title: 210_B5_EB_3D_3B_SC_Batch1


title: WGBS_EpiSC_WT_Rep2

status: Public on Jan 23 2023

type: SRA

city: Turin

first: Andrea

last: Lauria

address: Via Accademia Albertina, 13

department: Life Sciences and Systems Biology

country: Italy

institute: University of Turin

library_source: genomic

data_processing:
Following quality controls, sequencing reads were processed with Trim Galore! v0.5.0 to perform quality and adapter trimming (parameters: --stringency 3 –q 20 --paired). Trimmed reads were next aligned to the mouse reference genome (UCSC mm10/GRCm38) using Bismark v0.22.3.The bisulfite-converted genome was created using the bismark_genome_preparation utility (parameters: --genomic_composition --bowtie2). Read mapping was performed with the bismark command (parameters: --nucleotide_coverage). Duplicated alignments were removed with the deduplicate_bismark utility and methylation calling was carried out using the bismark_methylation_extractor utility (parameters: --ignore 1 --bedGraph --counts --gzip). Genome-wide cytosine methylation reports with the top and bottom strand methylation evidence pooled into a single CpG dinucleotide entity were obtained using the coverage2cytosine utility (parameters: --zero_based --gzip --merge_CpG).
Genome_build: mm10/GRCm38
Supplementary_files_format_and_content: BigWig files containing whole-genome CpG methylation scores at single-base resolution.

molecule: genomic DNA

organism: Mus musculus

source_name: Mouse Epiblast Stem Cells

growth_protocol: E14 mouse WT and mutant (Dnmt3a -/-   and Dnmt3b -/-) ESCs were cultured in high-glucose DMEM (Euroclone) supplemented with 15% FBS (Millipore Corp., Billerica, MA, USA), 0.1 mmol/l nonessential amino acids (Invitrogen), 1 mmol/l sodium pyruvate (Invitrogen), 0.1 mmol/l β-mercaptoethanol, 1500 U/ml Leukemia Inhibitory Factor (LIF; Millipore), 25 U/ml penicillin, and 25 μg/ml streptomycin. For EpiSC induction, a single-cell suspension was seeded onto Geltrex (A1413202 GIBCO)-coated plates at a density of 10,000 cells cm-2 in N2B27 medium supplemented with 20ng/ml ActivinA (PHC9564 GIBCO) and 12 ng/ml bFGF (PHG0026 GIBCO). The cells were passaged 1:3 as small clumps using Collagenase IV (17104019 GIBCO). EpiSCs were collected for DNA and RNA analyses after 14 days of induction followed by daily medium changes. N2B27 medium is composed by 50% advanced DMEM/F12 (12634028 GIBCO) and 50% Neurobasal medium (21103049 GIBCO), supplemented with 0.5% N2 Supplement (17502048 GIBCO), 1% B27 Supplement (17504044 GIBCO), 0.033% BSA solution (A9647 SIGMA), 50 uM β-mercaptoethanol (M3148 Sigma), 2mM Glutamax (35050038 GIBCO), 100U/ml penicillin and 100 ug/ml streptomycin (DE17-602E LONZA).

extract_protocol:
Genomic DNA was extracted from cells using the DNeasy Blood and Tissue kit (QIAGEN, 69506) following the manufacturer’s instructions.
The whole-genome bisulfite-seq library was prepared starting from 5 μg of sonicated genomic DNA for 2 runs of 10 cycles [30 sec ‘‘ON’’, 30 sec ‘‘OFF’’]. In order to obtain 200 nt fragments. Sonicated DNA was then end-repaired twice, dA-tailed, and ligated to adapters, using the Illumina TruSeq DNA Sample Prep Kit, following manufacturer’s instructions. Adapter-ligated DNA was loaded on an EGel Size select 2% agarose pre-cast gel (Invitrogen), and a fraction corresponding to fragments ranging from 25 0bp to 350bp was recovered. Purified DNA was then subjected to bisulfite conversion using the EpiTect Bisulfite Kit (QIAGEN) following the manufacturer’s instruction except for the double desulfonation after the conversion. Bisulfite-converted DNA was finally enriched by 15 cycles of PCR using Pfu Turbo Cx. HotStart Taq (Agilent). Libraries were sequenced on Illumina Novaseq 6000 System.

tag: strain

tag: cell type

value: ES-E14 derived EpiSC

tag: genotype

value: Wild type (WT)

tag: clone

value: WT

tag: Stage

value: EpiSC


title: WGBS_ESC_3BKO_B126

source_name: Mouse Embryonic Stem Cells

value: Dnmt3b -/-

value: ESC


title: 210_B3_EB_3D_3B_SC_Batch1

library_source: transcriptomic

data_processing:
Following quality controls (performed with FastQC v0.11.2 (https://www.bioinformatics.babraham.ac.uk/projects/fastqc)), sequencing reads were processed with Trim Galore! v0.5.0 (https://www.bioinformatics.babraham.ac.uk/projects/trim_galore) to perform quality and adapter trimming (parameters: --stringency 3 –q 20). Trimmed reads were next aligned to the mouse reference genome (mm10/GRCm38.p6) using STAR v2.7.1a with options: --outFilterMultimapNmax 10 --outFilterMultimapScoreRange 1 –outFilterMismatchNmax 999 –outFilterMismatchNoverLmax 0.04. Gene expression levels were quantified with featureCounts v1.6.1 (options: -t exon -g gene_name) using the GENCODE Release M23 annotation. Multi-mapped reads were excluded from quantification.
Genome_build: mm10/GRCm38.p6
Supplementary_files_format_and_content: Tab-delimited text file of raw read counts matrix per gene/cell.

molecule: total RNA

source_name: Mouse Embryoid Bodies

growth_protocol: E14 mouse WT and mutant (Dnmt3a -/-   and Dnmt3b -/-) ESCs were cultured in high-glucose DMEM (Euroclone) supplemented with 15% FBS (Millipore Corp., Billerica, MA, USA), 0.1 mmol/l nonessential amino acids (Invitrogen), 1 mmol/l sodium pyruvate (Invitrogen), 0.1 mmol/l β-mercaptoethanol, 1500 U/ml Leukemia Inhibitory Factor (LIF; Millipore), 25 U/ml penicillin, and 25 μg/ml streptomycin. To induce formation of EBs, Wild type and mutant ESCs were transferred using trypsin to low-attachment plates (CORNING) in Alpha-MEM (BE02-002F LONZA) supplemented with 10% KOSR (10828-028 GIBCO), 5% FBS (Millipore), 1% nonessential amino acids (Invitrogen), 1% sodium pyruvate (Invitrogen), 0.1 mmol/l β-mercaptoethanol, 25 U/ml penicillin and 25 μg/ml streptomycin. Medium was changed every 3 days.

extract_protocol:
Full length single cell RNA-seq was performed using a modified version of the Smart-seq2 protocol.
Briefly, individual cells are sorted into 96 well plates containing lysis buffer in presence of RNase inhibitor, dNTPs and OligodT. Reverse transcription of the polyadenylated RNA will be performed with SuperScriptII and Template Switching Oligos. The resulting cDNA will be amplified with 25 cycles of PCR and libraries will be prepared for sequencing with standard NexteraXT Illumina protocol. Libraries were sequenced on Illumina NextSeq 500 System (single-end 75bp reads), reaching a median of ~ 578,000 generated reads per cell.

value: Embryoid Bodies (EB)

value: Dnmt3b -/- (3BKO)

tag: batch


title: Sample 2_Aorta_HFD_Tie2Ctrl

city: Helsinki

first: Andrey

last: Anisimov

address: Haartmaninkatu 8

department: Faculty of medicine

country: Finland

institute: University of Helsinki

data_processing:
The Cell Ranger  mkfastq and count pipelines (10x Genomics, Pleasanton, CA) were used to demultiplex and convert Chromium single-cell 3’ RNA-sequencing barcodes and read data to FASTQ files and to generate aligned reads and gene-cell matrices.
Reads were aligned to mouse reference genome mm10
We used the Seurat R package (v3.1) for QC, filtering and analysis of the data (Butler et al. Nature Biotechnology 2018)
Cells were filtered based on the number of genes and the percentage of mitochondrial genes.
For further analysis, Seurat R package was used.
Genome_build: mm10
Supplementary_files_format_and_content: barcode, features, and matrix

molecule: polyA RNA

growth_protocol: In general, all mice are fed with regular chow diet until the age of 22 weeks (or 26 weeks, as detailed in treatment protocol). At this age of the mice the diet is changed with the high-fat (western) diet for 4 or 20 weeks, as detailed in the treatment protocol.

extract_protocol:
The aortas were harvested in PBS and their surrounding adipose tissues were removed. The aortas were then minced into pieces and digested in DPBS containing 1 mg/ml collagenases Type I (Gibco #17100-017), II (Gibco #17101-015) and IV (Gibco #17104-019) each for 30 minutes at 37oC water bath with mild mixing for a few times during the digestion. After the digestion, the cell suspension was filtered through a 70 μm cell strainer. The red blood cells were lysed with ACK Lysing buffer (Gibco # A10492-01) according to the instructions. Left lobes of the lungs were minced using scissors in a mixture of 0.5 mg/mL Collagenase H (Roche), 0.5 U/mL of Dispase II (Sigma), 1 U/mL DNase I (Thermo Scientific) and 1% (w/v) BSA (Biowest) in 4 mL of phosphate buffered saline (PBS) and incubated at 37ºC with constant agitation for 20 minutes. The slurry was then passed through a 70 µm nylon cell strainer (Corning) and the reaction was stopped with 10 mL of 10% heat-inactivated FBS in FACS-buffer (1% heat-inactivated FBS, 2 mM EDTA in PBS). Red blood cells were lysed by adding 1 mL of Red Blood Cell Lysis buffer (Sigma-Aldrich). The cells were incubated with Fc-blocker, as above, and the alive CD31+DAPI- cells were sorted using a BD Influx flow cytometer in single cell 1 drop setup with a 100 µm nozzle. After the FACS step,  lung ECs were resuspended in PBS containing 0.05% of 0.2 µm filter sterile BSA. Total bone marrow cells were isolated from both (left and right) femur of a mouse by flashing with the FACS buffer (PBS, supllemented with 0.05% BSA).
We used the Chromium Single-Cell 3′RNA-sequencing system (10x Genomics, Pleasanton, CA) with the Reagent Kit v3.1 according to the manufacturer’s instructions.
Sample libraries were sequenced on the Illumina NovaSeq 6000 system using S2 flow cell (Illumina)

treatment_protocol:
For sample 1-4, mice received tamoxifen via gavage at 10 weeks old. Pcsk9-AAV and HFD were started at 26 weeks of age for a total of 4 weeks.
For samples 5-6, mice received tamoxifen via gavage at 10 weeks old. Pcsk9-AAV and HFD were started at 22 weeks of age for a total of 20 weeks.

tag: tissue

value: Aorta


title: 210_B1_EB_3D_3B_SC_Batch1


title: WGBS_EpiSC_WT_Rep1


title: 210_B2_EB_3D_3B_SC_Batch1


title: GS-9/10_N1_lymph_node_met (CGH087_LK)

type: genomic

city: Munich

first: Kristian

last: Unger

address: Marchioninistr. 15

department: Department of radiation oncology

country: Germany

institute: LMU University Hospital

description:
macrodissected FFPE sample
CGH087_LK

hyb_protocol: Converted gDNA was hybridized on Infinium®MethylationEPIC BeadChips (Illumina, San Diego, CA, USA) according to the manufacturer's protocol.

scan_protocol: Bead Array Slides were scanned using the Illumina Bead Array Reader according to the manufacturer's protocol.

data_processing:
GenomeStudio (version 2011.1) with Methylation Module (version 1.9.0) was used to process the raw image data generated by the BeadArray Reader followed by import and preprocessing using the RnBeads R package. Beta values were normalized using the dasen method from the R watermelon package
matrix_proc_df_111222.txt:  normalized (dasen) Average Beta
signals_df_111222.txt:  Unmethylated and methylated signal intensities

label: Cy5 and Cy3

organism: Homo sapiens

label_protocol: DNA methylation profiling was performed using Illumina Infinium MethylationEPIC 850 k arrays. Processing of DNA Methylation arrays was performed as follows: Bisulfite conversion of 500ng of DNA was done using the EZ DNA Methylation Kit (Zymo Research, Orange, CA, USA)  according to the manufacturer's protocol.

growth_protocol: Serial sections (10 µm) were generated from available formalin-fixed and paraffin-embedded (FFPE) tumor tissue blocks (n=100), with the first and last sections of each series stained with hematoxylin and eosin (HE). The HE stained tissue sections were histopathologically reviewed by a pathologist and the tumor area was defined. If necessary, the annotated tumor area was macro-dissected.

extract_protocol: Simultaneous DNA and RNA extraction using the AllPrep DNA/RNA FFPE Kit (Qiagen, Hilden, Germany) according to the manufacturer's protocol. Isolated DNA was quantified using the Qubit 2.0 fluorometer.

