title: 293T-EMX1.All_rep2.EMX1-Detect-Extend-5

description:
EMX1.All_rep2.EMX1-Detect-Extend-5 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 293T-EMX1.All_rep2.EMX1-Detect-Extend-4

status: Public on Apr 08 2021

type: SRA

city: Beijing

first: Howard

last: MENG

state: Beijing

address: No.5 Yiheyuan Road Haidian District

department: School of Life Sciences

country: China

institute: Peking University

description:
EMX1.All_rep2.EMX1-Detect-Extend-4 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

library_source: genomic

data_processing:
Library strategy: Targeted amplicon sequencing
Cut adapter step: The raw FASTQ files are firstly but the sequencing adapters with cutadapt software with the following command: "cutadapt --times 1  -e 0.1  -O 3  --quality-cutoff 25  -m 100 -a  AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC  -A  AGATCGGAAGAGCGTCGTGTAGGGAAAGAGTGTAGATCTCGGTGGTCGCCGTATCATT -o {read1.out}  -p {read2.out}  {read1.in}  {read2.in}"
Alignment step: All FASTQ files are aligned by bwa mem (version = 0.7.15) with default parameters. More information please query www.detect-seq.com
Genome_build: hg38
Supplementary_files_format_and_content: Processed data contains the primer and experiment design information of all target amplicon sequencing data.

molecule: genomic DNA

organism: Homo sapiens

source_name: HEK293T cells

growth_protocol: The HEK293T and MCF7 is separately cultivated in DMEM and MEM with 10% FBS and 1% PS at 37°C cultivator with 5% CO2. The subculture of cells was performed every 2–3 days and only passage 3-10 were used for experiments. All the cells were routinely tested for Mycoplasma contamination with TransDetect PCR Mycoplasma Detection Kit (TRANSGEN BIOTECH, FM311-01).

extract_protocol:
Genomic DNA was freshly extracted using CWBIO universal genomic DNA kit (CWbiotech, CW2298M) and stored in TE buffer (10 mM Tris-HCl, 1 mM EDTA, pH 8.0) at -80 °C.
Regions flanking the targeting sites were selected for the design of primers, whose overhangs contained the paired Illumina adaptor sequences. In addition, a 10-nt barcode was also added into each primer pairs to lower the detection limit from 10-1 to 10-5 (Primers are listed in the Supplementary). The first round of PCR amplification was performed with NEBNext® Q5® Hot Start HiFi PCR Master Mix (NEB, M0543L) using ~10 ng of genomic DNA as an input template. After ~10 cycles of amplification, the PCR products were purified with 1 × Agencourt AMPure XP beads and eluted with ddH2O. Purified DNA samples were then subjected to the second round of amplification for ~15 cycles and assigned with different indexes followed by a purification with 0.8 × AMPure XP beads. The libraries were quantified with Qubit 2.0 Fluorometer (INVITROGEN) and pooled together for high-throughput sequencing by Illumina HiSeq X Ten (Genetron Health, Beijing).    All FASTQ files in this dataset, have already been grouped by barcode. Meanwhile, the Illumina sequencing errors were trying to be corrected as well (decrease error rate from ~0.1% to ~0.005%). The Python scripts used in this step are available by request to correspondents.

treatment_protocol: 6.4 × 106 of HEK293T cells or 2.5 × 106 of MCF-7 cells were seeded into 6-well culture plates (Corning) for 16 h growth. Adherent cells were transfected with 4 μg BE and 2720 ng sgRNA plasmids per well using lipofectamine LTX following the manufacturer’s protocol. Cells were then collected after 72 h of transfection.

tag: cell type

value: embryonic kidney derived epithelial cells

tag: genotype

value: Wild Type

tag: experiment

value: Targeted Amplicon Sequencing

value: BE4max + sgRNA (EMX1)


title: Vehicle RNA - 813_constant_rep3

city: Bristol

first: Benjamin

middle: P

last: Flynn

state: County (optional)

address: Dorothy Hodgkin Building, Whitson street

country: United Kingdom

institute: University of Bristol

description: RNA-Seq

library_source: transcriptomic

data_processing:
Basecalling RTA 1.13.48 with default filter and quality settings
Adapters removed by bbduk (BBMap v38.22) tpe tbo
Aligned to rn6 using STAR v020201, default parameters
strand specific read counts calculated by featureCounts v1.6.2, exonic boundaries identified in Ensemble Rnor 6.0.92 release
Genes filtered for counts > 10 across conditions and DESeq2 normalisation, differenrial analysis assesed by DESeq2 using defualt parameters (n=3)
AllFinalCounts.gene represents matrix of exonic read counts (featureCounts), genes filtered for counts >10 across conditions and differential results from DESeq2 analysis are formatted as Gene ID, base mean, log2 fold change, standard error of log2 fold change values, Wald statistic, Wald test p-value and Benjamin-Hochberg adjusted p-value for pulsatile VEH difference to constant VEH (ExonRNAData_PULSVEH_CONSVEH.txt), pulsatile CORT change to pulsatile VEH (ExonRNAData_PULSCORT_PLSVEH.txt) and constant CORT change to constant VEH (ExonRNAData_CNSCORT_CNSVEH.txt)
Genome_build: rn6

molecule: total RNA

organism: Rattus norvegicus

source_name: Liver

extract_protocol:
RNA_Seq  RNA extraction  Frozen liver sections were homogenised (Polytron, PT-2100, Kinematica, CH) in TRIzol (ThermoFisher Scientific, USA). The aqueous phase was separated via chloroform extraction (Sigma-Aldritch, UK) and centrifugation (10,000 rpm, 40C), total RNA extracted, and DNA digested (miRNAeasy mini, Qiagen, US).
RNA_Seq  Libraries were constructed from 1 µg total RNA (TrueSeq Stranded Total RNA kit, Illumina, USA) and filtered for a RIN score > 8 and < 10 (Agilent Tape Station, Bioanalyzer, UK) for sequencing (HiSeq 2500, Illumina UK).

tag: strain

value: Sprague Dawley

tag: Sex

value: Male

tag: tissue

value: Liver

tag: treatment

value: Vehicle infusion - constant - 180min


title: 1d_Male_H3K27ac_4

city: Tallahassee

first: Michelle

middle: N

last: Arbeitman

state: Florida

address: 1115 W. Call Street

department: Department of Biomedical Sciences

country: USA

institute: Florida State University

description: Sequential Chromatag-ChIP for fru P1 neuron chromatin

data_processing:
ChIP-seq reads were pre-processed using Trimmomatic’s ILLUMINACLIP (v. 0.33) option to remove Illumina adaptor sequences. Pre-processed ChIP-seq reads were aligned to the D. melanogaster dm6 UCSC genome assembly using Bowtie2 (v.2.2.5), allowing up to 6 mismatches. Only uniquely aligned reads were considered and individual replicate and pooled replicate (all replicates) BAM files were generated with SAMtools (v. 1.2) for downstream analyses.
MACS2 (v.2.1.0) peaks were called for each histone modification (H3K27ac, H3K36me3, H3K4me3, H3K9me3, H3K27me3) at each time point (48hr APF, 1-day adult, 10-12 day adult) for each sex in fru P1 or elav neurons, relative to sex and stage-specific input chromatin library controls for individual and all (pooled) replicates. We performed “narrow” peak calling for histone modifications with more compact enrichment patterns (H3K27ac and H3K4me3) and the “broad” peak option for modifications with diffuse enrichment (H3K36me3, H3K9me3, and H3K27me3) on all individual biological replicates and on all (pooled) replicate data using the following “callpeak” parameters: --gsize dm -p 1e-2 --call-summits --fix-bimodal.
Input normalized BigWig files for all histone modifications for all replicate (pooled) data were obtained using Deeptools bamCompare function to subtract input ChIP-seq signal from histone modification .BAM files obtained from concatinating individual replicate .BAM files.
Genome_build: UCSC dm6 (BDGP Release 6 plus ISO1 MT)

organism: Drosophila melanogaster

source_name: Whole head

growth_protocol: For 48hr APF stage: pupae were sex-sorted at white pre-pupal stage and aged to 48hr after Puparium formation (APF) on Bromophenol blue agar plates at 25˚C on a 12 hour light and 12 hour dark cycle. Adult stage flies were reared and maintained on standard cornmeal food at 25°C on a 12 12-h light/12-h dark cycle.

extract_protocol: The adult heads were separated from their bodies by vortexing frozen animals. The heads were sifted through liquid nitrogen-cooled mesh sieves #25 and #40 (with heads retained in the #40 sieve). The sequential ChIP methodology presented here is based on a previously published ChIP protocol (Negre et al. 2006). Frozen adult heads or frozen whole body pupae (~0.3g) were dounce homogenized in 5ml of Buffer A1 (60mM KCL, 15mM NaCl, 4mM MgCl2, 15mM HEPES (pH 7.6), 0.5% TritonX-100, 0.5mM DTT, 10mM sodium butyrate, and Roche cOmplete Protease Inhibitor cocktail EDTA-free, product 5056489001), with 1% methanol-free formaldehyde (Pierce, 20% solution product 28906) for crosslinking at room temperature for 10 min. The crosslinking reaction was quenched using 540\265l 2.5M Glycine and incubated for 5 minutes at room temperature. Cross-linked homogenates were centrifuged at 4000g for 5 minutes at 4\272C and the nuclear pellets were resuspended in Buffer A1. The nuclear pellets were washed 3 more times by centrifugation at 4000g for 5 minutes at 4\272C with resuspension in 3ml of buffer A1. The nuclear pellets were washed an additional time with 1x Lysis Buffer without SDS (140mM NaCl, 15mM HEPES (pH 7.6), 1mM EDTA (pH 8.0), 0.5mM EGTA, 1% TritonX-100, 0.5mM DTT, 0.1% sodium deoxycholate, 10mM sodium butyrate, and Roche cOmplete Protease Inhibitor Cocktail EDTA-free) to remove all traces of Buffer A1 before the next lysis step. After one additional spin at 4000g for 5 minutes at 4\272C, the washed nuclear pellet was resuspended in Lysis Buffer containing 1% SDS and 0.5% N-Laurosylsarcosine and incubated at 4\272C, rotating for 1 hour. Nuclear lysates were quickly spun down for 10 seconds at 2000g and supernatant transferred to sonication tubes (Covaris; product 520130) for shearing. Chromatin was sheared for 25 minutes using a Covaris E220 (duty cycle: 5.0, peak incident power: 140, cycles per burst: 200). The sheared chromatin was transferred to microcentrifuge tubes and rotated for 10 min. at 4\272C. Next, the sheared chromatin was centrifuged at 18,000g for 5 minutes at 16 \272C. The supernatant was transferred to a new microcentrifuge tube and the centrifugation step was repeated twice. The final supernatant was split into two 0.5ml aliquots (material from ~0.15g frozen tissue) and brought to a 1.5ml volume with 1x Lysis Buffer without SDS. The sheared chromatin was then aliquoted into 250\265l volumes and stored at -80\272C. To perform the sequential ChIP, 750\265l of 1x Lysis Buffer without SDS was added to the 250\265l of the thawed sheared chromatin. The sheared chromatin was incubated with 50\265l of magnetic protein A/G beads (Pierce, product 88803) and 5\265g of each respective antibody at 4\272C overnight. The magnetic protein A/G beads were then washed four times with 1ml of Lysis Buffer wash (140mM NaCl, 15mM HEPES (pH 7.6), 1mM EDTA (pH 8.0), 0.5mM EGTA, 1% TritonX-100, 0.05% SDS, 0.5mM DTT, 0.1% sodium deoxycholate, 10mM sodium butyrate, and Roche cOmplete Protease Inhibitor Cocktail EDTA-free) and twice with TE (10mM Tris-HCL (pH 8.0), 1mM EDTA) for 10 minutes each, rotating at 4 \272C. Beads were incubated in 100\265l ChIP elution buffer (100mM NaHCO3, 80mM NaOH, 1% SDS) at room temperature, rotating for 30-60 minutes to elute chromatin. The eluate was diluted 1:10 with Lysis Buffer without SDS and incubated on magnetic streptavidin beads (Invitrogen Dynabeads MyOne Streptavidin C1) for 4hrs-overnight rotating at 4\272C. The beads were washed as above and chromatin was reverse crosslinked on beads in ChIP elution buffer (with 0.04mg/ml proteinase K, 0.01mg/ml RNaseA) at 65 \272C overnight. DNA was purified with a DNA purification and concentration kit (Machery-Nagel, product 740609). ChIPseq libraries were prepared using the NEBNext ChIP-seq library preparation kit. Libraries were size selected using AMPure XP beads (Beckman) and clean up steps were performed using the Machery Nagel PCR cleanup kit. A final wash was performed using the AMpure XP beads according to kit protocol to remove primers.

treatment_protocol: Approximately 1000 sex-sorted flies that were 48hr after pupal formation (APF, +/- 1hr), 16-24hrs post eclosion, or 10-12 days post eclosion were used for each ChIP independent biological replicate (n=4 and n=3 for 10-12 day adult female libraries). Flies were collected and sex sorted at the white prepupal stage, 0-8hrs post-eclosion, or 0-48hrs post-eclosion for appropriate aging to their respective time points and snap frozen in liquid nitrogen between ZT1 and ZT2. All flies collected under anesthetization were allowed to recover for at least 16hrs before freezing took place. All frozen animals were stored at -80°C until they were processed for downstream experiments.

