title: Naive_primed_D30ASECRiAV_10XscRNAseq

description: 10X Genomics scRNAseq

data_processing:
Raw sequence reads were quality-checked using the FastQC software. The CellRanger version 4.0.0 was used to process, align and summarize unique molecular identifier (UMI) counts against the 10X Genomics pre-built human GRCh38 and mouse mm10 reference genome dataset (2020-A, July 7, 2020). Downstream analyses were performed on R using Seurat (v4.0.1) (Satija et al. 2015). Human cells were retained and mouse cells (MEFs) were filtered out by adjusting the number of counts per cell (nCount_RNA) and the number of mapped genes per cell (nFeature_RNA) to only keep cells that were mostly mapped to the human GRCh38 (hg38) genome. Cells with more than 25% of mitochondrial counts were filtered out. Count matrix was normalised with Seurat global-scaling normalization method “LogNormalize'' that normalizes the feature expression measurements for each cell by the total expression, multiplies this by a 10000 scale factor, and log-transforms the result.
Genome_build: Human GRCh38 (hg38) GENCODE v32/Ensembl 98 and mouse GRCm28 (mm10) GENCODE vM23/Ensembl 98 reference genome dataset (version 2020-A, July 7 2020).
Supplementary_files_format_and_content: count matrix includes counts per gene per cell (.csv) and annotations describes (.csv)
Supplementary_files_format_and_content: scRNA-seq_annotation.csv

growth_protocol:
growth protocol for human primed hPSCs: Human primed pluripotent stem cells (H9 hESCs and Sigma hiPSCs) were grown with or without feeder conditions as per the cell lines at normoxia conditions (5% CO2) and under humidified conditions at 37° C. In feeder-free conditions cells were cultured in pre-coated geltrex tissue culture treated plates in complete E8Flex medium (Stem cell technology). Cells were dissociated into smaller clumps every 5-6 days using veresene incubating 5 min at room temperature.
growth protocol for human naive hPSCs: Naive hPSCs (H9 hESCs and Sigma hiPSCs) were cultured on mitotically inactivated MEF feeders in 5% O2 and 5% CO2 incubator under humidified conditions at 37°C. All naive hPSCs were cultured in PXGL medium (Bredenkamp/Guo Stem cell reports 2019 (PMID: 31708477) which consists of 1:1 DMEM/F12 and Neurobasal, 0.5% N2-supplement, 1% B27-Supplement, 2 mM L-Glutamine, 0.1 mM β-mercaptoethanol, 1x penicillin-streptomycin, 1 μM PD0325901 (Axon Medchem and Wellcome-MRC Cambridge Stem Cell Institute), 2 μM XAV939 (Sigma), 2 μM Gö6983 (Tocris), 20 ng/mL human LIF (PeproTech and Wellcome-MRC Cambridge Stem Cell Institute) and 10 μM Y-27632 (Tocris and Cell Guidance Systems). Naive hPSCs were passaged every 4-5 days in a ratio 1:2 or 1:3 by single-cell dissociation with Accutase followed by filtering through a 40 μm cell strainer.
growth protocol for human TSCs: Human naive to trophoblast and EXMC conversion were done using the following previously described protocol (Dong et al. 2020; Cinkornpumin et al. 2020). Naive hPSCs were seeded in such a way that expecting at least 90% confluency by Day 2 on mitomycin C treated MEFs after dissociating in single-cell using TryplE (for 15 min at 37°C) in their respective naive culture conditions supplemented with 10μM Y-27632. The very next day the media was switched from naive to trophoblast stem cell medium (Okae et al. 2018) consisting of DMEM/F12 (Gibco) supplemented with 0.3% BSA, 0.2% FBS, 1% Penicillin-streptomycin, 1% insulin-transferrin-selenium-ethanolamine-X 100 supplement, 1.5 μg/ml L-ascorbic acid, 0.5 μM A83-01, 1μM SB431542, 50ng/ml hEGF, 2uM CHIR99021, 0.8 mM Valproic acid, 0.1 mM β-mercapto-EtOH (Gibco) and 5 μM Y-27632. The medium was changed every two days supplemented with 5 μM Y-27632. From passage 1 onwards cultures were cultured and maintained on 5 μg/ml Collagen IV coated tissue culture treated plates in hypoxia conditions (5% O2 and 5% CO2) and passaged every 4 days in 1:3 or 1:6 splitting ratios.

extract_protocol:
Cells were washed with 1X PBS before dissociation from the culture dish by TryplETM Express Enzyme (ThermoFisher, 12605-036) incubation for 8 minutes at 37°C. Single cell suspension was filtered through a Falcon 40 µm Cell Strainer and centrifuged at 200G for 5 min. After resuspension in 1X PBS with 0.04% BSA, the single cell suspension was counted with a Luna-FL automated Fluorescence Cell Counter (Logos Biosystems). TSCs were mixed with primed and naive hPSCs using the following ratio (H9 primed: Sigma primed: H9 naive: Sigma naive: Sigma TSCs = 1:1:1:1:2)
Cells were loaded onto the 10X Chromium Single Cell Platform (10X Genomics) targeting 2000 cells (Next GEM Single Cell 3’ library and Gel Bead Kit v3.1) according to the manufacturer’s protocol (10x User Guide; CG000204, Revision D). Generation of gel beads in emulsion (GEMs), barcoding, GEM-RT cleanup, complementary DNA amplification and library construction were all performed according to the manufacturer’s protocol. Individual sample quality was assessed using a Tapestation (Agilent). Qubit 2.0 (ThermoFisher Scientific) and KAPA Library Quantification Kit for Illumina Platform (KAPA Biosystems) were used for library quantification before pooling. The final library pool was sequenced on the NovaSeq6000 (Illumina) instrument using NovaSeq SP kit (Illumina) for 2 lanes of 100-base-pair paired-end reads.

value: mix of human naive pluripotent stem cells (H9 and Sigma), primed pluripotent stem cells (H9 and Sigma), trophoblast stem cells (Sigma) and extraembryonic mesoderm cells (Sigma), all samples were pooled in one.

value: H9 (primed, and naive hPSCs) and Sigma (primed hPSCs, naive hPSCs, day 30 ASECRiAV conversion (trophobast stem cells and extraembryonic mesoderm cells)


title: T47D_LTED3d_Rep2_RNAseq

status: Public on Aug 01 2022

type: SRA

city: Miami

first: Liliana

last: Garcia-Martinez

state: Florida

address: 1501 NW 10th Avenue

department: Human Genetics

country: USA

institute: University of Miami

library_source: transcriptomic

data_processing:
ChIP-seq: Single-end ChIP-seq fastq files were processed using the ENCODE Transcription Factor and Histone ChIP-Seq processing pipeline (https://github.com/ENCODE-DCC/chip-seq-pipeline2). Reads were trimmed using cutadapt v2.5. Reads were aligned to the hg19 genome using Bowtie2 v2.3.4.3, and the SAMtools v1.9 was used to convert the output file to the BAM format. Duplicates were removed using Picard Tools v2.20.7. Peaks were called/signal tracks were generated using MACS2 v2.2.4, and peaks with fold change > 3.5 compared to input and a FDR < 0.05 were used for subsequent analysis. Only default parameters were used.
RNA-seq: Single-end RNA-seq fastq files were processed using the LSF-RNAseq Pipeline from Diderote (https://github.com/diderote/LSF-RNAseq). Reads were trimmed using cutadapt v2.3 with the following parameters: -j 4 --netseq-trim=20 -m 18. Reads were aligned to the hg19 genome using STAR v2.6.1a along default parameters, and RSEM v1.3.0 was used to obtain the expected gene counts against the human refseq also along default parameters.
RNA-seq differential expression was determined using DESeq2 v1.18.1 and R (version 3.4.1) with q-value <0.05. Ruvseq v1.12.0 was used for factor analysis to remove count based on ERCC spike-in. GSEA analysis was performed using gsea-3.0.jar. These additional analysis steps were included in the Diderote LSF-RNAseq Pipeline.
Bedtools v2.29.0 intersect was used to determine peak overlaps and assign target genes.
Homer v4.11 was used for peak annotation and motif analysis (http://homer.ucsd.edu/homer/data/software/homer.v4.11.1.zip)
Genome_build: hg19
Supplementary_files_format_and_content: The processed data files are in bigWig format containing the indexed tag chromosome coordinates and score produced from MACS2 peak calling.

molecule: total RNA

organism: Homo sapiens

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were were culture in estrogen deprived conditions for 3 days. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL)

extract_protocol:
Total RNA was isolated from frozen cell pellets using the TRIzol reagent (Thermo Fisher Scientific cat# 15596018) according to manufacturer's instructions. RNA quantity and quality was measured on a nanodrop.2 ug of RNA were treated using the TURBO DNA-free Kit (Thermo Fisher AM1907) to remove DNA contamination and later on DNAse and divalent cations from the sample. 1ug of the treated RNA was used for library preparation. Quality was checked using a Bioanalyzer
Library preparation was perfromed on Dnase-treated RNA using the Illumina TruSeq Stranded Total RNA Library Prep Gold (Illumina 20020598). TruSeq RNA UD Indexes (Illumina 20022371) were used for the adapter ligation step. Samples were normalized to a concentration of ranging from 100-250ng for input, and 1 ul of ERCC spike-in Mix 1 (Thermo Fisher 4456740) diluted 1:200 was added to the samples.

tag: cell type

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site

tag: cell source

value: ATCC #HTB-133

tag: modifications

value: Parental estrogen deprived for 3 days

tag: chip antibody


title: D5CB_6

city: Lisbon

first: Mariana

middle: A.

last: Branco

state: Porto Salvo

address: Av. Prof. Doutor Cavaco Silva

department: Institute for Bioengineering and Biosciences

country: Portugal

institute: Instituto Superior Tecnico

data_processing:
Illumina NovaSeq 6000 system with a paired end 150 bp strategy
Raw reads were filtered by removing reads containing (1) adapters, (2) N > 10% (N represents base that could not be determined) and (3) low quality (when Qscore - Quality value - of over 50% bases of the read is <= 5). Paired-end clean reads were mapped to hg38 reference genome using HISAT2 software. Counts were obtained using Featurecounts.
Genome_build: hg38
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

source_name: cardiac progenitor cells

extract_protocol:
hPSC-derived mesendoderm progenitor cells were enzymaticly dissociated with accutase and total RNA was extracted using High Pure RNA Isolation Kit (Roche), according to manufacturer´s instructions.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: hiPSC-derived cardiac progenitor cells

tag: treatment

value: WNT and BMP signaling pathway activation

tag: differentiation days


title: GPX8-KO-Caki1, rep3

city: Seoul

first: Sunghyouk

last: Park

state: Not US or Canada

address: Bldg 143, Room 302 College of Pharmacy Seoul National University, Gwanak 1

department: Natural Products

country: South Korea

institute: Seoul National University

data_processing:
Data quality was checked using FastQC v0.11.7 (http://www.bioinformatics.babraham.ac.uk/projects/fastqc/).
Low quality ends with <Q30 were trimmed and reads shorter than 36 bases after trimming were discarded.
Further adapter trimming was performed using Trimmomatic 0.38.
Raw sequencing data were mapped to the human reference genome (GRCh37.73) with HISAT2 version 2.1.0 (https://ccb.jhu.edu/software/hisat2/index.shtml).
The raw counts of genes were normalized by Deseq2 package (R ver. 4.1.1) and further analyzed to generate the differentially expressed genes (DEGs).
Genome_build: human reference genome (GRCh37.73)
Supplementary_files_format_and_content: results_GPx8KO_vs_WT_caki1.csv (Normalized raw count)
Supplementary_files_format_and_content: Caki_1_GPx8_KO_200721.csv (Unnormalized raw count)

source_name: Clear cell renal cell carcinoma (ccRCC) cell

extract_protocol:
Total RNA was extracted using the RNA extraction kit (Intron, Easy-spin).
RNA libraries were prepared for sequencing using standard Illumina protocols
RNA-Seq, paired-end reads

tag: cell line

tag: genotype

value: GPX8 knockout


title: WT-Caki1, rep1

value: Wild type

value: Clear cell renal cell carcinoma (ccRCC) cell


title: sc_B32: BY4741, WT, 16-hour

city: Beijing

first: Jincheng

last: Wang

address: yiheyuan road No5

country: China

institute: BIOPIC, Peking University

library_source: transcriptomic single cell

data_processing:
Paired-end reads were mapped to the S288c Saccharomyces cerevisiae genome R64 version (www.yeastgenome.org) with ERCC spike-in sequences added using HISAT2 (version 2.1.0).
Resulting bam files were sorted and indexed using samtools (version 1.1).
Final read counts mapped to genes were extracted using FeatureCounts.
Assembly: S288c Saccharomyces cerevisiae genome R64.
Supplementary files format and content: tab-delimited text files include log2-normalized read counts for each Sample

organism: Saccharomyces cerevisiae

growth_protocol: BY4742 WT were grown on SD solid agar plates.