tag: disease state

value: prostate cancer

tag: gleason group

tag: lymph node status

tag: tumor type


title: WGBS_EpiSC_3BKO_B126


title: WGBS_ESC_3BKO_B77


title: GS-9/10_N1_lymph_node_met (CGH053_LK)

description:
macrodissected FFPE sample
CGH053_LK


title: Naive_ESC_WT_rep2

city: San Francisco

first: Robert

last: Blelloch

state: CA

address: 35 Medical Center Way

country: USA

institute: UCSF

description: 3'end capture

data_processing:
Adapters for sample FastQ files were trimmed using cutadapt v2.5 followed by alignment to the mm10 genome using STAR align 2.7.2b. Featurecounts from Subread v1.6.4 was used to generate a counts matrix of reads per gene. FastQC v0.11.8 and MultiQC were used to validate quality sequencing and mapping.
The gene count matrix was converted to CPM and filtered for genes greater than 1 cpm in at least two total samples. Samples were normalized using TMM. Next, Log2 CPM averages were calculated for replicates of each sample for scatterplot visualization and nearest neighbor TSS analysis. All transcriptomics analyses were conducted using CPM values from TMM normalization of all samples except for nearest neighbor analysis where WT and DKO were TMM normalized together. To conduct differential gene expression we performed DESeq2 v1.34.0 analysis using the raw gene count matrix as input for each desired comparison.
Assembly: mm10
Supplementary files format and content: csv file containing the average CPMs between replicates for each gene of filtered and TMM-normalized samples

extract_protocol:
Total RNA was extracted and purified from cells using Trizol followed by ethanol precipitation
RNA-seq libraries were generated using the QuantSeq 3’ mRNA-Seq Library Prep Kit FWD for Illumina (Lexogen, CAT#A01172) according to their protocol using 200ng of total RNA for input. We utilized the PCR Add-on kit for Illumina (Lexogen, CAT#M02096) to determine an appropriate number of PCR cycles to amplify libraries. Amplified libraries were quantified using Agilent Tapestation 4200. Libraries were pooled and sequenced using a HiSeq 4000 to obtain single end 50bp reads. At least 10 million mapped reads or more per sample were obtained.

tag: cell line


title: GS-6/7a_N1_primary_tumor (CGH063_PCA)

description:
macrodissected FFPE sample
CGH063_PCA


title: Naive_ESC_WT_rep3

description: 3'end capture


title: 210_B12_EB_3D_3A_SC_Batch2

value: Dnmt3a -/- (3AKO)


title: Naive_ESC_MLL3KO_MLL4fl_rep1

description: 3'end capture


title: WGBS_ESC_WT_Rep2


title: 210_B11_EB_3D_3A_SC_Batch2


title: GS-6/7a_N1_lymph_node_met (CGH063_LK)

description:
macrodissected FFPE sample
CGH063_LK


title: GS-6/7a_N1_primary_tumor (CGH004_PCA)

description:
macrodissected FFPE sample
CGH004_PCA


title: WGBS_ESC_WT_Rep1


title: Naive_ESC_WT_rep1

description: 3'end capture


title: GS-9/10_N1_primary_tumor (CGH086_PCA)

description:
macrodissected FFPE sample
CGH086_PCA


title: CD63 low, replicate 1

city: New York

first: Friederike

last: Dündar

state: New York

address: 1300 York Ave

department: Physiology & Biophysics

institute: Weill Cornell Medicine

data_processing:
The quality of the sequenced reads was assessed with FastQC and QoRTs.
Sequenced reads were aligned with default parameters to the mouse reference genome (mm10 from 10X "refdata-cellranger-mm10-3.0.0") using STAR v2.6.0c.
Gene expression estimates were obtained with featureCounts v1.6.2 using composite gene models from 10X pre-built mouse reference GTF ("refdata-cellranger-mm10-3.0.0").
Assembly: mm10
Supplementary files format and content: tab-delimited text files include raw abundance values for each sample as obtained from featureCounts

source_name: Pancreatic Islets

extract_protocol:
Mouse pancreatic islets from 10 weeks-old C57BL/6J male mice were isolated by perfusion of the pancreases with CIzyme (VitaCyte) through the common hepatic duct and digested at 37°C for 17 minutes. After gradient-based separation using Histopaque-1077 (Sigma), islets were then hand-picked to avoid exocrine contamination. Pancreatic islets were isolated and cultured overnight in RPMI 1640 supplemented with 10% FBS and penicillin/streptomycin. Then, islets were dissociated with 0.05% Trypsin (Corning) for 10 minutes at 37°C. Dissociated cells were passed through a 35 μm nylon mesh and resuspended in 1x PBS + 0.05% BSA. Then, we FACS-sorted beta cells based on CD63 expression. After RNA isolation, total RNA integrity was analyzed using an Agilent 2100 Bioanalyzer and concentrations were measured using the NanoDrop Spectrophotometer.
Preparation of the RNA sample library were performed by the Genomics Core Laboratory at Weill Cornell Medicine. cDNA was generated using the SMARTer v4 Ultra Low Input RNA Kit (Clontech, # 63488) and Nextera XT kit (Nextera XT DNA Library Preparation Kit (Illumina, San Diego, CA) was used to make cDNA libraries suitable for Illumina sequencing.

value: Pancreatic Islets

tag: cd63 expression

value: CD63 lo

value: beta cells

value: Wild type


title: GS-9/10_N1_primary_tumor (CGH053_PCA)

description:
macrodissected FFPE sample
CGH053_PCA


title: CD63 low, replicate 2


title: AR_Tie1CKO

data_processing:
The Cell Ranger  mkfastq and count pipelines (10x Genomics, Pleasanton, CA) were used to demultiplex and convert Chromium single-cell 3’ RNA-sequencing barcodes and read data to FASTQ files and to generate aligned reads and gene-cell matrices.
Reads were aligned to mouse reference genome mm10
We used the Seurat R package (v3.1) for QC, filtering and analysis of the data (Butler et al. Nature Biotechnology 2018)
Cells were filtered based on the number of genes and the percentage of mitochondrial genes.
For further analysis, Seurat R package was used.
Genome_build: mm10
Supplementary_files_format_and_content: filtered barcodes.tsv
Supplementary_files_format_and_content: filtered features.tsv
Supplementary_files_format_and_content: filtered matrix.mtx

extract_protocol:
The aortas were harvested in PBS and their surrounding adipose tissues were removed. The aortas were then minced into pieces and digested in DPBS containing 1 mg/ml collagenases Type I (Gibco #17100-017), II (Gibco #17101-015) and IV (Gibco #17104-019) each for 30 minutes at 37oC water bath with mild mixing for a few times during the digestion. After the digestion, the cell suspension was filtered through a 70 μm cell strainer. The red blood cells were lysed with ACK Lysing buffer (Gibco # A10492-01) according to the instructions.
For sample 1-6, we use the Chromium Single-Cell 3′RNA-sequencing system (10x Genomics, Pleasanton, CA) with the Reagent Kit v3.1 according to the manufacturer’s instructions. For sample 7-10, we use the Chromium Single-Cell 3′RNA-sequencing system (10x Genomics, Pleasanton, CA) with the Reagent Kit v2.1
Sample libraries were sequenced on the Illumina NovaSeq 6000 system using S2 flow cell (Illumina)

treatment_protocol: For sample 1-2, mice received tamoxifen via gavage at 9-10 weeks old.

value: Aorta (thoracic/abdominal)


title: CD63 high, replicate 2

value: CD63 hi


title: 210_A6_EB_3D_3B_SC_Batch1


title: 210_A5_EB_3D_3B_SC_Batch1


title: PBS- and LNP- treated murine liver cells

city: Atlanta

first: James

last: Dahlman

state: ga

address: 311 Ferst dr

institute: Georgia Institute of Technology

data_processing:
Library strategy: SENT-seq
R1 contains the cell barcodes and R2 contains the sample and LNP barcodes along with the mRNA sequences.
The data was processed using zUMIs (v 2.9.7) for RNA mapping and counting and Salmon Alevin (v1.5.2) for the DNA barcode and cell hashes. All samples were mapped to GRCm39 and only Exonic regions were counted. All output files were loaded into Seurat (v 4.0.4) and in summary, cells were log normalized to a scale factor of 10,000 then scaled using a linear transformation. This was followed by PCA dimensional reduction, t-SNE clustering and then exported using rBCS for further analysis in BBrowser2 (v2.9.23). Once in BBrowser2, the cell search tool was used to identify the cell types within each cluster and gene expression profiles were compared within cell types of interest. Barcode counts were combined with RNA counts in Seurat and treated in a similar manner to other multimodal datasets such as CITE-seq.
Genome_build: GRCm39
Supplementary_files_format_and_content: Seurat v4.0.4 RDS file. This file includes PBS vs LNP metadata and identities of the 19 distinct LNPs.

source_name: Murine liver  cells

extract_protocol:
Mice were sacrificed 1 day after administration of LNPs and immediately perfused with 20 mL of 1X PBS through the right atrium. The liver was isolated immediately following perfusion, minced with scissors, and then placed in a digestive enzyme solution with Collagenase Type I (Sigma Aldrich), Collagenase XI (Sigma Aldrich), and Hyaluronidase (Sigma Aldrich) at 37oC and 750 rpm for 45 minutes. Digested tissues were passed through a 70 μm filter and red blood cells were lysed. Cells were stained to identify specific cell populations and sorted using a BD FacsFusion cell sorter. To allow for pooling of samples into a single device, cells were stained with a streptavidin conjugated H-2 MHC class I (M1/42, BioLegend) antibody and biotinylated cell hash oligos were added at 0.5 uM final concentration after a single wash to remove unbound antibody.
Pooled microwell-seq barcoded beads were placed in a reverse transcription reaction containing 200 U M-MLV Reverse Transcriptase (Biochain Institute), 1x RT buffer, 20 U RNAse inhibitor (NEB), 1 M betine (Sigma), 6 mM MgCl2 (Sigma), 2.5 mM DTT (Thermo Fisher), 1mM dNTP (NEB), and 1 uM TSO primer. The beads were incubated for 90 minutes at 42 C for 90 minutes followed by a hold 4 C with constant shaking at 500 RPM. After the reverse transcriptase step, enzyme was removed with a wash of 1 x TE with 0.5% SDS , followed by a 1 x TE with 0.01% Tween 20 and finally a wash in 100 mM Tris-HCl pH 8.0. To remove any unused single stranded oligo from the beads they were treated with 200 U of Exonuclease I (NEB) in 1x ExoI buffer for 60 minutes at 37 C with 500 RPM shaking. Following the digestion, excess ExoI was removed using the previously described TE-SDS, TE-Tween 20 and Tris-HCL pH 8.0 washes. After removal of ExoI, the beads were resuspended in 200 ul of Platinum II hot-start master mix (Thermo Fisher) with IS-PCR, p7 Multi Barcode Rvs and Hash p7 Rvs primers and the first round PCR was performed using the following cycling conditions: one cycle at 94 C for 2 minutes, 12 cycles of 94 C for 15 seconds, 60 C for 15 seconds and 68 C for 2 minutes. The sample was pooled, the beads were removed and discarded, and the sample was purified using 0.6x SPRI beads. The long RNA fragments were collected on the SPRI beads, while the shorter barcode and hash reads remained in the PCR supernatant, these were subsequently purified using 2.0x SPRI beads and saved for use during the final round PCR. The RNA sample was then treated with TN5 transposase to fragment and add on sequencing handles for PCR. Both the DNA and fragmented RNA sample was then amplified using a final round of PCR with non-hot start Q5 high fidelity polymerase (NEB), P7 Nextera index adapters and the Microwell P5 primer using the following cycling conditions: one cycle at 70 C for 5 minutes, 12 cycles of 98 C for 30 seconds, 58 C for 30 secconds and 72 C for 90 seconds with a final extension at 72 C for 2 minutes. The samples were then purified using 0.8x SPRI beads and pooled at a 10:1 molar ratio of RNA to DNA and sequenced on a Illumina HiSeq paired-end 150 cycle run.

value: FACS sorted liver cells

tag: time

value: 18 hours after injection

tag: molecule subtype

value: mRNA, barcode DNA


title: GS-6/7a_N1_primary_tumor (CGH061_PCA)

description:
macrodissected FFPE sample
CGH061_PCA


title: GS-6/7a_N1_lymph_node_met (CGH004_LK)

description:
macrodissected FFPE sample
CGH004_LK


title: 210_A9_EB_3D_3A_SC_Batch2


title: CD63 high, replicate 1


title: AR_Tie1Ctrl


title: 210_A8_EB_3D_3B_SC_Batch1


title: GS-9/10_N1_lymph_node_met (CGH049_LK)

description:
macrodissected FFPE sample
CGH049_LK


title: AP_Tie1CKO

value: Aorta (aortic arch)


title: 210_B10_EB_3D_3A_SC_Batch2


title: AP_Tie2CKO


title: Pancreas cells of high-fat diet-fed mice

description: 10X Genomics

data_processing:
Demultiplexing, barcoded processing, gene counting and aggregation done with Cell Ranger software v2.1.0
Sample integration (batch correction), expression value normalization, clustering, cell type annotation and so on were done with SCTransform, scater, and scran. For more details and the cell type annotation information, see https://github.com/abcwcm/RubioNavarro_Pancreas

extract_protocol:
Mouse pancreatic islets were isolated by perfusion of the pancreases with CIzyme (VitaCyte) through the common hepatic duct and digested at 37°C for 17 minutes. After gradient-based separation using Histopaque-1077 (Sigma), islets were then hand-picked to avoid exocrine contamination. Pancreatic islets were isolated and cultured overnight in RPMI 1640 supplemented with 10% FBS and penicillin/streptomycin. Then, islets were dissociated with 0.05% Trypsin (Corning) for 10 minutes at 37°C. Dissociated cells were passed through a 35 μm nylon mesh and resuspended in 1x PBS + 0.05% BSA. Cellular viability was higher than 95% as determined by using Trypan Blue assay and doublets represented less than 5% of total cells.
Library prep was performed according to the manufacturer’s instructions (single cell 3' v2 protocol, 10x Genomics).

treatment_protocol: C57BL/6J wild-type mice  were fed a regular chow diet (RD, N=4) or a 60% high fat diet (HFD, N=5) (Research Diets) for 6 weeks after weaning.