tag: developmental stage

tag: chip antibody

value: fru P1


title: 293T-EMX1.All_rep2.EMX1-Detect-Extend-21

description:
EMX1.All_rep2.EMX1-Detect-Extend-21 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 1d_Male_H3K27ac_3


title: Vehicle RNA - 1134_constant_rep1

description: RNA-Seq


title: Vehicle RNA - 1089_pulsatile_rep3

description: RNA-Seq

value: Vehicle infusion - pulsatile - 180min


title: Vehicle RNA - 1081_pulsatile_rep1

description: RNA-Seq


title: Vehicle RNA - 1088_pulsatile_rep2

description: RNA-Seq


title: 293T-EMX1.All_rep2.EMX1-Detect-Extend-14

description:
EMX1.All_rep2.EMX1-Detect-Extend-14 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: Corticosterone RNA - 788_constant_rep2

description: RNA-Seq

value: Corticosterone infusion - constant - 180min


title: Corticosterone RNA - 787_constant_rep1

description: RNA-Seq


title: Corticosterone RNA - 798_pulsatile_rep3

description: RNA-Seq

value: Corticosterone infusion - pulsatile - 180min


title: 293T-EMX1.All_rep2.EMX1-Detect-Extend-1

description:
EMX1.All_rep2.EMX1-Detect-Extend-1 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: Corticosterone RNA - 790_constant_rep3

description: RNA-Seq


title: 1d_Male_H3K27ac_1


title: 1d_Male_H3K27ac_2


title: 293T-EMX1.All_rep2.Control-9

description:
EMX1.All_rep2.Control-9 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 293T-EMX1.All_rep2.EMX1-Detect-Extend-10

description:
EMX1.All_rep2.EMX1-Detect-Extend-10 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 1d_Female_H3K27ac_3

value: Female


title: Corticosterone RNA - 792_pulsatile_rep2

description: RNA-Seq


title: 1d_Female_H3K27ac_4


title: Corticosterone RNA - 791_pulsatile_rep1

description: RNA-Seq


title: 293T-EMX1.All_rep2.Control-8

description:
EMX1.All_rep2.Control-8 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: Vehicle RNA - 812_constant_rep2

description: RNA-Seq


title: 1d_Female_H3K27ac_2


title: 1d_Female_H3K27ac_1


title: 1d_Male_H3_4


title: 293T-EMX1.All_rep2.Control-7

description:
EMX1.All_rep2.Control-7 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 293T-EMX1.All_rep2.Control-6

description:
EMX1.All_rep2.Control-6 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: WT_2

city: Wuhan, Hubei, China

first: Jing

last: Wang

address: Qingnian Road, Qiaokou, Hankou

department: Department of Pediatric Surgery

institute: Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology

data_processing:
Illumina HiSeqTM 2500 used for basecalling.
specific steps were followed using the methods previously described (WEI L, CAO L, MIAO Y, et al. Transcriptome analysis of Spodoptera frugiperda 9 (Sf9) cells infected with baculovirus, AcMNPV or AcMNPV-BmK IT [J]. Biotechnol Lett, 2017, 39(8): 1129-39.)
The PossionDis algorithm was applied to perform differential gene detection and screened |log2 (FoldChange)| > 1 & qvalue<0.05 as the DEGs.
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited text files include MEAN TPM values for each Sample

organism: Danio rerio

source_name: Intestinal

extract_protocol:
intestinal tissues of embryos were dissected by microforceps. All samples were stored at −80°C until total RNA extraction. Total RNA extraction was performed with TRIZol reagents from Invitrogen following the manufacturer’s instructions. cDNA libraries were constructed and adopted the paired-end sequencing on an Illumina HiSeqTM 2500.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: Embryos of BP-3 exposure group were incubated in 2mg/L BP-3

value: zebrafish

value: intestinal

tag: age

value: Control


title: wt 30min ischemia 6hrs reperfusion

city: Indianapolis

first: Seth

last: Winfree

email: winfrees@iu.edu, swinfree@unmc.edu

state: Indiana

address: 950 W Walnut St

institute: Indiana University

description: generated from three kidenys from two mice

data_processing:
Cellranger (10X Genomic v.3.0.2)  was used to process raw sequencing data including demultiplexing (bcl2fastq) and generate fastq files (as provided).
STAR was used to align the output fastq filesto the mm10 reference genome.
Expression levels for genes were tallied for each cell by UMIs from the aligned reads.
expression matrices, gene and cell UMI files generated (as provided in processed data .zip files).
Genome_build: mm10
Supplementary_files_format_and_content: tar archives files contain barcode.tsv.gz, features.tsv.gz and matrix.mtx.gx for loading into analysis tool of choice

organism: Mus musculus

source_name: kidney

extract_protocol:
kidneys were removed and processed by a MACs cell isolation protocol which inlcuded mechanical and enzymatic dissolution and dead cell removal (Reference, PMID:33448928).
Cell suspension was loaded onto a Chromium Single Cell Instrument (10X Genomics)

value: 8-12 weeks

value: kidney

value: wild type

value: ischemic acute kidney injury


title: BP_3_3

value: BP-5 exposure


title: WT_1


title: WT_3


title: 293T-EMX1.All_rep2.Control-5

description:
EMX1.All_rep2.Control-5 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

value: embryonic kidney derived epithelial cells

value: Wild Type

value: Targeted Amplicon Sequencing

value: BE4max + sgRNA (EMX1)


title: BP_3_2

value: BP-4 exposure


title: wt sham kidney

description: generated from both kidneys from one mouse

value: sham


title: 1d_Male_H3_3

value: Male

value: fru P1


title: BP_3_1

value: BP-3 exposure


title: 293T-EMX1.All_rep2.Control-4

description:
EMX1.All_rep2.Control-4 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 293T-EMX1.All_rep2.Control-3

description:
EMX1.All_rep2.Control-3 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 1d_Male_H3_1


title: 293T-EMX1.All_rep2.Control-2

description:
EMX1.All_rep2.Control-2 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 1d_Male_H3_2


title: ChIP_ESC_wtRes_input

city: Shanghai

first: Chuan

last: Chen

address: 1239 Siping Road

institute: Tongji University

description: 2.5 μM 4-OHT was added into the culture medium 72 hr before harvesting.

data_processing:
Primary fastq data were generated with the Illumina Casava (v 1.8.2) pipeline.
Sequencing reads were trimmed by Trim Galore (v 0.6.4) with parameters: --phred33, --illumina, --clip_R1 9, --clip_R2 9, --paired to remove adapter sequences.
Trimmed reads were aligned to mm9 reference genome using Bowtie2 (v 2.2.9) with default parameters.
ChIP peaks were defined by the callpeak function of MACS2 (v 2.1.1) with parameters: -g mm, -f BAM, --BDG, --QVALUE 0.05, --SPMR.
Genome_build: mm9
Supplementary_files_format_and_content: Bigwig files containing piled up reads.  Scores represent normalized tag counts per million reads.

source_name: embryonic stem cells

growth_protocol: Cells were cultured in ES cell medium (DMEM, 15% FBS, 1×Nucleosides, 1 mM L-Glutamine, 1×Non Essential Amino Acids, 0.1 mM 2-Mercaptoethanol, 1000 U/ml LIF) supplemented with 3 mM CHIR-99021 and 1 mM PD0325901.

extract_protocol:
5×10^6 ES cells per reaction were fixed in 1% formaldehyde and neutralized by 125 mM glycine. The crosslinked cells were pelleted, lysed and subjected to sonication. 1/100 of the product was saved as the input, and the rest was mixed with antibody-bound Dynabeads Protein A/G for IP reaction. DNAs of input and IP samples were extracted using phenol chloroform coupled with isopropanol precipitation.
DNAs purified from input and IP samples were subjected to library generation using HyperPlus Library Preparation Kit.

treatment_protocol: At the time of passaging, 2.5 μM 4-OHT was added into the culture medium for control and Ythdc1 cKO ES cell lines, and cells were harvested  after 72-hr treatment of 4-OHT.

value: blastocyst-derived ES cells

value: Ythdc1 flox/flox; CreERT2; WT-YTHDC1 overexpression


title: 1d_Female_H3_4

value: Female


title: cxcr3.2mu_rep3

city: London

first: Vincenzo

last: Torraca

state: London

address: Keppel street

institute: LSHTM

data_processing:
Image analysis and base calling was done by the Illumina HCS version 1.15.1
Quality trimmed reads were aligned to the Ensembl zebrafish genome (GRCz10) using Bowtie
Reads were mapped to zebrafish transcripts using TopHat
Genome_build: GRCz10
Supplementary_files_format_and_content: tab separated value files of read counts for each sample

molecule: polyA RNA

source_name: FACS mpeg1: mCherryF positive cells

extract_protocol:
3 independent groups of 150-200 5dpf Tg (mpeg1: mCherryF cxcr3.2-/- and cxcr3.2+/+) larvae were dissociated and red fluorescent macrophages were FACS sorted. RNA was extracted using the miRNeasy mini kit (Qiagen) according to the manufacturers’ instructions. cDNA synthesis and amplification was done using the SMARTer® Universal Low Input RNA Kit for Sequencing (Clontech) following the manufacturer’s guidelines.
Approximately 10 Mreads were obtained, using Illimuna Single-Read runs. Reads were then aligned to GRCz10 Danio rerio genome.

value: 6 days post-fertilisation

value: macrophage


title: cxcr3.2wt_rep3


title: cxcr3.2wt_rep2


title: ChIP_ESC_wtRes_K9_2

value: anti-H3K9me3 (Abcam ab8898)


title: 1d_Female_H3_3


title: ChIP_ESC_wtRes_Kap1_2

value: anti-KAP1 (Abcam ab22553)


title: cxcr3.2mu_rep2


title: 293T-EMX1.All_rep2.Control-18

description:
EMX1.All_rep2.Control-18 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: ChIP_ESC_wtRes_Kap1_1


title: 1d_Female_H3_2

description: Sequential Chromatag-ChIP for fru P1 neuron chromatin


title: cxcr3.2wt_rep1


title: cxcr3.2mu_rep1


title: 1d_Male_Input_3

description: ChIP input chromatin

value: ChIP Input

value: Whole head


title: 1d_Male_Input_4

description: ChIP input chromatin


title: 293T-EMX1.All_rep2.Control-17

description:
EMX1.All_rep2.Control-17 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

value: embryonic kidney derived epithelial cells

value: Wild Type


title: 293T-EMX1.All_rep2.Control-16

description:
EMX1.All_rep2.Control-16 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

value: Targeted Amplicon Sequencing

value: BE4max + sgRNA (EMX1)


title: ChIP_ESC_W378A_input

value: Ythdc1 flox/flox; CreERT2; W378A-YTHDC1 overexpression


title: 1d_Female_H3_1


title: ChIP_ESC_wtRes_K9_1


title: ChIP_ESC_W378A_K9_2


title: Total RNA from BFTC 905/DMSO cell line

city: Chia-Yi

first: Yu Ming

last: Chuang

address: 168 University Road,Min Hsiung

department: Department of Biomedical Science

country: Taiwan

institute: National Chung Cheng University

description: BFTC 905 bladder cancer cell line.

data_processing:
Illumina MiSeq used for basecalling.
CLC Genomics Workbench 8.5.1 software was used.
The raw reads were cleaned by trimming of adaptor sequences.
Sequenced reads were mapped to hg19 genome using CLC Workbench default parameters.
Genome_build: hg19
Supplementary_files_format_and_content: RPKM

source_name: bladder cancer cell line

growth_protocol: BFTC 905 cells were maintained in RPMI-1640 medium supplemented with 10% FBS. Cells were incubated at 37C under a humidified atmosphere containing 5% CO2

extract_protocol: Total RNA was extracted from each sample using TRIzol reagent.

treatment_protocol: BFTC 905 cells were either treated with DMSO or 55µM CPH.