extract_protocol:
Yeast cells were lysed with ~4µl lysis buffer, containing 0.5%Triton, 2.5µM oligo-dT, 2.5mM dNTP (Invitrogen,1959189), 8000 molecules of external RNA control consortium (ERCC) spike in, 3 x 10-2U/µl zymolyase(ZYMO, E1004-A), 1U/µl Recombinant RNase Inhibitor(TaKaRa, AI41189A), and RNA released.
RNA libraries for RNA-seq were prepared using Smart-seq2 following manufacturer's protocols.

tag: strain

value: WT

tag: time

value: 16 hour


title: sc_B33: BY4741, WT, 16-hour


title: Mf_3w-ivc_MS238_204

city: Kyoto

first: Yukihiro

last: Yabuta

state: Kyoto

address: Yoshida-Konoe-cho, Sakyo-ku

department: Anatomy and Cell Biology

country: Japan

institute: Kyoto University, Graduate school of medicine

description: cDNAs amiplified from 1 ng total RNA extracted from cells using SC3-seq technology

data_processing:
The processing of the 3-prime-sequencing data for calculating the gene-expression levels was performed as described previously (Nakamura et al., 2015).  In brief, all reads were processed with cutadapt v1.9.1 to remove adaptors and poly-A sequences.
The reads of 30-bp or longer were mapped onto a mouse transcript reference, ref_macFas5, annotation 101, with 3'-end of transcripts, extended by up to 10kb, using Tophat v2.0.11/Bowtie2 v2.2.7.  Read counts per gene were calculated using HTSeq v0.9.1 with the default settings.  Read counts were converted to read counts per million mapped reads (RPM) and used for the analysis.
Genome_build: macFas5
Supplementary_files_format_and_content: Tab-delimited text files include RPM values for each sample.

molecule: polyA RNA

organism: Macaca fascicularis

source_name: An oocyte from 3wks-ivc rOvary

extract_protocol:
Fetal ovaries collected from cy fetuses at 8-18 wpf were carefully dissected out in chilled PBS.  Fetal ovaries at 8-14 wpf were minced with scissors in 0.05% trypsin/EDTA/PBS solution (GIBCO, 15400-054), then incubated for 15 min at 37℃ with gentle pipetting every 5 min.  Dissociation medium [DMEM (GIBCO, 10313021) containing 10% FBS, 2mM L-glutamine (GIBCO, 25030081), 1X penicillin/streptomycin (GIBCO, 15070063), and 100μg/mL DNaseI (MERCK, DN25)] was added for further dissociation. For fetal ovaries at 16 and 18 wpf, minced tissues were incubated in Accumax (Innovative Cell Technologies, AM105) for 40 min at 37℃ with gentle pipetting every 5 min.  Dissociation medium was added for further dissociation.  For SC3-seq analysis, cells were picked under the inverted microscope (OLYMPUS, IX71) to exclude multiplet cells.   For dissociating cy rOvaries, tissues were incubated with TryPLE Express (1X) (GIBCO, 12604021) for 5 min.  After adding dissociation medium, cells were processed for single-cell analyses. Dissociated germ cells from in vivo monkey fetal ovaries and monkey rOvaries were manually picked using glass capillaries under the stereo microscope (Leica, S8 APO).  Following cDNA synthesis and amplification methods were previously described (Nakamura et al., 2015).
The cDNA synthesis was performed according to the single-cell 3 prime RNA-seq (SC3-seq) method described previously (Nakamura et al., 2015).  The cDNA libraries for Nextseq500 (Illumina) were constructed as described previously (Ishikura et al., 2016).The sequencing was performed on an Illumina Nextseq500/550 platform with a Nextseq500/550 Hiqh output kit (Illumina).

tag: developmental stage

tag: xenotransplantation period

tag: in vitro culture period

value: 3 weeks

value: Germ cell


title: Mf_3w-ivc_MS263_202


title: Mf_3w-ivc_MS263_203


title: WT-Caki1, rep2


title: Mf_15w-ptp_MS238_065

source_name: An oocyte from 15w-ptp rOvary

value: 15 weeks


title: Mf_3w-ivc_MS263_201


title: sc_B36: BY4741, WT, 16-hour


title: sc_B29: BY4741, WT, 16-hour


title: D5CB_3


title: HMC3 cells_rep1

city: St. Louis

first: Andrew

middle: D.

last: Nguyen

state: MO

address: 1402 S Grand Blvd, Doisy Hall, R205

department: Internal Medicine

institute: Saint Louis University

data_processing:
Reads were aligned to hg19 using TMAP software
miRNA coverage per nucleotide was calculated using BEDtools genomecoverage
miRNA coverage was normalized by total coverage to yield total normalized nucleotide coverage for each RNA
Differential expression ratios and p values were calculated in Excel
Genome_build: hg19
Supplementary_files_format_and_content: normalized nucleotide coverage per gene, average values per cell type, and differential  analysis reported as HMC3 to H4 ratio with p-values

source_name: HMC3 cells

growth_protocol: H4 cells were cultured in Gibco DMEM supplemented with 10% FBS, 10 U/ml penicillin, and 10 µg/ml of streptomycin. HMC3 cells were cultured in Corning EMEM supplemented with 10% FBS, 10 U/ml penicillin, and 10 µg/ml of streptomycin. Both cell lines were passaged every 3-4 days using trypsin-EDTA.

extract_protocol:
The small RNA fractions were isolated from cells using the mirVana miRNA Isolation Kit (Invitrogen, AM1561)
Libraries were constructed using the Ion Total RNA-Seq Kit v2 (Life Technologies, 4475936)

value: ATCC #CRL-3304

value: Microglia cells

value: Untreated

tag: molecule subtype

value: RNA, small fraction


title: Mf_3w-ivc_MS238_206


title: Mf_12w-ptp_MS236_103

source_name: An oocyte from 12w-ptp rOvary

value: 12 weeks


title: sc_B27: BY4741, WT, 16-hour


title: sc_B31: BY4741, WT, 16-hour


title: D5CB_1


title: D3CB_5


title: sc_B28: BY4741, WT, 16-hour


title: Mf_12w-ptp_MS236_102


title: D3CB_3


title: Mf_12w-ptp_MS236_111


title: HMC3 cells_rep2


title: H4 cells_rep3

source_name: H4 cells

value: ATCC #HTB-148

value: Neuroglioma cells


title: sc_B30: BY4741, WT, 16-hour


title: sc_B26: BY4741, WT, 16-hour


title: D5N_5

value: hiPSC-derived cardiac progenitor cells

value: Without supplementation (Control)


title: H4 cells_rep2


title: Mf_12w-ptp_MS236_092


title: H4 cells_rep1


title: 3,4-DC3: spinal cord, injuried+3,4-DC, 3 days

city: wenzhou

first: zhang

middle: hao

last: jie

address: chashan

institute: wenzhou medical university

data_processing:
Through the Illumina platform, a large number of sample paired-end sequencing data were obtained. In view of the influence of the data error rate on the results, Trimmomatic software was used to preprocess the quality of the original data, and the number of reads in the whole quality control process was statistically summarized.
Quality preprocessing steps (1) to the adapter (Adaptor);(2) Remove low-quality reads; (3) Remove low-quality bases in different ways from the 3' and 5' ends; (4) Count the original sequencing quantity, effective sequencing quantity, Q30, GC content, and conduct a comprehensive evaluation.
Assembly: mm10
Supplementary files format and content: tab-delimited text files include RPKM values for each Sample

organism: Mus musculus

source_name: spinal cord tissue

extract_protocol:
A. Take an appropriate amount of tissue/cell sample, add 600 μL Lysis/Binding Buffer, and homogenize. Add 30 μL miRNA  Homogenate Additive, mix well. Ice bath for 10 min; B. Add an equal volume of phenol (acid phenol) chloroform - self-preparation. Centrifuge at maximum speed for 5 min at room temperature. Take the supernatant; C. Add 1.25 times the volume of 100% ethanol; D. Add the mixture to the column, (note: add up to 700 μL) at room temperature, 13000 rpm, 30 s, discard the supernatant; E. Add 350 μL of miRNA Wash Solution 1 to the spin column, 13000 rpm for 30 s. Discard the supernatant and leave  The stem is repositioned into the collection tube; F. Mix 10 μL DNase I and 70 μL Buffer RDD QIAGEN (#79254) Total volume: 80 μL, add  Put it on the membrane in the spin column and leave it at room temperature for 15 minutes; G. Add 350 μL of miRNA Wash Solution 1 to the spin column, 13000 rpm for 30 s. Discard the supernatant and leave  The stem is repositioned into the collection tube; H. 500 μL Wash Solution 2/3 was passed through the column twice, 13000 rpm, 30 s. Discard the supernatant and replace the spin column  Placed in the collection tube; I. Centrifuge the empty column for 1 minute. Place the spin column in a new collection tube and add 100 μL of pre-warmed 95°C to the center of the column.  Elution Solution for 2 min. Centrifuge at the highest speed for 20-30 seconds at room temperature, the liquid in the collection tube is the extracted liquid  Total RNA can be stored at -70°C.
1. Purification and fragmentation of mRNA 2.Synthesis of cDNA 1 strand 3.Synthesis of cDNA 2 strands 4.Plus A at the 3 prime end 5.The connection of the joint sequence 6.Enrichment of DNA fragments 7. Library quality inspection

tag: tissue

value: spinal cord tissue

value: spinal cord injury+3,4-DC treatment


title: 3,4-DC2: spinal cord, injuried+3,4-DC, 3 days


title: Mf_6w-ptp_MS238_213

source_name: An oocyte from 6w-ptp rOvary

value: 6 weeks


title: sc_B23: BY4741, WT, 16-hour

value: WT


title: Mf_6w-ptp_MS200_045


title: sc_B22: BY4741, WT, 16-hour

value: 16 hour


title: D3CB_6

value: WNT and BMP signaling pathway activation


title: D3N_3


title: SCI3: spinal cord, injuried, 3 days

value: spinal cord injury


title: sc_B25: BY4741, WT, 16-hour


title: D3N_1


title: Mf_6w-ptp_MS236_211

value: Germ cell


title: sc_B24: BY4741, WT, 16-hour


title: D5N_3


title: T47D_LTED_LSD1_K661A_K4me1_CUT_and_RUN

library_source: genomic

data_processing:
Library strategy: CUT_and_RUN
ATAC-seq: Paired-end ATAC-seq fastq files were processed using the ATAC-seq Pipeline from the Kundaje Lab (https://github.com/kundajelab/atac_dnase_pipelines). Reads were trimmed using cutadapt v1.9.1 using the following parameters: -m 5. Reads were aligned to the hg19 genome using Bowtie2 v2.2.6, and the SAMtools v1.7 was used to convert the output files into the BAM format using default parameters. Duplicate reads were removed using Picard tool v1.126 using default parameters. Shuf was used to subsample reads to 50 million. Peaks were called using MACS2 v2.1.0 using default parameters
Genome_build: hg19
Supplementary_files_format_and_content: The processed data files are in bigWig format containing the indexed tag chromosome coordinates and score produced from MACS2 peak calling.

molecule: genomic DNA

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations and split 1:3 every 3 days. Culture media: RPMI-1640(Lonza 12-167Q) supplemented with 10% FBS(Gemini 100-106), 1x penicillin/streptomycin(Thermo Fisher Scientific 15140-12, 10000U/mL), and 1x glutamax( Life Technologies 35050-061 ).

extract_protocol:
CUT&RUN was performed with CUTANATM ChIC/CUT&RUN kit according to the manufacturer’s protocol (EpiCypher, 14-1048). Briefly, 500,000 cells were gently harvested with a cell scraper (Corning, 3008) and bound to activated Concanavalin A beads. The cells bound to the beads were incubated with antibodies at 4℃ overnight. After extensive washing, cells were incubated with pAG-MNase (EpiCypher, 15-1016) for 10 min at room temperature and then chromatin digestion was performed with the chromatin digestion buffer for 2h at 4°C. After chromatin digestion, the stop buffer and E. coli spike-in DNA (EpiCypher, 18-1401) were added and chromatin fragments were purified with DNA cleanup columns.
Libraries were generated using the NEBNext Ultra II DNA Library Prep Kit for Illumina (NEB, E7370L) and sequenced on Illumina NovaSeq 6000 (Illumina).