value: Pancreatic islet cells

tag: treatment

value: Animals fed with high fat diet


title: AR_Tie2CKO


title: GS-9/10_N1_lymph_node_met (CGH086_LK)

description:
macrodissected FFPE sample
CGH086_LK


title: AP_Tie1Ctrl


title: 210_A4_EB_3D_3B_SC_Batch1

value: Embryoid Bodies (EB)


title: 210_A3_EB_3D_3B_SC_Batch1

value: Dnmt3b -/- (3BKO)


title: GS-6/7a_N1_primary_tumor (CGH002_PCA)

description:
macrodissected FFPE sample
CGH002_PCA


title: VE-PTPiKO-IR rep3 [S4723-IR-KO]

city: Chicago

first: Yalu

last: Zhou

state: IL

address: 303 E. Superior St,

department: Department of Biochemistry and Molecular Genetics

institute: Northwestern University

data_processing:
The quality of DNA reads was evaluated using FastQC
Adapters were trimmed and reads of poor quality or aligning to rRNA sequences were filtered. The cleaned reads were aligned to the Mus musculus genome (mm10) using STAR.
Read counts for each gene were calculated using HTSeq-count and a gene annotation file for mm10 obtained from UCSC (University of California Santa Cruz; http://genome.ucsc.edu)
Normalization and differential expression were determined using DESeq2
Genome_build: Mus musculus genome (mm10)
Supplementary_files_format_and_content: Normalization and differential expression

extract_protocol:
Total RNA was extracted from whole kidney using Trizol (Invitrogen) according to manufacturer’s instructions. The integrity and quantity of total RNA was determined using a Agilent 2100 Bioanalyzer
In total 1mg of total RNA was used to generate RNAseq libraries using TruSeq RNA sample preparation kit (Illumina) according to the manufacturer’s instructions.

tag: age

value: Kidney

tag: genotype/variation

value: VE-PTPiKO

value: ischemia-reperfusion (IR) injury


title: GS-9/10_N1_primary_tumor (CGH049_PCA)

description:
macrodissected FFPE sample
CGH049_PCA

value: prostate cancer


title: 210_A7_EB_3D_3B_SC_Batch1

tag: strain

tag: cell type


title: VE-PTPiKO-IR rep1 [S4716-IR-KO]


title: 210_A12_EB_3D_3A_SC_Batch2

tag: genotype

value: Dnmt3a -/- (3AKO)


title: GS-6/7a_N1_lymph_node_met (CGH061_LK)

description:
macrodissected FFPE sample
CGH061_LK


title: Pancreas cells of WT mice

description: 10X Genomics

value: Pancreatic Islets

value: Animals fed with chow diet


title: AR_Tie2Ctrl

value: Aorta (thoracic/abdominal)


title: VE-PTPiKO-IR rep2 [S4718-IR-KO]


title: WGBS_EpiSC_3BKO_B77

value: ES-E14 derived EpiSC

value: Dnmt3b -/-

tag: clone

tag: Stage

value: EpiSC


title: GS-9/10_N1_primary_tumor (CGH085_PCA)

description:
macrodissected FFPE sample
CGH085_PCA


title: 210_A10_EB_3D_3A_SC_Batch2

tag: batch


title: 209_H9_EB_3D_WT_SC_Batch2

value: WT


title: Aorta_Tie2CKO

tag: tissue

value: Aorta


title: 210_A11_EB_3D_3A_SC_Batch2


title: VE-PTPiKO-sham rep1 [S4715-sh-ko]

value: sham


title: VE-PTPiKO-sham rep3 [S4728-sh-ko]


title: GS-6/7a_N1_lymph_node_met (CGH002_LK)

description:
macrodissected FFPE sample
CGH002_LK

tag: disease state

tag: gleason group


title: AP_Tie2Ctrl

value: Aorta (aortic arch)


title: GS-9/10_N1_primary_tumor (CGH044_PCA)

description:
macrodissected FFPE sample
CGH044_PCA

tag: lymph node status

tag: tumor type


title: Aorta_Tie2Ctrl


title: 209_H8_EB_3D_WT_SC_Batch1

value: Embryoid Bodies (EB)


title: GS-9/10_N1_lymph_node_met (CGH085_LK)

description:
macrodissected FFPE sample
CGH085_LK


title: GS-9/10_N1_lymph_node_met (CGH044_LK)

description:
macrodissected FFPE sample
CGH044_LK


title: 209_H7_EB_3D_WT_SC_Batch1


title: Control-IR rep3 [S4719-IR-CON]

value: Control


title: GS-6/7a_N1_lymph_node_met (CGH030_LK)

description:
macrodissected FFPE sample
CGH030_LK


title: VE-PTPiKO-sham rep2 [S4724-sh-ko]

value: Kidney


title: GS-6/7a_N1_primary_tumor (CGH030_PCA)

description:
macrodissected FFPE sample
CGH030_PCA


title: CD4+Tcells-12hactivation-healthycontrol4 [C4_T12]

type: RNA

city: Homburg

first: Caroline

last: Diener

address: Kirrberger Str., Building 60

department: Institute of Human Genetics

institute: Saarland University

hyb_protocol: labeled samples were hybridized to Human SurePrint G3 Unrestricted miRNA 8 × 60K arrays (Release 21.0, G4872A, Agilent Technologies, Santa Clara, CA, USA) following the instructions from miRNA Complete Labeling and Hyb Kit (Agilent Technologies, Santa Clara, CA, USA).

scan_protocol: Array slides were scanned with a resolution of 3 μm.

data_processing: Raw data was extracted using Feature Extraction software (Agilent Technologies, Santa Clara, CA, USA). Raw expression values were processed using the limma R-package. A background correction (method = normexp, offset = 16) was conducted.

source_name: CD4+ T cells, 12 h activation, healthy control 4

label_protocol: Cyanine-3 (Cy3) labeling was performed using 75 ng total RNA and following the instructions from miRNA Complete Labeling and Hyb Kit (Agilent Technologies, Santa Clara, CA, USA).

extract_protocol: RNA was prepared using the miRNeasy Micro Kit (Qiagen, Hilden, Germany) following the manufacturer's recommendations.

treatment_protocol: CD4+ T cells were isolated from peripheral blood samples. The isolated cells were incubated in 25 mm flasks overnight (RPMI 1640 medium with 10 % v/v heat inactivated fetal bovine serum and 1 % v/v penicillin-streptomycin (100 U/ml)) . At the following day, the cells were seeded in a 96 well format (350,000 cells/well) and were in vitro activated by αCD2/αCD3/αCD28 beads. Cellular samples were collected from different wells at 0, 2, 4, 8, 12 and 24 h after activation for subsequent RNA extraction.

value: Healthy Control

value: CD4+ T cells

tag: gender

value: male

value: 0 (Healthy Control)

tag: time-point post activation


title: 209_H5_EB_3D_WT_SC_Batch1


title: Control-sham rep3 [S4729-sh-con]


title: 209_H6_EB_3D_WT_SC_Batch1


title: Control-IR rep2 [S4717-IR-CON]

value: ischemia-reperfusion (IR) injury


title: Control-IR rep1 [S1896-IR-CON]


title: CD4+Tcells-24hactivation-healthycontrol4 [C4_T24]

source_name: CD4+ T cells, 24 h activation, healthy control 4


title: CD4+Tcells-8hactivation-healthycontrol4 [C4_T8]

source_name: CD4+ T cells, 8 h activation, healthy control 4

value: 8 h


title: CD4+Tcells-4hactivation-healthycontrol4 [C4_T4]

source_name: CD4+ T cells, 4 h activation, healthy control 4

value: 4 h


title: GS-6/7a_N1_primary_tumor (CGH001_PCA)

description:
macrodissected FFPE sample
CGH001_PCA

value: prostate cancer


title: 209_H4_EB_3D_WT_SC_Batch1


title: Control-sham rep2 [S4727-sh-con]


title: 209_H2_EB_3D_WT_SC_Batch1


title: GS-6/7a_N1_lymph_node_met (CGH001_LK)

description:
macrodissected FFPE sample
CGH001_LK


title: 209_H3_EB_3D_WT_SC_Batch1


title: 209_H1_EB_3D_WT_SC_Batch1


title: Control-sham rep1 [S4725-sh-con]


title: IR-Veh

data_processing:
Cell Ranger (v4.0.0) was used for demultiplexing and counting
data were processed with the Seurat R package and SoupX package
Genome_build: refdata-gex-mm10-2020
Supplementary_files_format_and_content: matrix.mtx, features.tsv, barcodes.tsv

source_name: Kidney

extract_protocol:
Kidneys were minced with a sterile razor blade. Approximately 65 mg tissue was digested in liberase TM (0.3 mg/mL, Sigma, TM5401119001), hyaluronidase (10 μg/mL, Sigma, H4272), DNaseI (20 μg/mL, Roche, 0471672800) at 37°C for 40 min, followed by incubation with 0.25% trypsin EDTA at 37°C for 30 min. Trypsin was inactivated using 10% fetal bovine serum in PBS. Cells were then resuspended in PBS supplemented with 0.04% bovine serum albumin. After filtration through a 40 μm strainer, cell number and viability were analyzed using Nexcelom Cellometer Auto2000 with AOPI fluorescent staining method.
The cDNA and library were generated using the Chromium Next GEM Single Cell 3′ Reagent Kits v3.1 (10X Genomics, PN-1000286) and Single Index Kit T Set A (10X Genomics, PN-1000213) according to the manufacturer’s manual.

value: vehicle


title: CD4+Tcells-2hactivation-healthycontrol4 [C4_T2]

source_name: CD4+ T cells, 2 h activation, healthy control 4

value: 2 h


title: H4

first: Aarthi

last: Ramakrishnan

address: 1425 Madison Avenue

department: Neuroscience

institute: Icahn School of Medicine at Mount Sinai

data_processing:
Raw sequencing reads from mice were mapped to mm10 using HISAT2.
Counts of reads mapping to genes were obtained using featureCounts software of Subread package against Ensembl v90 annotation.
Differential expression was done using the DESeq2 package.
Assembly: mm10
Supplementary files format and content: tab-delimited text file containing counts

source_name: Brain

extract_protocol:
Bilateral medial prefrontal cortex (mPFC; 12-gauge), nucleus accumbens (NAc; 14-gauge), and ventral tegmental area (VTA; 16-gauge) punches were taken from each animal. For each region, punches from two animals were pooled per biological replicate. Total RNA was isolated with TRIzol and integrity was confirmed with an Agilent BioAnalyzer.
mRNA-Seq libraries were prepared using a TruSeq RNA sample preparation kit (Illumina RS-122-2001/2)

value: Brain

tag: Sex

value: Nac

value: SNI

tag: condition

value: oxy


title: 209_G9_EB_3D_WT_SC_Batch2

value: Embryoid Bodies (EB)

value: WT


title: C4

value: Sham


title: H3


title: IR-Hepta-ANG1

tag: treatment

tag: age


title: 209_G6_EB_3D_WT_SC_Batch1

tag: strain


title: CD4+Tcells-12hactivation-healthycontrol3 [C3_T12]

source_name: CD4+ T cells, 12 h activation, healthy control 3

value: Healthy Control

tag: cell type


title: CD4+Tcells-0hactivation-healthycontrol4 [C4_T0]

source_name: CD4+ T cells, 0 h activation, healthy control 4

value: CD4+ T cells

value: 0 h


title: CD4+Tcells-2hactivation-healthycontrol3 [C3_T2]

source_name: CD4+ T cells, 2 h activation, healthy control 3

value: male

value: 0 (Healthy Control)


title: CD4+Tcells-24hactivation-healthycontrol3 [C3_T24]

source_name: CD4+ T cells, 24 h activation, healthy control 3


title: CD4+Tcells-8hactivation-healthycontrol3 [C3_T8]

source_name: CD4+ T cells, 8 h activation, healthy control 3

value: 8 h


title: CD4+Tcells-0hactivation-healthycontrol3 [C3_T0]

source_name: CD4+ T cells, 0 h activation, healthy control 3


title: B4

tag: genotype

value: Sal


title: F4


title: CD4+Tcells-4hactivation-healthycontrol3 [C3_T4]

source_name: CD4+ T cells, 4 h activation, healthy control 3

value: 4 h


title: F3


title: 209_G7_EB_3D_WT_SC_Batch1

source_name: Mouse Embryoid Bodies


title: 209_G8_EB_3D_WT_SC_Batch1


title: C3


title: CD4+Tcells-24hactivation-healthycontrol2 [C2_T24]

source_name: CD4+ T cells, 24 h activation, healthy control 2


hyb_protocol: labeled samples were hybridized to Human SurePrint G3 Gene Expression Microarrays (V3, G4851C; Agilent Technologies, Santa Clara, CA, USA) following the inctructions from Gene Expression Hybridization Kit (Agilent Technologies)

data_processing: Raw data was extracted using Feature Extraction software (Agilent Technologies, Santa Clara, CA, USA). Raw expression values were processed using the limma R-package. A background correction (method = normexp, offset = 16) was conducted

label_protocol: Cyanine-3 (Cy3) labeling of cRNA was performed using 25 ng total RNA and following the instructions from Low Input Quick Amp Labeling Kit (One-Color) (Agilent Technologies, Santa Clara, CA, USA). The cRNA samples were purified by RNeasy Mini Kit (Qiagen, Hilden, Germany) and concentrations of cRNAS were determined using a NanoDrop™ 2000c Spectrophotometer (Thermo Fisher Scientific Inc., Waltham, MA, USA).