tag: cell line

value: BFTC 905

tag: agent

value: DMSO


title: 1d_Male_Input_2

description: ChIP input chromatin

value: Male


title: 293T-EMX1.All_rep2.Control-15

description:
EMX1.All_rep2.Control-15 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 293T-EMX1.All_rep2.Control-14

description:
EMX1.All_rep2.Control-14 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: ChIP_ESC_W378A_Kap1_2

value: blastocyst-derived ES cells


title: 1d_Male_Input_1

description: ChIP input chromatin


title: HF-2927_chia-pet

city: Seattle

first: Chia-Lin

last: Wei

state: WA

address: 3720 15th Ave NE

department: The Northwest Genomics Center (NWGC)

institute: University of Washington

data_processing:
ChIA-PET data was processed by ChIA-PET Utilities (CPU) (https://github.com/cheehongsg/CPU). CPU (options: stat -s -p -T 18 -t 16) removed the adaptors, and grouped the reads into three categaries: PET reads, single tag reads and no-linker reads. Tags identified (>=16bp) and reads without linkers were mapped to hg19 using BWA alignment and mem according to the tag/read length. CPU further separated the mapping results into uniquely-mapped,multiply-mapped, and unmapped classes with the pairing consideration (options: pair -S -t 16) and removed duplicates generated by PCR amplification (dedup -g -t 16). Next, the non-redundant uniquely-mapped reads from 3 categaries were merged and used for peak calling with MACS v2.1.1.20160309 (options: --keep-dup all --nomodel --extsize 250 -B --SPMR -g hs). CPU furhter clustered the non-redundant uniquely-mapped pair-end tags and grouped them as intra-chromosomal PET clusters (cis-interactions) and inter-chromosomal PET clusters (trans-interactions) (options: cluster -s 8000 -M -B 1000 -5 5,-20 -3 5,480 -t 16 -O). The cis-interaction significance was assessed with ChiaSigScaled (https://github.com/cheehongsg/ChiaSigScaled ; options: -c 3 -t 16 -p -m 8000). Cis-interactions with member size >=3 and FDR < 0.05 were reported. The complete list trans interaction was reported here while additional filtering were applied (e.g. exclude singletons ) for corresponding analyses in publication. (see details in publication).
Illumina RTA-1.18 software used for basecalling
Genome_build: hg19

source_name: Glioblastoma neurosphere cell suspensions

growth_protocol: Tumor specimens were dissociated and cultured as neurospheres in DMEM/F12 medium (11330-032, Gibco) supplemented with N-2 supplement (17502-048, Gibco) and growth factors (EGF and FGF-basic). Cells with passages between 15 and 26 were used for experiments.

extract_protocol:
10 million cells were dual-crosslinked with 1.5mM EGS (21565, Thermo Fisher) for 45 min followed by 1% formaldehyde (F8775, Sigma) for 20 min at room temperature and then quenched with 0.125 M Glycine (G8898, Sigma) for 10 min. The crosslinked cells were washed with PBS twice and lysed in 100 ul 0.55% SDS with incubation at room temperature, 62°C and 37°C sequentially for 10 min each, which was followed by 37°C for 30 min with addition of 25 ul 25% Triton-X 100 to quench SDS and 37°C overnight with addition of 50 ul AluI (R0137L, NEB), 50 μl 10 × CutSmart buffer and 275 μl H2O to fragmentize the chromatin. The pelleted digested nuclei were resuspended in 500 ul A-tailing solution containing 50 μl 10 × CutSmart buffer, 10 μl BSA (B9000S, NEB), 10 μl 10 mM dATP (N0440S, NEB), 10 μl Klenow (3’- 5’ exo-) (M0202L, NEB), and 420 μl H2O  with 1 hr incubation at room temperature and then subjected to proximity ligation by adding 200 μl 5 × ligation buffer (B6058S, NEB), 6 μl biotinylated bridge linker (200 ng/ul), 10 μl T4 DNA ligase (M0202L, NEB) and 284 μl H2O and incubating at 16°C overnight. The ligated chromatins were then sheared by sonication and immunoprecipitated with anti-RNApolII antibody (920102, Biolegend).
The ChIPped DNA were tagmented using the Nextera DNA Sample Preparation Kit (Illumina). The linker ligated ChIA-PET constructs were selected using streptavidin magnetic beads. The magnetic beads were blocked with sheared gDNA prior to the selection. The streptavidin selected constructs were amplified with PCR and purified.

tag: source

value: Glioblastoma

tag: ip antibody

value: RNA polymerase II (920102, Biolegend)


title: ChIP_ESC_W378A_K9_1


title: 1d_Female_Input_4

description: ChIP input chromatin

value: Female


title: 293T-EMX1.All_rep2.Control-1

description:
EMX1.All_rep2.Control-1 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 293T-EMX1.All_rep2.Control-13

description:
EMX1.All_rep2.Control-13 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 1d_Female_Input_3

description: ChIP input chromatin


title: RNA_ESC_Mettl3KO_3

data_processing:
Primary fastq data were generated with the Illumina Casava (v 1.8.2) pipeline.
Sequencing reads were trimmed by Trim Galore (v 0.6.4) with parameters: --phred33, --illumina, --clip_R1 9, --clip_R2 9, --paired to remove adapter sequences.
Trimmed reads were aligned to mm9 reference genome using Tophat2 (v2.1.1) with default parameters.
Expression levels of individual were quantified by FPKM using Cufflinks (v 2.2.1).
Genome_build: mm9
Supplementary_files_format_and_content: Tab-delimited text files containing FPKM values for each transcript.

extract_protocol:
ES cells were disrupted in TRIzol Reagent and total RNAs were isolated by chloroform extraction coupled with isopropanol precipitation.
1 μg RNA per reaction was subjected to rRNA elimination and library generation using VAHTS Total RNA-seq (H/M/R) Library Prep Kit.

value: Mettl3 KO


title: 293T-EMX1.All_rep2.Control-12

description:
EMX1.All_rep2.Control-12 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: M3

city: changsha

first: jiang

last: he

address: 87 Xiangya Road

institute: xiangya hospital

data_processing:
using lllumina Novaseq platform
index of the reference genome was built using Hisat2 v2.0.5
Genome_build: hg18
Supplementary_files_format_and_content: FPKM of each gene was calculated based on the length of the gene and reads count mapped to this gene

source_name: cell line

extract_protocol:
RNA was harvested using Trizol reagent. Illumina TruSeq RNA Sample Prep Kit was used with 1 ug of total RNA for the construction of sequencing libraries. Sequencing was completed by Novogene
RNA libraries were prepared for sequencing using standard Illumina protocols

value: nasopharyngeal carcinoma

value: stably expressing LMP1


title: M2


title: 293T-EMX1.All_rep2.Control-11

description:
EMX1.All_rep2.Control-11 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 293T-EMX1.All_rep2.Control-10

description:
EMX1.All_rep2.Control-10 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: RNA_ESC_Mettl3KO_2


title: M1


title: Total RNA from BFTC 905/CPH cell line

description: BFTC 905 bladder cancer cell line.

value: CPH


title: 1d_Female_Input_2

description: ChIP input chromatin

value: ChIP Input

value: Whole head


title: 1d_Female_Input_1

description: ChIP input chromatin


title: 293T-EMX1.All_rep1.EMX1-on-target

description:
EMX1.All_rep1.EMX1-on-target in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

value: embryonic kidney derived epithelial cells

value: Wild Type


title: ChIP_ESC_W378A_Kap1_1

value: anti-KAP1 (Abcam ab22553)


title: C2

value: stably expressing empty vector (EV)


title: C3


title: RNA_ESC_WT_2

value: WT


title: RNA_ESC_WT_3


title: 293T-EMX1.All_rep1.EMX1-Detect-pRBS-7

description:
EMX1.All_rep1.EMX1-Detect-pRBS-7 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

value: Targeted Amplicon Sequencing

value: BE4max + sgRNA (EMX1)


title: 48hrAPF_Male_H3K9me3_2

source_name: Whole pupae

value: Male

value: fru P1


title: 48hrAPF_Male_H3K9me3_4


title: 293T-EMX1.All_rep1.EMX1-Detect-pRBS-9

description:
EMX1.All_rep1.EMX1-Detect-pRBS-9 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 48hrAPF_Male_H3K9me3_3


title: C1


title: 293T-EMX1.All_rep1.EMX1-Detect-pRBS-4

description:
EMX1.All_rep1.EMX1-Detect-pRBS-4 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: RNA_ESC_WT_1

value: blastocyst-derived ES cells


title: 48hrAPF_Male_H3K9me3_1


title: 293T-EMX1.All_rep1.EMX1-Detect-pRBS-3

description:
EMX1.All_rep1.EMX1-Detect-pRBS-3 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 48hrAPF_Female_H3K9me3_4

value: Female


title: PC3_ecDNA_positive-PolII-ChIA-Drop_rep2

description: The PC3 was initiated from a bone metastasis of a grade IV prostatic adenocarcinoma from a 62-year-old male Caucasian.

data_processing:
Library strategy: ChIA-Drop
Illumina RTA-1.18 software used for basecalling.
We use the ChIA-DropBox (CDB), a data-processing and visualization pipeline for multiplex chromatin interaction analysis, to process the raw reads into chromatin complex interaction data. Briefly, CDB aligns the reads to the reference genome (hg19) using 10X Genomics Long Ranger pipeline (v2.1.5) and identify their corresponding droplet-specific barcode. The uniquely mapped reads with MAPQ ≥ 30 and read length ≥ 50 bp are extended by 500 bps from its 3’ end, and those with the same droplet-specific barcode within 3kb distance were merged. The collection of all merged reads with the same droplet-specific barcode represents the DNA fragments interacting simultaneously at a single-chromatin complex. 2D interaction heatmap of this data is generated by CDB via Juicer (v1.22.01) and JuiceBox (v1.11.08). Multiplex interactions are visualized with ChIA-view (v1.0).
Genome_build: hg19 (GRCh37)
Supplementary_files_format_and_content: *_processed.txt: Tab-delimited text files.

source_name: PC3 from NCI60

growth_protocol: PC3 cell lines are cultured in F-12K medium (30-2004, ATCC) with 10% FBS.

extract_protocol:
2 million cells were crosslinked, permeabilized and digested following the ChIA-PET protocol except the HindIII enzyme (R0104M, NEB) was used instead of AluI. The digested nuclei were collected for chromatin sonication followed by immunoprecipitation with anti-RNAPII antibody (920102, Biolegend). RNAPII-enriched chromatin was collected with EB Buffer and cleared through the Amicon Ultra-0.5 100 K column (Z740183-96EA, MilliporeSigma) following the manufacture’s instruction to remove small fragments.
The RNAPII-associated chromatin complexes were adjusted to a concentration of 0.5 ng/μl and proceeded to ChIA-Drop library construction using Chromium Genome v2 Library Kit & Gel Bead Kit (PN-120258, 10X Genomics) following the manufacturer’s protocol. The final libraries were sequenced on Illumina platform with 150 bp pair-end sequencing.

value: RNA polymerase II (920102, Biolegend)


title: Co-infection rep3

city: Fuzhou

first: Song

state: Fujian

address: Cangshan District

institute: Fujian Agriculture and Forestry University

data_processing:
Sequenced reads were mapped to GRCm38 whole genome using hisat2 V2.1.0
Differential gene expression was analyzed using DESeq2 V1.10.1
Goatools V0.6.5 was used for gene set analysis
Genome_build: GRCm38
Supplementary_files_format_and_content: tab-delimited files include tpm values
Supplementary_files_format_and_content: matrix table with raw gene counts
Supplementary_files_format_and_content: a FASTA file with the transcript sequences for all MSTRG* IDs

source_name: Lung tissue

extract_protocol:
Mice lungs were removed, flash frozen on dry ice, and RNA was harvested using Trizol reagent. Illumina TruSeq RNA Sample Prep Kit (Cat#FC-122-1001) was used with 1 ug of total RNA for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: Mice were mock infected or infected with H9N2 AIV or E. coli alone, or co-infected with these two pathogens for 48h.

value: BALB/c

value: lung

value: Seven-week-old


title: RNA_ESC_Mettl3KO_1

value: Mettl3 KO


title: Co-infection rep2


title: 293T-EMX1.All_rep1.EMX1-Detect-pRBS-21

description:
EMX1.All_rep1.EMX1-Detect-pRBS-21 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: Control_exp2_2

city: Moscow

first: Natalia

last: Klimenko

address: 34/5 Vavilova Street

country: Russia

institute: Institute of Gene Biology (IGB) of the Russian Academy of Sciences

data_processing:
Cutadapt software (Martin 2011) was used to remove the adapters and poly-A tails and trimming the low-quality ends (quality threshold was set to 20); the reads shorter than 20 bp post-trimming were discarded.
The processed reads were aligned to Homo sapiens genome (assembly GRCh38.p13) using the splice-aware STAR algorithm (Dobin et al. 2013).
The gene read counts were calculated using the STAR quantMode option (--quantMode GeneCounts).
Genome_build: hg38
Supplementary_files_format_and_content: Raw reads counts per gene obtained with STAR

source_name: melanoma cells

growth_protocol: Human melanoma A375 cell lines (from American Type Culture Collection, Manassas, VA) were propagated in Dulbecco modified Eagle's medium (DMEM; PanEco, Russia) with 10% fetal bovine serum (HyClone, Logan, UT) supplemented with 2 mM L-glutamine (Merck) and penicillin/streptomycin at 370C, 5% CO2 in a humidified atmosphere.

extract_protocol:
RNA was isolated from 3 millions of cell with TriReagent (MRC) according to the manufacturer's protocol.
Total RNA from A375 cells after knockdown of MYC or PHF10 (isolated as described above) was treated with DNase I (Thermo Fisher Sci.) at 370C for 15 min followed by phenol-chloroform extraction and 96% ethanol precipitation. For NGS library preparation, one µg of RNA and NEBNext Ultra II Directional RNA library prep kit for Illumina with NEBNext Poly(A) mRNA magnetic isolation module (NEB) were used according to the manufacturer’s protocol. Adaptor ligation was performed using NEBNext multiplex oligos for Illumina index primer set #1 (NEB). Quality of RNA and cDNA libraries was analyzed with Bioanalyzer RNA 6000 Nano Chip and Bioanalyzer DNA 1000 Chip kits (Agilent). RNA and DNA yields were assessed with Qubit assay kits (Thermo Fisher Sci.). Libraries were sequenced on an Illumina NovaSeq 6000 with single strand 100 bp length reads. Approximately 3x107 reads were obtained per one sample.

