplemented with 10% FBS (Sigma, F6178).

tag: cell line

value: 4sU labeling (100 μM 4hr) and TFEA treatment


title: K562-TFEA

value: TFEA treatment (without 4sU labeling)


title: M2B_K23R_SELEXseq_R2


title: M2B_K4E_SELEXseq_R1


title: M2B_K4E_SELEXseq_R2


title: M2B_K5E_SELEXseq_R2


title: M2B_SELEXseq_R0


title: K562-4sU

value: 4sU labeling (100 μM 4hr) without TFEA


title: M2B_K23R_SELEXseq_R1


title: K562-TFEA-2ndSS

value: TFEA treatment (without 4sU labeling) , and 2nd strand systhesis


title: K562-4sU-2ndSS

value: 4sU labeling (100 μM 4hr) without TFEA, and 2nd strand systhesis


title: NORM28A

city: Tampere

first: Mikko

last: Oittinen

address: Arvo Ylpön katu 34

department: Tampere Center for Child Health Research

country: Finland

institute: Tampere University

data_processing:
Raw data was de-multiplexed and FASTQ files for each sample were generated using the bcl2fastq software (Illumina inc.). FASTQ data were checked using the FastQC tool. Data analysis was performed using the QIAGEN GeneGlobe bioinformatics tool.
The UMI and cell index sequences are extracted from the R2 reads. A specific read structure is expected from the R2 reads. The expected R2 read structure from a NextSeq instrument is: <cell_index><UMI><ACG><poly-T>
If present, any 3’ poly-A region is trimmed off the 3’ end of the R1 reads. R1 reads with less than 25 base pairs are dropped.
The poly-A trimmed R1 reads from the above step are aligned to the reference genome using the splice aware aligner STAR. The reference genome used is human GRCh38, to which ERCC spike-in sequences are added. Gencode Release 23 for Human was used to annotate splice junctions when building the genome index with STAR. The same transcript models are used for annotating reads during UMI counting.
The aligned reads from the above step are annotated to gene regions. UMIs are then counted for each gene across different cell indices. Before counting UMIs, we drop the following reads: For targeted enrichment, any R1 read with <= 25 bp aligned after the primer region (i.e. endogenous); For targeted enrichment, any R1 read which does not begin with a primer sequence; For targeted enrichment, any R1 read for which the primer is not aligned to the design locus; For poly-A transcriptome, any R1 read which is mapped to multiple genome loci.
The analysis of differentially regulated genes was made using the R package DEseq2
Genome_build: GRCh38
Supplementary_files_format_and_content: umi_counts.txt; Tab delimited text file containing normalized UMI counts presenting transcription amount for all samples

source_name: duodenum small intestine

extract_protocol:
Around 100 sections were cut from each PAXgene-fixed paraffin block. RNA extraction was carried through PAXgene Tissue RNA kit (QIAGEN #765134) and QIAGEN QIAcube (#9001885) automated extracion system.
The library preparation was done using the QIAseq UPX 3’ Transcriptome Kit (QIAGEN). A total of 10ng purified RNA was converted into cDNA NGS libraries. During reverse transcription, each cell is tagged with a unique ID and each RNA molecule is tagged with a unique molecular index (UMI). Then RNA was converted to cDNA. The cDNA was amplified using PCR and during the PCR indices were added. After PCR the samples were purified. Library preparation QC was performed using TapeStation 4200 (Agilent) or Agilent Bioanalyzer. Based on quality of the inserts and the concentration measurements the libraries were pooled in equimolar ratios. The library pool(s) were quantified using qPCR. The library pool were then sequenced on a NextSeq500 sequencing instrument according to the manufacturer instructions.

value: duodenal biopsy

tag: disease state

value: healthy control

tag: diet

value: normal diet


title: NORM29B


title: NORM5B


title: 103-017EOS

value: coeliac disease

value: post gluten challenge


title: NORM7A


title: NORM8B


title: 103-008EOS


title: 103-001BL

value: gluten free diet


title: 102-026EOS


title: 102-026BL


title: 103-016EOS


title: 102-021BL


title: 102-007BL


title: 103-016BL


title: 102-021EOS


title: 103-017BL


title: 103-006EOS


title: 103-006BL


title: 103-008BL


title: 101-035EOS


title: 102-007EOS


title: 103-001EOS


title: 102-001BL


title: 102-001EOS


title: 101-027EOS


title: 101-035BL


title: 101-029EOS


title: 101-023EOS


title: 101-019EOS


title: 101-027BL


title: 101-019BL


title: 101-023BL


title: 101-002BL


title: 101-002EOS


title: 101-029BL


title: HeLa_shINTS11_dox_r3_RNAPII-Tyr1_ChIP-seq

data_processing:
Total RNA-seq and ChIP-seq: FASTQ data were processed with Trimmomatic v0.32 to remove adapters and low-quality reads. Small_RNA:  FASTQ data were processed with Cutadapt v1.14 adapter trimmed (AAAAAAA). Proseq: FASTQ data were trimmed by Cutadapt 1.14 and Trimmomatic v0.32.
ChIP-seq: reads were aligned to the hg19 human genome using STAR aligner v2.5.3a with default parameters. RNA-seq: reads were aligned to the hg19 human genome using STAR aligner v 2.5.3a with default parameters and RSEM v1.2.31 to obtain expected gene counts against the human Ensembl (release 87). ATAC-seq reads were aligned to the hg19 human genome using Bowtie2 v2.2.6.
ATAC-seq peaks were called using the MACS2.1 with the parameters: -g hs -p 0.01 --nomodel --shift -75 --extsize 150 and a cutoff of q-value <0.05.
Genome_build: hg19
Supplementary_files_format_and_content: The processed data files are in bigWig format : ChIP-seq/Total_RNA-seq/small_RNA-seq/CAGE-seq bigwigs were normalized by Count Per Million produced from DeepTools2. ATAC-seq: bigwigs weregenerate from NucleoATAC to define Nucleosome position.

value: HeLa

tag: cell type

value: cervical adenocarcinoma

tag: cell source

value: Tet-pLKO_shINTS11 stable cells

value: RPB1 Tyr1 [Active Motif  #61383]


title: NORM27A


title: HeLa_shINTS11_ctrl_r4_RNAPII-Tyr1_ChIP-seq


title: HeLa_shINTS11_dox_r4_RNAPII-Tyr1_ChIP-seq


title: HeLa_shINTS11_ctrl_r3_RNAPII-Tyr1_ChIP-seq


title: HeLa_shINTS11_ctrl_r2_SPT5_ChIP-seq

value: SPT5 [Millipore #MABE1803]


title: HeLa_shINTS11_dox_r1_NELF-E_ChIP-seq

value: NELF-E [Santa Cruz #sc-377052]


title: HeLa_shControl_dox_r1_ATAC-Seq

description: shControl

extract_protocol:
ATAC-seq experiments were performed as previously described (Corces,et al., 2017) with the following modification: 40,000 cells for each cell line were used to perform the transposon reaction and DNA libraries were quantified using Qubit dsDNA high sensitivity reagents (Thermo Fisher Scientific) and measured Qubit 2 fluorometer
ATAC-seq libraries were performed as previously described (Corces,et al., 2017)

value: Tet-pLKO_shControl stable cells


title: HeLa_shControl_ctrl_r2_ATAC-Seq

description: shControl


title: HeLa_shINTS11_ctrl_r1_NELF-E_ChIP-seq


title: HeLa_shControl_ctrl_r1_ATAC-Seq

description: shControl


title: HeLa_shINTS11_dox_r2_ATAC-Seq


title: HeLa_shControl_Total_RNA_dox_r1

description:
shGFP
processed data file: Total_RNA_shControl_expression_values.txt

growth_protocol: HeLa stable containing GFP shRNA cell lines were maintained at 37°C with 5% CO2 and passaged every 2-3 days at a 1:3 split ratio, according to ATCC recommendations. DMEM medium was supplemented with 10% FBS and puromycin (2 μg/ml). Knockdowns were induced by adding of 1 μg/ml oxycycline (Dox) into the culture medium daily for 3 days.