title: CD4+Tcells-12hactivation-healthycontrol2 [C2_T12]

source_name: CD4+ T cells, 12 h activation, healthy control 2


title: B3


source_name: CD4+ T cells, 12 h activation, healthy control 4


title: 209_G3_EB_3D_WT_SC_Batch1


title: 209_G5_EB_3D_WT_SC_Batch1


title: F2


title: 209_G4_EB_3D_WT_SC_Batch1


source_name: CD4+ T cells, 24 h activation, healthy control 4


title: 209_G2_EB_3D_WT_SC_Batch1


title: B2


title: 209_G1_EB_3D_WT_SC_Batch1


title: CD4+Tcells-8hactivation-healthycontrol2 [C2_T8]

source_name: CD4+ T cells, 8 h activation, healthy control 2


title: H2


title: 209_G11_EB_3D_WT_SC_Batch2


title: C2


title: F1


title: 209_G12_EB_3D_WT_SC_Batch2


title: 209_G10_EB_3D_WT_SC_Batch2


title: 209_F9_EB_3D_WT_SC_Batch2


title: CD4+Tcells-0hactivation-healthycontrol2 [C2_T0]

source_name: CD4+ T cells, 0 h activation, healthy control 2


title: CD4+Tcells-24hactivation-healthycontrol1replicate2 [C1_2_T24]

source_name: CD4+ T cells, 24 h activation, healthy control 1 replicate 2


title: H1


title: CD4+Tcells-12hactivation-healthycontrol1replicate2 [C1_2_T12]

source_name: CD4+ T cells, 12 h activation, healthy control 1 replicate 2


title: CD4+Tcells-2hactivation-healthycontrol2 [C2_T2]

source_name: CD4+ T cells, 2 h activation, healthy control 2


title: CD4+Tcells-4hactivation-healthycontrol2 [C2_T4]

source_name: CD4+ T cells, 4 h activation, healthy control 2


title: CD4+Tcells-8hactivation-healthycontrol1replicate2 [C1_2_T8]

source_name: CD4+ T cells, 8 h activation, healthy control 1 replicate 2


title: 209_F8_EB_3D_WT_SC_Batch1


title: 209_F7_EB_3D_WT_SC_Batch1


title: C1

source_name: Brain


source_name: CD4+ T cells, 2 h activation, healthy control 3


title: 209_F6_EB_3D_WT_SC_Batch1


title: 209_F5_EB_3D_WT_SC_Batch1


title: 209_F3_EB_3D_WT_SC_Batch1


title: B1


title: HeLa cells, isolated midbody RNA sample, Rep2

city: Aurora

first: Matthew

last: Taliaferro

state: CO

department: Biochemistry and Molecular Genetics

institute: University of Colorado Anschutz Medical Campus

data_processing:
Transcript quantification with salmon
Gene-level quantification with tximport
Differential enrichments quantified with DESeq2
Assembly: hg38
Supplementary files format and content: salmon-derived transcript quantifications

source_name: HeLa

growth_protocol: DMEM + 10% FBS

extract_protocol:
Midbodies isolated through centrifugation and sucrose gradients. RNA extracted with Trizol.
Truseq polyA library preparation
polyA enrichment

tag: cell line

value: HeLa

value: midbodies


title: CD4+Tcells-2hactivation-healthycontrol1replicate2 [C1_2_T2]

source_name: CD4+ T cells, 2 h activation, healthy control 1 replicate 2

tag: Stage

value: 2 h


title: HeLa cells, whole cell sample, Rep1

value: whole cell


title: HeLa cells, whole cell sample, Rep3


title: HeLa cells, whole cell sample, Rep2


title: CD4+Tcells-4hactivation-healthycontrol1replicate2 [C1_2_T4]

source_name: CD4+ T cells, 4 h activation, healthy control 1 replicate 2


title: CD4+Tcells-0hactivation-healthycontrol1replicate2 [C1_2_T0]

source_name: CD4+ T cells, 0 h activation, healthy control 1 replicate 2


title: CD4+Tcells-4hactivation-PDpatient3 [P3_T4]

source_name: CD4+ T cells, 4 h activation, PD patient 3

value: Parkinson's disease


title: HeLa cells, isolated midbody RNA sample, Rep1


source_name: CD4+ T cells, 12 h activation, healthy control 2


title: HeLa cells, isolated midbody RNA sample, Rep3


source_name: CD4+ T cells, 24 h activation, healthy control 2


title: CD4+Tcells-24hactivation-PDpatient3 [P3_T24]

source_name: CD4+ T cells, 24 h activation, PD patient 3


title: CD4+Tcells-8hactivation-PDpatient3 [P3_T8]

source_name: CD4+ T cells, 8 h activation, PD patient 3


title: 209_F4_EB_3D_WT_SC_Batch1

source_name: Mouse Embryoid Bodies

tag: batch


title: CD4+Tcells-2hactivation-PDpatient3 [P3_T2]

source_name: CD4+ T cells, 2 h activation, PD patient 3


source_name: CD4+ T cells, 0 h activation, healthy control 2


source_name: CD4+ T cells, 2 h activation, healthy control 2


source_name: CD4+ T cells, 4 h activation, healthy control 2


title: CD4+Tcells-8hactivation-healthycontrol2 [C2_T8]

source_name: CD4+ T cells, 8 h activation, healthy control 2


source_name: CD4+ T cells, 12 h activation, healthy control 1 replicate 2


title: 209_F1_EB_3D_WT_SC_Batch1


title: CD4+Tcells-0hactivation-PDpatient3 [P3_T0]

source_name: CD4+ T cells, 0 h activation, PD patient 3


title: Vacc_54

city: Guangzhou

first: Jinbao

last: Gu

address: No.1023, South Shatai Road

country: China

institute: Southern Medical University

data_processing:
Low-quality reads and reads without primers were discarded.
PCR and sequencing errors were corrected by unique molecular identifiers (UMIs)
TCRβ V, D, J gene and clonotype defined according to IMGT and Igblast
Assembly: IMGT/GENE-DB
Supplementary files format and content: tab-delimited text files include clone values for each Sample

source_name: whole blood

extract_protocol:
RNA was harvested using RNAiso Plus (Takra). 1 ug of total RNA was used for the construction of sequencing libraries.
RNA libraries for RNA-seq were prepared using SMARTER mRNA-Seq Library Prep Kit following manufacturer's protocols.

tag: tissue

value: whole blood

value: leukocyte


title: Vacc_53


title: 209_F12_EB_3D_WT_SC_Batch2

value: Embryoid Bodies (EB)

value: WT


title: 209_F2_EB_3D_WT_SC_Batch1


title: Vacc_52


title: Vacc_51


title: 209_F10_EB_3D_WT_SC_Batch2


title: 209_E8_EB_3D_WT_SC_Batch1


title: 209_E9_EB_3D_WT_SC_Batch2


title: 209_E7_EB_3D_WT_SC_Batch1


source_name: CD4+ T cells, 8 h activation, healthy control 1 replicate 2

tag: disease state

value: Healthy Control


title: CD4+Tcells-12hactivation-PDpatient3 [P3_T12]

source_name: CD4+ T cells, 12 h activation, PD patient 3

value: CD4+ T cells


title: 209_F11_EB_3D_WT_SC_Batch2


title: CD4+Tcells-24hactivation-PDpatient2 [P2_T24]

source_name: CD4+ T cells, 24 h activation, PD patient 2


title: CD4+Tcells-2hactivation-PDpatient2 [P2_T2]

source_name: CD4+ T cells, 2 h activation, PD patient 2


title: CD4+Tcells-12hactivation-PDpatient2 [P2_T12]

source_name: CD4+ T cells, 12 h activation, PD patient 2


title: Vacc_47


title: Vacc_49


source_name: CD4+ T cells, 4 h activation, healthy control 1 replicate 2


title: Vacc_50


title: 209_E6_EB_3D_WT_SC_Batch1


title: Vacc_48


title: CD4+Tcells-0hactivation-PDpatient2 [P2_T0]

source_name: CD4+ T cells, 0 h activation, PD patient 2

value: 0 h


title: Vacc_46


title: Vacc_45


title: CD4+Tcells-8hactivation-PDpatient2 [P2_T8]

source_name: CD4+ T cells, 8 h activation, PD patient 2

value: male


title: CD4+Tcells-2hactivation-healthycontrol1replicate2 [C1_2_T2]

source_name: CD4+ T cells, 2 h activation, healthy control 1 replicate 2

value: 0 (Healthy Control)


source_name: CD4+ T cells, 0 h activation, healthy control 1 replicate 2


source_name: CD4+ T cells, 24 h activation, PD patient 3


title: CD4+Tcells-24hactivation-PDpatient1 [P1_T24]

source_name: CD4+ T cells, 24 h activation, PD patient 1


title: 209_E4_EB_3D_WT_SC_Batch1

source_name: Mouse Embryoid Bodies


title: CD4+Tcells-4hactivation-PDpatient2 [P2_T4]

source_name: CD4+ T cells, 4 h activation, PD patient 2


title: CD4+Tcells-12hactivation-PDpatient1 [P1_T12]

source_name: CD4+ T cells, 12 h activation, PD patient 1


source_name: CD4+ T cells, 8 h activation, PD patient 3

value: 8 h


title: CD4+Tcells-8hactivation-PDpatient1 [P1_T8]

source_name: CD4+ T cells, 8 h activation, PD patient 1


title: Vacc_44

source_name: whole blood


title: 209_E3_EB_3D_WT_SC_Batch1


title: CD4+Tcells-4hactivation-PDpatient1 [P1_T4]

source_name: CD4+ T cells, 4 h activation, PD patient 1


title: Vacc_43


title: 209_E5_EB_3D_WT_SC_Batch1


title: 209_E2_EB_3D_WT_SC_Batch1


title: CD4+Tcells-4hactivation-PDpatient3 [P3_T4]

source_name: CD4+ T cells, 4 h activation, PD patient 3


title: 209_E1_EB_3D_WT_SC_Batch1


title: Vacc_41


title: 209_E12_EB_3D_WT_SC_Batch2


title: 209_E11_EB_3D_WT_SC_Batch2


title: Vacc_39


title: CD4+Tcells-24hactivation-healthycontrol1 [C1_T24]

source_name: CD4+ T cells, 24 h activation, healthy control 1


title: CD4+Tcells-2hactivation-PDpatient3 [P3_T2]

source_name: CD4+ T cells, 2 h activation, PD patient 3


source_name: CD4+ T cells, 12 h activation, PD patient 2


source_name: CD4+ T cells, 0 h activation, PD patient 3


title: CD4+Tcells-2hactivation-PDpatient1 [P1_T2]

source_name: CD4+ T cells, 2 h activation, PD patient 1


title: 209_E10_EB_3D_WT_SC_Batch2


title: Vacc_42


title: Vacc_38


title: 209_D9_EB_3D_WT_SC_Batch2


title: CD4+Tcells-0hactivation-PDpatient1 [P1_T0]

source_name: CD4+ T cells, 0 h activation, PD patient 1


title: CD4+Tcells-12hactivation-healthycontrol1 [C1_T12]

source_name: CD4+ T cells, 12 h activation, healthy control 1


title: 209_D7_EB_3D_WT_SC_Batch1


title: CD4+Tcells-24hactivation-PDpatient2 [P2_T24]

source_name: CD4+ T cells, 24 h activation, PD patient 2


title: CD4+Tcells-2hactivation-healthycontrol1 [C1_T2]

source_name: CD4+ T cells, 2 h activation, healthy control 1


source_name: CD4+ T cells, 8 h activation, PD patient 2


title: Vacc_40


source_name: CD4+ T cells, 0 h activation, PD patient 2


title: 209_D6_EB_3D_WT_SC_Batch1

source_name: Mouse Embryoid Bodies


title: Vacc_34


title: CD4+Tcells-0hactivation-healthycontrol1 [C1_T0]

source_name: CD4+ T cells, 0 h activation, healthy control 1


title: Vacc_37


title: CD4+Tcells-2hactivation-PDpatient2 [P2_T2]

source_name: CD4+ T cells, 2 h activation, PD patient 2


title: Vacc_33

source_name: whole blood


source_name: CD4+ T cells, 4 h activation, PD patient 2


title: 209_D8_EB_3D_WT_SC_Batch1


title: 209_D4_EB_3D_WT_SC_Batch1


title: 209_D3_EB_3D_WT_SC_Batch1


title: CD4+Tcells-4hactivation-healthycontrol1 [C1_T4]

source_name: CD4+ T cells, 4 h activation, healthy control 1


title: 209_D5_EB_3D_WT_SC_Batch1


title: Vacc_35


title: Vacc_31


source_name: CD4+ T cells, 4 h activation, PD patient 1


title: CD4+Tcells-24hactivation-PDpatient5 [P5_T24]

source_name: CD4+ T cells, 24 h activation, PD patient 5


title: CD4+Tcells-8hactivation-PDpatient5 [P5_T8]

source_name: CD4+ T cells, 8 h activation, PD patient 5


title: Vacc_36


title: CD4+Tcells-4hactivation-PDpatient5 [P5_T4]

source_name: CD4+ T cells, 4 h activation, PD patient 5


title: CD4+Tcells-8hactivation-healthycontrol1 [C1_T8]

source_name: CD4+ T cells, 8 h activation, healthy control 1


title: Vacc_30


title: CD4+Tcells-24hactivation-PDpatient1 [P1_T24]

source_name: CD4+ T cells, 24 h activation, PD patient 1


title: CD4+Tcells-8hactivation-PDpatient1 [P1_T8]

source_name: CD4+ T cells, 8 h activation, PD patient 1


title: CD4+Tcells-2hactivation-PDpatient5 [P5_T2]

source_name: CD4+ T cells, 2 h activation, PD patient 5


title: CD4+Tcells-12hactivation-PDpatient5 [P5_T12]

source_name: CD4+ T cells, 12 h activation, PD patient 5


source_name: CD4+ T cells, 2 h activation, PD patient 1


title: Vacc_27


title: Vacc_32


title: CD4+Tcells-24hactivation-healthycontrol1 [C1_T24]

source_name: CD4+ T cells, 24 h activation, healthy control 1


title: CD4+Tcells-12hactivation-PDpatient1 [P1_T12]