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site

value: ATCC #HTB-133

value: Parental estrogen deprived for 1 year, plko-shLSD1 estable (TRCN0000382379) rescued with pCR 2.1-TOPO expressing catalytic mutant LSD1

value: anti-H3K4me1 (Abcam, ab4729)


title: SCI1: spinal cord, injuried, 3 days


title: sc_B18: BY4741, WT, 16-hour


title: Mf_3w-ptp_MS236_210

source_name: An oocyte from 3w-ptp rOvary

value: 3 weeks


title: Mf_6w-ptp_MS238_212


title: 6WK WT (R6/2 Littermate) Glial Progenitor Cells Replicate 6

city: Rochester

first: John

last: Mariani

state: NY

address: 601 Elmwood Avenue

department: Neurology

institute: University of Rochester

data_processing:
Basecalling was performed with Illumina CASAVA
Raw FASTQ files were aligned to GRCh38 using Ensembl 92 gene annotations with STAR 2.5.2b.
Reads were quantified with RSEM 1.3.0
Genome_build: Genome_build: GRCh38
Supplementary_files_format_and_content: Expected gene count matrix - output from RSEM.

source_name: Glial Progenitor Cells

extract_protocol:
mRNA was extracted and purified with the Qiagen RNeasy kit, according to the manufacturer's instructions.
Purified mRNA was used to generate sequencing ibraries using the TruSeq RNA v2 kit according to manufacturer instructions.

treatment_protocol: Isolation of PDGFRA-EGFP glial progenitors from freshly dissociated striatal cel of wild-type, R6/2, or zQ175 mice was first carried out by euthanizing mice with carbon dioxide, perfusing transcardially with HBSS, and removing the brain.  . The brains were immersed in ice-cold sterile HBSS for about 5 minutes to facilitate the microdissection. Under a dissecting microscope, the sub-ventricular zone was removed and discarded and the striata from each mouse was dissected and placed in sterile HBSS on ice. The striata from all the mice of the litter were pooled together as per genotype. The striatal tissues were transferred to a petri dish containing sterile HBSS then chopped into small pieces using sterile disposable scalpels, transferred into a sterile tube and then incubated in a papain/DNase dissociation solution at 37oC for 50 minutes. Minimum essential media plus 0.5% BSA (MEM-BSA) containing 5% serum was then added to inactivate the papain. The tissue was triturated by repeated pipetting in order to achieve a single cell suspension. The cells were then pelleted, resuspended into MEM-BSA and overlaid first onto a 90% Percoll gradient followed by a second 30% Percoll gradient and the solution centrifuged. The myelin and debris were removed from the tube and the cell pellet was resuspended in MEM containing 20U/ml DNase.  For FACS of PEGFRA-EGFP GPCs, mouse striata were pooled together (n=3-4/group for R6/2 and 12 week zQ175 and WT controls; 4-8/group for 1 year-old zQ175 and WT).  All samples were resuspended in MEM containing 20U/ml DNase to a concentration of 1-1.5x106 cells/ml and then passed over a 35 µm tube top cell strainer prior to flow cytometry. DAPI was added at 1 µg/ml. Flow cytometry analysis and FACS was performed on a BD FACSAria IIIU (Becton Dickinson, San Jose, CA). The cells were analyzed by forward and side scatter, for EGFP fluorescence through a 530 ± 30 nm band-pass filter and for DAPI fluorescence through a 450 ± 50 nm band-pass. Non-fluorescent cells were used to set the background fluorescence; a false positive rate of 0.5% was accepted.

value: Striatum

tag: age


title: T47D_LTED_LSD1_4G4_K4me1_CUT_and_RUN

value: Parental estrogen deprived for 1 year, plko-shLSD1 estable (TRCN0000382379) rescued with pCR 2.1-TOPO expressing wildtype LSD1


title: sc_B19: BY4741, WT, 16-hour


title: Mf_3w-ptp_MS200_027


title: sc_B17: BY4741, WT, 16-hour


title: Mf_3w-ptp_MS200_026


title: D5N_2

value: hiPSC-derived cardiac progenitor cells


title: T47D_LTED_shLSD1_2E4_K4me1_CUT_and_RUN

value: Parental estrogen deprived for 1 year and plko-shLSD1 estable (TRCN0000382379)


title: 3,4-DC1: spinal cord, injuried+3,4-DC, 3 days

value: spinal cord tissue


title: 6WK WT (R6/2 Littermate) Glial Progenitor Cells Replicate 4


title: sc_B16: BY4741, WT, 16-hour


title: Mf_6w-ptp_MS236_212


title: Mf_6w-ptp_MS200_031


title: D3N_2

value: Without supplementation (Control)


title: T47D_LTED_shcontrol_K4me1_CUT_and_RUN

value: Parental estrogen deprived for 1 year and pLKO-shCtrl stable


title: 6WK WT (R6/2 Littermate) Glial Progenitor Cells Replicate 3

value: WT


title: sc_B15: BY4741, WT, 16-hour

value: 16 hour


title: Mf_3w-ptp_MS200_025


title: Mf_3w-ptp_MS200_024


title: 6WK WT (R6/2 Littermate) Glial Progenitor Cells Replicate 2


title: 6WK WT (R6/2 Littermate) Glial Progenitor Cells Replicate 1


title: sc_B14: BY4741, WT, 16-hour


title: sc_B13: BY4741, WT, 16-hour


title: Mf_3w-ptp_MS200_022


title: T47D_LTED_LSD1_KO_2E8_K4me1_CUT_and_RUN

value: Parental estrogen deprived for 1 year and LSD1 knockout


title: T47D_LTED_K4me1_CUT_and_RUN

value: Parental estrogen deprived for 1 year


title: 6WK R6/2 Glial Progenitor Cells Replicate 6


title: 6WK R6/2 Glial Progenitor Cells Replicate 5


title: Mf_3w-ptp_MS263_205

value: Germ cell


title: sc_B12: BY4741, WT, 16-hour


title: sc_B11: BY4741, WT, 16-hour


title: Mf_3w-ptp_MS200_019


title: T47D_FM_K4me1_CUT_and_RUN

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site

value: Parental


title: T47D_FM_LSD1_KO_2E8_K4me1_CUT_and_RUN

value: ATCC #HTB-133

value: LSD1 knockout


title: 6WK R6/2 Glial Progenitor Cells Replicate 4


title: Mf_3w-ptp_MS200_014

value: 3 weeks


title: sc_B10: BY4741, WT, 16-hour


title: T47D_LTED_LSD1_K661A_K27ac_CUT_and_RUN

value: Parental estrogen deprived for 1 year, plko-shLSD1 estable (TRCN0000382379) rescued with pCR 2.1-TOPO expressing catalytic mutant LSD1

value: Anti-H3K27ac (Diagenode, C15410194)


title: 6WK R6/2 Glial Progenitor Cells Replicate 3

value: Striatum


title: T47D_LTED_LSD1_4G4_K27ac_CUT_and_RUN

value: Parental estrogen deprived for 1 year, plko-shLSD1 estable (TRCN0000382379) rescued with pCR 2.1-TOPO expressing wildtype LSD1


title: sc_B9: BY4741, WT, 16-hour


title: 6WK R6/2 Glial Progenitor Cells Replicate 2


title: sc_B8: BY4741, WT, 16-hour


title: T47D_LTED_shLSD1_2E4_K27ac_CUT_and_RUN

value: Parental estrogen deprived for 1 year and plko-shLSD1 estable (TRCN0000382379)


title: T47D_LTED_shcontrol_K27ac_CUT_and_RUN


title: 6WK R6/2 Glial Progenitor Cells Replicate 1


title: Mf_3w-ptp_MS200_010


title: 12WK WT (zQ175 Littermate) Glial Progenitor Cells Replicate 6


title: Mf_3w-ptp_MS200_011


title: Mf_3w-ptp_MS200_009


title: 12WK WT (zQ175 Littermate) Glial Progenitor Cells Replicate 5


title: Mf_3w-ptp_MS200_007


title: T47D_LTED_K27ac_CUT_and_RUN


title: T47D_LTED_LSD1_KO_2E8_K27ac_CUT_and_RUN


title: sc_B6: BY4741, WT, 16-hour


title: Mf_3w-ptp_MS238_203


title: sc_B5: BY4741, WT, 16-hour


title: SCI2: spinal cord, injuried, 3 days

value: spinal cord tissue

value: spinal cord injury


title: 12WK WT (zQ175 Littermate) Glial Progenitor Cells Replicate 3

value: WT


title: 12WK WT (zQ175 Littermate) Glial Progenitor Cells Replicate 4


title: sc_B4: BY4741, WT, 16-hour

value: 16 hour


title: Mf_3w-ptp_MS200_006


title: sc_B3: BY4741, WT, 16-hour


title: sc_B7: BY4741, WT, 16-hour


title: Mf_3w-ptp_MS200_003


title: 12WK WT (zQ175 Littermate) Glial Progenitor Cells Replicate 2


title: Mf_3w-ptp_MS200_005


title: T47D_FM_K27ac_CUT_and_RUN


title: T47D_FM_LSD1_KO_2E8_K27ac_CUT_and_RUN


title: T47D_LTED_LSD1_K661A_IgG_CUT_and_RUN

value: anti-IgG (EpiCypher, 13-0042K)


title: T47D_LTED_LSD1_4G4_IgG_CUT_and_RUN


title: Mf_3w-ptp_MS200_002


title: sc_A38: BY4741, WT, 2-hour

value: 2 hour


title: sc_B1: BY4741, WT, 16-hour


title: sc_A37: BY4741, WT, 2-hour


title: 12WK zQ175 Glial Progenitor Cells Replicate 6


title: 12WK zQ175 Glial Progenitor Cells Replicate 5


title: 12WK WT (zQ175 Littermate) Glial Progenitor Cells Replicate 1


title: T47D_LTED_shLSD1_2E4_IgG_CUT_and_RUN


title: sc_A36: BY4741, WT, 2-hour


title: sc_A35: BY4741, WT, 2-hour


title: T47D_LTED_IgG_CUT_and_RUN

value: Parental estrogen deprived for 1 year


title: sc_B2: BY4741, WT, 16-hour


title: T47D_LTED_shcontrol_IgG_CUT_and_RUN

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site

value: Parental estrogen deprived for 1 year and pLKO-shCtrl stable


title: T47D_LTED_LSD1_KO_2E8_IgG_CUT_and_RUN

value: ATCC #HTB-133

value: Parental estrogen deprived for 1 year and LSD1 knockout


title: sc_A34: BY4741, WT, 2-hour


title: 12WK zQ175 Glial Progenitor Cells Replicate 3


title: 12WK zQ175 Glial Progenitor Cells Replicate 4


title: T47D_FM_LSD1_KO_2E8_IgG_CUT_and_RUN

value: LSD1 knockout


title: 12WK zQ175 Glial Progenitor Cells Replicate 2


title: Mf_Adult_MS192_026

source_name: An oocyte from adult ovary

value: Adult

value: Germ cell


title: Mf_Adult_MS192_028


title: Mf_3w-ptp_MS200_001

value: 3 weeks


title: sc_A33: BY4741, WT, 2-hour


title: Mf_Adult_MS192_015


title: T47D_FM_IgG_CUT_and_RUN

value: Parental


title: Mf_Adult_MS192_013


title: Mf_Adult_MS192_011


title: 12WK zQ175 Glial Progenitor Cells Replicate 1

value: Striatum


title: Mf_Adult_MS192_009


title: 1YR WT (zQ175 Littermate) Glial Progenitor Cells Replicate 4


title: sc_A32: BY4741, WT, 2-hour


title: T47D_LTED_LSD1_K661A_Rep2_ATACseq

data_processing:
ATAC-seq: Paired-end ATAC-seq fastq files were processed using the ATAC-seq Pipeline from the Kundaje Lab (https://github.com/kundajelab/atac_dnase_pipelines). Reads were trimmed using cutadapt v1.9.1 using the following parameters: -m 5. Reads were aligned to the hg19 genome using Bowtie2 v2.2.6, and the SAMtools v1.7 was used to convert the output files into the BAM format using default parameters. Duplicate reads were removed using Picard tool v1.126 using default parameters. Shuf was used to subsample reads to 50 million. Peaks were called using MACS2 v2.1.0 using default parameters
Genome_build: hg19
Supplementary_files_format_and_content: The processed data files are in bigWig format containing the indexed tag chromosome coordinates and score produced from MACS2 peak calling.

extract_protocol:
ATAC-seq experiments were performed as previously described (Corces et al., 2017) with minor modifications. 50,000 viable cells were used to perform the transposition reaction.The cleanup of the transposed fragments was performed using Zymo DNA Clean and Concentrator-5 Kit (Zymo catalog number D4014) and DNA was eluted in 21 µL of provided elution buffer
The ATAC-seq library construction was performed as previously described (Corces et al, 2017). To determine the average fragment size of each library, samples were run through a high sensitivity DNA screentape (Agilent Technologies #5067–5584) following the manufacturer’s instructions on an Agilent Technologies 2200 TapeStation machine. Concentration of each library was measured using Qubit dsDNA high sensitivity reagents (Thermo Fisher Scientific #Q32851) following the manufacturer’s instructions on a Qubit 3 fluorometer. .