treatment_protocol: Knockdown of MYC and PHF10 in A375 was performed with 2 µl of siRNA (100pMol) and 10 µl Metafecten Pro (Biontex) per one 60 mm Petri dish. Sequences of MYC-siRNA, PHF10-siRNA and control-siRNA are listed in Supplementary material. siRNA and Metafecten Pro were mixed with 0.5 ml Opti-MEM media (Gibco) for 60 mm dish (1.3 ml for 90 mm dish) separately. Then the mixtures were pooled, incubated for 15 min at room temperature, and added to cells in the antibiotic free DMEM. Over four hours the media was replaced by new DMEM supplemented with 10% FBS. Cells were harvested 72 h after transfection.

tag: sirna

value: no treatment


title: Co-infection rep1


title: Control_exp2_1


title: 48hrAPF_Female_H3K9me3_2

description: Sequential Chromatag-ChIP for fru P1 neuron chromatin


title: 48hrAPF_Female_H3K9me3_3


title: H9N2 rep2


title: cMYC_exp2_2

value: cMyc knockdown


title: 293T-EMX1.All_rep1.EMX1-Detect-pRBS-15

description:
EMX1.All_rep1.EMX1-Detect-pRBS-15 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

value: embryonic kidney derived epithelial cells

value: Wild Type


title: H9N2 rep1


title: 293T-EMX1.All_rep1.EMX1-Detect-pRBS-14

description:
EMX1.All_rep1.EMX1-Detect-pRBS-14 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 293T-EMX1.All_rep1.EMX1-Detect-pRBS-20

description:
EMX1.All_rep1.EMX1-Detect-pRBS-20 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 48hrAPF_Female_H3K9me3_1


title: cMYC_exp2_1


title: E. coli rep3


title: 48hrAPF_Male_H3K4me3_4


title: 293T-EMX1.All_rep1.EMX1-Detect-pRBS-11

description:
EMX1.All_rep1.EMX1-Detect-pRBS-11 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: Control_exp1_2


title: 48hrAPF_Male_H3K4me3_3


title: 293T-EMX1.All_rep1.EMX1-Detect-pRBS-10

description:
EMX1.All_rep1.EMX1-Detect-pRBS-10 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: Control_exp1_1


title: PC3_ecDNA_positive-PolII-ChIA-Drop_rep1


title: E. coli rep2


title: E. coli rep1


title: 48hrAPF_Male_H3K4me3_2


title: 293T-EMX1.All_rep1.EMX1-Detect-Potential-2

description:
EMX1.All_rep1.EMX1-Detect-Potential-2 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: H9N2 rep3


title: PHF10_total_exp1_4

value: PHF10 knockdown


title: 293T-EMX1.All_rep1.EMX1-Detect-Potential-1

description:
EMX1.All_rep1.EMX1-Detect-Potential-1 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 48hrAPF_Male_H3K4me3_1


title: Mock rep3


title: 48hrAPF_Female_H3K4me3_4


title: Mock rep2


title: PHF10_total_exp1_1


title: Mock rep1


title: 293T-EMX1.All_rep1.EMX1-Detect-Extend-6

description:
EMX1.All_rep1.EMX1-Detect-Extend-6 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: PHF10_total_exp1_2


title: 48hrAPF_Female_H3K4me3_3


title: N2-iN-rep2

city: Taipei

first: Yu-Hui

last: Wong

address: No.155, Sec.2, Linong Street

institute: National Yang-Ming University

data_processing:
N1-iMG and N2-iMG libraries were sequenced using the Illumina HiSeq 4000 system; N2-iPSC and N2-iN libraries were sequenced using the Illumina NovaSeq 6000 system. Image analysis and base calling were conducted by the Control Software + OLB + GAPipeline-1.6 (Illumina) on the instrument.
Reads were first mapped to a reference genome of human (GRCh37; hg19) using HISAT2 (version 2.0.1).
Per-gene expression levels (read counts) were quantitated using HTSEQ (v 0.6.1).
According to the read counts quantitated by HTSEQ, the fragments per kilobase of transcript per million mapped reads (FPKM) was calculated by the formula “total exon fragments / (mapped reads [millions] x exon length [KB])”.
FPKM from different experiments was then normalized using Median of ratios method (DESeq2, v 1.6.3).
Genome_build: hg19
Supplementary_files_format_and_content: tab-delimited text files including FPKM values of each gene for each sample

source_name: induced neurons

growth_protocol:
[iMG] HiPSCs that had been infected with both FUW-rtTA and pTetO-CEBPA-T2A-SPI1-T2A-Puro were seeded on day -1.  On day 0, the cells had reached at least 70% confluency; at this point the culture medium was replaced with DMEM/F12 (Gibco) supplemented with N2 (Gibco) and non-essential amino acids (NEAA, Gibco). The medium also contained human BMP4 (50 ng/ml, PeproTech), basic FGF (bFGF/FGF2 50 ng/ml, PeproTech) and activin-A (20 ng/ml, PeproTech).  Doxycycline (2 μg/ml, Sigma) was added to promote TetO downstream gene expression and retained in the medium until the end of the experiment.  On day 1, the medium was replaced with DMEM/F12/N2/NEAA containing human VEGF (50 ng/ml, PeproTech), SCF (50 ng/ml, PeproTech) and FGF2 (20 ng/ml).  Puromycin (1 μg/ml, Gibco) was also added for 24 hours to select for successfully infected cells.  On day 2, the medium was replaced with DMEM/F12/N2/NEAA containing human IL-34 (10 ng/ml, PeproTech), M-CSF (10 ng/ml, PeproTech) and TGFβ-1 (10 ng/ml, PeproTech).  Two days later, the medium was replaced with DMEM/F12/N2/NEAA containing human IL-34 (100 ng/ml), M-CSF (20 ng/ml) and TGFβ-1 (20 ng/ml).  After day 4, half medium was removed and replaced with fresh DMEM/F12/N2/NEAA containing human IL-34 (100 ng/ml), M-CSF (20 ng/ml), GM-CSF (20 ng/ml, PeproTech) and TGFβ-1 (20 ng/ml); this was repeated every 3 days from this point onwards. Cells were assayed on days 9.
[iPSCs] Human iPSCs were routinely maintained in Essential 8™ (E8) medium (Gibco, USA) on vitronectin (VTN-N, Gibco, USA) coated dishes at 37°C in a 5% CO2 incubator.
[iN] HiPSC-derived neurons were generated using a published protocol (Zhang et al., 2013) with modifications.  HiPSCs that had been infected with FUW-rtTA and pTetO-Ngn2-puro were seeded on day -1.  On day 0, when the cells reached at least 70% confluency, the culture medium was replaced with DMEM/F12 supplemented with N2, NEAA, human BDNF (10 ng/ml, PeproTech), human NT-3 (10 ng/ml, PeproTech) and mouse laminin (0.2 μg/ml, Gibco).  Doxycycline (2 μg/ml) was added to induce TetO-regulated Ngn2 expression and retained in the medium until the end of the experiment.  On day 1, puromycin (1 μg/ml) was added to kill any uninfected cells.  On day 2, puromycin was removed and the cells were replated in Neurobasal medium supplemented with B-27 supplement (Gibco), GlutaMAX (Gibco), mouse laminin (0.2 μg/ml), human BDNF (10 ng/ml) and NT-3 (10 ng/ml) on poly-L-lysine and laminin coated dishes (~105 cells/mL).  On day 4, cytosine arabinoside (Ara-C, 2 μM, Sigma) was added to the medium to inhibit non-neuronal cell proliferation and was then removed on day 6.  After day 6, 50% of the medium was changed every 4 days. Cells were assayed on days 20.

extract_protocol:
Total RNA was isolated using a Tissue Total RNA Mini Kit (Geneaid, Taiwan) by following the manufacturer’s instructions. 2 μg of total RNA with a RIN value above 9.7 was used to prepare the appropriate libraries.
Next generation sequencing library preparation was constructed according to the manufacturer’s protocol (NEBNext® Ultra™ RNA Library Prep Kit for Illumina®). The poly(A) mRNA isolation was performed using  a NEBNext Poly(A) mRNA Magnetic Isolation Module (NEB).  The mRNA fragmentation and priming was performed using NEBNext RNA First Strand Synthesis Reaction Buffer and NEBNext Random Primers.  First strand cDNA was synthesized using ProtoScript II Reverse Transcriptase and the second-strand cDNA was synthesized using Second Strand Synthesis Enzyme Mix.  The purified double-stranded cDNA obtained by AxyPrep Mag PCR Clean-up (Axygen) was then treated with End Prep Enzyme Mix to repair both ends and add a dA-tail; this was done in a single reaction, and was followed by a T-A ligation to add adaptors to both ends.  Size selection of Adaptor-ligated DNA was then performed using AxyPrep Mag PCR Clean-up (Axygen), and fragments of ~360 bp (with the approximate insert size being 300 bp) were recovered.  Each sample was then amplified by PCR for 11 cycles using the P5 and P7 primers, with both primers carrying sequences that are able to anneal with the flow cell in order to perform bridge PCR and P7 primer carrying a six-base index allowing for multiplexing.  The PCR products were cleaned up using AxyPrep Mag PCR Clean-up (Axygen), validated using an Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA, USA), and quantified by Qubit 2.0 Fluorometer (Invitrogen, Carlsbad, CA, USA).

value: Neurons derived from iPSC


title: 48hrAPF_Female_H3K4me3_2


title: 293T-EMX1.All_rep1.EMX1-Detect-Extend-4

description:
EMX1.All_rep1.EMX1-Detect-Extend-4 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

value: Targeted Amplicon Sequencing


title: N2-iN-rep1


title: 293T-EMX1.All_rep1.EMX1-Detect-Extend-5

description:
EMX1.All_rep1.EMX1-Detect-Extend-5 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

value: BE4max + sgRNA (EMX1)


title: 48hrAPF_Female_H3K4me3_1


title: differentiated

city: West Lafayette

first: Nadia

middle: Marie

last: Atallah

state: IN

address: 201 S. University St

department: Center for Cancer Research

institute: Purdue University

data_processing:
FastQC v.0.11.3 was used to observe read quality
Cutadaptversion 2.2 to trim data
Bowtie 2.3.3 to align data
Genome_build: GRCm38.93
Supplementary_files_format_and_content: TSV of methylated sites

source_name: whole cell

growth_protocol: ESCs were cultured and maintained in gelatin-coated tissue culture plates

extract_protocol: Genomic DNA was isolated using standard phenol:chloroform extraction, followed by ethanol precipitation. DNA from various samples was digested with FspEI for 4 hours at 37°C and subjected to electrophoresis in a 2% agarose gel. FspEI recognizes CmC and makes double stranded break at N12/N16 position from the modified cytosine. At a CpG sites FspEI cleaves bidirectionally releasing 31 or 32-bp fragments. The DNA around 30-bp was excised out of the agarose gel and purified using modified freeze and squeeze technique (Figure S1A)42. In brief, the excised gel pieces were placed inside a 0.65 mL micro centrifuge tube with a small hole in the bottom of the tube which was made using a 20-gauge needle. The 0.65 mL tube containing the gel piece was then placed in a 1.5 mL micro centrifuge tube and flash frozen in liquid nitrogen. The samples were then thawed at room temperature and centrifuged at 15,000 rpm for 10 minutes. This extrudes the DNA along with the buffer into the 1.5 mL tube leav  ing behind agarose in the small tube. The DNA was extracted by ethanol precipitation, resuspended in TE and quantified by nanodrop. This DNA was further used for adaptor ligation at 4°C overnight22. The ligated DNA was PCR amplified with index primers, purified using QIAquick PCR purification kit and sequenced using Novaseq 6000.

treatment_protocol: cell type

value: embryonic stem cells

tag: state

value: differentiated


title: 293T-EMX1.All_rep1.EMX1-Detect-Extend-21

description:
EMX1.All_rep1.EMX1-Detect-Extend-21 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: N2-iPSC-rep2

source_name: human induced pluripotent stem cells

value: Human induced pluripotent stem cells


title: 293T-EMX1.All_rep1.EMX1-Detect-Extend-1

description:
EMX1.All_rep1.EMX1-Detect-Extend-1 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

source_name: HEK293T cells


title: PHF10_total_exp1_3


title: 48hrAPF_Male_H3K36me3_3

value: Male

value: fru P1


title: differentiated treated with PG


title: N2-iPSC-rep1


title: 293T-EMX1.All_rep1.EMX1-Detect-Extend-14

description:
EMX1.All_rep1.EMX1-Detect-Extend-14 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: undifferentiated

value: undifferentiated


title: 48hrAPF_Male_H3K36me3_2


title: 293T-EMX1.All_rep1.EMX1-Detect-Extend-10

description:
EMX1.All_rep1.EMX1-Detect-Extend-10 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: RNA_ICM_Ythdc1KO_2

data_processing:
Primary fastq data were generated with the Illumina Casava (v 1.8.2) pipeline.
Sequencing reads were trimmed by Trim Galore (v 0.6.4) with parameters: --phred33, --illumina, --clip_R1 9, --clip_R2 9, --paired to remove adapter sequences.
Trimmed reads were aligned to mm9 reference genome using Tophat2 (v2.1.1) with default parameters.
Expression levels of individual genes were quantified by FPKM using Cufflinks (v 2.2.1).
Genome_build: mm9
Supplementary_files_format_and_content: Tab-delimited text files include FPKM values for each transcript.

