title: XAUT1-POOL-SCG5

description:
XAUT1-POOL-SCG5_pool2: XAUT1-HS1-CLL1 XAUT1-HS2-CLR1 XAUT1-HS3-CLL1 XAUT1-HS4-CLR1 XAUT1-HS5-CLL1 XAUT1-HS6-CLR1 XAUT1-HS7-CLL1 XAUT1-HS9-CLL1 XAUT1-HS10-CLR1 XAUT1-HS11-CLL1 XAUT1-HS12-CLR1
biopsy_RNA.rds
biopsy_RNA.h5ad


title: Control093

status: Public on Jan 22 2024

type: SRA

city: Kiel

first: Florian

last: Uellendahl-Werth

address: Rosalind-Franklin-Straße 12

department: Institute of Clinical Molecular Biology (IKMB)

country: Germany

institute: Christian-Albrechts Universität zu Kiel

description: 3'-biased mRNA

library_source: transcriptomic

data_processing:
Bases were called and samples demultiplexed with an Illumina 1.8+ pipeline.
Pipeline rnaseq v1.3 https://zenodo.org/record/2610144
Alignment with STAR v2.6.1d
Feature counting with FeatureCounts from the subread package v1.6.4
for more details, see pipeline documentation
Genome_build: GRCh37
Supplementary_files_format_and_content: Comma-separated values of raw feature counts results

molecule: total RNA

organism: Homo sapiens

source_name: Whole Blood

extract_protocol:
PAXgene Blood RNA Kit; Pellets from PAXgene Blood RNA Tubes are washed, resuspended, and incubated in optimized buffers together with proteinase K to digest proteins. Lysates are applied to a PAXgene 96 Filter Plate and centrifuged to remove residual cell debris. Ethanol is added to the flow-through to adjust binding conditions, and the lysates are applied to a PAXgene 96 RNA Plate. RNA is selectively bound to the PAXgene 96 RNA membrane as contaminants pass through. Residual DNA is removed through a DNase I digestion on the PAXgene 96 membrane. Remaining contaminants are removed in three efficient wash steps and RNA is eluted in Buffer BR5. (from manufacturer’s guide)
Lexogen QuantSeq 3’ mRNA-Seq Library Prep Kit with 50 ng RNA input. Optional “Globin Block Module” used in all samples. Poly(A) RNA is reverse transcribed with poly(T)-Primers. RNA is digested and the second strand is synthesized by random priming. cDNA is purified. Illumina adapters are added. The PCR was run with 15 cycles.

tag: disease

value: Control

tag: age

tag: sequencing plate number (used for batch correction)


title: Control092

description: 3'-biased mRNA


title: Kouiki_I_140811_G08

type: RNA

city: Hitachi

first: Mine

last: Nose

state: Ibaraki

address: 3809-1 Ishi, Juo

country: Japan

institute: Forest Tree Breeding Center

hyb_protocol: 600 ng of Cy3-labelled cRNA was fragmented at 60°C for 30 minutes in a reaction volume of 25 ul containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 25 ul of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent SurePrint G3 Gene Expression Custom 8×60K Array (G4102A) for 17 hours at 65°C in a rotating Agilent hybridization oven.  After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent).

scan_protocol: Slides were scanned immediately after washing on SureScan Microarray Scanner G4900DA using one color scan setting for 8x60k array slides (AgilentG3_HiSen_GX_1Color).

data_processing: The scanned images were analyzed with Feature Extraction Software 11.5.1.1 (Agilent) using default parameters (Protocol: GE1_1105_Oct12 and Grid: 069025_D_F_20140811) to obtain background subtracted and spatially detrended Processed Signal intensities.

organism: Cryptomeria japonica

source_name: Ibaraki, Aug-14,  replicate 2

label_protocol: Cyanine-3 (Cy3) labeled cRNA was prepared from  200 ng total RNA using the One-Color Low Input Quick Labeling kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA).  Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.

growth_protocol: The cuttings of Japanese cedar were planted in three regions (Yamagata, Ibaraki and Kumamoto, Japan) in 2013.

extract_protocol: The total RNA was extracted from 500 mg samples as Gehrig et al. (2000) reported using an RNeasy Plant Mini Kit (Qiagen, Hilden, Germany), and a DNase digestion was performed on-column using an RNase-free DNase Set (Qiagen).

treatment_protocol: All samples were immediately frozen in liquid nitrogen and stored at -80 °C until use.