title: HeLa_shINTS11_dox_r1_SPT5_ChIP-seq


title: HeLa_shINTS11_dox_r1_ATAC-Seq


title: HeLa_E203Q_Proseq_dox_r1

data_processing:
Library strategy: PRO-seq
Proseq: FASTQ data were trimmed by Cutadapt 1.14 and Trimmomatic v0.32.
Proseq: Reads were aligned on hg19 or dm3 genome by bowtie 1.1.2 26. Strand-specific single nucleotide ends of aligned reads were generated by Bedtools v2.26 with genomecov as bedgraph format. Bedgraph data were normalized by the number of reads mapped to spike-in dm3 genome, and then converted to bigwig files.
Genome_build: hg19
Supplementary_files_format_and_content: The processed data files are in bigWig format: Proseq: bigwigs were with Bedtools v2.26 with genomecov.

source_name: HeLa

growth_protocol: HeLa stable containing INTS11 shRNA and overexpression E203Q mutant INTS11 cell lines were maintained at 37°C with 5% CO2 and passaged every 2-3 days at a 1:3 split ratio, according to ATCC recommendations. DMEM medium was supplemented with 10% FBS, puromycin (2 μg/ml) and G418 (200 μg/ml). Knockdowns were induced by adding of 1 μg/ml oxycycline (Dox) into the culture medium daily for 3 days

extract_protocol:
PRO-seq experiments were performed as described previously (Mahat, D. B. et al. 2016). Nuclei were isolated by Dounce homogenizer with loose pestle. 1x107 nuclei were subjected to nuclear run-on (30oC, 3min) in the presence of 25 μM Biotin-11-ATP/GTP/CTP/UTP (PerkinElmer). Total RNA was extracted and hydrolyzed in 0.2 M NaOH (on ice, 10 min).
PRO-seq  libraries used biotinylated nascent RNAs were purified by Dynabeads M-280 streptavidin (Invitrogen). Following adaptor ligation as described in previously (Mahat, D. B. et al. 2016), cDNA synthesis, and PCR amplification 140–350 bp libraries were size-selected by Pippin HT with 2% gel cassette 20B (Sage Science).


title: HeLa_E203Q_Proseq_r1


title: HeLa_WT_Proseq_dox_r1

description: WT

growth_protocol: HeLa stable containing INTS11 shRNA and overexpression WT INTS11 cell lines were maintained at 37°C with 5% CO2 and passaged every 2-3 days at a 1:3 split ratio, according to ATCC recommendations. DMEM medium was supplemented with 10% FBS, puromycin (2 μg/ml) and G418 (200 μg/ml). Knockdowns were induced by adding of 1 μg/ml oxycycline (Dox) into the culture medium daily for 3 days.


title: HeLa_WT_Proseq_dox_r2

description: WT


title: HeLa_E203Q_Proseq_r2


title: HeLa_WT_Proseq_ctrl_r1

description: WT


title: HeLa_WT_Proseq_ctrl_r2

description: WT


title: HeLa_shControl_Proseq_dox_r2

description: shGFP


title: HeLa_shINTS11_dox_r2_NELF-E_ChIP-seq


title: HeLa_shINTS11_ctrl_r2_NELF-E_ChIP-seq


title: HeLa_shINTS11_dox_r2_SPT5_ChIP-seq


title: HeLa_shINTS11_ctrl_r1_SPT5_ChIP-seq


title: HeLa_shControl_dox_r2_ATAC-Seq

description: shControl


title: HeLa_shControl_Proseq_dox_r1

description: shGFP


title: HeLa_shControl_Proseq_ctrl_r1

description: shGFP


title: HeLa_shINTS11_Proseq_dox_r1


title: HeLa_shControl_Proseq_ctrl_r2

description: shGFP


title: HeLa_shINTS11_Proseq_dox_r2


title: HeLa_shControl_Total_RNA_dox_r2


title: HeLa_shINTS11_ctrl_r2_INTS11_ChIP-seq

data_processing:
Total RNA-seq and ChIP-seq: FASTQ data were processed with Trimmomatic v0.32 to remove adapters and low-quality reads. Small_RNA:  FASTQ data were processed with Cutadapt v1.14 adapter trimmed (AAAAAAA). Proseq: FASTQ data were trimmed by Cutadapt 1.14 and Trimmomatic v0.32.
ChIP-seq: reads were aligned to the hg19 human genome using STAR aligner v2.5.3a with default parameters. RNA-seq: reads were aligned to the hg19 human genome using STAR aligner v 2.5.3a with default parameters and RSEM v1.2.31 to obtain expected gene counts against the human Ensembl (release 87). ATAC-seq reads were aligned to the hg19 human genome using Bowtie2 v2.2.6. Proseq: Reads were aligned on hg19 or dm3 genome by bowtie 1.1.2 26. Strand-specific single nucleotide ends of aligned reads were generated by Bedtools v2.26 with genomecov as bedgraph format. Bedgraph data were normalized by the number of reads mapped to spike-in dm3 genome, and then converted to bigwig files.
ATAC-seq peaks were called using the MACS2.1 with the parameters: -g hs -p 0.01 --nomodel --shift -75 --extsize 150 and a cutoff of q-value <0.05.
Genome_build: hg19
Supplementary_files_format_and_content: The processed data files are in bigWig format : ChIP-seq/Total_RNA-seq/small_RNA-seq/CAGE-seq bigwigs were normalized by Count Per Million produced from DeepTools2. Proseq: bigwigs were with Bedtools v2.26 with genomecov. ATAC-seq: bigwigs weregenerate from NucleoATAC to define Nucleosome position.


title: HeLa_shINTS11_Proseq_ctrl_r1


title: HeLa_shINTS11_ctrl_r1_INTS11_ChIP-seq

value: INTS11 [Sigma#HPA029025]


title: HeLa_shINTS11_dox_r1_RNAPII-Ser5_ChIP-seq

value: RPB1 Ser-5 [Abcam#5131]


title: HeLa_shINTS11_ctrl_r2_RNAPII-Ser5_ChIP-seq


title: HeLa_E203Q_Proseq_dox_r2


title: HeLa_shINTS11_ctrl_r1_RNAPII-Ser5_ChIP-seq


title: HeLa_shINTS11_ctrl_r2_RNAPII_ChIP-seq

value: RPB1 NTD [Cell Signaling Technology #14958]


title: HeLa_shINTS11_dox_r2_RNAPII_ChIP-seq


title: HeLa_shINTS11_ctrl_r2_ATAC-Seq

description:
shINTS11
atac_shints11_ctrl_merge_peaks.txt


title: HeLa_shINTS11_ctrl_r1_ATAC-Seq


title: HeLa_shINTS11_ctrl_r1_RNAPII_ChIP-seq


title: HeLa_shControl_CAGE_ctrl_r1

description:
Total RNA (CAGE)
shGFP
annotated_cluster_CAGE_shGFP_ctrl_toppeaks.bed

growth_protocol: HeLa stable containing GFP shRNA cell lines were maintained at 37°C with 5% CO2 and passaged every 2-3 days at a 1:3 split ratio, according to ATCC recommendations. DMEM medium was supplemented with 10% FBS and puromycin (2 μg/ml).