value: Parental estrogen deprived for 1 year, plko-shLSD1 estable (TRCN0000382379) rescued with pCR 2.1-TOPO expressing catalytic mutant LSD2


title: T47D_LTED_LSD1_4G4_Rep1_ATACseq

value: Parental estrogen deprived for 1 year, plko-shLSD1 estable (TRCN0000382379) rescued with pCR 2.1-TOPO expressing wildtype LSD1


title: Mf_18wpf_MS192_082

source_name: An oocyte from 18 wpf fetal ovary

value: 18 wpf


title: T47D_LTED_LSD1_K661A_Rep1_ATACseq

value: Parental estrogen deprived for 1 year, plko-shLSD1 estable (TRCN0000382379) rescued with pCR 2.1-TOPO expressing catalytic mutant LSD1


title: 1YR WT (zQ175 Littermate) Glial Progenitor Cells Replicate 3

value: WT


title: T47D_LTED_LSD1_4G4_Rep2_ATACseq


title: sc_A30: BY4741, WT, 2-hour

value: 2 hour


title: 1YR WT (zQ175 Littermate) Glial Progenitor Cells Replicate 5


title: 1YR WT (zQ175 Littermate) Glial Progenitor Cells Replicate 2


title: Mf_18wpf_MS192_079


title: 1YR zQ175 Glial Progenitor Cells Replicate 6


title: HMC3 cells_rep3

value: ATCC #CRL-3304

value: Microglia cells

value: Untreated

value: RNA, small fraction


title: sc_A28: BY4741, WT, 2-hour


title: sc_A27: BY4741, WT, 2-hour


title: sc_A29: BY4741, WT, 2-hour


title: Mf_18wpf_MS192_081


title: 1YR zQ175 Glial Progenitor Cells Replicate 4


title: Mf_18wpf_MS192_078


title: Mf_18wpf_MS192_077


title: 1YR zQ175 Glial Progenitor Cells Replicate 5


title: sc_B21: BY4741, WT, 16-hour

value: 16 hour


title: sc_A31: BY4741, WT, 2-hour


title: T47D_LTED_shLSD1_2E4_Rep2_ATACseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 5 months. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL)

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site

value: ATCC #HTB-133

value: Parental estrogen deprived for 1 year and plko-shLSD1 estable (TRCN0000382379)


title: T47D_LTED_shcontrol_Rep2_ATACseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 4 months. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and (Thermo Fisher Scientific 15140-12, 10000U/mL

value: Parental estrogen deprived for 1 year and pLKO-shCtrl stable


title: Mf_18wpf_MS192_076

value: Germ cell


title: T47D_LTED_shLSD1_2E4_Rep1_ATACseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 5 months. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin(Thermo Fisher Scientific 15140-12, 10000U/mL)


title: T47D_LTED_shcontrol_Rep1_ATACseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 4 months. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL)


title: Mf_18wpf_MS192_074


title: sc_A25: BY4741, WT, 2-hour


title: sc_A24: BY4741, WT, 2-hour


title: Mf_18wpf_MS192_073


title: sc_A23: BY4741, WT, 2-hour


title: sc_A22: BY4741, WT, 2-hour


title: 1YR zQ175 Glial Progenitor Cells Replicate 3

value: Striatum


title: sc_A20: BY4741, WT, 2-hour


title: 12WK WT (R6/2 Littermate) Glial Progenitor Cells Replicate 3


title: 12WK WT (R6/2 Littermate) Glial Progenitor Cells Replicate 6


title: 12WK WT (R6/2 Littermate) Glial Progenitor Cells Replicate 4


title: sc_A21: BY4741, WT, 2-hour


title: 12WK WT (R6/2 Littermate) Glial Progenitor Cells Replicate 7


title: T47D_LTED_LSD1_KO_2E10_Rep2_ATACseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 3 months. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL)

value: Parental estrogen deprived for 1 year and LSD1 knockout


title: sc_A19: BY4741, WT, 2-hour


title: 1YR zQ175 Glial Progenitor Cells Replicate 2


title: sc_A26: BY4741, WT, 2-hour


title: Mf_18wpf_MS192_072


title: sc_A18: BY4741, WT, 2-hour


title: sc_A17: BY4741, WT, 2-hour


title: T47D_FM_LSD1_KO_2E8_Rep2_ATACseq

value: LSD1 knockout


title: Mf_18wpf_MS192_071

value: 18 wpf


title: Mf_18wpf_MS192_067


title: T47D_LTED_LSD1_KO_2E10_Rep1_ATACseq


title: Mf_18wpf_MS192_066


title: T47D_LTED_Rep2_ATACseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 1 month. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL)

value: Parental estrogen deprived for 1 year


title: T47D_FM_Rep1_ATACseq

value: Parental


title: Mf_18wpf_MS192_070


title: Mf_18wpf_MS192_065


title: Lymphoma_SLL_1

city: Shanghai

first: Jiahao

last: Shi

state: Shanghai

address: No. 12, Lane 833, Zhizaoju Road, Huangpu District,

department: Department of Ophthalmology

institute: Shanghai 9th Peoples Hospital Affiliated to Shanghai Jiaotong University School of Medicine

data_processing:
Illumina bcl2fastq software used for basecalling and original sequenced reads conversion, called Raw Data. And fastQC software were used for base quality control.
Sequenced reads were trimmed off primer/adaptor contaminated reads,and masked for  Low quality reads and N overtop reads,called clean reads
The clean reads were mapped onto the reference gene set by using Bowtie1 software
Use Stringtie software to calculate the reads count and convert it to the FPKM of the transcript.And then the low-abundance transcripts,  were removed
The differential expressed genes were identified by using limma R package after normalization. A |log2FC|>=1 and p-Value<=0.05 were set as the threshold for DEGs
Genome_build: GRCh38 (Genome Reference Consortium Human Build 38)
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

source_name: ocular adnexal lymphoma

extract_protocol:
Total RNA were isolated from tissues by using the Trizol Reagent according to the manufacture’s protocol
The poly-A containing mRNA molecules were purified from the 3 μg of total RNA by using poly-T oligo-attached magnetic beads. The cleaved RNA fragments were reversely transcribed into first strand cDNA using random hexamers, following by second strand cDNA synthesis using DNA Polymerase I and RNase H. The cDNA fragments were purified, end blunted, 'A' tailed, and adaptor ligated. PCR was used to selectively enrich those DNA fragments that have adapter molecules on both ends and to amplify the amount of DNA in the library. The number of PCR cycles was minimized to avoid skewing the representation of the library. The library were qualified by Agilent 2100 bioanalyzer and quantified by Qubit and qPCR.

value: lymphoma specimen

tag: lymphoma subtype

value: chronic lymphocytic leukemia/small lymphocytic lymphoma


title: T47D_FM_Rep2_ATACseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations and split 1:3 every 3 days. Culture media: RPMI-1640(Lonza 12-167Q) supplemented with 10% FBS(Gemini 100-106), 1x penicillin/streptomycin(Thermo Fisher Scientific 15140-12, 10000U/mL) and Glutamax (Life Technologies 35050-061 ).

tag: cell type


title: T47D_FM_LSD1_KO_2E8_Rep1_ATACseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in estrogen deprived conditions for 3 days. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin(Thermo Fisher Scientific 15140-12, 10000U/mL)

tag: cell source


title: Mf_18wpf_MS192_063


title: sc_A16: BY4741, WT, 2-hour

value: WT

value: 2 hour


title: T47D_LTED_Rep1_ATACseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 1 month. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS (Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL)

tag: modifications


title: sc_A15: BY4741, WT, 2-hour


title: Lymphoma_MALT_8

value: mucosa-associated lymphoid tissue lymphoma; extranodal marginal zone B-cell lymphoma


title: sc_B20: BY4741, WT, 16-hour

value: 16 hour


title: Mf_18wpf_MS192_062


title: sc_A14: BY4741, WT, 2-hour


title: Mf_18wpf_MS192_068


title: Lymphoma_MALT_9


title: Mf_18wpf_MS192_064


title: Lymphoma_MALT_6


title: 12WK WT (R6/2 Littermate) Glial Progenitor Cells Replicate 5


title: Lymphoma_MALT_3


title: sc_A12: BY4741, WT, 2-hour


title: 12WK WT (R6/2 Littermate) Glial Progenitor Cells Replicate 1


title: Mf_18wpf_MS192_061


title: sc_A13: BY4741, WT, 2-hour


title: 12WK R6/2 Glial Progenitor Cells Replicate 4


title: 12WK R6/2 Glial Progenitor Cells Replicate 5


title: 12WK R6/2 Glial Progenitor Cells Replicate 6


title: 12WK R6/2 Glial Progenitor Cells Replicate 7


title: 12WK WT (R6/2 Littermate) Glial Progenitor Cells Replicate 2


title: Lymphoma_MALT_4


title: sc_A11: BY4741, WT, 2-hour


title: sc_A10: BY4741, WT, 2-hour


title: Mf_18wpf_MS192_060


title: Lymphoma_MALT_28


title: 6wpi_c

city: Bethesda

first: Kaya

last: Matson

state: MD

address: 35 Convent Drive

department: NINDS

institute: NIH

description:
6 weeks post injury lumbar spinal cord
F-14

data_processing:
Reads were aligned using Cellranger v.2.2.0
Exon and intron counts were tabulated using Velocyto
Data were analyzed using Seurat v3
Genome_build: mm10-1.2.0 (GRCm38.93)
Supplementary_files_format_and_content: tab-delimited text files include UMI values for each cell with tab delimeters

source_name: Lumbar spinal cord

extract_protocol:
Single nucleus dissociation was completed with a modified protocol based on our previous work64. Briefly, animals were euthanized by isoflurane inhalation and cervical dislocation. The thoracic SCI site was rapidly dissected and frozen on dry ice. Spinal cords were dounced in 500 µl sucrose buffer (0.32 M sucrose, 10 mM HEPES [pH 8.0], 5 mM CaCl2, 3 mM Mg-acetate, 0.1 mM EDTA, 1 mM DTT) and 0.1% Triton X-100 with the Kontes Dounce Tissue Grinder. 2 mL of sucrose buffer was added and filtered through a µm cell strainer. The lysate was subsequently centrifuged at 3200 g for 10 min at 4°C. The supernatant was decanted, and 3 mL of sucrose buffer added to the pellet and incubated for 1 min. The pellet was homogenized using an Ultra-Turrax and 12.5 mL of density buffer (1 M sucrose, 10 mM HEPES [pH 8.0], 3 mM Mg-acetate, 1 mM DTT) was added below the nuclei layer. The tube was centrifuged at 3200 g at 4°C and supernatant immediately poured off. The nuclei on the bottom half of the tube wall were collected with 100 µl PBS with 0.04% BSA and 0.2 U/µl RNase inhibitor. Resuspended nuclei were filtered through a 30 µm strainer. The nuclei suspension was finally adjusted to 1000 nuclei/µl.
Library preparation was carried out with 10X Genomics Chromium Single Cell Kit Version 2. The nuclei suspension was added to the Chromium RT mix to achieve loading numbers of 5000. For downstream cDNA synthesis (13 PCR cycles), library preparation and sequencing, the manufacturer’s instructions were followed.