source_name: preimplantation embryo

growth_protocol: Embryos were cultured in G-1 PLUS medium.

extract_protocol:
300 ICM cells per reaction were disrupted in TRIzol Reagent and total RNAs were isolated by chloroform extraction coupled with isopropanol precipitation.
RNAs extracted from ICM cells were subjected to rRNA-free library generation using SMARTer Stranded Total RNA-Seq Kit.

treatment_protocol: To isolate ICM cells, blastocysts were incubated in calcium-free CZB medium for 30 min to disrupt the cell-cell junction. ICM cells were then distinguished and collected according to their sizes and shapes.

tag: development stage

value: E4.5 inner cell mass

value: Ythdc1 knockout


title: RNA_ICM_Ythdc1KO_1


title: 293T-EMX1.All_rep1.Control-8

description:
EMX1.All_rep1.Control-8 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

data_processing:
Library strategy: Targeted amplicon sequencing
Cut adapter step: The raw FASTQ files are firstly but the sequencing adapters with cutadapt software with the following command: "cutadapt --times 1  -e 0.1  -O 3  --quality-cutoff 25  -m 100 -a  AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC  -A  AGATCGGAAGAGCGTCGTGTAGGGAAAGAGTGTAGATCTCGGTGGTCGCCGTATCATT -o {read1.out}  -p {read2.out}  {read1.in}  {read2.in}"
Alignment step: All FASTQ files are aligned by bwa mem (version = 0.7.15) with default parameters. More information please query www.detect-seq.com
Genome_build: hg38
Supplementary_files_format_and_content: Processed data contains the primer and experiment design information of all target amplicon sequencing data.

extract_protocol:
Genomic DNA was freshly extracted using CWBIO universal genomic DNA kit (CWbiotech, CW2298M) and stored in TE buffer (10 mM Tris-HCl, 1 mM EDTA, pH 8.0) at -80 °C.
Regions flanking the targeting sites were selected for the design of primers, whose overhangs contained the paired Illumina adaptor sequences. In addition, a 10-nt barcode was also added into each primer pairs to lower the detection limit from 10-1 to 10-5 (Primers are listed in the Supplementary). The first round of PCR amplification was performed with NEBNext® Q5® Hot Start HiFi PCR Master Mix (NEB, M0543L) using ~10 ng of genomic DNA as an input template. After ~10 cycles of amplification, the PCR products were purified with 1 × Agencourt AMPure XP beads and eluted with ddH2O. Purified DNA samples were then subjected to the second round of amplification for ~15 cycles and assigned with different indexes followed by a purification with 0.8 × AMPure XP beads. The libraries were quantified with Qubit 2.0 Fluorometer (INVITROGEN) and pooled together for high-throughput sequencing by Illumina HiSeq X Ten (Genetron Health, Beijing).    All FASTQ files in this dataset, have already been grouped by barcode. Meanwhile, the Illumina sequencing errors were trying to be corrected as well (decrease error rate from ~0.1% to ~0.005%). The Python scripts used in this step are available by request to correspondents.

tag: cell type


title: RNA_ICM_Ctrl_2

tag: genotype

value: wild type


title: 48hrAPF_Male_H3K36me3_4


title: RNA_ICM_Ctrl_1


title: D546 MDDC NF110b WT

first: Aderem

last: Lab

address: 307 Westlake Avenue North, Suite 500

institute: Seattle Children's Research Institute

description:
rawCounts.tsv
D546-NF110b-Mock

data_processing:
Reads with more that 67% identical bases were discarded prior to alignment.
The remaining read pairs were aligned to the human genome (hg19, GRCh37 Genome Reference Consortium Human Reference 37 (GCA_000001405.1)) using the gsnap aligner (v. 2016-08-24) allowing for novel splicing.
Concordantly mapping read pairs (average 17-million / sample) that aligned uniquely were assigned to exons using the subRead program and gene definitions from GRCh38.87.
Genes with low expression were filtered using the filterByExpr function in the edgeR package from Bioconductor.org resulting in a total of 13,125 genes in the final dataset.
Differential expression was calculated using the edgeR package.
Genome_build: hg19
Supplementary_files_format_and_content: rawCounts.tsv contains the raw counts for each sample generated by subRead, tab-separated

source_name: Monocyte-Derived Dendritic Cells

growth_protocol: 2x10^5 CD14+ magnetically-isolated monocytes were incubated in RPMI containing 10% heat-inactivated fetal bovine serum, 50 U/mL penicillin, 50 μg/mL streptomycin, 10 mM HEPES, 2-Mercaptoethanol, and 2 mM L-glutamine, in the presence of recombinant human GM-CSF at 10 ng/mL and IL-4 at 50 ng/mL. Virus-like particules containing VPX were provided in trans to allow for transduction with either control (LKO), NF110b WT, or NF110ΔDZF lentivirus (10uL). Cells were fed the next day with half volume of IL-4/GM-CSF. Cells taken for RNA isolation on Day 5 post-transduction.

extract_protocol:
RNA was harvested on Day 5 post-transduction using Trizol reagent.Illumina TruSeq stranded mRNA sequencing (100-PE w/base trimming after 150 PE ~40M reads 6Gb production target) was performed by Psomagen (Rockville,MD) was used with 500 ng of total RNA for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols by Psomagen.

value: Monocyte-Derived Dendritic Cells

tag: overexpression

value: NF110b WT

tag: donor


title: 48hrAPF_Female_H3K36me3_3

data_processing:
ChIP-seq reads were pre-processed using Trimmomatic’s ILLUMINACLIP (v. 0.33) option to remove Illumina adaptor sequences. Pre-processed ChIP-seq reads were aligned to the D. melanogaster dm6 UCSC genome assembly using Bowtie2 (v.2.2.5), allowing up to 6 mismatches. Only uniquely aligned reads were considered and individual replicate and pooled replicate (all replicates) BAM files were generated with SAMtools (v. 1.2) for downstream analyses.
MACS2 (v.2.1.0) peaks were called for each histone modification (H3K27ac, H3K36me3, H3K4me3, H3K9me3, H3K27me3) at each time point (48hr APF, 1-day adult, 10-12 day adult) for each sex in fru P1 or elav neurons, relative to sex and stage-specific input chromatin library controls for individual and all (pooled) replicates. We performed “narrow” peak calling for histone modifications with more compact enrichment patterns (H3K27ac and H3K4me3) and the “broad” peak option for modifications with diffuse enrichment (H3K36me3, H3K9me3, and H3K27me3) on all individual biological replicates and on all (pooled) replicate data using the following “callpeak” parameters: --gsize dm -p 1e-2 --call-summits --fix-bimodal.
Input normalized BigWig files for all histone modifications for all replicate (pooled) data were obtained using Deeptools bamCompare function to subtract input ChIP-seq signal from histone modification .BAM files obtained from concatinating individual replicate .BAM files.
Genome_build: UCSC dm6 (BDGP Release 6 plus ISO1 MT)

extract_protocol: The adult heads were separated from their bodies by vortexing frozen animals. The heads were sifted through liquid nitrogen-cooled mesh sieves #25 and #40 (with heads retained in the #40 sieve). The sequential ChIP methodology presented here is based on a previously published ChIP protocol (Negre et al. 2006). Frozen adult heads or frozen whole body pupae (~0.3g) were dounce homogenized in 5ml of Buffer A1 (60mM KCL, 15mM NaCl, 4mM MgCl2, 15mM HEPES (pH 7.6), 0.5% TritonX-100, 0.5mM DTT, 10mM sodium butyrate, and Roche cOmplete Protease Inhibitor cocktail EDTA-free, product 5056489001), with 1% methanol-free formaldehyde (Pierce, 20% solution product 28906) for crosslinking at room temperature for 10 min. The crosslinking reaction was quenched using 540\265l 2.5M Glycine and incubated for 5 minutes at room temperature. Cross-linked homogenates were centrifuged at 4000g for 5 minutes at 4\272C and the nuclear pellets were resuspended in Buffer A1. The nuclear pellets were washed 3 more times by centrifugation at 4000g for 5 minutes at 4\272C with resuspension in 3ml of buffer A1. The nuclear pellets were washed an additional time with 1x Lysis Buffer without SDS (140mM NaCl, 15mM HEPES (pH 7.6), 1mM EDTA (pH 8.0), 0.5mM EGTA, 1% TritonX-100, 0.5mM DTT, 0.1% sodium deoxycholate, 10mM sodium butyrate, and Roche cOmplete Protease Inhibitor Cocktail EDTA-free) to remove all traces of Buffer A1 before the next lysis step. After one additional spin at 4000g for 5 minutes at 4\272C, the washed nuclear pellet was resuspended in Lysis Buffer containing 1% SDS and 0.5% N-Laurosylsarcosine and incubated at 4\272C, rotating for 1 hour. Nuclear lysates were quickly spun down for 10 seconds at 2000g and supernatant transferred to sonication tubes (Covaris; product 520130) for shearing. Chromatin was sheared for 25 minutes using a Covaris E220 (duty cycle: 5.0, peak incident power: 140, cycles per burst: 200). The sheared chromatin was transferred to microcentrifuge tubes and rotated for 10 min. at 4\272C. Next, the sheared chromatin was centrifuged at 18,000g for 5 minutes at 16 \272C. The supernatant was transferred to a new microcentrifuge tube and the centrifugation step was repeated twice. The final supernatant was split into two 0.5ml aliquots (material from ~0.15g frozen tissue) and brought to a 1.5ml volume with 1x Lysis Buffer without SDS. The sheared chromatin was then aliquoted into 250\265l volumes and stored at -80\272C. To perform the sequential ChIP, 750\265l of 1x Lysis Buffer without SDS was added to the 250\265l of the thawed sheared chromatin. The sheared chromatin was incubated with 50\265l of magnetic protein A/G beads (Pierce, product 88803) and 5\265g of each respective antibody at 4\272C overnight. The magnetic protein A/G beads were then washed four times with 1ml of Lysis Buffer wash (140mM NaCl, 15mM HEPES (pH 7.6), 1mM EDTA (pH 8.0), 0.5mM EGTA, 1% TritonX-100, 0.05% SDS, 0.5mM DTT, 0.1% sodium deoxycholate, 10mM sodium butyrate, and Roche cOmplete Protease Inhibitor Cocktail EDTA-free) and twice with TE (10mM Tris-HCL (pH 8.0), 1mM EDTA) for 10 minutes each, rotating at 4 \272C. Beads were incubated in 100\265l ChIP elution buffer (100mM NaHCO3, 80mM NaOH, 1% SDS) at room temperature, rotating for 30-60 minutes to elute chromatin. The eluate was diluted 1:10 with Lysis Buffer without SDS and incubated on magnetic streptavidin beads (Invitrogen Dynabeads MyOne Streptavidin C1) for 4hrs-overnight rotating at 4\272C. The beads were washed as above and chromatin was reverse crosslinked on beads in ChIP elution buffer (with 0.04mg/ml proteinase K, 0.01mg/ml RNaseA) at 65 \272C overnight. DNA was purified with a DNA purification and concentration kit (Machery-Nagel, product 740609). ChIPseq libraries were prepared using the NEBNext ChIP-seq library preparation kit. Libraries were size selected using AMPure XP beads (Beckman) and clean up steps were performed using the Machery Nagel PCR cleanup kit. A final wash was performed using the AMpure XP beads according to kit protocol to remove primers.

tag: Sex

value: Female


title: 48hrAPF_Female_H3K36me3_4


title: 293T-EMX1.All_rep1.Control-9

description:
EMX1.All_rep1.Control-9 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

value: embryonic kidney derived epithelial cells

value: Wild Type

tag: experiment


title: D545 NF110bΔDZF

description:
rawCounts.tsv
D545-NF110-DZFm-Mock


title: D545 NF110b WT

description:
rawCounts.tsv
D545-NF110b-Mock


title: D545 MDDC LKO

description:
rawCounts.tsv
D545-LKO-Mock

value: Control (LKO)


title: 48hrAPF_Female_H3K36me3_1

description: Sequential Chromatag-ChIP for fru P1 neuron chromatin


title: 293T-EMX1.All_rep1.Control-6

description:
EMX1.All_rep1.Control-6 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

value: Targeted Amplicon Sequencing


title: 293T-EMX1.All_rep1.Control-5

description:
EMX1.All_rep1.Control-5 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

value: BE4max + sgRNA (EMX1)


title: 48hrAPF_Male_H3K27me3_4


title: RIP_ESC_Ythdc1_1

data_processing:
Primary fastq data were generated with the Illumina Casava (v 1.8.2) pipeline.
Sequencing reads were trimmed by Trim Galore (v 0.6.4) with parameters: --phred33, --illumina, --clip_R1 9, --clip_R2 9, --paired to remove adapter sequences.
Trimmed reads were aligned to mm9 reference genome using Tophat2 (v 2.1.1) with default parameters.
RIP peaks were defined by the callpeak function of MACS2 (v 2.1.1) with parameters: -g mm, -f BAM, --BDG, --QVALUE 0.05, --SPMR.
Genome_build: mm9
Supplementary_files_format_and_content: Bigwig files containing piled up reads.  Scores represent normalized tag counts per million reads.