source_name: CD4+ T cells, 12 h activation, PD patient 1


title: 209_C9_EB_3D_WT_SC_Batch2

source_name: Mouse Embryoid Bodies


title: 209_D1_EB_3D_WT_SC_Batch1


title: CD4+Tcells-8hactivation-PDpatient4 [P4_T8]

source_name: CD4+ T cells, 8 h activation, PD patient 4


title: Vacc_29

source_name: whole blood


title: Vacc_25


title: 209_D2_EB_3D_WT_SC_Batch1


title: CD4+Tcells-4hactivation-PDpatient4 [P4_T4]

source_name: CD4+ T cells, 4 h activation, PD patient 4


title: Vacc_24


source_name: CD4+ T cells, 2 h activation, healthy control 1


title: CD4+Tcells-0hactivation-PDpatient1 [P1_T0]

source_name: CD4+ T cells, 0 h activation, PD patient 1


title: 209_D12_EB_3D_WT_SC_Batch2


source_name: CD4+ T cells, 0 h activation, healthy control 1


title: CD4+Tcells-0hactivation-PDpatient4 [P4_T0]

source_name: CD4+ T cells, 0 h activation, PD patient 4


title: Vacc_23


title: Vacc_28


title: 209_D10_EB_3D_WT_SC_Batch2


title: 209_D11_EB_3D_WT_SC_Batch2


title: CD4+Tcells-12hactivation-healthycontrol1 [C1_T12]

source_name: CD4+ T cells, 12 h activation, healthy control 1


title: CD4+Tcells-12hactivation-PDpatient4 [P4_T12]

source_name: CD4+ T cells, 12 h activation, PD patient 4


title: CD4+Tcells-4hactivation-healthycontrol1 [C1_T4]

source_name: CD4+ T cells, 4 h activation, healthy control 1


title: Vacc_22


title: CD4+Tcells-24hactivation-PDpatient4 [P4_T24]

source_name: CD4+ T cells, 24 h activation, PD patient 4


title: Vacc_26


title: Vacc_21


title: belt_deletion

city: Hong Kong

first: Duc

middle: Trung

last: Nguyen

address: Clear Water Bay

department: Life Science

country: Hong Kong

institute: The Hong Kong University of Science and Technology

data_processing:
First, cutdapt were used to remove adapters from both ends of raw reads (-a TGGAATTCTCGGGTGCCAAGG -A GATCGTCGGACTGTAGAACTCTGAAC). Next, fastq-join were applied to join paired-end reads (-p 5). The low quality joined reads and duplicated reads were removed using fastq_quality_filter (-q 20 -p 90) and fastx_collapser, respectively. Then, the randomized barcodes in both ends of remaining reads were trimmed by cutadapt (-u 4 -u -6: 4 nt in 5p-end and 6 nt in 3p-end for reads in control samples, -u 4 -u -4: 4 nt in both ends for reads in product samples). Next, bowtie2 were used to map the trimmed reads the build-in reference which contains the sequences of 137 C. elegans pri-miRNAs (--local --norc -p 4 -k 5). The pri-miRNA sequences contain 30- and 25-nt extending from 5p- and 3p-ends of pre-miRNA sequences, respectively. Additionally, GGG also were appended at the 5p-end of pri-miRNA reference sequences to represent the truly pri-miRNAs generated from in vitro transcription. The unique mapped reads were collected. The 132 out of 137 pri-miRNAs with more than 20 read counts in control sample were used for further analysis. For each sample, the raw counts were normalized as counts per million.
Assembly: Pri-miRNA construct sequences
Supplementary files format and content: reference_sequences.txt: Tab-delimited text file contains the reference sequences of 137 C. elegans pri-miRNAs
Supplementary files format and content: control_rawcount.txt: Tab-delimited text file contains the raw count of 137 C. elegans pri-miRNAs in the control sample
Supplementary files format and content: product_rawcount.txt: Tab-delimited text file contains the raw count of cleaved products for each pri-miRNAs in product samples

organism: Caenorhabditis elegans

source_name: In vitro cleavage

extract_protocol:
Illumina adapter sequences were incorporated to the libraries using PCR.
We collected 145 C. elegans pri-miRNAs from MirGeneDB v2.0. We omitted 7 duplicated pri-miRNAs sharing the same miRBase ID. Cel-pri-mir-229 were excluded since its length is longer than 150 nt.   The C. elegans pri-miRNAs were pooled into a substrate mixture.  To clone the cleaved products, the gel pieces containing cleaved products sizing from 50 - 70 nt were sliced out, gel purified, precipitated by IPA, 80% ethanol washed and air dried. The RNAs were dissolved in 10 μL distilled water and ligated with 10 pmol of 3ꞌ-adapter (4N-RA3, /5rApp/NN NNT GGA ATT CTC GGG TGC CAA GG/3ddC/). The ligation was performed in a total 20 μL reaction containing 1 μL  T4 RNA Ligase 2, truncated KQ (NEB, M0373L), 2 μL 10x T4 RNA ligase buffer, 6 μL of 50% PEG8000, and 0.5 μL of SUPERase•In RNase inhibitor (20 U/μL). The mixture was incubated at 25°C for 16 h. After that, it was treated with 20 μL 2x TBE-Urea sample buffer, denatured at 75 °C for 5 min, and loaded onto a pre-run 12% Urea-PAGE. The 3ꞌ-adapter ligated RNAs were sliced and gel purified. The ligated RNAs were dissolved in distilled water and subjected to the 5ꞌ-adapter ligation reaction. For this second ligation reaction, 10 pmol of 5ꞌ-adapter (RA5-4N, GUU CAG AGU UCU ACA GUC CGA CGA UCN NNN), 1 μL T4 RNA ligase I (NEB, M0204L), 2 μL 10x T4 RNA ligase buffer, 6 μL of 50% PEG8000, and 0.5 μL of SUPERase•In RNase inhibitor (20 U/μL) were used. The reaction was kept at 25°C for 16 h and directly reverse transcribed with R-RA3 primer (TTG GCA CCC GAG AAT TCC A) using Superscript IV reverse transcriptase (Invitrogen, 18090050). The resulting cDNA was PCR amplified using a pair of sequencing primers RP1 (AAT GAT ACG GCG ACC ACC GAG ATC TAC ACG TTC AGA GTT CTA CAG TCC GA), and each of the specific primers, RPIx.   To clone the uncleaved substrate, 10 pmol of the substrate mixture was ligated with 30 pmol of  3ꞌ-adapter as described above. The ligated RNA was reverse transcribed with the cirRTP primer (/5Phos/NNN NNN GA TCG TCG GAC TGT AGA ACT CTG AAC /iSp18/CC TTG GCA CCC GAGAAT TCC A) using Superscript IV reverse transcriptase (Invitrogen, 18090050). The cDNA was treated with 0.1mM NaOH at 95°C for 5 mingel to eliminate the RNAs and gel eluted. The purified cDNA was circularized using the CircLigase™ ss- DNA Ligase (Epicentre, CL4115K) and loaded onto a pre-run 18% Urea-PAGE. The circular DNA were gel purified and PCR amplified using sequencing primers RP1 (AAT GAT ACG GCG ACC ACC GAG ATC TAC ACG TTC AGA GTT CTA CAG TCC GA), and a specific primer, RPIx. The resulting DNA libraries were sequenced with  Illumina NovaSeq 6000 system in 150 bp paired-end mode.

tag: protocol

value: In vitro cleavage

tag: type sample

value: belt_deletion product


title: CD4+Tcells-8hactivation-PDpatient5 [P5_T8]

source_name: CD4+ T cells, 8 h activation, PD patient 5


title: CD4+Tcells-2hactivation-PDpatient4 [P4_T2]

source_name: CD4+ T cells, 2 h activation, PD patient 4


title: 209_C3_EB_3D_WT_SC_Batch1


title: 209_C8_EB_3D_WT_SC_Batch1


title: dsRBD_deletion

value: dsRBD_deletion product


title: beta_deletion

value: beta_deletion product


title: CD4+Tcells-0hactivation-PDpatient5 [P5_T0]

source_name: CD4+ T cells, 0 h activation, PD patient 5


title: 209_C7_EB_3D_WT_SC_Batch1


title: 209_C4_EB_3D_WT_SC_Batch1


title: 209_C2_EB_3D_WT_SC_Batch1


title: CD4+Tcells-24hactivation-PDpatient5 [P5_T24]

source_name: CD4+ T cells, 24 h activation, PD patient 5


title: 209_C6_EB_3D_WT_SC_Batch1

source_name: Mouse Embryoid Bodies


source_name: CD4+ T cells, 12 h activation, PD patient 5


title: Vacc_18

source_name: whole blood


title: Vacc_19


source_name: CD4+ T cells, 2 h activation, PD patient 5

value: 2 h


title: 209_C12_EB_3D_WT_SC_Batch2


title: 209_C5_EB_3D_WT_SC_Batch1


title: Vacc_17


title: Vacc_20


title: cMP

value: cMP product


title: Vacc_16


source_name: CD4+ T cells, 8 h activation, PD patient 4

value: Parkinson's disease


title: cDrosha

value: cDrosha product


title: control

source_name: In vitro transcription

value: In vitro transcription

value: control


title: 288 PRE

city: Bari

first: Simona

last: De Summa

state: (Click to Select US State)

address: Via Orazio Flacco, 65

institute: IRCCS-Istituto Tumori "Giovanni Paolo II"

hyb_protocol: Firstly, poly (A) tailing was carried out at 37°C for 15 min in a volume of 15 ml reaction mix that  contained 1 ml Reaction Buffer, 1.5 ml MgCl2 (25 mM), 1 ml ATP Mix diluted 1:500 and 1 ml PAP enzyme. Subsequently, Flash Tag Ligation was performed at room temperature for 30 min by adding 4 ml of 5 Flash Tag Ligation Mix Biotin and 2 ml T4 DNA Ligase into 15 ml of reaction mix. Next, 2.5 ml of Stop Solution was added to stop the reaction. Each sample was hybridized on the array, washed, stained with the Affymetrix Fluidics Station 450

scan_protocol: Affymetrix Gene Chip Scanner 3000 7G using the Command Console software (Affymetrix).

data_processing: Raw data were normalized with the Robust Multiarray Average (RMA) method to remove systematic variations.

label: biotin

source_name: plasma

label_protocol: For each sample, 500 ng of total RNA was labelled using the 3 DNA Array Detection Flash Tag RNA Labelling Kit (http://www.genisphere.com), according to the manufacturer’s instructions.

extract_protocol: Total RNA was isolated from plasma using mirVana™ miRNA Isolation Kit (Ambion, Austin, TX, USA), according to the manufacturer’s recommendations.

treatment_protocol: Eight milliliters of peripheral blood were collected into Cell-Free DNA BCT (Streck Corporate, La Vista, NE) at each time-point and was centrifuged at 3000 rpm at 4 °C for 15. The obtained plasma samples were further centrifuged at 16 000 g at 4 °C for 10 min to remove cell debris. The separated plasma was stored at 80 °C until nucleic acid extraction.

value: plasma

tag: diet yes/no

value: yes


title: Vacc_14

value: whole blood

value: leukocyte


title: Tag16: articular cartilage and fibrous tissue

city: Suita

first: Noriyuki

last: Tsumaki

state: Select One

address: 2-2 Yamadaoka

department: Department of Biochemistry and Molecular Biology

country: Japan

institute: Osaka University, Graduate School of Medicine

library_source: transcriptomic single cell

data_processing:
Pair-end Fastq files (R1: cell barcode reads, R2: RNA reads) were processed as follows. Adapter trimming of sequencing data was performed using cutadapt 3.4. Filtered reads were chunked into 16 parts for parallel processing by using Seqkit 0.14.0. Filtered cell barcode reads were annotated by Python script provided by BD Biosciences with minor modification for compatibility to Python 3.7. Reference RNA sequences were built by concatenating cDNA and ncRNA fasta files of the Ensembl database (build Macaca_fascicularis_6.0 for macaca fascicularis data, and GRCh38 release-101 for human data). Associated cDNA reads were mapped to reference RNA using bowtie2-2.4.2 by the following parameters: -p 2 --very-sensitive-local -N 1 -norc -seed 656565 -reorder. Then, cell barcode information of each read was added to the bowtie2-mapped BAM files by the python script and pysam 0.15.4, and read counts of each gene in each cell barcode were counted using mawk. Resultant count data was converted to a single-cell gene-expression matrix file by using R-3.6.3. The inflection point of the knee-plot (total read count versus the rank of the read count) was detected using DropletUtils package in R-3.6.3. Cells of which total read count was over inflection point were considered as valid cells.
Assembly: Macaca_fascicularis_6.0 for macaca fascicularis data
Supplementary files format and content: tab-delimited text files include raw tag-count values for each sample
Supplementary files format and content: each cell was named as follows: [hashtag ID]_[experiment ID]_[cell barcode]
Supplementary files format and content: Fig5.matrix.txt.gz (cyiPSC, pre-transplant cyiPS-Cart)
Supplementary files format and content: Fig6.matrix.txt.gz (pre-transplant cyiPS-Cart, post-transplant cyiPS-Cart, cyAC, cyFT)

organism: Macaca fascicularis

source_name: articuar cartilage and fibrous tissue formed in chondral defect

extract_protocol:
The cyiPS-Cart (pre-transplant cyiPS-Cart), intact articular cartilage (cyAC), fibrous tissue formed in chondral defects in the empty group (cyFT), and cyiPS-Cart in chondral defects in the transplantation group (post-transplant cyiPS-Cart) were minced into 1-2 mm pieces. Next, we digested these pieces with Liberase solution (for NP, AC, and post-implant hiPS-Cart: RPMI-1640 (Nacalai Tesque) supplemented with 0.2% FBS, 10 mM HEPES pH 7.2-7.4, 0.2-0.4 mg/mL Liberase TM (Roche), and 2 kU/mL DNase I (Merck); for pre-implant hiPS-Cart: DMEM with 1% FBS, 1% ITS-X, 50 µg/mL ascorbic acid, 1 mM sodium pyruvate, 1% nonessential amino acids, 1% penicillin-streptomycin, 10 ng/mL TGF-β1, 10 ng/mL GDF5, 10 ng/mL BMP2, 0.2 mg/mL Liberase TM (Roche), and 2 kU/mL DNase I (Merck)) at 37 °C, 5% CO2 for 120-210 minutes with continuous shaking. The cells were mixed 10 times using a 1,000 μL blue tip-fitted pipette and then passed through a cell strainer (70 μm pore size; BD Biosciences), centrifuged at 4 °C for 300 × g for 5 min, and the supernatant was discarded. The cells were resuspended in RPMI-1640 medium supplemented with 0.2% FBS and 10 mM HEPES.
For cell hashing, we biotinylated cell surface proteins using EZ-Link Sulfo-NHS-Biotin (Thermo Scientific), and then stained them with 0.6 μg/mL Totalseq (A0951-A0955, and A0436 (BioLegend)) (Details of the method will be described elsewhere.) We selected live cells stained with A0951-A0955 using a FACS Aria II flow cytometer (BD Biosciences) and suspended in a sample buffer (BD Biosciences).
TAS-Seq libraries were generated by Immunogeneteqs Inc. (Noda city, Chiba, Japan) according to a previous report: S. Shichino, S. Ueha, S. Hashimoto, T. Ogawa, H. Aoki, W. Bin, . . . K. Matsushima, TAS-Seq: a robust and sensitive amplification method for beads-based scRNA-seq, bioRxiv  (2021).