extract_protocol:
Total RNA was extracted using Trizol reagent (Thermo Fisher Scientific, #15596026) according to the manufacturer’s instructions. Genomic DNA was removed by Turbo DNAse treatment (Invitrogen, #AM1907).
CAGE library preparation were performed by DNAFORM (Kanagawa, Japan), the CAGE library prep is based on nAnTiCAGE protocol (Murata, M. et al. 2014)


title: HeLa_shINTS11_total_RNA_dox_r2

description:
shINTS11
Total_RNA_shINTS11_expression_values.txt


title: HeLa_shControl_CAGE_ctrl_r2


title: HeLa_shINTS11_total_RNA_dox_r1


title: HeLa_shINTS11_total_RNA_ctrl_r1


title: HeLa_shINTS11_total_RNA_ctrl_r2


title: HeLa_E203Q_nuclear_smRNA_dox_r1

molecule: nuclear RNA

extract_protocol:
Total RNA was extracted using Trizol reagent (Thermo Fisher Scientific, #15596026) according to the manufacturer’s instructions. Genomic DNA was removed by Turbo DNAse treatment (Invitrogen, #AM1907).
Small RNA libraries were prepared using the SMARTer smRNA-seq kit (Takara, #635030) with 750 ng of nuclear-enriched total RNA


title: HeLa_E203Q_nuclear_smRNA_dox_r2


title: HeLa_shINTS11_Proseq_ctrl_r2


title: HeLa_shINTS11_dox_r2_RNAPII-Ser5_ChIP-seq


title: HeLa_E203Q_nuclear_smRNA_r2


title: HeLa_WT_nuclear_smRNA_dox_r2

description: WT


title: HeLa_E203Q_nuclear_smRNA_r1


title: HeLa_WT_nuclear_smRNA_ctrl_r2

description: WT


title: HeLa_WT_nuclear_smRNA_ctrl_r1

description: WT


title: HeLa_WT_nuclear_smRNA_dox_r1

description: WT


title: HeLa_shControl_nuclear_smRNA_ctrl_r2

description: shGFP


title: HeLa_shControl_nuclear_smRNA_dox_r1

description: shGFP


title: HeLa_shControl_nuclear_smRNA_dox_r2

description: shGFP


title: HeLa_shINTS11_dox_r1_RNAPII_ChIP-seq


title: HeLa_shINTS11_nuclear_smRNA_ctrl_r2


title: HeLa_shINTS11_nuclear_smRNA_dox_r2


title: Mix-4sU-TFEA

data_processing:
Paired-end sequencing reads of Drop-seq/scNT-seq were processed using publicly available the Drop-Seq Tools v1.12 software. Briefly, each mRNA read (read2) was tagged with the cell barcode (bases 1 to 12 of read 1) and unique molecular identifier (UMI, bases 13 to 20 of read 1), trimmed of sequencing adaptors and poly-A sequences, and aligned using STAR v 2.5.2a to the mouse (mm10, Gencode release vM13). The intronic reads were also retained for downstream analysis. A custom Perl script was implemented in the Drop-Seq Tools pipeline to retrieve both exonic and intronic reads mapped to predicted strands of annotated genes. Uniquely mapped reads were grouped by cell barcodes. Cell barcodes were corrected for possible bead synthesis errors, using the DetectBeadSynthesisErrors program from the Drop-Seq Tools v1.12 software. To generate digital expression matrix, a list of UMIs in each gene (as rows), within each nucleus (as columns), was assembled, and UMIs within ED = 1 were merged together. The total number of unique UMI sequences was counted, and this number was reported as the number of transcripts of that gene for a given nucleus.
Genome_build: A concatenation of the mouse and human reference genome (mm10 and GRCh38)
Supplementary_files_format_and_content: Gene expression matrix containing read counts for each gene (rows) in each cell (columns)

organism:
Homo sapiens
Mus musculus

source_name: Cell lines (mESC & K562 mix)

extract_protocol:
After 4sU-labeling, K562 cells were pelleted and washed once with DPBS. mESCs were rinsed once with DPBS and dissociated with TrypLE-Express (Gibco, 12605010) for 5 min at 37Â°C. After neutralizing with culture medium, cells were pelleted at 1,000 rpm for 3 min. The cells were resuspended with DPBS-0.01% BSA. After cell counting with Countess II system, the single cell suspension was diluted to 100 cells/uL with DPBS containing 0.01% BSA and mixed at 1:1 ratio (mESCs:K562) for Drop-seq or scNT-seq analysis.
Single cells, polyA-containing barcoded beads (ChemGenes), and droplet generation oil (Bio-Rad) were each flowed into individual input ports on a microfluidic device (uFluidix). Cells were lysed and hybridized to beads in droplets. Beads were isolated and subjected to reverse transcription (Thermo Fisher) and exonuclease I (NEB) treatment, followed by PCR of full length transcripts (Kapa). Final libraries were created by tagmentation of cDNA, amplified by PCR (Illumina, Nextera xT DNA Library Preparation kit) and purified by SPRI beads. For scNT-seq, the beads were treated with either TimeLapse chemistry (TFEA) or SLAM-seq chemistry (IAA) to convert 4sU to cytidine-analog before reverse transcription. For second-strand synthesis, after exonuclease I treatment, the beads were washed once with TE-SDS buffer and twice with TE-TW buffer. After aspiration of TE-TW wash, the beads were resuspended in 500 uL 0.1 M NaOH and incubated for 5 min at room temperature with rotation. Then 500 uL 0.2 M Tris-HCl (pH 7.5) was added to neutralize the solution. The beads were washed once with TE-TW buffer and once with 10 mM Tris-HCl (pH 8.0), then 200 uL 2nd-strand synthesis mix (1x Blue buffer (Enzymatics), 4% Ficoll PM-400, 1 mM dNTPs (Clontech), 2.5 uM Template Switch Oligo-GAATG (TSO-GAATG: /5SpC3/AAGCAGTGGTATCAACGCAGAGTGAATG), 5 uM TSO-N9 (TSO-N9: /5SpC3/AAGCAGTGGTATCAACGCAGAGTGAAT(N1:25252525)(N1) (N1)(N1)(N1)(N1)(N1)(N1)(N1)), and 1.25 U/ uL Klenow exo- (Enzymatics)) were added [N1 stands for mixture of A,T,C,G at 25:25:25:25 ratio]. The reaction was incubated at room temperature for 10 min, followed by incubation at 37°C for 1 hour with end-over-end rotation. The reaction was stopped by washing the beads once with TE-SDS buffer and twice with TE-TW buffer.

tag: mouse sample

value: mESC-J1 (ATCC, SCRC-1010

tag: human sample

tag: genotype

value: Wild-type

tag: treatment

value: TFEA

tag: datatype

value: scNT-seq


title: HeLa_shINTS11_nuclear_smRNA_dox_r1

tag: chip antibody


title: Mix-4sU-IAA

value: IAA


title: MDA-MB-231, input

city: wuxi

first: ming

last: chen

address: zhongshan road

country: China

institute: nanjing medical university

data_processing:
Sequence reads were generated from Illumina HiSeq 4000, image analysis and base calling were performed using Off-Line Basecaller software (OLB V1.8.0).
After passing Solexa CHASTITY quality filter, the clean reads were aligned to Human genome (HG19) using BOWTIE software (V2.1.0).
Genome_build: HG19
Supplementary_files_format_and_content: The BED file containing peaks in each sample can be found from the *_peaks.bed file

source_name: breast cancer cells

extract_protocol:
The purity and concentration of DNA samples were determined with Qubit® Fluorometer.
DNA samples were end-repaired, A tailed, and adaptor ligated using TruSeq Nano DNA Sample Prep Kit (#FC-121-4002, Illumina), following the manufacturer’s instructions. ~200-1500 bp fragments were size selected using AMPure XP beads. The final size of the library was confirmed by Agilent 2100 Bioanalyzer.The samples were diluted to a final concentration of 8 pM and cluster generation was performed on the Illumina cBot using HiSeq 3000/4000 PE Cluster Kit (#PE-410-1001, Illumina), following manufacturer’s instructions.

tag: antibody


title: M2B_WT_SELEXseq_R0

data_processing:
Library strategy: SELEX-seq
Reads were trimmed with cutadapt v1.12 (cutadapt -u -54 -o) to obtain the variable region of reads +/- 3 bases on each side
Relative affinity and kmer enrichment were obtained using the SELEX package available under Bioconductor bioconductor.org/packages/SELEX
Supplementary_files_format_and_content: relative affinity tables were generated using the SELEX package

source_name: in vitro binding

extract_protocol:
Protein-DNA binding reactions were loaded onto a polyacrylamide gel and subject to electrophoretic mobility assay (EMSA), allowing for separation of bound and unbound fragments. Bound fragments were isolated from the gel and subject to phenol:chloroform extraction for DNA extraction, followed by ethanol precipitation.
Libraries were prepared for sequencing based on the Illumina TruSeq small RNA kit sequences. The DNA library 5’-GAGTTCTACAGTCCGACGATCCGC[N16]CCTGGAATTCTCGGGTGCCA-3’ containing  fixed regions compatible with Illumina sequencing was used in a 4-cycle PCR with primers RP1 5’AATGATACGGCGACCACCGAGATCTACACGTTCAGAGTTCTACAGTCCGA  and RPI# 5’CAAGCAGAAGACGGCATACGAGAT[Index]GTGACTGGAGTTCCTTGGCACCCGAGAATTCCA to add final Illumina adapter regions and indexes for sample multiplexing.