value: spinal cord

tag: condition

value: paralyzed


title: Mf_18wpf_MS192_057


title: Mf_18wpf_MS192_059


title: 12WK R6/2 Glial Progenitor Cells Replicate 1


title: Mf_18wpf_MS192_058


title: sc_A9: BY4741, WT, 2-hour


title: Lymphoma_MALT_25


title: 6wpi_b

description:
6 weeks post injury lumbar spinal cord
F-13


title: 12WK R6/2 Glial Progenitor Cells Replicate 3


title: 12WK R6/2 Glial Progenitor Cells Replicate 2


title: Lymphoma_MALT_27


title: Mf_18wpf_MS192_056


title: Lymphoma_MALT_24


title: sc_A8: BY4741, WT, 2-hour


title: 6wpi_a

description:
6 weeks post injury lumbar spinal cord
F-12


title: Lymphoma_MALT_23


title: sc_A7: BY4741, WT, 2-hour


title: 3wpi_b

description:
3 weeks post injury lumbar spinal cord
D-26


title: Lymphoma_MALT_22


title: Lymphoma_MALT_21


title: 3wpi_a

description:
3 weeks post injury lumbar spinal cord
D-25


title: 3wpi_c

description:
3 weeks post injury lumbar spinal cord
D-29


title: Mf_18wpf_MS192_054


title: sc_A6: BY4741, WT, 2-hour


title: sc_A5: BY4741, WT, 2-hour


title: Mf_18wpf_MS192_053


title: sc_A4: BY4741, WT, 2-hour


title: Lymphoma_MALT_20


title: Mf_18wpf_MS192_052


title: 1wpi_c

description:
1 week post injury lumbar spinal cord
C-44


title: Mf_18wpf_MS192_051


title: Lymphoma_MALT_2


title: Mf_18wpf_MS192_055


title: sc_A2: BY4741, WT, 2-hour


title: Lymphoma_MALT_18


title: sc_A1: BY4741, WT, 2-hour


title: 1wpi_a

description:
1 week post injury lumbar spinal cord
C-41


title: Mf_18wpf_MS192_050


title: 1dpi_c

description:
1 day post injury lumbar spinal cord
B-51


title: Mf_18wpf_MS192_049


title: Lymphoma_MALT_17


title: NT2 [ H7YWTCCX2_5_200424_FD07688460]

city: Darlinghurst

first: Alexander

last: Swarbrick

state: NSW

address: 384 Victoria St

department: Cancer Ecosystems Program

country: Australia

institute: Garvan Institute of Medical Research

data_processing:
Mapped to mouse genome reference, CNV analysis with cn.mops (R, v1.4.2) with a read window length of 10kb.
Assembly: mm10 (GRCm38.p4)
Library strategy: WGS

source_name: cell line

extract_protocol:
QIAGEN DNeasy blood and tissue kit, according to manufacturers instructions
Roche KAPA PCR-free library preparation kit
seuqned n the HiSeqX to a depth of ~30x

value: cell line

value: cancer epithlial cell

value: Balb/c

tag: treatment


title: Mf_18wpf_MS192_048

value: Germ cell


title: Lymphoma_MALT_16


title: 1dpi_b

description:
1 day post injury lumbar spinal cord
B-50


title: NT1 [H7YWTCCX2_8_200424_FD07688484]

description: Roche KAPA PCR-free library preparation kit


title: 1dpi_a

description:
1 day post injury lumbar spinal cord
B-49


title: IE2 [H7YWTCCX2_7_200424_FD07688477]


title: Lymphoma_MALT_14


title: IE1 [H7YWTCCX2_6_200424_FD07688475]


title: 1wpi_b

description:
1 week post injury lumbar spinal cord
C-42


title: Mf_18wpf_MS192_046


title: Uninjured_c

description:
Uninjured lumbar spinal cord
A-48

value: uninjured


title: Lymphoma_MALT_13


title: Mf_18wpf_MS192_045


title: Mf_18wpf_MS192_044


title: Veh_rep3

city: La Jolla

first: Ye Ji

last: Shin

address: 9500 Gilman Dr

institute: University of California, San Diego

data_processing:
Adapter sequences were trimmed from sequenced reads using Trimmomatic v0.38 and low quality bases also trimmed.
Reads were aligned to mm10 using Bowtie2 v2.3.4.1. parameter set --maxins 5000 --very-sensitive --no-discordant --no-mixed --no-unal --mm
Mitochondrial genome and duplicated reads were removed using Samtools and MarkDuplicates in Picard.
Peak calling and Annotation of peaks were identified using HOMER v4.11 with default parameter
Assembly: mm10
Supplementary files format and content: bedGraph files were generated using HOMER for each sample

source_name: bone marrow

extract_protocol:
Neutrophils were isolated with Percoll gradient and 10 μM of PGE2 or vehicle were treated for 24 hours. After that, nuclei from 50,000 cells were isolated using Countess II Automated Cell Counter and microscopy.
Libraries were prepared using Nextera DNA Flex kit

value: Bone marrow

value: wild type

value: Vehicle


title: Uninjured_b

description:
Uninjured lumbar spinal cord
A-46


title: sc_A3: BY4741, WT, 2-hour


title: Uninjured_a

description:
Uninjured lumbar spinal cord
A-45


title: Mf_18wpf_MS192_043

value: 18 wpf


title: Mf_18wpf_MS192_042


title: Lymphoma_MALT_11

value: lymphoma specimen


title: Lymphoma_MCL_2

value: mantle cell lymphoma


title: Veh_rep2


title: Veh_rep1


title: Lymphoma_MCL_1


title: Lymphoma_MALT_15


title: PGE2_rep3

value: 10 uM PGE2


title: Mf_18wpf_MS192_047


title: Mf_18wpf_MS190_137


title: Mf_18wpf_MS192_041


title: Lymphoma_DLBCL_8

value: diffuse large B cell lymphoma


title: PGE2_rep2


title: Lymphoma_DLBCL_7


title: merged replicates AID ChIP

city: London

first: David

middle: Jonathan

last: Fear

address: 5th Floor, Tower Wing, Great Maze Pond

department: Respiratory Medicine and Allergy

country: United Kingdom

institute: Kings College London

data_processing:
Basecalls performed using CASAVA version 1.4
ChIP-seq reads were aligned to the hg19 genome build using Bowtie2 (Galaxy)
PCR duplicates, non-uniquely mapped sequences and prro quality reads were removed
peaks were called using MACS and SICER peak callers (Galaxy)
Following peak calling, regions with >2x average input read depth were removed. Peaks falling in known blacklisted regions (Duke blacklist) and regions of high input depth (top 0.1 of genome - UCSC) were excluded. Only peaks >4.5 fold enriched (3 fold for SICER) were used in subsequent analyses
Genome_build: hg19
Supplementary_files_format_and_content: bigwig files were generated by MACS

source_name: total (CD20+) B cells_AID ChIP

growth_protocol: Primary Human B cells were isolated from the tonsils of donors undergoing routine tonsilectomy and culture in the presence of anti-CD40 antibody and rhIL-4 for 48 hours

extract_protocol:
Lysates were clarified from Micrococcal treated and sonicated nuclei and histone-DNA complexes isolated with an antibody.
Libraries were prepared according to Illumina's instructions accompanying the CHIP-Seq sample prep kit. Libraries were sequenced on the Genome Analyzer II or HiSEq2000 following the manufacturer's protocols.

treatment_protocol: After 48hours B cells were crosslinked and nuclei collected

tag: subject status

value: donors undergoing routine tonsilectomy

tag: donor id

value: donor 1,2 and 3

value: tonsil

value: Primary Human B cells

tag: chip antibody

value: AID (Abcam, ab5197)


title: Mf_18wpf_MS190_135


title: Lymphoma_DLBCL_5


title: PGE2_rep1

tag: strain

tag: genotype


title: Mf_18wpf_MS190_134

tag: developmental stage


title: Lymphoma_DLBCL_4


title: Lymphoma_DLBCL_3


title: PKI3

city: Hangzhou

first: Xudong

last: Zhu

address: #2318 Yuhangtang Road

institute: Hangzhou Normal University

data_processing:
Illumina Casava software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to mm9 whole genome using bowtie v0.12.2 with parameters -q -p 4 -e 100 -y -a -m 10 --best --strata
FPKM
Genome_build: mm9
Supplementary_files_format_and_content: Differentially expressed genes (DEGs) between each genotype were calculated by a standard bioinformatic analysis package and the hierarchical clustering for DEGs between samples was generated based on DEGs

source_name: left ventricular tissue

extract_protocol:
Hearts were removed, flash frozen using liquid nitrogen, 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

value: heart

tag: genotype/variation

value: PGC1alpha overexpressed (PKI)


title: PKI2


title: rep 3 AID ChIP

value: donor 3


title: PKI1


title: Lymphoma_DLBCL_2


title: Mf_18wpf_MS190_126

tag: xenotransplantation period

tag: in vitro culture period

value: Germ cell


title: Mf_18wpf_MS190_130


title: WT3

tag: tissue

value: wild-type


title: Mf_18wpf_MS190_128


title: Lymphoma_DLBCL_1


title: rep 3 Input

source_name: total (CD20+) B cells_input


title: WT1

tag: age


title: Mf_18wpf_MS190_132


title: Mf_18wpf_MS190_124


title: T47D_LTED9M_H3K4me2_ChIPseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 9 months. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL)

extract_protocol:
Cells were grown to 80% confluence on 150cm plates and fixed with 11% solution containing formaldehyde (Thermo Fisher #28908), 50mM HEPES-KOH(pH 7.5), 100mM Nacl, 0.5M EDTA, 0.5M EGTA for 10 min. 1/20 of the plates volume equivalent of freshly prepared Glycine 2.5M(stock) was added to quench the reaction for 5min . The supernatant was aspirated, and the cells were washed twice with 5 ml PBS and harvested on ice using cell scrapers . Cells were pelleted at 2950 x g for 10 min at 4°C, washed 1x with 10 ml cold PBS, and pelleted once more. Cell pellets were flash freeze in liquid nitrogen and store at -80°C until processing day. Cell pellet collected from 6 plates of cells was resuspended in 5 ml of Lysis Buffer 1 (50 mM HEPES-KOH, pH 7.5; 140 mM NaCl, 1mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100), allowed to rotate for 10 min at 4°C, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 5 ml of Lysis Buffer 2 (10 mM Tris-HCL pH 8.0, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA), allowed to rotate for 10 min at room temperature, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 3 ml of Lysis Buffer 3 (10 mM Tris-HCl pH 8, 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine) and then transfer to 15ml sonication tubes (Diagenode cat# C01020031). Cells were sonicated on a Bioruptor Pico at 4°C for 20min of 30’’ ON-OFF cycles with 1.2 g of sonication beads. After sonication, samples were transferred to 2 separate 1.5 ml microtube and centrifuged at 20,000 x g for 15 minutes at 4°C. The supernatant was transferred to new 15 ml tubes. To check sonication efficiency, 50µl of sonicated samples was transferred to a new microtube with 50µl PBS and incubated at 65°C for 3hr on an Eppendorf Thermomixer shaking at 1000rpm to decrosslink. DNA was purified with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H20. 1 ug of decrosslinked chromatin from each sample were run in a 1% agarose gel at 100V for 45min and imaged on a Bio-rad ChemiDoc XRS+. DNA concentration in the sonicated decrosslinked samples were measured via qubit, and this concentration was then used to estimate the concentration of chromatin in the un-decrosslinked samples. 2ug of Antibody (see Key Resources table for list of antibodies) along with 0.4ug of Spike-in antibodies (Active Motif 61686) were bound to 50 ul of Dynabeads Protein G (Thermo Fisher 10004D), previously washed with 1 ml of 0.5% BSA in 1x PBS three times, and rotated end-to-end overnight at 4°C (Eppendorf cat# 0030108051). After binding of the antibodies, beads were washed with 1 ml of 0.5% BSA in 1x PBS three times. 10ug of chromatin and 20 ng of Spike-in chromatin (Active Motif 53083) were added to 1.5ml LoBind tube (Eppendorf cat# 0030108051) containing pre-bound antibodies beads and brought up to 500µl final volume with Lysis Buffer and rotated end-to-end overnight at 4°C.  5 µl was removed as input material (1%) and placed in a separate microtube at 4°C completing the volume to 50uL with Lysis Buffer 3.  Beads were washed 6 times with 1 ml Wash Buffer (50 mM Hepes-KOH, pH 7.6; 500 mM LiCl, 1 mM EDTA, 1% NP-40; 0.5% Na-Deoxycholate) and once with 1 ml TE containing 50 mM NaCl. Beads were dried after the last wash by removing as much TE as possible. 210 ul of Elution Buffer (50 mM Tris-HCl, pH 8; 10 mM EDTA, 1% SDS) was added to each sample and  150 ul elution buffer were added to each input sample that was previously set aside. To elute immunocomplexes from beads, samples were incubated at 65°C for 15 minutes on an Eppendorf Thermomixer shaking at 1000rpm. Tubes were centrifuged for 3min at 960xg at RT and 200µl of supernatant was transferred to a new tube and reverse crosslinked overnight at 65°C. Next day, 200 ul of TE was added to each decrosslinked sample and input, and the samples were treated sequentially with 0.2 mg/ml RNAse A (Sigma R5503) at 37°C and 0.2 ug/ml Proteinase K (NEB P8107S) at 55°C for two hours each. DNA purification was performed with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H2O and quantified by Qubit.
Immunoprecipitated DNA was used to generate libraries using the NEBNext Ultra DNA Library Prep Kit for Illumina (New England Biolabs cat# 7370) following the manufacturer’s instructions.