treatment_protocol: cyiPS-Cart was created from cyiPS cells. Allogeneic cyPS-Cart was transplanted into chondral defects in knee articular cartilage of cynomolgus monkeys for 4 months.

value: 3-4 years old

value: wild type

tag: samples

value: cyAC, cyFT


title: 209_B8_EB_3D_WT_SC_Batch1

data_processing:
Following quality controls (performed with FastQC v0.11.2 (https://www.bioinformatics.babraham.ac.uk/projects/fastqc)), sequencing reads were processed with Trim Galore! v0.5.0 (https://www.bioinformatics.babraham.ac.uk/projects/trim_galore) to perform quality and adapter trimming (parameters: --stringency 3 –q 20). Trimmed reads were next aligned to the mouse reference genome (mm10/GRCm38.p6) using STAR v2.7.1a with options: --outFilterMultimapNmax 10 --outFilterMultimapScoreRange 1 –outFilterMismatchNmax 999 –outFilterMismatchNoverLmax 0.04. Gene expression levels were quantified with featureCounts v1.6.1 (options: -t exon -g gene_name) using the GENCODE Release M23 annotation. Multi-mapped reads were excluded from quantification.
Genome_build: mm10/GRCm38.p6
Supplementary_files_format_and_content: Tab-delimited text file of raw read counts matrix per gene/cell.

extract_protocol:
Full length single cell RNA-seq was performed using a modified version of the Smart-seq2 protocol.
Briefly, individual cells are sorted into 96 well plates containing lysis buffer in presence of RNase inhibitor, dNTPs and OligodT. Reverse transcription of the polyadenylated RNA will be performed with SuperScriptII and Template Switching Oligos. The resulting cDNA will be amplified with 25 cycles of PCR and libraries will be prepared for sequencing with standard NexteraXT Illumina protocol. Libraries were sequenced on Illumina NextSeq 500 System (single-end 75bp reads), reaching a median of ~ 578,000 generated reads per cell.

value: Embryoid Bodies (EB)

value: WT


title: WTA16: articular cartilage and fibrous tissue

value: articuar cartilage and fibrous tissue formed in chondral defect


title: 249 POST


title: Vacc_13


title: 289 POST


source_name: CD4+ T cells, 4 h activation, PD patient 4

value: CD4+ T cells

tag: gender

value: male

tag: time-point post activation

value: 4 h


title: WTA15: post-transplant cyiPS-Cart

source_name: cyiPSC-derived cartilage after transplantation

value: post-transplant cyiPS-Cart


title: Tag15: post-transplant cyiPS-Cart

value: cyiPSC-derived cartilage after transplantation


title: Vacc_12


title: 209_B7_EB_3D_WT_SC_Batch1


title: 209_C1_EB_3D_WT_SC_Batch1


title: 288 POST


title: Vacc_11


title: 209_C10_EB_3D_WT_SC_Batch2


title: Vacc_15


title: 366 PRE


source_name: CD4+ T cells, 0 h activation, PD patient 4

value: Parkinson's disease

value: 0 h


title: 209_B5_EB_3D_WT_SC_Batch1


title: 209_C11_EB_3D_WT_SC_Batch2


title: 209_B9_EB_3D_WT_SC_Batch2


title: 500 PRE

value: plasma


title: 289 PRE

value: yes


title: WTA12: cyiPSCs and pre-transplant cyiPS-Cart

source_name: cyiPSCs and cyiPSC-derived cartilage before transplantation

value: EGFP reporter gene

value: cyiPSC, pre-transplant cyiPS-Cart


title: Vacc_9

value: whole blood

value: leukocyte


source_name: CD4+ T cells, 12 h activation, PD patient 4


title: Tag12: cyiPSCs and pre-transplant cyiPS-Cart

value: cyiPSCs and cyiPSC-derived cartilage before transplantation


title: 366 POST


title: Formative_ESC_MLL3CD_MLL4CD_H3K4m1_rep1

description:
CUT&RUN
Form_dCD_k4m1_merged.cpm.bw
supermatrix_k4m1.20220426_geo

data_processing:
Fastq files for CUT&RUN samples were processed using Nextflow and the nf-core CUT&RUN pipeline v1.0.0 beta. In brief, adapters were trimmed using Trim Galore. Paired-end alignment was performed using Bowtie2 and peaks were called using SEACR with a peak threshold of 0.05. We did not normalize peak calls to IgG. Spike in normalization was performed using the E.coli genome K12. However, spike-in normalization resulted in batch effects between replicates. These batch effects were greatly diminished when normalized by CPM. Therefore, all downstream analysis was performed using CPM normalized samples.
Assembly: mm10
Supplementary files format and content: supermatrix for H3K4m1 contains the intersection of naive and formative WT H3K4m1 peaks. Columns 1-3 H3K4m1 peak coordinates, Columns 12-19 Log2CPM RNAseq, Columns 5-10 Diffbind results for H3K4m1 peaks between Naive and Form, Columns 26-37 Log2 Peak densities for H3K4m1 and H3K27a
Supplementary files format and content: bigwig files are generated with deeptools by averaging bam files of replicate
Library strategy: CUT&RUN

extract_protocol:
CUT&RUN was conducted using the protocol from Skene et al. 2018 with the following modifications: Freshly trypsinized cells were bound to activated Concanavalin A beads (Bang Laboratories, CAT#BP531) at a ratio of 2e5 cells/10ul beads in CR Wash buffer (20mM HEPES, 150mM NaCl, 0.5mM Spermidine with protease inhibitors at room temp. An input of 2e5 cells were used per target. Bead-bound cells were then incubated rotating overnight at 4C in CR Antibody buffer (CR Wash with 0.05% Digitonin, 2mM EDTA) containing primary antibody. We used the following antibodies for CUT&RUN: 1:100 Rabbit anti-H3K4m1 (Abcam, ab4729), Rabbit anti-MLL4 1:100 (provided by Kai Ge), and 1:100 Rabbit IgG isotype control (Abcam, ab171870). After primary, we washed 3 times 5 minutes each with cold CR Dig-wash buffer (CR Wash with 0.05% Digitonin) and incubated with pA-MNase (1:100 of 143ug/mL provided by Steve Henikoff) for 1 hour rotating at 4C. After MNase binding, we washed 3 times 5 each with cold CR Dig-wash buffer, and chilled cells down to 0C using a metal tube rack partially submerged in an ice water slurry. MNase digestion was induced by adding CaCl2 at a final concentration of 2mM. After 30 minutes of digestion, the reaction was quenched using Stop Buffer containing 340mM NaCl, 20mM EDTA, 4mM EGTA, 0.05% Digitonin, 100ug/mL RNAse A, 50ug/mL Glycogen, and approximately 2pg/mL Yeast spike-in DNA (provided by Steve Henikoff). The digested fragments for each sample were then extracted using a phenol chloroform extraction.
Library preparation on samples was conducted using manufacturer’s protocols for NEBNext Ultra II Dna Library Prep Kit (New England BioLabs, CAT#E7645) and NEB Multiplex Dual Index oligos (New England BioLabs, CAT#E7600, #E7780) with the following modifications. We input approximately 10ng of sample for half reactions, we diluted the NEBNext Illumina adaptor 1:25, we used the following PCR cycling conditions: 1 cycle of Initial Denaturation at 98C for 30 seconds, 12+ cycles of Denaturation at 98C for 10 seconds then Annealing/Extension at 65C for 10 seconds, and 1 final cycle of extension at 65C for 5 minutes. Following library preparation, double size selection was performed using Ampure beads and quality and concentration of libraries were determined by an Agilent 4200 Tapestation with High-Sensitivity D1000 reagents before pooling for sequencing.

value: LIF2i withdrawal, 63hrs

tag: target antibody

value: anti-H3K4m1 (Abcam, #ab8895)


title: 497 POST


title: Naive_ESC_WT_IgG

description:
IgG control for H3K4m1 C&R
CR2_IgG_merged.cpm.bw
supermatrix_k4m1.20220426_geo

value: WT


value: CD4+ T cells

value: 2 h


title: 209_B2_EB_3D_WT_SC_Batch1

value: Embryoid Bodies (EB)


title: 209_B6_EB_3D_WT_SC_Batch1


title: 500 POST


title: Formative_ESC_MLL3KO_MLL4KO_H3K4m1_rep2

description:
CUT&RUN
Form_DKO_k4m1_merged.cpm.bw
supermatrix_k4m1.20220426_geo

tag: treatment


title: Formative_ESC_MLL3KO_MLL4fl_H3K4m1_rep2

description:
CUT&RUN
Form_CKO_k4m1_merged.cpm.bw
supermatrix_k4m1.20220426_geo


title: Vacc_7


title: 209_B4_EB_3D_WT_SC_Batch1


title: 497 PRE


title: Vacc_8


title: 248 POST


title: 249 PRE


title: Vacc_10


title: Formative_ESC_MLL3KO_MLL4fl_H3K4m1_rep1


title: 209_B3_EB_3D_WT_SC_Batch1


title: 490 POST


title: Formative_ESC_WT_H3K4m1_rep2

description:
CUT&RUN
Form_WT_k4m1_merged.cpm.bw
supermatrix_k4m1.20220426_geo


title: 490 PRE


title: Vacc_6


title: 247 PRE


title: Formative_ESC_MLL3KO_MLL4KO_H3K4m1_rep1


title: Vacc_5


title: 247 POST


title: 209_B12_EB_3D_WT_SC_Batch2


title: Formative_ESC_MLL3CD_MLL4CD_H3K4m1_rep2


title: Vacc_2


title: Vacc_3


title: 209_A8_EB_3D_WT_SC_Batch1


title: 209_B10_EB_3D_WT_SC_Batch2


title: 209_A9_EB_3D_WT_SC_Batch2


title: Naive_ESC_MLL3CD_MLL4CD_H3K4m1_rep2

description:
CUT&RUN
Naive_dCD_k4m1_merged.cpm.bw
supermatrix_k4m1.20220426_geo


title: 209_A7_EB_3D_WT_SC_Batch1


title: 248 PRE


title: 209_B1_EB_3D_WT_SC_Batch1


title: MDA_1_N_1

city: Baltimore

first: Ines

last: Godet

state: MD

address: 1650 Orleans Street, CRB1, room 128

department: Oncology

institute: Johns Hopkins University

data_processing:
Preliminary analysis was performed by Novogene. The reads were then mapped to Homo Sapiens(GRCh38/hg38) using STAR (v2.5) with the parameter mismatch =2.
Quantification was done using HTSeq (v0.6.1) software with the parameter -m union.
Differentially expressed genes were identified using EdgeR (v3.16.5) with padj<0.005 and [log2(FoldChange)]>1.
Supplementary files format and content: Comma delimited text file containing gene-level counts

source_name: human cell line

growth_protocol: MDA-MB-231 and MCF7 cells obtained directly from the ATCC and the SUM159 cell line was given as a gift from Dr. Sara Sukumar's laboratory.

extract_protocol:
Total RNA was extracted from cells using TRIzol (Invitrogen) and purified using Direct-zol RNA mini kit (Zymo Research) with DNase I treatment.
Following RNA purification, samples were confirmed to have a RIN value >9.0 when measured on an Agilent Bioanalyzer. Total RNA qualification, mRNA enrichment, double-stranded cDNA synthesis, end repair, poly-A and adaptor addition, fragments selection and PCR, library quality assessment and Illumina sequencing. Sequencing was performed on a NovaSeq 6000 system with 150-bp paired-end run by Novogene (Sacramento, CA).

treatment_protocol: Cell lines were treated with 1 uM of Mebendazole or DMSO vehicle control for 72 hrs for SUM159 or 48 hrs for MDA-MB-231 and MCF7.