tag: round

tag: variable region

value: WT


title: HeLa_shINTS11_nuclear_smRNA_ctrl_r1

tag: modification


title: HeLa_shControl_nuclear_smRNA_ctrl_r1

description: shGFP


title: M2B_WT_SELEXseq_R2


title: M2B_WT_SELEXseq_R1


title: Cardiomyocytes_DNA_rep2-control treated_24hrs

city: Cincinnati

first: Mario

last: Medvedovic

state: OH

address: 3223 Eden Av. ML 56

department: Department of Environmental Health

institute: University of Cincinnati

description: DNA submitted for methyl-seq

data_processing:
Quality control of sequencing reads was performed using FastQC
Reads were trimmed with trim_galore in pair-end mode, steps included: removing sequencing artifacts, trimmed 5’ or 3’ ends with low scores, removed adaptor contamination, and filtered low quality reads.
The cleaned reads were then aligned to the Mus musculus reference genome (Genome Reference Consortium 37, mm10) using bismark.
Bismark_methylation_extractorwere used to infer the methylation state of all cytosines in a CpG, CHH, or CHG context and to compute the percentage of methylation for each CpG site.
BedGraph files were generated using bismark_methylation_extractor module of bismark.
Genome_build: mm10
Supplementary_files_format_and_content: BedGraph files were generated using bismark_methylation_extractor module of bismark.
Supplementary_files_format_and_content: cov file - Bismark Coverage report with chromosome, start position, end position, methylation Percentage, count Methylated, and count Unmethylated.

source_name: Cardiomyocytes

growth_protocol: Undifferentiated C57BL/6N-C2 ES cells were maintained in ES medium containing high-glucose Dulbecco’s minimal essential medium (DMEM; Gibcol Carlsbad, CA) supplemented with 15% ES cell qualified serum (Knockout Serum Replacement; Gibco), 2 mM glutamine, 1% nonessential amino acids, 1% sodium pyruvate, 100U/mL penicillin, 100 µg/mL streptomycin and 0.1 mM β-mercaptoethanol, and 1,000 U/mL ESGRO leukemia inhibitory factor (LIF; Bioscience Research Reagents, Temucula, CA). Cells were seeded on 0.1% gelatin-coated plates, incubated at 37OC (95% humidity, 5% CO2) and passaged every second or third day. The establishment and maintenance of the G418-resistant Nkx2.5-ES cell line containing the pNkx2.5PuroIRES2eGFP plasmid were performed as follows. Briefly, cells were transfected by the pNkx2.5PuroIRES2eGFP plasmid bearing as well a G418-resistance marker regulated by the CMV promoter, seeded on 0.1% gelatin-coated plates and ES medium, and allowed to attach for 24 hours before adding 600 µg/ml G418. The transfected cells were selected for 3-5 passages to insure all cells not resistant to G418 were removed. The media was then replaced with ES medium without G418. After 24 hours, cells were trypsinized and seeded into wells of gelatin-coated 6-well plates with 5x104 cells in ES medium. After 24 hours, ES medium was replaced with differentiation medium containing LIF-free DMEM supplemented with 15% non-ES qualified fetal bovine serum and subjected to differentiation treatment sequentially as follows: 150 ng/ml Noggin for 1 day, 5 ng/ml BMP4 for the next 2 days, and then 3 µg/ml puromycin for 4 days. Thereafter, Nkx2.5¬ positive cells were selected by treatment with 3µg/ml puromycin every 2-3 days and examined by microscopy until day 14 for the presence of beating cardiomyocytes.

extract_protocol:
DNA was extracted after each treatment timepoint using the DNeasy Blood & Tissue Kit (Qiagen, Hilden, Germany) according to the manufacturer’s instructions.
Methyl-seq was performed by Genomics, Epigenomics and Sequencing Core at the University of Cincinnati. SureSelect Methyl-Seq Target Enrichment System for mouse (Agilent, Santa Clara, CA) was used to prepare the sequencing library. A total of 1 μg high quality genomic DNA quantified by Qubit assay (Thermo Fisher, Waltham, MA) was sheared by Covaris S2 focused-ultrasonicator (Covaris, Woburn, MA) to about 200 bp under the recommended settings, and validated by 2100 Bioanalyzer (Agilent, Santa Clara, CA). Next, the DNA fragments were end prepared and ligated to the methylated adaptor. The size of the ligated libraries was then validated by Bioanalyzer, followed by hybridization with biotin-labeled RNA-baits to capture the regions where methylation is known to impact gene regulation. After hybridization, libraries were captured with streptavidin beads and bisulfite modified with EZ DNA Methylation-Gold kit (Zymo, Irvine, CA), and enriched with 8 cycles of PCR. These individually amplified libraries were labeled with unique indices by 6 cycles of PCR. After purification and size selection of the indexed libraries using AMPure XP beads (Beckman Coulter, Indianapolis, IN), the quality and quantity of the libraries were assessed by Bioanalyzer High Sensitivity DNA assay. To quantify the library concentration for the clustering generation, the libraries were qPCR analyzed by NEBNext Library Quant Kit (New England BioLabs, Ipswich, MA) using QuantStudio 5 Real-Time PCR System (Thermo Fisher, Waltham, MA). The sequencing was performed using Illumina Nextseq 550 sequencer under the sequencing setting of PE 2x150 bp to generate ~60M pass filter reads per sample.

treatment_protocol: Cardiomyocytes were treated with TCDD at a concentration of 5 nM, (a concentration commonly used for tissue culture studies of the high-affinity AHR of C57BL/6 mice), for 24, 72, and 96 hours. TCDD was dissolved in DMSO, which never exceeded 0.05% of the volume of medium in either treatment or vehicle control. Undifferentiated, untreated parental ES cells and differentiated untreated cardiomyocytes served as controls.

value: ES-derived cardiomyocytes

tag: strain

tag: time (hrs)

value: DMSO


title: Cardiomyocytes_DNA_rep2-TCDD treated_72hrs

description: DNA submitted for methyl-seq

value: TCDD


title: prostate-biopsy-replicate_RIB0006995

city: Leipzig

first: Kristin

last: Reiche

address: Perlickstr. 1

institute: Fraunhofer Institute for Cell Therapy and Immunology

description: Gene expression of prostate biopsy prior to RP (FFPE)

data_processing:
Processing of primary sequencing data was done using the workflow management software UAP (https://github.com/yigbt/uap).
We demultiplexed FASTQ files allowing for one mismatch with CASAVA 1.8.2. We clipped adapter sequences from the reads by utilizing AdapterRemoval v2.2.02 with parameters minquality 20 and minlength 20, respectively.
For de novo transcript assembly, we mapped all reads to the human genome version GRCh37/hg19 with TopHat v.2.0.8 setting RNA library type to fr-secondstrand and all other parameters to default values, and used cufflinks v.2.2.1 with default parameters and library type specification fr-secondstrand to assemble transcripts. We compared all assembled transcripts to Gencode v24 lifted to the human genome reference assembly GRCh37/hg19 [https://www.gencodegenes.org/human/release_24lift37.html] utilizing cuffmerge and cuffcompare with default parameters in order to identify novel transcripts. We received a comprehensive transcriptome annotation for prostate tissue for all subsequent analyzes by extending the Gencode annotation v24lift37 by all novel transcripts classified as “u” (unknown intergenic).
For gene expression quantification, we mapped reads to the human genome version GRCh37/hg19 excluding unannotated (Un) chromosomes and chromosome 6 MHC haplotypes using segemehl v.0.2.0. We utilized segemehl in split read mode (option -S) with additional parameters -H 1 -D 0 to report all alignments with no indels or mutations in the initial seed and passing the default minimal alignment accuracy. We used segemehl for expression quantification, since it outperforms TopHat in terms of sensitivity and specificity. Read counts for known (Gencode v24lift37) and novel transcripts contained in the comprehensive transcript assembly were determined using HTSeqCount v0.6.1 with parameters mode intersection-strict, stranded yes, idattr gene_id, and type exon.
Raw read counts were analyzed using the R-package DESeq2. Reads were normalized and variance stabilized (vst transformation).
The primary prognostic endpoint of the study was DoD, defined as the time from RP to the death because of PCa. For each gene, a univariate Cox regression was applied to standardized expression values for each cohort individually. The estimated log hazard ratio (logHR) and the respective standard error (SE) for each gene were passed to a meta-analysis using R-package meta v4.9-0. We applied the fixed effect model to determine the overall prognostic effect for each gene individually. We classified genes as prognostic markers using the tail area-based false discovery rate Fdr provided by fdrtool and selected all genes with Fdr<0.05.
Genome_build: GRCh37/hg19
Supplementary_files_format_and_content: Processed data files contain raw read counts (HTSeqCount v0.6.1 with parameters mode intersection-strict, stranded yes, idattr gene_id, and type exon) and gene identifiers according to the custom assembly (de-novo transcript assembly using cufflinks v.2.2.1 merged with Gencode v24 lifted to the human genome reference assembly GRCh37/hg19 [https://www.gencodegenes.org/human/release_24lift37.html]).