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site

value: ATCC #HTB-133

value: Parental estrogen deprived for 9 months

value: Histone H3K4me2 antibody (pAb)   Active Motif 39141


title: T47D_LTED9M_LSD1_KO_H3K4me2_ChIPseq

value: Parental long term estrogen deprived  with CRISPR/Cas9 LSD1 KO


title: ATACseq

first: Andrea

last: Streit

address: Centre for Craniofacial & Regenerative Biology, King's College London

data_processing:
Basecalls performed using CASAVA version 1.8
For ATACseq, low quality reads and adaptors were removed with cutadapt version 1.8.1 with the following conditions: cutadapt -q 10 -a CTGTCTCTTATA. For ChIP-seq low quality reads were were removed with cutadapt with following conditions:  cutadapt -q 10.
Filtered reads were aligned to the gg4 genome assembly using Novaalign version 2.07.11 with the following configurations: novoalign -F STDFQ -i PE 30-2000 -l 30 -H -k -c 32
Genome_build: gg4
Supplementary_files_format_and_content: Bigwig files were generated using deepTools. All reads were normalized to RPKM. For ChIPseq, the input was subtracted

organism: Gallus gallus

source_name: otic ectoderm

extract_protocol:
Otic ectoderm was micro-dissected from chicken embryo at 8-9 somite stage. Nuclei were released with dounce homogenizer and sonicated to get fragmented chromatin.  Specific chromatin was isolated with antibody.
For ChIPseq, libraries were prepared according to Nano-Chip protocol (Adli M & Bernstein B E, 2011) followed by NEB's instructions accompanying the library preparation kit (E6040S). Briefly, DNA was pre-amplified with sequenase for 4 cycles with primers containing BciVI restriction sites on both ends. After BciVI digestion, 3’-A overhangs were produced for direct ligation with Illumina adaptors. After purification, the library was amplified for 14 cycles with Illumina primers.The amplified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the HiSeq2000 following the manufacturer's protocols. For ATACseq, the library was prepared following the published protocol (Buenrostro J D et al, 2015) with 9 cycles of amplification.
ChIP-Seq and other

value: Bovans Brown

value: otic ectoderm

value: 8-9 somite stage

tag: chip antibodies


title: H3K27me3 ChIPseq

department: Centre for Craniofacial & Regenerative Biology

value: H3K27me3(Millipore, 07449)


title: H3K27ac ChIPseq

value: H3K27ac(Abcam, ab4729)


title: T47D_LTED9M_LSD1_KO_cjun_ChIPseq

data_processing:
ChIP-seq: Single-end ChIP-seq fastq files were processed using the ENCODE Transcription Factor and Histone ChIP-Seq processing pipeline (https://github.com/ENCODE-DCC/chip-seq-pipeline2). Reads were trimmed using cutadapt v2.5. Reads were aligned to the hg19 genome using Bowtie2 v2.3.4.3, and the SAMtools v1.9 was used to convert the output file to the BAM format. Duplicates were removed using Picard Tools v2.20.7. Peaks were called/signal tracks were generated using MACS2 v2.2.4, and peaks with fold change > 3.5 compared to input and a FDR < 0.05 were used for subsequent analysis. Only default parameters were used.
RNA-seq: Single-end RNA-seq fastq files were processed using the LSF-RNAseq Pipeline from Diderote (https://github.com/diderote/LSF-RNAseq). Reads were trimmed using cutadapt v2.3 with the following parameters: -j 4 --netseq-trim=20 -m 18. Reads were aligned to the hg19 genome using STAR v2.6.1a along default parameters, and RSEM v1.3.0 was used to obtain the expected gene counts against the human refseq also along default parameters.
RNA-seq differential expression was determined using DESeq2 v1.18.1 and R (version 3.4.1) with q-value <0.05. Ruvseq v1.12.0 was used for factor analysis to remove count based on ERCC spike-in. GSEA analysis was performed using gsea-3.0.jar. These additional analysis steps were included in the Diderote LSF-RNAseq Pipeline.
Bedtools v2.29.0 intersect was used to determine peak overlaps and assign target genes.
Homer v4.11 was used for peak annotation and motif analysis (http://homer.ucsd.edu/homer/data/software/homer.v4.11.1.zip)
Genome_build: hg19
Supplementary_files_format_and_content: The processed data files are in bigWig format containing the indexed tag chromosome coordinates and score produced from MACS2 peak calling.

extract_protocol:
Cells were grown to 80% confluence on 150cm plates and fixed with 11% solution containing formaldehyde (Thermo Fisher #28908), 50mM HEPES-KOH(pH 7.5), 100mM Nacl, 0.5M EDTA, 0.5M EGTA for 10 min. 1/20 of the plates volume equivalent of freshly prepared Glycine 2.5M(stock) was added to quench the reaction for 5min . The supernatant was aspirated, and the cells were washed twice with 5 ml PBS and harvested on ice using cell scrapers . Cells were pelleted at 2950 x g for 10 min at 4°C, washed 1x with 10 ml cold PBS, and pelleted once more. Cell pellets were flash freeze in liquid nitrogen and store at -80°C until processing day. Cell pellet collected from 6 plates of cells was resuspended in 5 ml of Lysis Buffer 1 (50 mM HEPES-KOH, pH 7.5; 140 mM NaCl, 1mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100), allowed to rotate for 10 min at 4°C, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 5 ml of Lysis Buffer 2 (10 mM Tris-HCL pH 8.0, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA), allowed to rotate for 10 min at room temperature, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 3 ml of Lysis Buffer 3 (10 mM Tris-HCl pH 8, 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine) and then transfer to 15ml sonication tubes (Diagenode cat# C01020031). Cells were sonicated on a Bioruptor Pico at 4°C for 20min of 30’’ ON-OFF cycles with 1.2 g of sonication beads. After sonication, samples were transferred to 2 separate 1.5 ml microtube and centrifuged at 20,000 x g for 15 minutes at 4°C. The supernatant was transferred to new 15 ml tubes. To check sonication efficiency, 50µl of sonicated samples was transferred to a new microtube with 50µl PBS and incubated at 65°C for 3hr on an Eppendorf Thermomixer shaking at 1000rpm to decrosslink. DNA was purified with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H20. 1 ug of decrosslinked chromatin from each sample were run in a 1% agarose gel at 100V for 45min and imaged on a Bio-rad ChemiDoc XRS+. DNA concentration in the sonicated decrosslinked samples were measured via qubit, and this concentration was then used to estimate the concentration of chromatin in the un-decrosslinked samples.  10ug of Antibody (see Key Resources table for list of antibodies) along with 2ug of Spike-in antibodies (Active Motif 61686) were bound to 50 ul of Dynabeads Protein G (Thermo Fisher 10004D), previously washed with 1 ml of 0.5% BSA in 1x PBS three times, and rotated end-to-end overnight at 4°C (Eppendorf cat# 0030108051). After binding of the antibodies , beads were washed with 1 ml of 0.5% BSA in 1x PBS three times. 30ug of chromatin and 50 ng of Spike-in chromatin (Active Motif 53083) were added to 1.5ml LoBind tube (Eppendorf cat# 0030108051) containing pre-bound antibodies beads and brought up to 500µl final volume with Lysis Buffer and rotated end-to-end overnight at 4°C.  5 µl was removed as input material (1%) and placed in a separate microtube at 4°C completing the volume to 50uL with Lysis Buffer 3.  Beads were washed 6 times with 1 ml Wash Buffer (50 mM Hepes-KOH, pH 7.6; 500 mM LiCl, 1 mM EDTA, 1% NP-40; 0.5% Na-Deoxycholate) and once with 1 ml TE containing 50 mM NaCl. Beads were dried after the last wash by removing as much TE as possible. 210 ul of Elution Buffer (50 mM Tris-HCl, pH 8; 10 mM EDTA, 1% SDS) was added to each sample and  150 ul elution buffer were added to each input sample that was previously set aside. To elute immunocomplexes from beads, samples were incubated at 65°C for 15 minutes on an Eppendorf Thermomixer shaking at 1000rpm. Tubes were centrifuged for 3min at 960xg at RT and 200µl of supernatant was transferred to a new tube and reverse crosslinked overnight at 65°C. Next day, 200 ul of TE was added to each decrosslinked sample and input, and the samples were treated sequentially with 0.2 mg/ml RNAse A (Sigma R5503) at 37°C and 0.2 ug/ml Proteinase K (NEB P8107S) at 55°C for two hours each. DNA purification was performed with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H2O and quantified by Qubit.
Immunoprecipitated DNA was used to generate libraries using the NEBNext Ultra DNA Library Prep Kit for Illumina (New England Biolabs cat# 7370) following the manufacturer’s instructions.

tag: cell type

value: Anti-c-Jun antibody (ab31419)


title: H3K4me3 ChIPseq

value: H3K4me3(Diagenode, A5051-001P)


title: Input for ChIPseq


title: G3 PGC1aOE 3

data_processing:
Illumina Casava software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to mm9 whole genome using bowtie v0.12.2 with parameters -q -p 4 -e 100 -y -a -m 10 --best --strata
FPKM (expected number of Fragments Per Kilobase of transcript sequence per Millions base pairs sequenced) were calculated using a protocol from Trapnell, et al., Nature Biotechnology, 2010.
Genome_build: mm9
Supplementary_files_format_and_content: FPKM and counts

value: G3 PGC1aOE

value: heart


title: T47D_LTED9M_LSD1_KO_LSD1_ChIPseq

tag: cell source

value: Anti-KDM1 / LSD1 antibody - ChIP Grade (ab17721)


title: G3 PGC1aOE 2


title: Mf_18wpf_MS190_123

value: 18 wpf

value: Germ cell


title: WT2

value: wild-type


title: Mf_18wpf_MS190_122


title: G3 PGC1aOE 1


title: G3 4


title: 786-O FOXD1null_1

city: New York

first: Leif

last: Oxburgh

address: 310 E 67th Street

institute: The Rogosin Institute

data_processing:
Quality was assessed per base sequence, and mean quality was calculated. Quality scores greater than 30 were deemed acceptable for further downstream analysis.
Primer and adaptor sequences were trimmed using Trimmomatic v.0.36, and only high-quality reads with >= 30 bp over a 4 bp sliding window based on the phred algorithm were accepted.
Reads were aligned to the human genome build hg38 using STAR aligner v.2.5.2b using default settings for mammalian genomes.
Unique gene hit counts were calculated using the featureCounts function from the Subread RNA-seq package v.1.5.2., and did not include intron or intergenic regions, and used Gene ID as the identifier.
Differential gene expression was calculated using DESEQ2 and exported to Excel.
Assembly: Homo sapiens GRCh38
Supplementary files format and content: triplicate samples 786-O vs 786-OFOXD1null; Differential Expression Analysis - Excel sheet
Supplementary files format and content: Ensembl Gene ID; log2FoldChange; pvalue; adjusted pvalue (padj); Delta value

growth_protocol: Cells were maintained in RPMI-1640 containing 10% FBS, 1% GlutaMax (ThermoFisher), and 1% Penicillin-Streptomycin and grown to 70% confluency on tissue culture treated plates before RNA extraction.

extract_protocol:
Qiagen REeasy Microkit with on-column DNase digestion (Qiagen). 2ug of total RNA was used for the construction of sequencing libraries.
RNA libraries for RNA-seq were prepared using Illumina Standard mRNA Library Prep Kit following manufacturer's protocols, and sequenced using Illmina HiSeq 2000 in 2x150 bp sequencing mode

tag: cell line

value: ccRCC Cell Line

value: FOXD1 null


title: 786-O FOXD1null_3


title: Mf_18wpf_MS190_125


title: 786-O FOXD1null_2


title: 24 h uncompressed AsPC-1 cells

type: RNA

city: Sapporo

first: Tatsuya

last: Nihei

state: Hokkaido

address: N10-W8, Kita-ku

institute: Hokkaido university

hyb_protocol: Biotin-labeled cDNA was injected into the GeneChipTM Human Gene 2.0 ST Array and hybridized with the probe using a GeneChip hybridization device (Hybridization Oven 645, Affymetrix, Inc.). GeneChips were washed using GeneChipTM Hybridization, Wash, and Stain Kit and an automatic GeneChip washing / staining device (Fluidics Station 450, Affymetrix, Inc.), and stained with phycoerythrin.

scan_protocol: GeneChips were scanned using the GeneChip scanner (Scanner 3000 7G、Affymetrix, Inc.). Using the analysis software GeneChip® Command Console® (Affymetrix, Inc.), and obtained a CEL file.

data_processing: The analysis software Affymetrix® Expression ConsoleTM (Affymetrix, Inc.) was used to normalize the signal values in the CEL file and create a CHP file. The normalization algorithm used was "RMA". Using the created CHP file, the expression difference was analyzed by the data mining software Transcriptome Analysis Console (Thermo Fisher Scientific Inc.).