value: 1 uM MBZ

tag: oxygen


title: Naive_ESC_MLL3CD_MLL4CD_H3K4m1_rep1


title: 209_B11_EB_3D_WT_SC_Batch2

tag: strain


title: 243 PRE


title: 209_A6_EB_3D_WT_SC_Batch1

tag: cell type


title: Vacc_1


title: Formative_ESC_WT_H3K4m1_rep1

tag: genotype


title: MDA_0_N_1

tag: cell line

value: 0 uM MBZ


title: 243 POST


title: MDA_0_H_1


title: 209_A4_EB_3D_WT_SC_Batch1

tag: Stage


title: MDA_1_H_1


title: Naive_ESC_MLL3KO_MLL4fl_H3K4m1_rep2

description:
CUT&RUN
Naive_CKO_k4m1_merged.cpm.bw
supermatrix_k4m1.20220426_geo


title: Vacc_4


title: 241 POST


title: 209_A3_EB_3D_WT_SC_Batch1

tag: batch


title: 241 PRE

tag: tissue


title: 209_A12_EB_3D_WT_SC_Batch2


title: 209_A5_EB_3D_WT_SC_Batch1


title: Naive_ESC_MLL3KO_MLL4KO_H3K4m1_rep2

description:
CUT&RUN
Naive_DKO_k4m1_merged.cpm.bw
supermatrix_k4m1.20220426_geo


title: 209_A11_EB_3D_WT_SC_Batch2


title: 242 POST


title: MCF7_0_H_1


title: Naive_ESC_WT_H3K4m1_rep1

description:
CUT&RUN
Naive_WT_k4m1_merged.cpm.bw
supermatrix_k4m1.20220426_geo


title: Naive_ESC_MLL3KO_MLL4KO_H3K4m1_rep1


title: Naive_ESC_MLL3KO_MLL4fl_H3K4m1_rep1


title: MCF7_1_H_1


title: 250 PRE


title: SMN_0M_1


title: Meso_T48_WT_Rep1

data_processing:
NOT PROVIDED; REQUESTED
Genome_build: mm10/GRCm38
Supplementary_files_format_and_content: Tab-delimited text file of normalized read counts per gene/samples.

source_name: Mouse Meso-Endodermal Progenitors

growth_protocol: E14 mouse WT and mutant (Dnmt3a -/-   and Dnmt3b -/-) ESCs were cultured in high-glucose DMEM (Euroclone) supplemented with 15% FBS (Millipore Corp., Billerica, MA, USA), 0.1 mmol/l nonessential amino acids (Invitrogen), 1 mmol/l sodium pyruvate (Invitrogen), 0.1 mmol/l β-mercaptoethanol, 1500 U/ml Leukemia Inhibitory Factor (LIF; Millipore), 25 U/ml penicillin, and 25 μg/ml streptomycin. For EpiSC induction, a single-cell suspension was seeded onto Geltrex (A1413202 GIBCO)-coated plates at a density of 10,000 cells cm-2 in N2B27 medium supplemented with 20ng/ml ActivinA (PHC9564 GIBCO) and 12 ng/ml bFGF (PHG0026 GIBCO). The cells were passaged 1:3 as small clumps using Collagenase IV (17104019 GIBCO). EpiSCs were collected for DNA and RNA analyses after 14 days of induction followed by daily medium changes. N2B27 medium is composed by 50% advanced DMEM/F12 (12634028 GIBCO) and 50% Neurobasal medium (21103049 GIBCO), supplemented with 0.5% N2 Supplement (17502048 GIBCO), 1% B27 Supplement (17504044 GIBCO), 0.033% BSA solution (A9647 SIGMA), 50 uM β-mercaptoethanol (M3148 Sigma), 2mM Glutamax (35050038 GIBCO), 100U/ml penicillin and 100 ug/ml streptomycin (DE17-602E LONZA). For Meso-endoderm directed lineage specific differentiation, EpiSCs were plated as small clumps onto Geltrex-coated plates in EpiSCs medium for 24 hours. The day after, medium was replaced with N2NB27 medium consisted of 50% advanced DMEM/F12 (12634028 GIBCO) and 50% Neurobasal medium (21103049 GIBCO), supplemented with 0.5% N2 Supplement (17502048 GIBCO), 1% B27 supplement minus Vitamin A (12587010 GIBCO) and 3 uM iGSK3β (CHIR99021 SIGMA). Cells were fed daily until the end of differentiation.

extract_protocol:
Total RNA was extracted by using TRIzol reagent (Invitrogen), according to manufacturer’s protocol.
Quantity and quality of the starting RNA were checked by Qubit and Bioanalyzer (Agilent). 1 μg of total RNA was subjected to poly(A) selection, and libraries were prepared using the TruSeq RNA Sample Prep Kit (Illumina) following the manufacturer’s instructions. Libraries were sequenced on Illumina NextSeq 500 System (single-end 75 bp reads).

tag: strain background

value: Wild type (WT)

value: EpiSC derived ME progenitors

value: ME T48h

tag: clone


title: SMN_1M_1


title: 250 POST

value: plasma

value: yes


title: 240 PRE


title: LCM ventricular zone, PTU, replicate 4

city: Research Triangle Park

first: Katherine

middle: Lynn

last: O'Shaughnessy

state: NC

address: 109 TW Alexander Drive

institute: United States Environmental Protection Agency

data_processing:
Samples were demultiplexed and trimmed using Trimmomatic (Bolger et al., 2014).
Downstream processing of RNA-Seq data was performed in Partek Flow Bioinformatics Software (Chesterfield, MO).
Samples were aligned to the Rattus norvegicus reference genome (mRatBN7.2/rn7) using STAR 2.7.8a.
Counts were quantified using the Partek Expectation/Maximization (E/M) model and the rat RefSeq annotation (GCF_015227675.2) and normalized using median ratio. Differential expression was identified by comparing N=6 PTU exposed samples to N=7 controls using DESeq2 (Love et al., 2014)
Assembly: mRatBN7.2/rn7
Supplementary files format and content: tab-delimited text file includes DESeq2 median ratio normalization

organism: Rattus norvegicus

source_name: Long Evans

extract_protocol:
RNA was isolated using Qiagen’s Rneasy Micro kit according to the manufacturer’s protocol, including on-column DNA digestion.
Total RNA-Sequencing libraries were prepared using the SMARTer® Stranded Total RNA-Seq Kit v3 - Pico Input Mammalian kit according to the manufacturer’s protocol. This included sample barcoding and rRNA depletion.

treatment_protocol: Pregnant rats were exposed to 0 (control) or 3 ppm PTU through their drinking water through pregnancy or lactation. This exposure induced mild hypothyroidism in the mothers, and hypothyroidism in the offspring. On postnatal day 2 (PN2), the posterior ventricular zone was microdissected from 1 male pup/litter for RNA-Seq.

value: Long Evans

tag: Sex

value: Male

tag: cell population

value: microdissected ventricular zone, posterior

tag: developmental stage

value: postnatal day 2 (PN2)

value: maternal PTU exposure (PTU)

tag: physiological stage

value: Hypothyroid


title: Naive_ESC_WT_H3K4m1_rep2

value: WT

value: anti-H3K4m1 (Abcam, #ab8895)


title: 242 PRE

tag: diet yes/no


title: SMH_1M_1


title: LCM ventricular zone, PTU, replicate 6


title: 239 PRE


title: 209_A10_EB_3D_WT_SC_Batch2


title: MCF7_1_N_1


title: 240 POST


title: Meso_T48_B126_Rep2

value: Dnmt3b -/- (3BKO)


title: Meso_T48_B77_Rep2


title: Meso_T48_WT_Rep2


title: LCM ventricular zone, PTU, replicate 1


title: Meso_T48_B126_Rep1


title: Transfection of pXG-SRSF7 and pcDNA3-pri-mir-142_WT repeat1

data_processing:
We trimmed adapters, joined the paired-end reads, removed low-quality reads, and mapped reads to build-in reference for ilime-seq samples as described in the previous study (Le et al., 2020).
Assembly: Pri-miRNA construct sequences
Supplementary files format and content: reference_sequences.txt: Tab-delimited text file contains the reference sequences of different pre-miR-142 isoforms
Supplementary files format and content: rawcount.txt: Tab-delimited text file contains the raw count of pre-miR-142 in different samples
Library strategy: Ilime-seq

source_name: DICER-KO HCT116 cell line

extract_protocol:
Illumina adapter sequences were incorporated to the libraries using PCR.
1.5 μg of pXG-empty vector, pXG-SRSF7, or pXG-SRSF3 was co-tranfected with 1 μg of pcDNA3-pri-mir-142_WT into one well in the 6-well-plate of HCT116-DICER KO cells. The cells were also harvested two days later. All the total RNAs were extracted using TRIzol™ Reagent (Thermo Fisher, Cat. 15596018). We used iLIME-seq to clone and sequence pre-mir-142 from these total RNAs (Le et al., 2022). In short, 2.5 μg of total RNA was incubated with 16 units of T4 RNA ligase 1 (NEB, Cat. M0437M) in the ligation buffer containing 50 mM Tris-HCl (pH 7.5), 10 mM MgCl2, 1 mM DTT, 16 units of SUPERase•In RNase inhibitor (Thermo Fisher, Cat. AM2694), and 20% PEG 8000. The mixtures were incubated at 25°C for 2 h. The ligated RNA products were then subjected to reverse transcription reactions using SuperScript™ IV Reverse Transcriptase at 50°C for 20 min. The resulting cDNAs were amplified the first time by pre-mir-142 sequence-specific primers and the second time by sequencing primers, both by Phusion Hot Start II DNA Polymerase (Thermo Fisher, Cat. F549L). The final DNA products from the second PCR were subjected to next-generation sequencing using the Illumina Novaseq 6000.

value: DICER-KO

tag: transfection

value: pXG-SRSF7 + pcDNA3-pri-mir-142_WT

tag: repeat

value: repeat 1


title: 373 PRE

value: no


title: SMH_0M_1

tag: treatment

value: 0 uM MBZ

tag: oxygen


title: LCM ventricular zone, control, replicate 7

tag: strain

value: vehicle control (Control)

value: Euthyroid


title: Meso_T24_WT_Rep2

tag: genotype

value: Wild type (WT)

tag: cell type

value: EpiSC derived ME progenitors

value: ME T24h


title: Meso_T24_B77_Rep2

tag: Stage


title: 239 POST

value: plasma

value: yes


title: MCF7_0_N_1

tag: cell line


title: Meso_T48_B77_Rep1

value: ME T48h


title: LCM ventricular zone, PTU, replicate 3

value: Long Evans

value: Male

value: microdissected ventricular zone, posterior

value: postnatal day 2 (PN2)

value: maternal PTU exposure (PTU)

value: Hypothyroid


title: Meso_T24_B77_Rep1

value: Dnmt3b -/- (3BKO)


title: LCM ventricular zone, control, replicate 6


title: 499 POST

tag: tissue

value: no


title: Transfection of pXG-SRSF7 and pcDNA3-pri-mir-142_WT repeat3

value: DICER-KO

value: pXG-SRSF7 + pcDNA3-pri-mir-142_WT

value: repeat 3


title: 373 POST


title: 236 POST


title: Transfection of pXG-SRSF7 and pcDNA3-pri-mir-142_WT repeat2

value: repeat 2


title: LCM ventricular zone, PTU, replicate 2


title: Transfection of pXG-SRSF3 and pcDNA3-pri-mir-142_WT repeat3

value: pXG-SRSF3 + pcDNA3-pri-mir-142_WT


title: 499 PRE


title: Meso_T24_WT_Rep1

tag: strain background

value: Wild type (WT)

value: EpiSC derived ME progenitors

value: ME T24h

value: WT


title: 236 PRE

value: plasma

value: yes


title: Transfection of pXG-SRSF3 and pcDNA3-pri-mir-142_WT repeat2


title: Transfection of pXG and pcDNA3-pri-mir-142_nonCNNC repeat3

value: pXG + pcDNA3-pri-mir-142_nonCNNC


title: LCM ventricular zone, control, replicate 4

value: Long Evans

value: Male

value: microdissected ventricular zone, posterior

value: postnatal day 2 (PN2)

value: vehicle control (Control)

value: Euthyroid


title: Transfection of pXG-SRSF3 and pcDNA3-pri-mir-142_WT repeat1

value: DICER-KO

value: repeat 1


title: Meso_T24_B126_Rep1

value: Dnmt3b -/- (3BKO)

tag: clone


title: LCM ventricular zone, PTU, replicate 5

tag: Sex

value: maternal PTU exposure (PTU)

value: Hypothyroid


title: LCM ventricular zone, control, replicate 2

tag: cell population


title: Transfection of pXG and pcDNA3-pri-mir-142_nonCNNC repeat2

value: repeat 2


title: Meso_T24_B126_Rep2

value: EpiSC derived ME progenitors

value: ME T24h


title: 364 PRE

value: plasma

value: no


title: LCM ventricular zone, control, replicate 3

tag: developmental stage


title: Transfection of pXG and pcDNA3-pri-mir-142_WT repeat3

value: pXG + pcDNA3-pri-mir-142_WT

value: repeat 3


title: 301 PRE


title: ES_WT_Rep2

source_name: Mouse Embryonic Stem Cells

value: Wild type (WT)

value: ESC

value: WT


title: LCM ventricular zone, control, replicate 5

value: Long Evans

value: Male

value: microdissected ventricular zone, posterior

value: postnatal day 2 (PN2)

value: vehicle control (Control)

tag: physiological stage


title: 364 POST

tag: diet yes/no


title: ES_B77_Rep1

value: Dnmt3b -/- (3BKO)


title: 299 PRE


title: Transfection of pXG and pcDNA3-pri-mir-142_WT repeat1

value: DICER-KO

tag: transfection

value: repeat 1


title: ES_WT_Rep1


title: ES_B77_Rep2


title: ES_B126_Rep2


title: 301 POST


title: NCI-H524_suspension

city: Bethesda

first: Lorinc

middle: Sandor

last: Pongor

state: Maryland

address: 37 Convent drive

department: NCI

institute: NIH

data_processing:
Raw reads were trimmed using trimmomatic (v0.36), followed by alignment to the hg19 genome using BWA MEM.
Aligned reads were sorted using samtools, duplicate reads were marked using picard-tools.
Normalized sequencing depth tracks were generated using BAMscale
Assembly: hg19
Supplementary files format and content: bw: normalized sequencing depth
Library strategy: WGS

source_name: Lung

growth_protocol: NCI-H524 cell lines was obtained from ATCC (www.atcc.org) and tested for mycoplasmas (Lonza, LT07-418). Human H524 cells were grown at 37 °C in a 5% CO2 atmosphere in RPMI 1640 medium (ThemoFisher, 11875119), supplemented with 10% fetal calf serum (GeminiBio, 100-106). Cells were passage by splitting cells 1:3 to 1:4, keeping cells below 0.5 x 105/mL. Adherent NCI-H524 cells were selected by removing three quarters of the floating cells at regular intervals to avoid the medium to turn yellow. Adherent NCI-H524 were first collected by trypsinization and transferred to new flasks when ~ 50% confluency was achieved.