source_name: prostate biopsy

extract_protocol:
From up to five FFPE tissue slices of 10 µm we isolated total RNA following the truXTRAC FFPE RNA Kit (Covaris, Brighton, UK) instructions, using the Covaris M222 instrument for 5 min with duty factor 20 %, power 70 Watts, 200 cycles per burst at a temperature of 20°C. After subsequent proteinase K digestion and formalin-de-crosslinking, we transferred the samples onto RNeasy MinElute spin columns. We used the miRNeasy FFPE kit on the QIAcube following the manufacturer's instruction for the final RNA purification. To eliminate all traces of genomic DNA, we performed DNAse-digestion (TURBO DNA free Kit, Ambion, Austin, TX) twice in each sample. The RNA was concentrated using a standard precipitation protocol if necessary. Finally, we quantified extracted RNA using a Qubit RNA-Kit and the Qubit 2.0 instrument (Life Technologies, Carlsbad, CA). We analyzed the quality of RNA on a Bioanalyzer 2100 instrument and calculated the DV200 values (the percentage of fragments >200 nucleotides), ranging between 12 % and 29 % (mean = 20 %).
For RNA-sequencing of FFPE specimens we used 200 ng RNA per sample. We prepared RNA libraries using TruSeq-Stranded Total RNA Sample Prep kit (Illumina) including an rRNA depletion step according to manufacturers’ protocol, i.e. Ribo-Zero Gold rRNA Removal Kit. Following Illumina's recommendation, we skipped the fragmentation step for all samples. We analyzed the quality and quantity of each prepared library with the Qubit 2.0 instrument (Qubit DNA-Kit) and with the Bioanalyzer 2100, calculated the molarity of each library and pooled equal amounts which were used for subsequent sequencing (12 pM). We sequenced 2 x 126-bp paired-end reads using SBS V4 chemistry with a HiSeq 2500 (Illumina)

tag: gleason score

tag: gender

value: male

tag: time to dod in months


title: prostate-biopsy-replicate_RIB0006813


title: prostate-biopsy-replicate_RIB0006841


title: prostate-biopsy-replicate_RIB0006872


title: prostate-biopsy-replicate_RIB0006944


title: Cardiomyocytes_DNA_rep1-control treated_24hrs

description: DNA submitted for methyl-seq

tag: cell type


title: Cardiomyocytes_DNA_rep2-TCDD treated_24hrs

description: DNA submitted for methyl-seq


title: Cardiomyocytes_DNA_rep2-TCDD treated_96hrs

description: DNA submitted for methyl-seq


title: Cardiomyocytes_DNA_rep1-TCDD treated_96hrs

description: DNA submitted for methyl-seq


title: Cardiomyocytes_DNA_rep1-TCDD treated_72hrs

description: DNA submitted for methyl-seq


title: MDA-MB-231

tag: cell line


title: Cardiomyocytes_DNA_rep1-TCDD treated_24hrs

description: DNA submitted for methyl-seq


title: Mix-4sU

tag: mouse sample


title: prostate-biopsy-replicate_RIB0006640


title: ES cells_DNA_rep2

description: DNA submitted for methyl-seq

source_name: ES cells

value: ES cells

value: untreated


title: ES cells_DNA_rep1

description: DNA submitted for methyl-seq


title: prostate-biopsy-replicate_RIB0006646


title: Sequence3 (mixed samples)

first: Jes

last: Alexander

address: 3720 15th Ave NE, Foege Building S-210

department: Genome Sciences

description:
Mixed sample (non-demultiplexed):
No IR Brain 2 (Normal brain from a PDGF-driven mouse model of glioma)
No IR Brain 3 (Normal brain from a PDGF-driven mouse model of glioma)
IR 72hr Brain 3 (Normal brain from a PDGF-driven mouse model of glioma harvested at 72 hours after 10 Gy of ionizing radiation)
IR 72hr Tumor 2 (Tumor from a PDGF-driven mouse model of glioma harvested at 72 hours after 10 Gy of ionizing radiation)

Processed data files:
MouseGliomaCDS.rds
MouseGliomaCells.tsv
MouseGliomaGenes.tsv
MouseGliomaMatrix.mtx
Sequence3Barcodes.tsv

data_processing:
Demultiplexing was performed using bcl2fastq v2.16.  Barcodes used are in Sequence1Barcodes.tsv for Sequence1_S0_R1_001.fastq.gz and Sequence1_S0_R2_001.fastq.gz, Sequence2Barcodes.tsv for Sequence2_S0_R1_001.fastq.gz and Sequence2_S0_R2_001.fastq.gz, Sequence3Barcodes.tsv for Sequence3_S0_R1_001.fastq.gz and Sequence3_S0_R2_001.fastq.gz, and Sequence4Barcodes.tsv for Sequence4_S0_R1_001.fastq.gz and Sequence4_S0_R2_001.fastq.gz.
Poly-A tails and adaptors were clipped using trim_galore v 0.4.1
Using STAR v2.5.2b, trimmed reads were aligned to a reference genome including human (hg19) and mouse (mm10) genomes with annotations from GENCODE V19 for human and VM11 for mouse
Reads not mapping uniquely and duplicates based on unique molecular index (UMIs), reverse transcription index, and rear 2 end coordinate were removed.
Reads were mapped to cells by further demultiplexing using the reverse transcription index (RTBarcodes.tsv).
A digital gene expression matrix was created by counting, in a strand-specific manner, the number of unique sequences based on UMIs that map to either exonic or intronic regions of each gene using bedtools (version 2.26.0)
RCAS virus reads were added back to the matrix by first aligning previously unaligned reads to the RCAS LTR reference sequence using bowtie v2.2.3
Aligned reads were deduplicated based on the UMIs, reverse transcription index, and rear 2 end coordinates.
Counts were then added for the RCAS LTR transcript (RCAS, ENSMUSG00000000LTR.1) to the appropriate cell in the digital gene expression matrix based on the barcodes.
Cells with < 90% of UMIs mapping to a single species and cells with < 100 UMIs total were filtered out.
Genome_build: hg19 and mm10, RCAS LTR: >RCAS_ASLV_LTR  AATGTAGTCTTATGCAATACTCTTGTAGTCTTGCAACATGGTAACGATGAGTTAGCAACATGCCTTACAAGGAGAGAAAAAGCACCGTGCATGCCGATTGGTGGAAGTAAGGTGGTACGATCGTGCCTTATTAGGAAGGCAACAGACGGGTCTGACATGGATTGGACGAACCACTGAATTCCGCATTGCAGAGATATTGTATTTAAGTGCCTAGCTCGATACAATAAACGCCATTTGACCATTCACCACA
Supplementary_files_format_and_content: MouseGliomaCDS.rds: R data file (.rds) containing a Monocle 3alpha CellDataSet Object with labeled mouse glioma data after removing cells with <= 100 transcripts and < 90% mouse transcripts.
MouseGliomaCells.tsv: Tab-separated value (.tsv) file containing the names of cells (i.e., barcodes representing cells) for the mouse glioma dataset.
MouseGliomaGenes.tsv: Tab-separated value (.tsv) file containing the Ensembl IDs for the mouse glioma dataset.
MouseGliomaMatrix.mtx: Matrix Market (.mtx) file containing transcript counts for the mouse glioma dataset (cells are columns and Genes are rows).
BarnyardCDS.rds: R data file (.rds) containing a Monocle 3alpha CellDataSet Object with labeled 293T/NIH3T3 mixed cell data after removing cells with <= 100 transcripts.
BarnyardCells.tsv: Tab-separated value (.tsv) file containing the names of cells (i.e., barcodes representing cells) for the barnyard (293T/NIH3T3 mix) dataset.
BarnyardGenes.tsv: Tab-separted value (.tsv)  file containing the Ensembl IDs for the barnyard (293T/NIH3T3 mix) dataset.
BarnyardMatrix.mtx: Matrix Market (.mtx) file containing transcript counts for the  barnyard (293T/NIH3T3 mix) dataset.  Cells are columns and Genes are rows.