label: biotin

source_name: 24 h uncompressed AsPC-1 cells

label_protocol: 5.5 µg of single-stranded cDNA is fragmentated and labeled with biotin according to the manufacturer's instructions (GeneChip™ WT PLUS Reagent Kit Manual Target Preparation for GeneChip™Whole Transcript (WT) Expression Arrays).

growth_protocol: AsPC-1 cells were were cultured in collagen gel overlay conditions as a 3D culture, and uncompressed or compressed by 2.9 mmHg for 24 h.

extract_protocol: Cells were lysed with TriPure Isolation Reagent (Sigma) for total RNA extraction according to the manufacturer's instructions.

treatment_protocol: AsPC-1 cells were cultured in RPMI 1640 (Thermo) supplemented with 10% FBS (Biowest SAS) and 1% antibiotics (Sigma). The cells were cultured in a humidified incubator at 37 ℃ with 5% CO2.

value: AsPC-1, human pancreatic adenocarcinoma cell line

value: uncompression


title: T47D_LTED9M_LSD1_KO_input_ChIPseq

first: Liliana

last: Garcia-Martinez

address: 1501 NW 10th Avenue

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site

value: ATCC #HTB-133

tag: modifications

value: Parental long term estrogen deprived  with CRISPR/Cas9 LSD1 KO


title: 786-O parent_3

value: parental


title: 24 h compressed AsPC-1 cells

source_name: 24 h compressed AsPC-1 cells

tag: treatment

value: compression


title: merged replicates Input

tag: subject status

value: donors undergoing routine tonsilectomy

value: donor 1,2 and 3

value: tonsil

value: Primary Human B cells


title: 786-O parent_2


title: G3 3

value: heart


title: T47D_LTED9M_2x_linked_input_ChIPseq

extract_protocol:
Cells were grown to 80% confluence on 150cm plates and fixed using dual crosslinking approach. First protein-protein interactions were preserved with ChIP-gold crosslinking reagent (Diagenode C01019027) according to manufacturer's instructions followed by  DNA-protein crosslink with 11% solution containing formaldehyde (Thermo Fisher #28908), 50mM HEPES-KOH(pH 7.5), 100mM Nacl, 0.5M EDTA, 0.5M EGTA for 10 min. 1/20 of the plates volume equivalent of freshly prepared Glycine 2.5M(stock) was added to quench the reaction for 5min . The supernatant was aspirated, and the cells were washed twice with 5 ml PBS and harvested on ice using cell scrapers . Cells were pelleted at 2950 x g for 10 min at 4°C, washed 1x with 10 ml cold PBS, and pelleted once more. Cell pellets were flash freeze in liquid nitrogen and store at -80°C until processing day. Cell pellet collected from 6 plates of cells was resuspended in 5 ml of Lysis Buffer 1 (50 mM HEPES-KOH, pH 7.5; 140 mM NaCl, 1mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100), allowed to rotate for 10 min at 4°C, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 5 ml of Lysis Buffer 2 (10 mM Tris-HCL pH 8.0, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA), allowed to rotate for 10 min at room temperature, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 3 ml of Lysis Buffer 3 (10 mM Tris-HCl pH 8, 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine) and then transfer to 15ml sonication tubes (Diagenode cat# C01020031). Cells were sonicated on a Bioruptor Pico at 4°C for 20min of 30’’ ON-OFF cycles with 1.2 g of sonication beads. After sonication, samples were transferred to 2 separate 1.5 ml microtube and centrifuged at 20,000 x g for 15 minutes at 4°C. The supernatant was transferred to new 15 ml tubes. To check sonication efficiency, 50µl of sonicated samples was transferred to a new microtube with 50µl PBS and incubated at 65°C for 3hr on an Eppendorf Thermomixer shaking at 1000rpm to decrosslink. DNA was purified with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H20. 1 ug of decrosslinked chromatin from each sample were run in a 1% agarose gel at 100V for 45min and imaged on a Bio-rad ChemiDoc XRS+. DNA concentration in the sonicated decrosslinked samples were measured via qubit, and this concentration was then used to estimate the concentration of chromatin in the un-decrosslinked samples.  10ug of Antibody (see Key Resources table for list of antibodies) along with 2ug of Spike-in antibodies (Active Motif 61686) were bound to 50 ul of Dynabeads Protein G (Thermo Fisher 10004D), previously washed with 1 ml of 0.5% BSA in 1x PBS three times, and rotated end-to-end overnight at 4°C (Eppendorf cat# 0030108051). After binding of the antibodies , beads were washed with 1 ml of 0.5% BSA in 1x PBS three times. 30ug of chromatin and 50 ng of Spike-in chromatin (Active Motif 53083) were added to 1.5ml LoBind tube (Eppendorf cat# 0030108051) containing pre-bound antibodies beads and brought up to 500µl final volume with Lysis Buffer and rotated end-to-end overnight at 4°C.  5 µl was removed as input material (1%) and placed in a separate microtube at 4°C completing the volume to 50uL with Lysis Buffer 3.  Beads were washed 6 times with 1 ml Wash Buffer (50 mM Hepes-KOH, pH 7.6; 500 mM LiCl, 1 mM EDTA, 1% NP-40; 0.5% Na-Deoxycholate) and once with 1 ml TE containing 50 mM NaCl. Beads were dried after the last wash by removing as much TE as possible. 210 ul of Elution Buffer (50 mM Tris-HCl, pH 8; 10 mM EDTA, 1% SDS) was added to each sample and  150 ul elution buffer were added to each input sample that was previously set aside. To elute immunocomplexes from beads, samples were incubated at 65°C for 15 minutes on an Eppendorf Thermomixer shaking at 1000rpm. Tubes were centrifuged for 3min at 960xg at RT and 200µl of supernatant was transferred to a new tube and reverse crosslinked overnight at 65°C. Next day, 200 ul of TE was added to each decrosslinked sample and input, and the samples were treated sequentially with 0.2 mg/ml RNAse A (Sigma R5503) at 37°C and 0.2 ug/ml Proteinase K (NEB P8107S) at 55°C for two hours each. DNA purification was performed with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H2O and quantified by Qubit.
Immunoprecipitated DNA was used to generate libraries using the NEBNext Ultra DNA Library Prep Kit for Illumina (New England Biolabs cat# 7370) following the manufacturer’s instructions.

value: Parental estrogen deprived for 9 months


title: Mf_18wpf_MS190_099

data_processing:
The processing of the 3-prime-sequencing data for calculating the gene-expression levels was performed as described previously (Nakamura et al., 2015).  In brief, all reads were processed with cutadapt v1.9.1 to remove adaptors and poly-A sequences.
The reads of 30-bp or longer were mapped onto a mouse transcript reference, ref_macFas5, annotation 101, with 3'-end of transcripts, extended by up to 10kb, using Tophat v2.0.11/Bowtie2 v2.2.7.  Read counts per gene were calculated using HTSeq v0.9.1 with the default settings.  Read counts were converted to read counts per million mapped reads (RPM) and used for the analysis.
Genome_build: macFas5
Supplementary_files_format_and_content: Tab-delimited text files include RPM values for each sample.

extract_protocol:
Fetal ovaries collected from cy fetuses at 8-18 wpf were carefully dissected out in chilled PBS.  Fetal ovaries at 8-14 wpf were minced with scissors in 0.05% trypsin/EDTA/PBS solution (GIBCO, 15400-054), then incubated for 15 min at 37℃ with gentle pipetting every 5 min.  Dissociation medium [DMEM (GIBCO, 10313021) containing 10% FBS, 2mM L-glutamine (GIBCO, 25030081), 1X penicillin/streptomycin (GIBCO, 15070063), and 100μg/mL DNaseI (MERCK, DN25)] was added for further dissociation. For fetal ovaries at 16 and 18 wpf, minced tissues were incubated in Accumax (Innovative Cell Technologies, AM105) for 40 min at 37℃ with gentle pipetting every 5 min.  Dissociation medium was added for further dissociation.  For SC3-seq analysis, cells were picked under the inverted microscope (OLYMPUS, IX71) to exclude multiplet cells.   For dissociating cy rOvaries, tissues were incubated with TryPLE Express (1X) (GIBCO, 12604021) for 5 min.  After adding dissociation medium, cells were processed for single-cell analyses. Dissociated germ cells from in vivo monkey fetal ovaries and monkey rOvaries were manually picked using glass capillaries under the stereo microscope (Leica, S8 APO).  Following cDNA synthesis and amplification methods were previously described (Nakamura et al., 2015).
The cDNA synthesis was performed according to the single-cell 3 prime RNA-seq (SC3-seq) method described previously (Nakamura et al., 2015).  The cDNA libraries for Nextseq500 (Illumina) were constructed as described previously (Ishikura et al., 2016).The sequencing was performed on an Illumina Nextseq500/550 platform with a Nextseq500/550 Hiqh output kit (Illumina).

value: 18 wpf

value: Germ cell


title: T47D_LTED9M_2x_linked_SMARCC1_ChIPseq

value: SMARCC1/BAF155 (D7F8S) Rabbit mAb #11956 Cell Signaling Technology


title: Mf_18wpf_MS190_101


title: T47D_LTED9M_2x_linked_RCOR1_ChIPseq

value: Anti-CoREST Antibody 07-455 Sigma Aldrich


title: 786-O parent_1

value: ccRCC Cell Line


title: A2780 cells, CA-3

city: nanjing

first: xiaogang

last: wang

address: Hankou road 22

institute: nanjing university

data_processing:
illumina Novaseq™ 6000 software used for basecalling.
Cutadapt software was used to remove the reads that contained adaptor contamination,(command line: ~cutadapt -a ADAPT1 -A ADAPT2 -o out1.fastq -p out2.fastq in1.fastq in2.fastq -O 5 -m 100). And After removed the low quality bases and undetermined bases ,we used HISAT2 software to map reads to the genome hg19, (command line: ~hisat2 -1 R1.fastq.gz -2 R1.fastq.gz -S sample_mapped.sam).
The mapped reads of each sample were assembled using StringTie with default parameters (command line: ~stringtie -p 4 -G genome.gtf -o output.gtf -l sample input.bam). Then, all transcriptomes from all samples were merged to reconstruct a comprehensive transcriptome using gffcompare software
After the final transcriptome was generated, StringTie and ballgown were used to estimate the expression levels of all transcripts and perform expression level for mRNAs by calculating FPKM (FPKM = [total_exon_fragments / mapped_reads(millions) × exon_length(kB)]),(command line: ~stringtie -e -B -p 4 -G merged.gtf -o samples.gtf samples.bam)
Genome_build: hg19
Supplementary_files_format_and_content: tab-delimited text file includes count and FPKM values

source_name: A2780 cells, CA

extract_protocol:
Total RNA was extracted from each sample using the Trizol method
RNA libraries were prepared for sequencing using standard Illumina protocols

value: citric acid (CA)


title: WT_plusN_Shoot_3

first: Ying

last: Li

address: 12 Waverly Place

department: Biology

institute: New York University

data_processing:
The raw sequencing reads were trimmed by the Purdue Genomics Core to remove adaptors and low-quality reads.
trimmed reads were mapped against the tomato genome (build 3.0, assessed on June 5, 2019) using BBmap
mapped reads were used to generate a count for each gene feature in the genome, using the tomato gene models ITAG3.2 and the annotated miRNA loci based on miRbase using FeatureCounts
differentially expressed genes were detected using DESeq2 (Love et al., 2014) with the design (~Genotype+Treatment+Genotype:Treatment), for sdg33 and sdg34, separately.
Genome_build: tomato genome (build 3.0, assessed on June 5, 2019)
Supplementary_files_format_and_content: normalized expression level of genes in txt format

organism: Solanum lycopersicum

source_name: shoot tissues

growth_protocol: Seeds were surface sterilized with 20% sodium hypochlorite for 20 minutes and rinsed with distilled water. The seeds were then germinated on filter paper in the dark until the radicle emerged and transferred into black sand for the seedlings to be established for one week. Seedlings were then grown hydroponically in foil tapped plastic containers with one litre nutrient medium, which consisted of 1.2 mM KNO3, 0.8mM Ca(NO3)2, 0.2mM NH4H2PO4, 0.2mM MgSO4, 50μM KCl, 12.5 μM H3BO3, 1 μM MnSO4, 1 μMZnSO4, 0.5 μMCuSO4, 0.1 μM H2MoO4, 0.1 μM NiSO4 and 10 μM Fe-EDDHA. Plants were grown in the greenhouse with a photoperiod of 16-h light and 8-h dark at 24 °C. Two plants were grown in one container and aeration was provided for one hour daily to increase the oxygen content of the nutrient medium. The nutrient medium was refreshed every three days.