extract_protocol:
Genomic DNA samples from five million cells were prepared using the Qiagen DNeasy blood and tissue kit (69504).
Cells were lysed in preparation for in-situ contact generation. Isolated nuclei were permeabilized, restriction digestion was carried out for 2hrs at 37C with MboI (New England Biolabs). Restriction sites filed with dNTP’s using Biotin-14-dATP (Jena Biosciences) for 1hr at 37C. The filled ends were then ligated together using a T4 ligase at room temperature for 4 hr with before nuclei were lysed, sonicated (using Covaris) and then H3k27ac (Abcam, ab4729) immunoprecipitation was carried out overnight using antibodies. The morning after, 30ul of beads were added to collect the chromatin-antibody complex; then the ChIP DNA was collected, washed and crosslinks reversed overnight using Proteinase K. ChIP DNA was eluted and samples purified using the DNA Clean up kit. The DNA was quantified using a Qubit before biotin ligation junction capture using Streptavidin C-1 beads. Samples were washed and taken forward for Tn5 Tagmentation. Tagmentation and PCR amplification was performed as described8.
NA samples were sequenced using the Novaseq S1 sequencing system (150bp, paired-end reads) to obtain more than 300 million total reads

value: Lung

value: suspension population


title: Transfection of pXG and pcDNA3-pri-mir-142_WT repeat2

tag: repeat

value: repeat 2


title: Transfection of pXG and pcDNA3-pri-mir-142_nonCNNC repeat1

value: pXG + pcDNA3-pri-mir-142_nonCNNC


title: 294 POST

value: plasma

value: no


title: 303 POST


title: BAS cells - paclitaxel resistant - biological sample 2

city: London

first: Ernesto

last: Yague

address: Du Cane Road

department: Division of Cancer

country: United Kingdom

institute: Imperial College London

description: Total RNA isolated from cells in culture

hyb_protocol: Following fragmentation and labeling, ss-cDNA were hybridized for 16 hr at 45°C to Clariom S Human Arrays. GeneChips were washed and stained in the GeneChip® Fluidics Station 450 (Affymetrix, P/N 00-0079).

scan_protocol: GeneChips were scanned using the GeneChip® Scanner 3000 7G.

data_processing: The raw CEL files were normalized using RMA background correction with quantile normalization, log2 transformation and mean probe-set summarization with adjustment for GC content. Differentially expressed genes were considered significant if p-value with FDR ≤ 0.05 and fold-change of >2 or < -2.

label: Biotin

source_name: BAS cells - paclitaxel resistant

label_protocol: Affymetrix GeneChipTM WT PLUS Reagent Kit

growth_protocol: HS578T cells were grown in low-glucose DMEM containing 10% fetal calf serum, glutamine and insulin. BAS cells were grown in keratynocyte serum-free medium supplemented with human recombinant epidermal growth factor and bovine pituitary extract

extract_protocol: Qiagen Rneasy total RNA extraction

value: breast cancer

value: BAS

tag: resistance

value: paclitaxel-resistant


title: 294 PRE


title: 299 POST


title: NCI-H524_adherent

value: adherent population


title: 265 POST


title: EpiSC_WT_Rep2

city: Turin

first: Andrea

last: Lauria

address: Via Accademia Albertina, 13

institute: University of Turin

source_name: Mouse Epiblast Stem Cells

value: Wild type (WT)

value: ES-E14 derived EpiSC

value: EpiSC

value: WT


title: BAS cells - paclitaxel resistant - biological sample 3


title: BAS cells - paclitaxel resistant - biological sample 1


title: DMS-273_scCNV

description: targeted amplicons

data_processing:
Targeted amplicons were designed to cover amplified ecDNA regions. The design covered a total of 199 sites
Sequencing run was demultiplexed using Bcl2fastq v2.20. The fastq files were processed through Mission Bio’s Tapestri pipeline v2.0.1 with default parameters.
the adapter sequences were trimmed from raw reads using Cutadapt v1.18. Reads that are too short with less than 30bps were discarded.
Trimmed reads were mapped to the reference genome hg19 using BWA-MEM v0.7.17.
The CellFinder module extracted the barcodes from the mapped reads to identify captured cells. Barcodes that passed a total reads cutoff (the number of amplicons in the panel x 8 reads) are selected from at least 80% of good-performing amplicons (with the number of reads more than 0.2 x the mean of all amplicon reads).
In the next step, the GATK HaplotypeCaller was used to call variants individually on each cell, and then GATK GenotypeGVCFs was applied to jointly genotype all cells using genotype likelihoods from the previous step based on the GATK Best Practices.
The unfiltered VCF was parsed into an HDF5 file, which contains single-cell variants and per-amplicon read count matrices compatible with downstream analysis.
Raw amplicon quantifications for the cell lines were analyzed using R. Data was normalized using the Seurat package, using the NormalizeData() function, setting the method to “RC”, minimum number of cells to 3, and minimum features to 25. Since many of the amplicons targeted ecDNA regions, normalization was first performed on background amplicons and amplicons targeting tumor suppressor genes, from which the scaling factors were obtained. The normalized read counts for all amplicons were calculated using the obtained scaling factors. Normalization was performed for each of the cell lines separately (DMS-273, NCI-H889 and the patient derived cells together), to exclude any variation in copy number of the non-ecDNA regions.
Assembly: hg19
Supplementary files format and content: read count table
Supplementary files format and content: scCNV amplicon design
Library strategy: scCNV

growth_protocol: Cells were maintained in RPMI media  supplemented with 5% heat deactivated fetal bovine serum without antibiotics at 37 °C in a humidified atmosphere containing 5% CO2 and 95% air.

extract_protocol:
Briefly, cells grown at 70% confluency were detached, cross-linked with formaldehyde for 10mins and quenched with 125mM Glycine.
Cryopreserved cells were quickly thawed by gently swirling the vials in the 37C water bath. Cells were suspended and washed with 1X DPBS at room temperature. Cells were then resuspended in Mission Bio cell buffer at a concentration of 3,000-4,000 cells/ul. Afterwards, the cells were microfluidically encapsulated, lysed, and barcoded in Tapestri platform. Barcoded samples were then simultaneously PCR-amplified in droplets with a custom 199-amplicons panel covering 16 genes. Amplified products were purified and ligated with Illumina specific P5 and P7 indices. In the next step, libraries were quantified with an Agilent Bioanalyzer for quality control, pooled and sequenced on an Illumina NovaSeq platform.
Pooled libraries were sequenced on NovaSeq 6000 SP with paired-end sequencing (2 x 151 cycles).


title: BAS cells - doxorubicin resistant - biological sample 3

source_name: BAS cells - doxorubicin resistant

value: doxorubicin-resistant


title: 303 PRE


title: ES_B126_Rep1

value: Dnmt3b -/- (3BKO)

value: ESC


title: EpiSC_B77_Rep2


title: EpiSC_B77_Rep1


title: LCM ventricular zone, control, replicate 1

value: Long Evans

value: Male

value: microdissected ventricular zone, posterior

value: postnatal day 2 (PN2)

tag: treatment

value: vehicle control (Control)

value: Euthyroid


title: NCI-H524_HiChIP_H3K27ac

data_processing:
Sequenced reads were processed using the HicPro pipeline (v3.1.0), aligning the data to the hg19 genome version and MboI motif for the analysis.
The resulting valid read pairs were converted to a contact matrix “.hic” format with the hicpro2juicebox.sh tool from the Juicebox suite.
Bigwig files were generated using the BAMscale tool on the sorted genome alignment BAM files.
Assembly: hg19
Supplementary files format and content: hic: contact file
Supplementary files format and content: bw: normalized sequencing depth
Library strategy: HiChIP

extract_protocol:
Briefly, cells grown at 70% confluency were detached, cross-linked with formaldehyde for 10mins and quenched with 125mM Glycine.
Cells were lysed in preparation for in-situ contact generation. Isolated nuclei were permeabilized, restriction digestion was carried out for 2hrs at 37C with MboI (New England Biolabs). Restriction sites filed with dNTP’s using Biotin-14-dATP (Jena Biosciences) for 1hr at 37C. The filled ends were then ligated together using a T4 ligase at room temperature for 4 hr with before nuclei were lysed, sonicated (using Covaris) and then H3k27ac (Abcam, ab4729) immunoprecipitation was carried out overnight using antibodies. The morning after, 30ul of beads were added to collect the chromatin-antibody complex; then the ChIP DNA was collected, washed and crosslinks reversed overnight using Proteinase K. ChIP DNA was eluted and samples purified using the DNA Clean up kit. The DNA was quantified using a Qubit before biotin ligation junction capture using Streptavidin C-1 beads. Samples were washed and taken forward for Tn5 Tagmentation. Tagmentation and PCR amplification was performed as described8.
Libraries were size selected to 200–700 bp and sequenced on the NovaSeq 600 using paired-end sequencing.

value: Lung


title: EpiSC_WT_Rep1


title: BAS cells - doxorubicin resistant - biological sample 1

value: breast cancer

value: BAS


title: BAS cells - biological sample 3

source_name: BAS cells

value: parental naive


title: BAS cells - doxorubicin resistant - biological sample 2


title: 293 POST

source_name: plasma

value: plasma

value: no


title: 293 PRE


title: MBT06 MRD Control

city: Hamilton

first: Sheila

last: Singh

state: ON

address: 1280 Main Street West

department: Biochemistry and Biomedical Sciences

country: Canada

institute: McMaster University

description: Cell line engrafted in NSG mice and grown as orthotopic tumors.

data_processing:
FASTQ files were first filtered using a bespoke Perl script to remove reads shorter than 36bp.
Filtered FASTQ files were aligned to Gencode v25 gene annotations and hg38 genome build using the STAR short-read aligner (v2.4.2a)
Read count files generated by STAR were merged with gene annotations in R
Genome_build: hg38
Supplementary_files_format_and_content: tab-delimited text read counts

source_name: Glioblastoma multiforme cell line

growth_protocol: Neurospheres derived from minimally-cultured human GBM samples were plated on polyornithine- laminin coated plates for adherent growth. Adherent cells were replated in low-binding plates and cultured as tumorspheres, which were maintained as spheres upon serial passaging in vitro. These cells retained their self-renewal potential and were capable of in vivo tumor formation.

extract_protocol:
Total RNA was isolated from human GBM cells sorted from mouse brains using Norgen total RNA purification kit following the manufacturer's instructions.
Sequencing libraries were prepared using Illumina stranded totalRNA kits following the manufacturer's protocol.

treatment_protocol:
1E6 GBM cells (BT428, BT799, BT935, BT954 and MBT06) were injected in NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mouse brains as follows. Mice were anesthetized using gas anesthesia (Isoflurane: 2.5%). Using a 15-blade scalpel a 1.5 cm vertical midline incision was made on top of the skull. A small burr hole was then made (2-3 mm anterior to the coronal suture, 3 mm lateral to midline) using a drill held perpendicular to the skull. A Hamilton syringe was used to inject 10μL of cell suspension of GBM cells into the frontal lobe. The syringe was inserted through the burr hole at a 30 degree angle to a depth of 5mm. The incision was closed using interrupted stitches and sutures were sealed with a tissue adhesive. Mice were identified using ear notches and placed in recovery cages. Mice were monitored weekly for signs of illness and sacrificed at endpoint.
Once tumor engraftment was confirmed using MRI, mice were randomly assigned to control or treatment groups and chemoradiotherapy treatment was started. For mice in the combined chemoradiotherapy arm, 50 mg/kg TMZ was oral gavaged one hour before cranial irradiation was administered to mice. The mouse shield is designed such that only the head of the mice is irradiated.

value: Glioblastoma multiforme cell line

tag: tumor stage

value: Miniminal residual disease


title: EpiSC_B126_Rep1

tag: genotype

value: ES-E14 derived EpiSC

value: EpiSC


title: BAS cells - biological sample 4


title: BT954 MRD Control

tag: cell type


title: NCI-H889_HiChIP_H3K27ac


title: 295 PRE


title: EpiSC_B126_Rep2

value: Dnmt3b -/- (3BKO)

tag: Stage


title: 291 POST


title: Normal keratinocytes

city: Cincinnati

first: Mike

last: Adam

state: Ohio

address: 240 Albert Sabin Way

department: CBDI

institute: Cincinnati Children's Hospital Research Foundation

data_processing:
Raw sequencing data was processed through 10X Genomics CellRanger v3.0.2 using default parameters to obtain the gene expression matrices
Genome_build: Modified hg19 including annotated HPV16 genome
Supplementary_files_format_and_content: CellRanger output files (barcodes, genes, matrix)

source_name: cell culture

extract_protocol:
Single-cell suspensions were processed through Chromium 10X to generate single-cell cDNA libraries attached to microbeads
Microbeads were counted, and amplified by PCR, and the 3' end of the cDNA prepared for sequencing using a modified Nextera XT protocol

value: immortalized keratinocytes

tag: infection

value: control