source_name: Brain

growth_protocol: Ink4a/Arf-/- mice were injected intracranially as pups with RCAS-PDGFB virus to generate glioma-like tumors.

extract_protocol:
Nuclei were extracted and fixed using the single-cell combinatorial indexed RNA sequencing protocol (Cao et al. Science. 2017. 357:661-7.)
Library construction was as per the single-cell combinatorial indexed RNA sequencing protocol (Cao et al. Science. 2017. 357:661-7.)

treatment_protocol: At symptom onset, tumor and normal brain was immediately harvested or mice were either irradiated with 10Gy of ioninzing radiation (X-rays) and tumor and normal brain was harvested at 8 +/- hours or 72 hours after radiation.

tag: tissue

value: mixed sample (Normal Brain, Tumor)

value: mixed sample (IR, No IR)

tag: hours after ionizing radiation (ir)

value: mixed sample (72hr after IR, No IR)


title: prostate-biopsy-replicate_RIB0006633


title: prostate-biopsy-replicate_RIB0006637


title: prostate-biopsy-replicate_RIB0024945


title: prostate-biopsy-replicate_RIB0024973


title: prostate-biopsy-replicate_RIB0006936


title: prostate-biopsy-replicate_RIB0006663


title: prostate-biopsy-replicate_RIB0006919


title: DM-RNA-IP

city: Birmingham

first: Anna

middle: E

last: Thalacker-Merc er

state: Alabama

address: 1720 University Blvd

institute: University of Alabama at Birmingham

description:
3d Differentaition Replicate 1
DM-RNA-IP-peaks.txt

data_processing:
The 5’, 3’-adaptor trimmed reads were aligned to the mm10 reference genome using HISAT2 (v2.1.0).
ExomePeak was used for MeRIP peak calling and statistically significant MeRIP enriched regions (MeRIP peaks) were identified at the sample level with a significance threshold of P≤0.05
ExomePeak was also used to compare transcripts which were differentially m6A-modified between the GM and 3d DM samples with significance set at P≤0.05
For RNA-seq - The input control RNA-sequencing results were subjected to differential gene expression analysis using edgeR
Genome_build: mm10
Supplementary_files_format_and_content: tab-delimited text files include TMM values for RNA-seq output and peak values for MeRIP-seq

source_name: C2C12 myoblasts

growth_protocol: C2C12 myoblasts were seeded on collagen (Type 1 rat tail) coated dishes.

extract_protocol:
RNA was isolated using an Omega E.Z.N.A.® Total RNA Kit I. mRNA was isolated using Arraystar Seq-StarTM poly(A) mRNA Isolation Kit .
Lysates were clarified from sonicated nuclei and histone-DNA complexes were isolated with antibody.
The KAPA Stranded mRNA-seq Kit (Illumina, San Diego, CA, USA) was used to prepare the RNA-seq libraries for m6A immunoprecipitated mRNA and input control mRNA libraries. Quality of the libraries was determined using an Agilent 2100 Bioanalyzer. The HiSeq 3000/4000 PE Cluster Kit (Illumina, San Diego, CA, USA) was used to generate clusters from the prepared libraries following the manufacturer’s instructions. The clustered libraries were sequenced on an Illumina HiSeq 4000 system.
MeRIP-seq or RNA-seq

treatment_protocol: Proliferation samples were taken when C2C12 myoblasts were 50% confluent and maintained in a medium of high glucose DMEM supplemented with 10% fetal bovine serum. 3d Differentiation samples were taken after C2C12 myoblasts had reached >90% confluence and were switched to a high glucose DMEM supplemented with 2% horse serum for 3 days.

tag: m6a antibody

value: anti-m6A (Cat:202 003, Synaptic Systems, Goettingen, Germany)


title: DM-IP-2

description:
3d Differentiation Replicate 2
DM-IP_2-peaks.txt


title: GM-IP-2

description:
Proliferation Replicate 2
GM-IP_2-peaks.txt


title: prostate-biopsy-replicate_RIB0006912


title: GM-RNA-IP

description:
Proliferation Replicate 1
GM-RNA-IP-peaks.txt


title: DM-Input-2

description:
3d Differentiation Replicate 2
normalizedCounts_TMM.txt


title: prostate-biopsy-replicate_RIB0006999


title: GM-RNA-Input

description:
Proliferation Replicate 1
normalizedCounts_TMM.txt


title: GM-Input-2

description:
Proliferation Replicate 2
normalizedCounts_TMM.txt


title: FFPE prostate-tissue-H-dt-replicate_RIB0034918

description: Gene expression of prostate RP tissue specimen (FFPE)

source_name: prostate tissue (FFPE)

extract_protocol:
From up to five FFPE tissue slices of 10 µm we isolated total RNA following the truXTRAC FFPE RNA Kit (Covaris, Brighton, UK) instructions, using the Covaris M222 instrument for 5 min with duty factor 20 %, power 70 Watts, 200 cycles per burst at a temperature of 20°C. A pathologist evaluated HE-stained flanking tissue slices and only FFPE specimen with high tumor content underwent RNA isolation. After subsequent proteinase K digestion and formalin-de-crosslinking, we transferred the samples onto RNeasy MinElute spin columns. We used the miRNeasy FFPE kit on the QIAcube following the manufacturer's instruction for the final RNA purification. To eliminate all traces of genomic DNA, we performed DNAse-digestion (TURBO DNA free Kit, Ambion, Austin, TX) twice in each sample. The RNA was concentrated using a standard precipitation protocol if necessary. Finally, we quantified extracted RNA using a Qubit RNA-Kit and the Qubit 2.0 instrument (Life Technologies, Carlsbad, CA). We analyzed the quality of RNA on a Bioanalyzer 2100 instrument and calculated the DV200 values (the percentage of fragments >200 nucleotides), ranging between 12 % and 29 % (mean = 20 %).
For RNA-sequencing of FFPE specimens we used 200 ng RNA per sample. We prepared RNA libraries using TruSeq-Stranded Total RNA Sample Prep kit (Illumina) including an rRNA depletion step according to manufacturers’ protocol, i.e. Ribo-Zero Gold rRNA Removal Kit. Following Illumina's recommendation, we skipped the fragmentation step for all samples. We analyzed the quality and quantity of each prepared library with the Qubit 2.0 instrument (Qubit DNA-Kit) and with the Bioanalyzer 2100, calculated the molarity of each library and pooled equal amounts which were used for subsequent sequencing (12 pM). We sequenced 2 x 126-bp paired-end reads using SBS V4 chemistry with a HiSeq 2500 (Illumina)

tag: risk group

value: H-dt


title: DM-RNA-Input

description:
3d Differentaition Replicate 1
normalizedCounts_TMM.txt


title: FFPE prostate-tissue-L-replicate_RIB0034489

value: L


title: FFPE prostate-tissue-Md-replicate_RIB0034731

value: Md


title: FFPE prostate-tissue-H-dt-replicate_RIB0034385


title: Sequence4 (mixed samples)

description:
Mixed sample (non-demultiplexed):
No IR Brain 2 (Normal brain from a PDGF-driven mouse model of glioma)
No IR Brain 3 (Normal brain from a PDGF-driven mouse model of glioma)
IR 72hr Brain 3 (Normal brain from a PDGF-driven mouse model of glioma harvested at 72 hours after 10 Gy of ionizing radiation)
IR 72hr Tumor 2 (Tumor from a PDGF-driven mouse model of glioma harvested at 72 hours after 10 Gy of ionizing radiation)