extract_protocol:
Three independent plants were pooled to make one replication and three replications per treatment per genotype were used for RNA extraction. RNA for transcriptome profiling was extracted from the root and shoot samples harvested five hours after N treatments. Total RNA was extracted from the shoots and roots using Trizol (Invitrogen) according to the manufacturer’s instructions.  After extraction, total RNA was treated with DNase (New England Biolabs) according to the manufacturer’s instructions. RNA was then precipitated with 3M sodium acetate and three-times volume of 100% ethanol and resuspended in DEPC treated water. RNA was then concentrated using ZYMO RNA clean and concentrator kit (ZYMO Research) according to the manufacturer’s instructions.  The integrity of RNA was accessed by the Agilent Bioanalyzer (Agilent technologies).
RNA samples were submitted to the Purdue Genomics Core for RNA-Seq library preparation and sequencing on an Illumina Novaseq platform with paired-end 50bp format.

treatment_protocol: After two weeks of growth in the nutrient medium, plants were transferred into starvation medium, which is identical to the nutrient medium except for that 1.2 mM KNO3, 0.8mM Ca(NO3)2, 0.2mM NH4H2PO4 were replaced by 0.6mM K2SO4, 0.8mM CaSO4, 0.2mM KH2PO4 respectively. After two days in the starvation medium, plants were transferred into two treatment media: (1) with N (+N); and (2) with control (-N). The +N treatments medium was the same as the growth medium and the -N treatment medium was the same as the starvation solution.

value: WT

tag: condition

value: plusN

tag: organ

value: Shoot


title: T47D_LTED9M_cjun_ChIPseq

city: Miami

institute: University of Miami

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site

value: ATCC #HTB-133

tag: chip antibody

value: Anti-c-Jun antibody (ab31419)


title: A2780 cells, CA-2


title: Mf_16wpf_MS173_183

source_name: An oocyte from 16 wpf fetal ovary

value: 16 wpf


title: Mf_18wpf_MS238_219


title: Mf_16wpf_MS173_181


title: A2780 cells, CA-1


title: Mf_18wpf_MS190_102


title: WT_plusN_Shoot_2

tag: genotype


title: WT_plusN_Shoot_1


title: Mf_16wpf_MS173_179

tag: developmental stage


title: T47D_LTED9M_H3K27ac_ChIPseq

value: Anti-Histone H3 (acetyl K27) antibody - ChIP Grade (ab4729)


title: T47D_LTED9M_RNApol_ChIPseq

extract_protocol:
Cells were grown to 80% confluence on 150cm plates and fixed with 11% solution containing formaldehyde (Thermo Fisher #28908), 50mM HEPES-KOH(pH 7.5), 100mM Nacl, 0.5M EDTA, 0.5M EGTA for 10 min. 1/20 of the plates volume equivalent of freshly prepared Glycine 2.5M(stock) was added to quench the reaction for 5min . The supernatant was aspirated, and the cells were washed twice with 5 ml PBS and harvested on ice using cell scrapers . Cells were pelleted at 2950 x g for 10 min at 4°C, washed 1x with 10 ml cold PBS, and pelleted once more. Cell pellets were flash freeze in liquid nitrogen and store at -80°C until processing day. Cell pellet collected from 6 plates of cells was resuspended in 5 ml of Lysis Buffer 1 (50 mM HEPES-KOH, pH 7.5; 140 mM NaCl, 1mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100), allowed to rotate for 10 min at 4°C, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 5 ml of Lysis Buffer 2 (10 mM Tris-HCL pH 8.0, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA), allowed to rotate for 10 min at room temperature, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 3 ml of Lysis Buffer 3 (10 mM Tris-HCl pH 8, 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine) and then transfer to 15ml sonication tubes (Diagenode cat# C01020031). Cells were sonicated on a Bioruptor Pico at 4°C for 20min of 30’’ ON-OFF cycles with 1.2 g of sonication beads. After sonication, samples were transferred to 2 separate 1.5 ml microtube and centrifuged at 20,000 x g for 15 minutes at 4°C. The supernatant was transferred to new 15 ml tubes. To check sonication efficiency, 50µl of sonicated samples was transferred to a new microtube with 50µl PBS and incubated at 65°C for 3hr on an Eppendorf Thermomixer shaking at 1000rpm to decrosslink. DNA was purified with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H20. 1 ug of decrosslinked chromatin from each sample were run in a 1% agarose gel at 100V for 45min and imaged on a Bio-rad ChemiDoc XRS+. DNA concentration in the sonicated decrosslinked samples were measured via qubit, and this concentration was then used to estimate the concentration of chromatin in the un-decrosslinked samples. 5ug of Antibody (see Key Resources table for list of antibodies) along with 1ug of Spike-in antibodies (Active Motif 61686) were bound to 50 ul of Dynabeads Protein G (Thermo Fisher 10004D), previously washed with 1 ml of 0.5% BSA in 1x PBS three times, and rotated end-to-end overnight at 4°C (Eppendorf cat# 0030108051). After binding of the antibodies , beads were washed with 1 ml of 0.5% BSA in 1x PBS three times. 30ug of chromatin and 50 ng of Spike-in chromatin (Active Motif 53083) were added to 1.5ml LoBind tube (Eppendorf cat# 0030108051) containing pre-bound antibodies beads and brought up to 500µl final volume with Lysis Buffer and rotated end-to-end overnight at 4°C.  5 µl was removed as input material (1%) and placed in a separate microtube at 4°C completing the volume to 50uL with Lysis Buffer 3.  Beads were washed 6 times with 1 ml Wash Buffer (50 mM Hepes-KOH, pH 7.6; 500 mM LiCl, 1 mM EDTA, 1% NP-40; 0.5% Na-Deoxycholate) and once with 1 ml TE containing 50 mM NaCl. Beads were dried after the last wash by removing as much TE as possible. 210 ul of Elution Buffer (50 mM Tris-HCl, pH 8; 10 mM EDTA, 1% SDS) was added to each sample and  150 ul elution buffer were added to each input sample that was previously set aside. To elute immunocomplexes from beads, samples were incubated at 65°C for 15 minutes on an Eppendorf Thermomixer shaking at 1000rpm. Tubes were centrifuged for 3min at 960xg at RT and 200µl of supernatant was transferred to a new tube and reverse crosslinked overnight at 65°C. Next day, 200 ul of TE was added to each decrosslinked sample and input, and the samples were treated sequentially with 0.2 mg/ml RNAse A (Sigma R5503) at 37°C and 0.2 ug/ml Proteinase K (NEB P8107S) at 55°C for two hours each. DNA purification was performed with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H2O and quantified by Qubit.
Immunoprecipitated DNA was used to generate libraries using the NEBNext Ultra DNA Library Prep Kit for Illumina (New England Biolabs cat# 7370) following the manufacturer’s instructions.

value: Rpb1 NTD (D8L4Y) Rabbit mAb #14958 Cell Signaling Technology


title: A2780 cells, Control-3

source_name: A2780 cells, Control

value: Control


title: G3 2

tag: genotype/variation

tag: strain

value: heart


title: sdg34_plusN_Shoot_3


title: A2780 cells, Control-2


title: G3 1


title: T47D_LTED9M_shcjun_cjun_ChIPseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 9 months.After transduction with pLKO-shcjun, cells were selected and maintained in media containing 2 ug/ml puromycin.Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL)

value: Parental estrogen deprived for 9 months, plko-shcjun estable SHCLNG-NM_002228 (TRCN0000010366) Sigma Aldrich


title: A2780 cells, Control-1


title: Mf_16wpf_MS173_178

tag: xenotransplantation period

tag: in vitro culture period

value: Germ cell


title: sdg34_plusN_Shoot_2


title: T47D_FM_RCOR1_ChIPseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations and split 1:3 every 3 days. Culture media: RPMI-1640(Lonza 12-167Q) supplemented with 10% FBS(Gemini 100-106), 1x penicillin/streptomycin(Thermo Fisher Scientific 15140-12, 10000U/mL), and 1x glutamax( Life Technologies 35050-061).

value: Parental


title: T47D_FM_ER_ChIPseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations and split 1:3 every 3 days. Culture media: RPMI-1640(Lonza 12-167Q) supplemented with 10% FBS(Gemini 100-106), 1x penicillin/streptomycin(Thermo Fisher Scientific 15140-12, 10000U/mL), and 1x glutamax( Life Technologies 35050-061 ).

extract_protocol:
Cells were grown to 80% confluence on 150cm plates and fixed with 11% solution containing formaldehyde (Thermo Fisher #28908), 50mM HEPES-KOH(pH 7.5), 100mM Nacl, 0.5M EDTA, 0.5M EGTA for 10 min. 1/20 of the plates volume equivalent of freshly prepared Glycine 2.5M(stock) was added to quench the reaction for 5min . The supernatant was aspirated, and the cells were washed twice with 5 ml PBS and harvested on ice using cell scrapers . Cells were pelleted at 2950 x g for 10 min at 4°C, washed 1x with 10 ml cold PBS, and pelleted once more. Cell pellets were flash freeze in liquid nitrogen and store at -80°C until processing day. Cell pellet collected from 6 plates of cells was resuspended in 5 ml of Lysis Buffer 1 (50 mM HEPES-KOH, pH 7.5; 140 mM NaCl, 1mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100), allowed to rotate for 10 min at 4°C, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 5 ml of Lysis Buffer 2 (10 mM Tris-HCL pH 8.0, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA), allowed to rotate for 10 min at room temperature, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 3 ml of Lysis Buffer 3 (10 mM Tris-HCl pH 8, 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine) and then transfer to 15ml sonication tubes (Diagenode cat# C01020031). Cells were sonicated on a Bioruptor Pico at 4°C for 20min of 30’’ ON-OFF cycles with 1.2 g of sonication beads. After sonication, samples were transferred to 2 separate 1.5 ml microtube and centrifuged at 20,000 x g for 15 minutes at 4°C. The supernatant was transferred to new 15 ml tubes. To check sonication efficiency, 50µl of sonicated samples was transferred to a new microtube with 50µl PBS and incubated at 65°C for 3hr on an Eppendorf Thermomixer shaking at 1000rpm to decrosslink. DNA was purified with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H20. 1 ug of decrosslinked chromatin from each sample were run in a 1% agarose gel at 100V for 45min and imaged on a Bio-rad ChemiDoc XRS+. DNA concentration in the sonicated decrosslinked samples were measured via qubit, and this concentration was then used to estimate the concentration of chromatin in the un-decrosslinked samples.  5ug of Antibody (see Key Resources table for list of antibodies) along with 1ug of Spike-in antibodies (Active Motif 61686) were bound to 50 ul of Dynabeads Protein G (Thermo Fisher 10004D), previously washed with 1 ml of 0.5% BSA in 1x PBS three times, and rotated end-to-end overnight at 4°C (Eppendorf cat# 0030108051). After binding of the antibodies , beads were washed with 1 ml of 0.5% BSA in 1x PBS three times. 30ug of chromatin and 50 ng of Spike-in chromatin (Active Motif 53083) were added to 1.5ml LoBind tube (Eppendorf cat# 0030108051) containing pre-bound antibodies beads and brought up to 500µl final volume with Lysis Buffer and rotated end-to-end overnight at 4°C.  5 µl was removed as input material (1%) and placed in a separate microtube at 4°C completing the volume to 50uL with Lysis Buffer 3.  Beads were washed 6 times with 1 ml Wash Buffer (50 mM Hepes-KOH, pH 7.6; 500 mM LiCl, 1 mM EDTA, 1% NP-40; 0.5% Na-Deoxycholate) and once with 1 ml TE containing 50 mM NaCl. Beads were dried after the last wash by removing as much TE as possible. 210 ul of Elution Buffer (50 mM Tris-HCl, pH 8; 10 mM EDTA, 1% SDS) was added to each sample and  150 ul elution buffer were added to each input sample that was previously set aside. To elute immunocomplexes from beads, samples were incubated at 65°C for 15 minutes on an Eppendorf Thermomixer shaking at 1000rpm. Tubes were centrifuged for 3min at 960xg at RT and 200µl of supernatant was transferred to a new tube and reverse crosslinked overnight at 65°C. Next day, 200 ul of TE was added to each decrosslinked sample and input, and the samples were treated sequentially with 0.2 mg/ml RNAse A (Sigma R5503) at 37°C and 0.2 ug/ml Proteinase K (NEB P8107S) at 55°C for two hours each. DNA purification was performed with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H2O and quantified by Qubit.
Immunoprecipitated DNA was used to generate libraries using the NEBNext Ultra DNA Library Prep Kit for Illumina (New England Biolabs cat# 7370) following the manufacturer’s instructions.

value: Anti-Estrogen Receptor alpha antibody (ab3575)


title: sdg34_plusN_Shoot_1


title: Mf_16wpf_MS173_177