extract_protocol:
5×10^6 control ES cells per reaction were cross-linked in 0.3% formaldehyde and neutralized by 1×glycine. The nuclei were isolated and IP reaction was performed using Magna Nuclear RNA-Binding Protein Immunoprecipitation Kit. RNAs of input and IP samples were extracted using TRIzol Reagent and chloroform coupled with isopropanol precipitation.
RNAs extracted from input and IP samples were subjected to library generation using VAHTS Total RNA-seq (H/M/R) Library Prep Kit.

tag: strain

value: blastocyst-derived ES cells

value: Ythdc1 flox/flox

tag: rip antibody

value: anti-YTHDC1 (Abcam ab122340)


title: 48hrAPF_Female_H3K36me3_2

tag: developmental stage

tag: chip antibody


title: RIP_ESC_Ythdc1_2


title: 293T-EMX1.All_rep1.Control-4

description:
EMX1.All_rep1.Control-4 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: D546 MDDC NF110bΔDZF

description:
rawCounts.tsv
D546-NF110-DZFm-Mock


title: D544 MDDC NF110bΔDZF

description:
rawCounts.tsv
D544-NF110-DZFm-Mock


title: ChIP_ICM_Ythdc1KO_K9_2

extract_protocol:
500 ICM cells per reaction were lysed and chromatin was sheared by MNase. 1/20 of the product was saved as the input, and the rest was mixed with antibody-bound Dynabeads Protein A/G for IP reaction. DNAs of input and IP samples were extracted using phenol chloroform coupled with isopropanol precipitation.
DNAs purified from input and IP samples were subjected to library generation using HyperPlus Library Preparation Kit.

tag: genetype

value: anti-H3K9me3 (Abcam ab8898)


title: D546 MDDC LKO

description:
rawCounts.tsv
D546-LKO-Mock


title: 48hrAPF_Male_H3K36me3_1

value: Male

value: fru P1


title: ChIP_ICM_Ythdc1KO_input

value: E4.5 inner cell mass


title: 293T-EMX1.All_rep1.Control-7

description:
EMX1.All_rep1.Control-7 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 48hrAPF_Male_H3K27me3_3


title: RIP_ESC_input


title: ChIP_ICM_Ythdc1KO_K9_1

value: Ythdc1 knockout


title: D544 MDDC LKO

description:
rawCounts.tsv
D544-LKO-Mock


title: 293T-EMX1.All_rep1.Control-2

description:
EMX1.All_rep1.Control-2 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: D543 MDDC NF110bΔDZF

description:
rawCounts.tsv
D543-NF110-DZFm-Mock


title: 48hrAPF_Male_H3K27me3_1


title: 48hrAPF_Male_H3K27me3_2


title: 48hrAPF_Female_H3K27me3_3


title: D543 MDDC NF110b WT

description:
rawCounts.tsv
D543-NF110b-Mock


title: 293T-EMX1.All_rep1.Control-16

description:
EMX1.All_rep1.Control-16 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 48hrAPF_Female_H3K27me3_4


title: 293T-EMX1.All_rep1.Control-18

description:
EMX1.All_rep1.Control-18 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 293T-EMX1.All_rep1.Control-15

description:
EMX1.All_rep1.Control-15 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 48hrAPF_Female_H3K27me3_1


title: PC3_ecDNA_negative_En-circles_RNAseq_Rep2

description:
PC3_ecDNA_negative_En-circles_vs_Ctrl-circles.gene_exp.diff
mRNA-seq

data_processing:
The raw sequencing reads were trimmed using Trim Galore version 0.4.3 (options: --stringency 3 -q 20 -e .20 --length 15 --paired) and aligned to the hg19 genome using hisat 2.1.0 (options: --dta-cufflinks). The transcripts were assembled using Cufflinks 2.2.1 and the differential gene expression analysis was performed using Cuffdiff (options: --library-type fr-firststrand).
Illumina RTA-1.18 software used for basecalling
Genome_build: hg19

source_name: PC3(CRL1435, ATCC)

growth_protocol: The circular enhancer DNA (En-circles) or negative control (Ctrl-circles) were purified and pooled at equal molar ratio. 500 ng of En-circles or Ctrl-circles were transfected separately into ecDNA-negative PC3 cells in triplicate using Lipofectamine 3000 (Invitrogen) as per manufacturer’s protocol. Briefly, ecDNA-negative PC3 cells were plated to obtain 70-80% confluence on the following day and were allowed to attach overnight. On the next day, En-circles plasmid DNA was diluted in Opti-MEM reduced serum medium along with P3000 transfection reagent and mixed with Opti-MEM medium containing Lipofectamine 3000. DNA-lipid complexes were allowed to form at room temperature for 15 minutes and mixtures were added to cells. Cells were incubated at standard conditions and harvested 24 hours post-transfection for RNAseq analysis.

extract_protocol:
Total RNA was isolated in biological replicates using AllPrep DNA/RNA Mini Kit (80204, QIAGEN).
Strand-specific RNA libraries were generated from 300 ng of total RNA following the manuals of KAPA Stranded mRNA Sequencing Kit (KK8502, KAPA Biosystems). Libraries were sequenced on Illumina platform.

value: The PC3 was initiated from a bone metastasis of a grade IV prostatic adenocarcinoma from a 62-year-old male Caucasian.


title: D543 MDDC LKO

description:
rawCounts.tsv
D543-LKO-Mock

value: Monocyte-Derived Dendritic Cells


title: ChIP_ICM_Ctrl_input

data_processing:
Primary fastq data were generated with the Illumina Casava (v 1.8.2) pipeline.
Sequencing reads were trimmed by Trim Galore (v 0.6.4) with parameters: --phred33, --illumina, --clip_R1 9, --clip_R2 9, --paired to remove adapter sequences.
Trimmed reads were aligned to mm9 reference genome using Bowtie2 (v 2.2.9) with default parameters.
ChIP peaks were defined by the callpeak function of MACS2 (v 2.1.1) with parameters: -g mm, -f BAM, --BDG, --QVALUE 0.05, --SPMR.
Genome_build: mm9
Supplementary_files_format_and_content: Bigwig files containing piled up reads.  Scores represent normalized tag counts per million reads.

value: wild type


title: 48hrAPF_Male_H3K27ac_4


title: ChIP_ICM_Ctrl_K9_2


title: PC3_ecDNA_negative_Ctrl-circles_RNAseq_Rep3


title: ChIP_ESC_Ythdc1KO_Kap1_2

description: 2.5 μM 4-OHT was added into the culture medium 72 hr before harvesting.

extract_protocol:
5×10^6 ES cells per reaction were fixed in 1% formaldehyde and neutralized by 125 mM glycine. The crosslinked cells were pelleted, lysed and subjected to sonication. 1/100 of the product was saved as the input, and the rest was mixed with antibody-bound Dynabeads Protein A/G for IP reaction. DNAs of input and IP samples were extracted using phenol chloroform coupled with isopropanol precipitation.
DNAs purified from input and IP samples were subjected to library generation using HyperPlus Library Preparation Kit.

value: Ythdc1 flox/flox; CreERT2

value: anti-KAP1 (Abcam ab22553)


title: ChIP_ESC_Ythdc1KO_K9_2


title: 293T-EMX1.All_rep1.Control-17

description:
EMX1.All_rep1.Control-17 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

value: embryonic kidney derived epithelial cells

value: Wild Type


title: 48hrAPF_Male_H3K27ac_2


title: ChIP_ICM_Ctrl_K9_1


title: 48hrAPF_Female_H3K27me3_2

value: Female


title: D544 MDDC NF110b WT

description:
rawCounts.tsv
D544-NF110b-Mock

value: NF110b WT


title: PC3_ecDNA_negative_En-circles_RNAseq_Rep3


title: PC3_ecDNA_negative_Ctrl-circles_RNAseq_Rep1


title: 48hrAPF_Male_H3K27ac_1


title: 48hrAPF_Female_H3K27ac_4


title: 293T-EMX1.All_rep1.Control-10

description:
EMX1.All_rep1.Control-10 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

value: Targeted Amplicon Sequencing

value: BE4max + sgRNA (EMX1)


title: ChIP_ESC_Ythdc1KO_input

value: blastocyst-derived ES cells


title: ChIP_ESC_ff_K9_2

value: Ythdc1 flox/flox

value: anti-H3K9me3 (Abcam ab8898)


title: 293T-EMX1.All_rep1.Control-1

description:
EMX1.All_rep1.Control-1 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: ChIP_ESC_ff_K9_1


title: PC3_ecDNA_negative_En-circles_RNAseq_Rep1


title: 293T-EMX1.All_rep1.Control-14

description:
EMX1.All_rep1.Control-14 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: 293T-EMX1.All_rep1.Control-3

description:
EMX1.All_rep1.Control-3 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: ChIP_ESC_Ythdc1KO_K9_1


title: MCF7-sgRNADel-PD_rep1

description:
replicate 1
Detect-seq sgRNADel Prep1 in MCF-7 cells

data_processing:
Library strategy: Detect-seq
Detect-seq raw FASTQ files are remove Illumina sequencing adapters with cutadapt software (version 1.18). Working command and key parameters are: cutadapt --times 1 -e 0.1 -O 3 --quality-cutoff 25 -m 50. After adapter removal, FASTQ files are mapped to the reference genome (hg38) with Bismark(version 0.22.3) aligner without special settings. Then the unmapped and low MAPQ (MAPQ < 20) FASTQ reads are re-mapped by BWA MEM (version 0.7.17) with default behavior. The Bismark-generated BAM and BWA-generated BAM are merged, then sorted by reference coordinate with samtools sort command (version 1.9). Sored BAM files are removed duplications by Picard MarkDuplicates (version 2.0.1) with default settings. Considering of CBE usually generate indel byproducts, BAM files are needed to be re-aligned with GATK RealignerTargetCreator and IndelRealigner (version 3.8.1) with default settings, and SNP know sites are download from NCBI dbSNP.
After Detect-seq mapping steps, several computation and analysis steps should perform to achieve final Detect-seq off-targets. While making it an easy-achieve analysis pipeline, we offer Detect-seq tools on GitHub (https://github.com/menghaowei/Detect-seq), which contains several useful Python scripts. And those tools can help to perform Detect-seq analysis like off-targets finding, sgRNA alignment, and result visualization.
Genome_build: hg38
Supplementary_files_format_and_content: The bed files are the final off-target results generated by the Detect-seq protocol

source_name: MCF-7 cells

extract_protocol:
Genomic DNA was freshly extracted using CWBIO universal genomic DNA kit (CWbiotech, CW2298M) and stored in TE buffer (10 mM Tris-HCl, 1 mM EDTA, pH 8.0) at -80 °C.
Extracted genomic DNA was fragmented into ~300 bp through Covaris Focused-ultrasonicator™ Instrument (ME220), followed by a purification using DNA Clean & Concentrator-5 (Vistech, DC2005). ~5 ug DNA fragments and 10 pg spike-in model sequences were subjected to end repair with NEBNext End Repair Module (NEB, E6050), and 1 μl E. coli ligase (NEB, M0205) was added to repair nicks in DNA. Then hydroxylamine protection of endogenous 5fC was performed in 100 mM MES buffer (pH 5.0), 10 mM O-ethylhydroxylamine (EtONH2, Aldrich, 274992) at 37 °C for 6 h. The product underwent 5fC protection was diluted 2.5 times and purified with DNA Clean & Concentrator-5. Next, dA-tailing procedure was carried out using NEBNext dA-Tailing Module (NEB, E6053) followed by a DNA purification with 1.8 × AMPure XP beads.   DNA damage that may interfere the subsequent labeling step was repaired in a mixture of 2 μl endonuclease IV (Endo IV, NEB, M0304), 1 μl Bst full-length polymerase (NEB, M0328), 2 μl Taq ligase (NEB, M0208), 1 μl NAD+ (NEB, B9007), 1 μl dNTP (2.5 mM each) in NEBuffer 3 for 1 h at 37 °C and 1 h at 45 °C. DNA was purified and subjected to in vitro BER labeling reaction containing a mixture of 1 μl UDG (NEB, M0280), 1.5 μl Endo IV, 0.8 μl Bst full-length polymerase, 1.7 μl Taq ligase, 1 μl NAD+, 200 nM biotin-dUTP, 800 nM d5fCTP, 200 nM dATP and 200 nM dGTP in NEBuffer 3 for 40 min at 37 °C. The DNA was incubated with 75 mM of malononitrile in 10 mM Tris-HCl (pH 7.0) at 37°C for 20 h in an enclosed thermomixer (Eppendorf, 850 rpm), away from light. The reaction product was then purified with 1.8 × AMPure XP beads.  Labeled fragments were enriched by streptavidin C1 beads (Invitrogen) following the manufacturer's instruction. Y adaptor (see Supplementary Note 1) was ligated (NEBNext Quick Ligation Module, E6056) to double-stranded DNA on streptavidin C1 beads so that free adaptor can be removed by three-times washing of 1×B&W buffer (5 mM Tris-HCl, pH 7.5, 0.5 mM EDTA, 1 M NaCl, 0.05% Tween-20). After washing once with 1× SSC buffer (pH 7.0), beads with biotinylated DNA strands were resuspended in freshly prepared 0.15 M NaOH and incubated at room temperature for 10 min, followed by a sequential washing of 1× SSC buffer (7.0) and 1× TE buffer (pH 7.5) for once. The DNA was eluted from C1 beads using deionized water after heating at 95 °C for 3 min.   Eluted DNA was finally subjected to library amplification with indexed primers (NEB). To obtain sufficient DNA template, 2 rounds of amplification were first performed by using MightyAmp DNA polymerase (Takara), which was proved to read through regions containing biotin-dU and d5fC-malononitrile sites more easily. The PCR product was recovered via DNA Clean & Concentrator-5 and then PCR amplified for ~12 cycles using NEBNext® Q5® Hot Start HiFi PCR Master Mix (NEB, M0543). The constructed libraries were checked using the Agilent 2100 bioanalyzer. Sequencing was performed by Illumina Hiseq X Ten.