Processed data files:
MouseGliomaCDS.rds
MouseGliomaCells.tsv
MouseGliomaGenes.tsv
MouseGliomaMatrix.mtx
Sequence4Barcodes.tsv

tag: treatment


title: FFPE prostate-tissue-Md-replicate_RIB0034485


title: FFPE prostate-tissue-H+dt-replicate_RIB0034370

value: H+dt


title: FFPE prostate-tissue-V-replicate_RIB0034483

value: V


title: FFPE prostate-tissue-H+st-replicate_RIB0034303

value: H+st


title: FFPE prostate-tissue-H+st-replicate_RIB0034830


title: FFPE prostate-tissue-H+dt-replicate_RIB0034827


title: prostate-tissue-H+st-replicate_RIB0034235

description: Gene expression of prostate RP tissue specimen

source_name: prostate tissue

extract_protocol:
Stacks of consecutive cryosections were generated from frozen RP tissue samples and immediately transferred to 1ml Qiazol (Qiagen, Hilden, Germany). A pathologist evaluated HE-stained cryosections flanking the stacks concerning tumor and stroma cell content. Only PCa tissue stacks, which were flanked on either side by sections containing at least 50% tumor cells underwent RNA isolation. In case of insufficient RNA quantity, we combined multiple tissue stacks with suitable tumor cell content. We isolated total RNA using the miRNeasy Mini Kit on the QIAcube (Qiagen) followed by manual DNase I digestion. After determining the RNA concentration with a NanoDrop 1000 (Peqlab, Erlangen, Germany), we verified the RNA integrity on an Agilent Bioanalyzer 2100 (Agilent Technologies, Palo Alto, CA). RNA samples with an RNA-Integrity-Number (RIN) of at least six passed for further processing.
We depleted 1 µg of total RNA of ribosomal RNA using the Ribo-Zero rRNA Removal Kit (Epicentre, Madison, WI) and processed 50 ng of rRNA-depleted RNA with ScriptSeq v2 RNA-Seq Library Preparation Kit (Epicentre). After ten cycles of PCR, to incorporate index barcodes for sample multiplexing and to amplify the cDNA libraries, we determined the quality and concentration of the amplified libraries using the Bioanalyzer 2100. 4 ng each of 8 samples were pooled and size-selected on 2% agarose gels using agarose gel electrophoresis. We gel-excised the sample ranging between 150 bp and 600 bp and purified it with the MinElute Gel Extraction Kit (Qiagen), according to manufacturer's instructions. To quantify the purified libraries we used the Bioanalyzer 2100. Then we loaded every purified and size-selected library pool onto an Illumina HiSeq2000 flow cell, distributing it among all lanes. Finally, we performed sequencing with 2 x 101-bp paired-end reads using SBS V3 chemistry on a HiSeq2000 (Illumina, Inc, San Diego, CA).


title: prostate-tissue-V-replicate_RIB0034299


title: prostate-tissue-H+dt-replicate_RIB0034276


title: prostate-tissue-H-dt-replicate_RIB0034239


title: prostate-tissue-L-replicate_RIB0034258


title: FFPE prostate-tissue-H+st-replicate_RIB0034434


title: FFPE prostate-tissue-L-replicate_RIB0034486


title: prostate-tissue-L-replicate_RIB0034271


title: FFPE prostate-tissue-Ms-replicate_RIB0034702

value: Ms


title: prostate-tissue-H-dt-replicate_RIB0034918


title: prostate-tissue-Ms-replicate_RIB0034214


title: prostate-tissue-Md-replicate_RIB0034731


title: prostate-tissue-Ms-replicate_RIB0034497


title: prostate-tissue-H+dt-replicate_RIB0034913


title: prostate-tissue-H-dt-replicate_RIB0034986


title: prostate-tissue-L-replicate_RIB0034489


title: prostate-tissue-V-replicate_RIB0034942


title: prostate-tissue-H-st-replicate_RIB0034224

value: H-st


title: prostate-tissue-L-replicate_RIB0034798


title: prostate-tissue-H+st-replicate_RIB0034434


title: prostate-tissue-Ms-replicate_RIB0034702


title: prostate-tissue-H-st-replicate_RIB0034471


title: prostate-tissue-H-dt-replicate_RIB0034491


title: prostate-tissue-V-replicate_RIB0034496


title: TAM-MG-WT74

city: London

first: Silvia

last: Marino

address: 4, Newark St

country: United Kingdom

institute: Queen Mary University of London

description:
WT_74
Tam_mg_raw_counts.csv, Tam_mg_tpm_counts.csv

data_processing:
RNA-Seq data were processed using two separate pipelines. For hierarchical clustering and principal component analysis, transcript-level expression was estimated directly using the pseudoalignment package Salmon (Patro, Duggal et al., 2017). The results were then aggregated to obtain gene level expression estimates in units of transcripts per million (TPM), which are normalized for library size and transcript length. For the purpose of identifying deregulated genes, gene counts were estimated using the gapped alignment software STAR (Dobin, Davis et al., 2013)
Genome_build: Ensembl GRCm38
Supplementary_files_format_and_content: Processed files are of two types: raw count tables representing the expression of all genes obtained with STAR; TPM expression tables obtained with Salmon.

source_name: TAM-MG

growth_protocol: GL261 cells were cultures in DMEM supplemented with FBS, pen-stre and glutamine

extract_protocol:
Total RNA was extracted from sorted cells, and FFPE tissue. TAM-MG were sorted by gating on single live CD11b+ CD45low cells, and RNA was extracted from sorted cells using TRizol and the Qiagen RNA extraction Microkit. GL261 bulk tumor RNA was extracted from tumor tissue scrapped off FFPE slides, using the FFPE RNA/DNA Purification Plus Kit (NORGEN, 54300).
RNA samples were sent for sequencing to Oxford Genomic centre. They performed a SmartSeq2 and NexteraXT library preparation. Sequencing run were performed with the HiSeq4000 75bp pair end (Illumina).

treatment_protocol: Intracranial injections of GL261 cells in Cx3cr1-cre;Rheb1-loxp mice 3 weeks post-tamoxifen injections. GL261 cells were injected at a concentration of 2x10e4 cells/5µl in PBS at 6 weeks of age. samples were collected  3 weeks post tumor initiation. Mouse models have been previously reported (Bowman et al., 2016, Yona, Kim et al., 2013, Zou, Zhou et al., 2011)