tag: cell line

value: MCF-7 cells

value: breast derived epithelial cells

value: sgRNADel-PD


title: 293T-EMX1.All_rep1.Control-13

description:
EMX1.All_rep1.Control-13 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: PC3_ecDNA_negative_Ctrl-circles_RNAseq_Rep2

tag: source

value: The PC3 was initiated from a bone metastasis of a grade IV prostatic adenocarcinoma from a 62-year-old male Caucasian.


title: 293T-EMX1.All_rep1.Control-12

description:
EMX1.All_rep1.Control-12 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx


title: MCF7-RNF2-All-PD_rep1


title: 48hrAPF_Male_H3_4

description: Sequential Chromatag-ChIP for fru P1 neuron chromatin

value: Male

value: fru P1


title: MCF7-Empty-PD_rep1

description:
replicate 1
Detect-seq Empty PD rep1 in MCF-7 cells

value: Empty-PD


title: RNA_ESC_shKap1_2

description: Successfully infected ES cells were screened by FACS.

source_name: embryonic stem cells

value: Kap1 shRNA expressed


title: 293T-EMX1.All_rep1.Control-11

description:
EMX1.All_rep1.Control-11 in HEK293T cell
20200615-TargetSeq_experiment_design_processed_data.xlsx

value: embryonic kidney derived epithelial cells


title: ChIP_ESC_Ythdc1KO_Kap1_1

value: Ythdc1 flox/flox; CreERT2

value: anti-KAP1 (Abcam ab22553)


title: ChIP_ESC_ff_input


title: 48hrAPF_Male_H3K27ac_3


title: PC3_ecDNA_positive_ChIP_Input.umi

description: ChIP-seq

data_processing:
The raw reads were quality trimmed using Trim Galore version 0.4.3 (options: --stringency 3 -q 30 -e .20 --length 15) and mapped to the hg19 genome using BWA 0.7.12 (command: mem). Reads with mapping quality greater than 30 were retained, de-duplicated. Only reads share the same mapping coordinate and UMI barcodes will be removed. The high quality non-redundant reads were used to call peaks with MACS2.1.0.20151222 (macs2 callpeak options: --nomodel --extsize 250 -B --SPMR -g hs --keep-dup all). Fold enrichment was computed with MACS2 bdgcmp and the output bedgraph file was converted to bigWig with bedGraphToBigWig.
Illumina RTA-1.18 software used for basecalling
Genome_build: hg19

extract_protocol:
2 million cells were crosslinked and lysed in the same way as ChIA-PET. After lysis, the nuclei pellets were sonicated and immunoprecipitated with anti-RNApolII antibody (920102, Biolegend) or anti-H3K27ac antibody (39133, Active Motif).
4 ng of ChIPped DNA or input DNA were subjected to end-repair, A-tailing and adaptor ligation (IDT xGen Dual Index UMI Adapters) with the KAPA Hyper Prep Kit (KK8505, KAPA Biosystems). Adaptor-ligated DNA fragments were PCR amplified with the KAPA Library Amplification ReadyMix (KK2612, Kapa Biosystems) and sequenced on Illumina platform.

tag: ip antibody


title: 293T-HEK4-All-Input_rep2

description:
replicate 2
Detect-seq HEK4 All Irep2 in HEK293T cells

value: HEK293T cells

value: Wild Type


title: 48hrAPF_Female_H3K27ac_3

value: Female


title: 48hrAPF_Female_H3K27ac_2


title: ChIP_ESC_ff_Kap1_2

value: blastocyst-derived ES cells


title: 293T-HEK4-All-Input_rep1

description:
replicate 1
Detect-seq HEK4 All Irep1 in HEK293T cells


title: ChIP_ESC_ff_Kap1_1

value: Ythdc1 flox/flox


title: HF-3035_ChIP_Input.umi

description: ChIP-seq

value: Glioblastoma


title: 48hrAPF_Female_H3K27ac_1


title: PC3_ecDNA_negative_ChIP_Input.umi

description: ChIP-seq

growth_protocol: PC3 cell lines are cultured in F-12K medium (30-2004, ATCC) with 10% FBS


title: 48hrAPF_Male_H3_3


title: HF-3016_ChIP_Input.umi

description: ChIP-seq


title: RNA_ESC_shCtrl_1

extract_protocol:
ES cells were disrupted in TRIzol Reagent and total RNAs were isolated by chloroform extraction coupled with isopropanol precipitation.
1 μg RNA per reaction was subjected to rRNA elimination and library generation using VAHTS Total RNA-seq (H/M/R) Library Prep Kit.

value: control shRNA expressed


title: RNA_ESC_wtRes_2

value: Ythdc1 flox/flox; CreERT2; W378A Ythdc1 overexpression


title: RNA_ESC_shKap1_1


title: HF-3177_ChIP_Input.umi

description: ChIP-seq


title: 48hrAPF_Male_H3_2


title: HF-2927_ChIP_Input.umi

description: ChIP-seq


title: MCF7-HEK4-All-PD_rep1

value: MCF-7 cells

value: breast derived epithelial cells


title: HF-2354_ChIP_Input.umi

description: ChIP-seq


title: RNA_ESC_shCtrl_2


title: PC3_ecDNA_negative_ChIP_H3K27ac.umi

description: ChIP-seq

value: The PC3 was initiated from a bone metastasis of a grade IV prostatic adenocarcinoma from a 62-year-old male Caucasian.

value: H3K27ac (39133, Active Motif)


title: RNA_ESC_wtRes_1


title: HF-3016_ChIP_H3K27ac.umi

description: ChIP-seq


title: 48hrAPF_Male_H3_1

value: Male

value: fru P1


title: RNA_ESC_W378A_3

value: Ythdc1 flox/flox; CreERT2; WT Ythdc1 overexpression


title: 293T-HN-Empty-PD_rep1

description:
replicate 1
Detect-seq HN Empty Prep1 in HEK293T cells

value: embryonic kidney derived epithelial cells

value: HN-Empty-PD


title: PC3_ecDNA_positive_ChIP_H3K27ac.umi

description: ChIP-seq


title: 48hrAPF_Female_H3_4


title: 293T-VEGFA-Empty-PD_rep2

description:
replicate 2
Detect-seq VEGFA Emptrep2 in HEK293T cells

value: HEK293T cells

value: VEGFA-Empty-PD


title: 293T-NT-All-PD_rep2

description:
replicate 2
Detect-seq NT All PD rep2 in HEK293T cells

value: Wild Type

value: NT-All-PD


title: RNA_ESC_W378A_2

description: 2.5 μM 4-OHT was added into the culture medium 72 hr before harvesting.

value: blastocyst-derived ES cells


title: HF-3177_ChIP_H3K27ac.umi

description: ChIP-seq

value: Glioblastoma


title: 48hrAPF_Male_Input_3

description: ChIP input chromatin

value: ChIP Input

value: whole pupae


title: 293T-NT-All-PD_rep1

description:
replicate 1
Detect-seq NT All PD rep1 in HEK293T cells


title: 293T-HEK4-All-PD_rep2

description:
replicate 2
Detect-seq HEK4 All Prep2 in HEK293T cells


title: 293T-HN-Empty-PD_rep2

description:
replicate 2
Detect-seq HN Empty Prep2 in HEK293T cells


title: HF-3035_ChIP_H3K27ac.umi

description: ChIP-seq


title: 48hrAPF_Female_H3_2

value: Female


title: 48hrAPF_Female_H3_1


title: RNA_ESC_Ythdc1KO_2

value: Ythdc1 flox/flox; CreERT2


title: 48hrAPF_Male_Input_4

description: ChIP input chromatin


title: 48hrAPF_Female_Input_4

description: ChIP input chromatin


title: 48hrAPF_Female_Input_3

description: ChIP input chromatin


title: HF-2927_ChIP_H3K27ac.umi

description: ChIP-seq


title: RNA_ESC_wtRes_3

value: Ythdc1 flox/flox; CreERT2; W378A Ythdc1 overexpression


title: 293T-HEK4-sgRNADel-PD_rep2

description:
replicate 2
Detect-seq HEK4 sgRNArep2 in HEK293T cells


title: RNA_ESC_ff_2

value: Ythdc1 flox/flox


title: HF-2354_ChIP_H3K27ac.umi

description: ChIP-seq

value: H3K27ac (39133, Active Motif)


title: 48hrAPF_Female_H3_3

value: fru P1


title: 293T-HEK4-sgRNADel-PD_rep1

description:
replicate 1
Detect-seq HEK4 sgRNArep1 in HEK293T cells


title: RNA_ESC_W378A_1

value: Ythdc1 flox/flox; CreERT2; WT Ythdc1 overexpression


title: RNA_ESC_Ythdc1KO_3


title: 48hrAPF_Male_Input_2

description: ChIP input chromatin

value: Male


title: 48hrAPF_Male_Input_1

description: ChIP input chromatin


title: PC3_ecDNA_positive_ChIP_PolII.umi

description: ChIP-seq

value: The PC3 was initiated from a bone metastasis of a grade IV prostatic adenocarcinoma from a 62-year-old male Caucasian.

value: RNA polymerase II (920102, Biolegend)


title: 48hrAPF_Female_Input_1

description: ChIP input chromatin


title: 293T-EMX1-sgRNADel-PD_rep1

description:
replicate 1
Detect-seq EMX1 sgRNArep1 in HEK293T cells

value: HEK293T cells

value: embryonic kidney derived epithelial cells


title: RNA_ESC_Ythdc1KO_1


title: 10d_Female_TRAP_5

data_processing:
RNA-seq reads were aligned using STAR (v. 2.7.2a) to the D. melanogaster dm6 UCSC genome assembly. Aligned reads were sorted using the sort function in SAMtools (v.1.2).
Exon-level count tables were generated with FeatureCounts on the meta-feature level excluding multimapping as follows: -s 0 -O -t exon -g exon_id.
Expression was then quantified for each exon using Reads Per Million Mapped Reads (RPM/CPM) in EdgeR using cpm function.
Genome_build: UCSC dm6 (BDGP Release 6 plus ISO1 MT)
Supplementary_files_format_and_content: Processed data files for exon counts are in the .CSV format and contain following columns: exon id, chromosome , exon start, exon end, strand, exon length, gene_id, number of reads in exon region.
Supplementary_files_format_and_content: Supplementary file contains Reads Per Million (RPM) for all exons listed in first column and all samples in data set in subsequent columns.

source_name: RNA from fru P1-expressing neurons

growth_protocol: Flies were reared on standard cornmeal food at 25°C on a 12 12-h light/12-h dark cycle.

extract_protocol:
Translated mRNA purification was performed as described in Newell et al. 2016 with the following modifications: the 40µl of lysate supernatant (input) and TRAP (IP) fractions were placed into/resuspended in RA1 lysis buffer (Machery-Nagel Nucleospin RNA) for total RNA isolation according to manufacturer’s protocol.
The isolated total RNA from for TRAP and input libraries underwent poly(A)+ selection using the NEBNext Poly(A) mRNA isolation module. TRAP and input libraries were prepared with the NEBNext Ultra RNA library Prep Kit for Illumina.

treatment_protocol: Approximately 1000 10-12 day post eclosion sex-sorted flies were collected for fru P1 TRAP experiments, (n=4). Flies were collected and sex sorted 0-48hrs post-eclosion for appropriate aging to 10-12 days and snap frozen in liquid nitrogen between ZT1 and ZT2. All frozen animals were stored at -80°C until they were processed for downstream experiments.

tag: tissue

value: Whole head

value: 10-12day Adult


title: HF-2927_ChIP_PolII.umi

description: ChIP-seq

value: Glioblastoma


title: 293T-HEK4-All-PD_rep1

description:
replicate 1
Detect-seq HEK4 All Prep1 in HEK293T cells

value: Wild Type


title: RNA_ESC_ff_1

data_processing:
Primary fastq data were generated with the Illumina Casava (v 1.8.2) pipeline.
Sequencing reads were trimmed by Trim Galore (v 0.6.4) with parameters: --phred33, --illumina, --clip_R1 9, --clip_R2 9, --paired to remove adapter sequences.
Trimmed reads were aligned to mm9 reference genome using Tophat2 (v2.1.1) with default parameters.
Expression levels of individual were quantified by FPKM using Cufflinks (v 2.2.1).
Genome_build: mm9
Supplementary_files_format_and_content: Tab-delimited text files containing FPKM values for each transcript.

value: blastocyst-derived ES cells


title: HF-2354_ChIP_PolII.umi

description: ChIP-seq


title: HF-3177_ChIP_PolII.umi

description: ChIP-seq


title: HF-3035_ChIP_PolII.umi

description: ChIP-seq


title: 293T-EMX1-UGIDel-PD_rep1

description:
replicate 1
Detect-seq EMX1 UGIDerep1 in HEK293T cells


title: HF-3035_ChIP-free-chia-pet

description: ChIAPET

data_processing:
ChIA-PET data was processed by ChIA-PET Utilities (CPU) (https://github.com/cheehongsg/CPU). CPU (options: stat -s -p -T 18 -t 16) removed the adaptors, and grouped the reads into three categaries: PET reads, single tag reads and no-linker reads. Tags identified (>=18bp) and reads without linkers were mapped to hg19 using BWA alignment and mem according to the tag/read length. CPU further separated the mapping results into uniquely-mapped,multiply-mapped, and unmapped classes with the pairing consideration (options: pair -S -t 16) and removed duplicates generated by PCR amplification (dedup -g -t 16). Next, the non-redundant uniquely-mapped reads from 3 categaries were merged. CPU furhter clustered the non-redundant uniquely-mapped pair-end tags and grouped them as intra-chromosomal PET clusters (cis-interactions) and inter-chromosomal PET clusters (trans-interactions) (options: cluster -s 8000 -M -B 1000 -5 5,-20 -3 5,480 -t 16 -O).
Illumina RTA-1.18 software used for basecalling
Genome_build: hg19

extract_protocol:
10 million cells were dual-crosslinked with 1.5mM EGS (21565, Thermo Fisher) for 45 min followed by 1% formaldehyde (F8775, Sigma) for 20 min at room temperature and then quenched with 0.125 M Glycine (G8898, Sigma) for 10 min. The crosslinked cells were washed with PBS twice and lysed in 100 ul 0.55% SDS with incubation at room temperature, 62°C and 37°C sequentially for 10 min each, which was followed by 37°C for 30 min with addition of 25 ul 25% Triton-X 100 to quench SDS and 37°C overnight with addition of 50 ul AluI (R0137L, NEB), 50 μl 10 × CutSmart buffer and 275 μl H2O to fragmentize the chromatin. The pelleted digested nuclei were resuspended in 500 ul A-tailing solution containing 50 μl 10 × CutSmart buffer, 10 μl BSA (B9000S, NEB), 10 μl 10 mM dATP (N0440S, NEB), 10 μl Klenow (3’- 5’ exo-) (M0202L, NEB), and 420 μl H2O  with 1 hr incubation at room temperature and then subjected to proximity ligation by adding 200 μl 5 × ligation buffer (B6058S, NEB), 6 μl biotinylated bridge linker (200 ng/ul), 10 μl T4 DNA ligase (M0202L, NEB) and 284 μl H2O and incubating at 16°C overnight. The ligated chromatins were then sheared by sonication and decrosslinked.
The decrosslinked DNA were tagmented using the Nextera DNA Sample Preparation Kit (Illumina). The linker ligated ChIA-PET constructs were selected using streptavidin magnetic beads. The magnetic beads were blocked with sheared gDNA prior to the selection. The streptavidin selected constructs were amplified with PCR and purified.


title: 48hrAPF_Female_Input_2

description: ChIP input chromatin

value: Female

value: ChIP Input

value: whole pupae


title: PC3_ecDNA_negative_ChIP_PolII.umi

description: ChIP-seq


title: 293T-EMX1-sgRNADel-PD_rep2

description:
replicate 2
Detect-seq EMX1 sgRNArep2 in HEK293T cells


title: HF-3016_ChIP_PolII.umi

description: ChIP-seq


title: HF-2354_ChIP-free-chia-pet

description: ChIAPET


title: Kdm6_RNA34

city: Easton

first: Ezra

last: Lencer

state: PA

address: 746 High Street

department: Department of Biology

institute: Lafayette College

data_processing:
Alignment to human genome (hg38) using STAR aligner (v2.7.3a) and computing counts using quant mode.
Genome_build: hg38
Supplementary_files_format_and_content: Processed data are raw per-gene counts from STAR quant mode in tab-delimited format

source_name: Human lymphoblastoid cell line from KDM6 patient

growth_protocol: Standard

extract_protocol:
Zymo Research Direct-zol RNA MiniPrep Ki
Nugen mRNA Kit

treatment_protocol: KMT2d and KDM6A patient derived cell lines

value: Human lymphoblastoid cell line

value: KDM6A Mutation


title: 293T-EMX1-UGIDel-PD_rep2

description:
replicate 2
Detect-seq EMX1 UGIDerep2 in HEK293T cells


title: 293T-EMX1-All-Input_rep2

description:
replicate 2
Detect-seq EMX1 All Irep2 in HEK293T cells


title: 293T-EMX1-All-Input_rep1

description:
replicate 1
Detect-seq EMX1 All Irep1 in HEK293T cells


title: TAC JQ1 withdrawn scRNA rep 2

city: San Francisco

first: Pawel

last: Przytycki

state: California

address: 1650 Owens Street

department: GIDB

institute: Gladstone Institutes

data_processing:
cellranger (version 3.0.2) mkfastq with default parameters to demultiplex raw base calls
cellranger (version 3.0.2) count with default parameters for alignment, filtering, barcode counting, UMI counting, and cell calling. See https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/algorithms/overview for details
Genome_build: Mm10
Supplementary_files_format_and_content: We provide barcodes.tsv files, matrix.mtx files and features.tsv files (for single cell RNA seq) or peaks.bed files (for single cell ATAC seq) generated by the 10x Genomics cellranger command. We provide BigWig files for PROseq and 4C. We provide bed files and bigWig files for ChIPseq. We provide excel files for differential expression analysis with Reads Per Kilobase of exon per Megabase of library size (RPKM) for every gene.

source_name: Heart tissue (non-cardiomyocyte cells)

extract_protocol:
Briefly, after proper anesthesia level was reached, thoracotomy was performed, and mouse heart was isolated. The isolated heart was cannulated and perfused with perfusion buffer (120.4 mM NaCl, 14.7 mM KCl, 0.6 mM KH2PO4, 0.6 mM Na2HPO4, 1.2 mM MgSO4, 10 mM Na-HEPES, 4.6 mM NaHCO3, 30 mM taurine, 10 mM 2,3-butanedione monoxime, and 5.5 mM glucose, pH 7.0) in a Langendorff perfusion system (Radnoti 120108EZ) for 5-10 minutes at 37 °C. The cannulated heart was then digested by digestion buffer (perfusion buffer with 300 units/mL collagenase II (Worthington Biochemical) and 50 μM CaCl2) for about 10 min at 37 °C. At the end of digestion, the atria and great vessels were removed and the ventricular tissue was transferred to and gently teased into small pieces in stop buffer (perfusion buffer with 10% fetal bovine serum) at 37 °C. After gently pipetting, cell suspension was passed through a 250uM strainer in a falcon tube and then at 30xg for 3 minutes at room temperature (RT). Then, the supernatant - containing most of the non-cardiomyocytes (CMs) - was divided from the pellet (containing the CM fraction). The non-CM fraction was centrifuged again at 30xg for 3 minutes at RT and the supernatant kept. The supernatant was then filtered with a cell strainer (70um) and finally centrifuged at 400xg for 3 minutes at RT for eliminating debris. The non-CM pellet was finally resuspended in 1mL cold PBS 0.5% BSA. 30k cells were counted with trypan blue using a hemocytometer and then used for subsequent 10X Genomics Chromium single cell RNAseq preparation. For single cell ATAC, 500k isolated and purified non-CMs were resuspended in 100uL lysis buffer (Tris-HCl 10mM pH 7.4, NaCl 10mM, MgCl2 3mM, Tween-20 0.1%, P40 0.1%, Digitonin 0.01%, BSA 1% in Nuclease-free water), pipetted 10 times and kept on ice for 5 minutes. Nuclei were then washed 1mL 1X PBS with 1% BSA and then centrifuged at for 5 minutes at 4C. The nuclei pellet was resuspended in 1mL 1X PBS with 1% BSA and filtered with a 10uM strainer (pluriSelect #43-50010-00). Nuclei were then counted with DAPI using a hemocytometer and finally 30k non-CM nuclei used for subsequent 10X Genomics Chromium single cell ATACseq preparation. The CM-fraction was, after the first centrifugation, centrifugated again at 30xg for 3 minutes at RT in stopping buffer and the supernatant was discarded. The CM pellet was finally centrifuged at 400xg for 3 minutes in stopping buffer at RT and after discarding the supernatant, the cell pellet was lysed in Qiazol (miRNeasy kit - Qiagen) for subsequent RNA extraction.
Single-cell droplet libraries from the non-CM cell suspension (2x Sham, 2x TAC-Veh, 2x TAC-JQ1 and 2x TAC-JQ1 withdrawn) were generated in the 10X Genomics Chromium controller according to the manufacturer’s instructions in the Chromium Single Cell 3′ Reagent Kit v.2 User Guide. Additional components used for library preparation include the Chromium Single Cell 3′ Library and Gel Bead Kit v.2 (PN-120237) and the Chromium Single Cell 3′ Chip kit v.2 (PN-120236). Libraries were prepared according to the manufacturer’s instructions using the Chromium Single Cell 3′ Library and Gel Bead Kit v.2 (PN-120237) and Chromium i7 Multiplex Kit (PN-120262). Final libraries were sequenced on the NextSeq 500 (Illumina) for a quality control run and then on the NovaSeq (Illumina) for deeper sequencing. All the 8 samples were pooled and sequenced in one single lane. Sequencing parameters were selected according to the Chromium Single Cell v.2 specifications. All libraries were sequenced to a mean read depth of at least 50,000 total aligned reads per cell. Raw sequencing reads were processed using the Cell Ranger v.2.2.0 pipeline from 10X Genomics. In brief, reads were demultiplexed, aligned to the mouse mm10 genome and UMI counts were quantified per gene per cell to generate a gene-barcode matrix. Data from the 8 samples were aggregated and normalized to the same sequencing depth, resulting in a combined gene-barcode matrix of all samples.

tag: heart surgery

value: TAC

tag: treatment

value: JQ1 withdrawn

tag: type of experiment

value: Single cell RNA seq

value: Heart tissue (non-cardiomyocyte cells)


title: 10d_Female_Input_3

source_name: total RNA input


title: 293T-Empty-PD_rep2

first: Howard

address: No.5 Yiheyuan Road Haidian District

institute: Peking University

description:
replicate 2
Detect-seq Empty PD rep2 in HEK293T cells

tag: cell type

tag: genotype

value: Empty-PD


title: 293T-EMX1-APODel-PD_rep2

description:
replicate 2
Detect-seq EMX1 APODerep2 in HEK293T cells

value: HEK293T cells


title: PC3_ecDNA_positive-chia-pet

description: ChIAPET

extract_protocol:
10 million cells were dual-crosslinked with 1.5mM EGS (21565, Thermo Fisher) for 45 min followed by 1% formaldehyde (F8775, Sigma) for 20 min at room temperature and then quenched with 0.125 M Glycine (G8898, Sigma) for 10 min. The crosslinked cells were washed with PBS twice and lysed in 100 ul 0.55% SDS with incubation at room temperature, 62°C and 37°C sequentially for 10 min each, which was followed by 37°C for 30 min with addition of 25 ul 25% Triton-X 100 to quench SDS and 37°C overnight with addition of 50 ul AluI (R0137L, NEB), 50 μl 10 × CutSmart buffer and 275 μl H2O to fragmentize the chromatin. The pelleted digested nuclei were resuspended in 500 ul A-tailing solution containing 50 μl 10 × CutSmart buffer, 10 μl BSA (B9000S, NEB), 10 μl 10 mM dATP (N0440S, NEB), 10 μl Klenow (3’- 5’ exo-) (M0202L, NEB), and 420 μl H2O  with 1 hr incubation at room temperature and then subjected to proximity ligation by adding 200 μl 5 × ligation buffer (B6058S, NEB), 6 μl biotinylated bridge linker (200 ng/ul), 10 μl T4 DNA ligase (M0202L, NEB) and 284 μl H2O and incubating at 16°C overnight. The ligated chromatins were then sheared by sonication and immunoprecipitated with anti-RNApolII antibody (920102, Biolegend).
The ChIPped DNA were tagmented using the Nextera DNA Sample Preparation Kit (Illumina). The linker ligated ChIA-PET constructs were selected using streptavidin magnetic beads. The magnetic beads were blocked with sheared gDNA prior to the selection. The streptavidin selected constructs were amplified with PCR and purified.

value: The PC3 was initiated from a bone metastasis of a grade IV prostatic adenocarcinoma from a 62-year-old male Caucasian.

value: RNA polymerase II (920102, Biolegend)


title: TAC JQ1 withdrawn scRNA rep 1


title: 10d_Female_TRAP_4

value: Whole head

value: 10-12day Adult