value: CD11b+ CD45low TAM-MG

tag: genotype

value: WT


title: TAM-MG-KO65

description:
KO_Rheb_65
Tam_mg_raw_counts.csv, Tam_mg_tpm_counts.csv

value: Rheb1 KO


title: TAM-MG-WT78

description:
WT_78
Tam_mg_raw_counts.csv, Tam_mg_tpm_counts.csv


title: prostate-tissue-L-replicate_RIB0034486

value: male


title: prostate-tissue-Md-replicate_RIB0034448


title: Bulk-KO26

description:
KO26
Bulk_raw_counts.csv; Bulk_tpm_counts.csv

source_name: Bulk tumor

value: GL261 bulk tumor cells


title: Bulk-KO37

description:
KO37
Bulk_raw_counts.csv; Bulk_tpm_counts.csv


title: prostate-tissue-V-replicate_RIB0034457


title: TAM-MG-KO63

description:
KO_Rheb_63
Tam_mg_raw_counts.csv, Tam_mg_tpm_counts.csv


title: Bulk-WT110

description:
WT110
Bulk_raw_counts.csv; Bulk_tpm_counts.csv


title: prostate-tissue-Md-replicate_RIB0034485


title: TAM-MG-KO54

description:
KO_Rheb_54
Tam_mg_raw_counts.csv, Tam_mg_tpm_counts.csv


title: prostate-tissue-V-replicate_RIB0034321


title: prostate-tissue-H-dt-replicate_RIB0034385


title: prostate-tissue-V-replicate_RIB0034483


title: Bulk-WT38

description:
WT38
Bulk_raw_counts.csv; Bulk_tpm_counts.csv


title: prostate-tissue-L-replicate_RIB0034383


title: Bulk-KO36

description:
KO36
Bulk_raw_counts.csv; Bulk_tpm_counts.csv


title: Bulk-WT35

description:
WT35
Bulk_raw_counts.csv; Bulk_tpm_counts.csv


title: prostate-tissue-L-replicate_RIB0034340


title: prostate-tissue-Ms-replicate_RIB0034372


title: prostate-tissue-H+st-replicate_RIB0034303


title: prostate-tissue-V-replicate_RIB0034226


title: prostate-tissue-L-replicate_RIB0034373


title: prostate-tissue-H-st-replicate_RIB0034477


title: prostate-tissue-H+dt-replicate_RIB0034370


title: prostate-tissue-Ms-replicate_RIB0034207


title: prostate-tissue-V-replicate_RIB0034227


title: prostate-tissue-H+dt-replicate_RIB0034827


title: RAD21_CTRL_rep-1

description:
Rad21CTRL_Input_optimal_IDR_thresholded_peaks_mm10.narrowPeak
Rad21CTRL_nodup_pooled_signal_p_value_mm10.bw
Rad21CTRL_nodup_pooled_fold_change_over_imput_mm10.bw

data_processing:
Data were processed with ENCODE pipeline
Single reads were aligned to mm10 using bowtie
Insert size was assessed by spp
MACS2 was used to call differentiated peaks between treatments on pooled samples.
Genome_build: mm10
Supplementary_files_format_and_content: narrowPeak, bigWig

source_name: Mouse Hepatoma Cell Line

growth_protocol: Cells were grown in Alpha-MEM medium (Gibco) supplemented with 5% FBS and 1% Penicillin-Streptomycin solution and passaged at approximately 80% confluency.

extract_protocol:
Cells were fixed with 1% formaldehyde and the reaction subsequently quenched using 0.125 M glycine.  After fixation, cells were lysed to release nuclei, which were washed and re-suspended in nuclei lysis buffer before sonicating the chromatin using a Diagenode Bioruptor.  Fragmented chromatin was diluted and separated into equivalent aliquots prior to antibody application and complex retrieval.
A ChIP-seq script was selected for sequencing library preparation using PrepX DNA Library kit and Apollo 324 NGS automatic library prep system (WaferGen, Fremont, CA). Overhangs in samples of double-strand DNA fragments (~5 ng) were first blunt-ended, and adenylated at the 3' ends for TA ligation to the sequencing adaptors. The ligated library was enriched by 8 cycles of PCR using index-specific primers, followed by automated AMPure XP beads (Beckman Coulte, Brea CA) purification. To check the quality and yield of the library, one µl of purified PCR library was analyzed by Bioanalyzer with DNA High Sensitivity chip. To accurately quantify the library concentration for cluster generation, the library was 1:104 diluted with dilution buffer (10 mM Tris-HCl, pH 8.0 with 0.05% Tween 20), and qPCR analyzed with NEBNext Library Quant Kit (New England Biolabs, Ipswich, MA) using ABI 9700HT real-time PCR system (Lifetech, Grand Island, NY). Individually indexed libraries were proportionally pooled for clustering in the cBot system (Illumina, San Diego, CA). Libraries at the final concentration of 16 pM were clustered onto a flow cell using Illumina TruSeq SR Cluster kit v3, and sequenced for 50 cycles using TruSeq SBS kit on Illumina HiSeq system. According to Illumina, for studies targeting transcription factors, ~10M reads per sample were generated.

treatment_protocol: Cells were treated with filter-sterilized potassium chromate dissolved in MilliQ-grade water for 90 mins prior to fixation and processing.

value: 0 µM chromium (Control)

tag: chip antibody

value: Anti-Rad21 antibody - ChIP Grade (abcam, ab992)

tag: pulldown target


title: STAG1_CTRL_rep-2

description:
STAG1CTRL_Input_optimal_IDR_thresholded_peaks_mm10.narrowPeak
STAG1CTRL_nodup_pooled_signal_p_value_mm10.bw
STAG1CTRL_nodup_pooled_fold_change_over_imput_mm10.bw

value: SA1 Antibody (Bethyl Laboratories, A300-157A)


title: STAG1_Cr1_rep-2

description:
STAG1Cr1_Input_optimal_IDR_thresholded_peaks_mm10.narrowPeak
STAG1Cr1_nodup_pooled_signal_p_value_mm10.bw
STAG1Cr1_nodup_pooled_fold_change_over_imput_mm10.bw

value: 1 µM chromium


title: RAD21_Cr1_rep-1

description:
Rad21Cr1_Input_optimal_IDR_thresholded_peaks_mm10.narrowPeak
Rad21Cr1_nodup_pooled_signal_p_value_mm10.bw
Rad21Cr1_nodup_pooled_fold_change_over_imput_mm10.bw


title: RAD21_Cr1_rep-2


title: prostate-tissue-H+st-replicate_RIB0034830


title: STAG1_Cr1_rep-1


title: H3K9me3_CTRL_rep-1

description:
H3K9me3CTRL_Input_optimal_IDR_thresholded_peaks_mm10.narrowPeak
H3K9me3CTRL_nodup_pooled_signal_p_value_mm10.bw
H3K9me3CTRL_nodup_pooled_fold_change_over_imput_mm10.bw

value: Anti-Histone H3 (tri methyl K9) antibody - ChIP Grade (abcam, ab8898)


title: Active CD4 T1h Cyto BruChase-Seq

city: Washington DC

first: Weiqun

last: Peng

state: Washington DC

address: 800 22nd St NW

department: Physics

institute: George Washington University

data_processing:
Library strategy: Bru-Seq
The sequencing quality of Bru-Seq and BruChase-Seq libraries was assessed by FastQC (http://www.bioinformatics.babraham.ac.uk/projects/fastqc/).
Bru-Seq and BruChase-Seq libraries were mapped to human genome using Tophat (v2.1.0, Trapnell et al., Bioinformatics). UCSC genes for human hg19 assembly from iGenome (http://support.illumina.com/sequencing/sequencing_software/igenome.html) were used for gene annotation.
Genome_build: hg19
Supplementary_files_format_and_content: [.wig file] ERCC normalized expression levels

source_name: CD4 T cells

extract_protocol:
Human resting CD4+ T cells were sort-purified and then activated by CD3/CD28. Bromouridine (Bru) was added to the media of resting and activated CD4+ T cells. The Bru-labeled RNA was isolated from the total or cytoplasmic RNA of resting and activated CD4+ T cells.
Libraries were prepared for sequencing using stardard Illumina protocol

value: wild type

value: Activated CD4 T cells

tag: cell subset

tag: molecule subtype

value: Bru-labeled cytoplasmic RNA


title: Active CD4 T0.5h Cyto BruChase-Seq


title: H3K9me3_Cr1_rep-2

description:
H3K9me3Cr1_Input_optimal_IDR_thresholded_peaks_mm10.narrowPeak
H3K9me3Cr1_nodup_pooled_signal_p_value_mm10.bw
H3K9me3Cr1_nodup_pooled_fold_change_over_imput_mm10.bw


title: Rest CD4 T2h Cyto BruChase-Seq

value: Resting CD4 T cells


title: Active CD4 T0h Cyto Bru-Seq


title: H3K9me3_Cr1_rep-1


title: Rest CD4 T1h Cyto BruChase-Seq


title: Rest CD4 T0.5h Cyto BruChase-Seq


title: H3K4me3_CTRL_rep-1

description:
H3K4me3CTRL_Input_optimal_IDR_thresholded_peaks_mm10.narrowPeak
H3K4me3CTRL_nodup_pooled_signal_p_value_mm10.bw
H3K4me3CTRL_nodup_pooled_fold_change_over_imput_mm10.bw

value: Anti-trimethyl-Histone H3 (Lys4) Antibody (Millipore Sigma, 07-473)


title: Rest CD4 T0h Cyto Bru-Seq