tag: tissue

value: shoot

tag: day of sampling

tag: geographical location

value: Ibaraki


title: GL78_087_WN-S2-P1_C12_AGM_E10-5_SP_32-33_Gfi1_HE__S87

city: Barcelona

first: Anna

last: Bigas

state: Barcelona

address: Dr. Aiguader 88

department: Cancer Research

country: Spain

institute: Institut Hospital del Mar d'Investigacions Mèdiques

library_source: transcriptomic single cell

data_processing:
Raw data was aligned to the reference mouse genome using STAR aligner tool (v2.7.3)
Gene counts were extracted from BAM files using HTSeq software (v0.9.1)
Python (v3.7.3) and scanpy (v1.4.4) for data pre-processing
As QC, cells with less than 2,500 expressed genes and less than 50,000 counts were discarded, and no more than 5% mitochondrial gene expression.
Assembly: mm10
Supplementary files format and content: Tab-separated value file: scRNAseq_Cell_annotation.txt: cell sample metadata table. For each cell (row, 860 in total): cell full name, corresponding plate/well, embryo developmental stage, cell population, batch name, FACS index data for (in this order) DLL4, NOTCH2, CKIT, GFI1, CD41, NOTCH1, CD31, JAG1 and CD45. Additionally, for those cells passing the QC, UMAP coordinates and annotated cluster (Louvain method, see publication for details) are included.
Supplementary files format and content: Tab-separated value file: scRNAseq_Raw_counts.txt: full-length RNAseq (SMART-Seq2).Raw expression levels for each gene (46,170 rows) in each single cell (860 columns) as calculated using HTseq. This data is prior to conducting any QC.
Supplementary files format and content: Tab-separated value file: scRNAseq_Norm_counts_filtered.txt. Normalized expression levels after QC. Matrix comprises 30,362 rows (genes) per 775 columns (cells). Counts were normalized per cell with pp.normalize_total function from scanpy (target sum of 1e4) and values logarithmize with pp.log1p scanpy function.

organism: Mus musculus

source_name: AGM

growth_protocol: E10.5 and E11.5 AGMs were prepared for flow sorting

extract_protocol:
Single cells were sorted into well plates containing lysis buffer and snap frozen. Libraries were prepared using a modified version of the Smart-Seq2 protocol. Briefly, cDNA was prepared using a Mantis platform (Formulatrix) and quantified with quantIT picogreen reagent (Thermo Fisher).
Dual indexed sequencing libraries were prepared from 0.1ng cDNA using an Echo525 automation system (Labcyte) in miniaturized reaction volumes. The library pool was quantified by qPCR using a Library Quantification Kit for Illumina sequencing platforms (Kapa Biosystems).

treatment_protocol: no treatments, only FACS staining

tag: cell type

value: Hemogenic endothelium cells

value: AGM

tag: strain

value: CD1/Gf1+ (Thambyrajah et al 2015)

value: 6-8 weeks old

tag: developmental stage

tag: population

tag: batch


title: Kouiki_I_140610_G06

source_name: Ibaraki, Jun-14,  replicate 2


title: Kouiki_I_140610_G04

source_name: Ibaraki, Jun-14,  replicate 1


title: Kouiki_I_140514_G03

source_name: Ibaraki, May-14,  replicate 2


title: XAUT1-POOL-SCG4

city: San Francisco

first: Michael

middle: George

last: Kattah

state: CA

address: 513 Parnassus Ave

department: Medicine-Gastroenterology

country: USA

institute: UCSF

description:
XAUT1-POOL-SCG4_pool1: XAUT1-HS1-CLR1 XAUT1-HS2-CLL1 XAUT1-HS3-CLR1 XAUT1-HS4-CLL1 XAUT1-HS5-CLR1 XAUT1-HS6-CLL1 XAUT1-HS7-CLR1 XAUT1-HS8-CLL1 XAUT1-HS9-CLR1 XAUT1-HS11-CLR1
biopsy_RNA.rds
biopsy_RNA.h5ad

data_processing:
Briefly, sequencer obtained bcl files were demultiplexed into individual library fastq trios using the mkfastq program from the Cellranger 3.0.2 suite of tools (https://support.10xgenomics.com). Feature-barcode matrices were obtained for each sample by aligning the raw fastqs to GRCh38 reference genome (annotated with Ensembl v85) using the Cellranger count. Raw feature-barcode matrices were loaded into Seurat 3.1.5 and low-quality cells (with fewer than 100 features), and features in 3 or fewer cells were dropped from the dataset. The remaining events were assessed for inter-sample doublet detection (generated due to libraries containing samples pooled prior to loading) using Freemuxlet (https://github.com/statgen/popscle), the genotype-free version of Demuxlet.
Clusters of cells belonging to the same patient were identified via SNP concordance to a “truth set” generated by bulk RNASeq.
Clusters of cells belonging to a unique sample were mapped to patients using their individual Freemuxlet-generated genotype, and ground truth genotypes per patient identified via bulk RNA-seq. The pairwise discordance between inferred and ground-truth genotypes was assessed using the bcftools gtcheck command.
Assembly: GRCh38
Supplementary files format and content: Freemuxlet format: clust1.samples.gz
Supplementary files format and content: Seurat object (.rds) and anndata (.h5ad)

molecule: polyA RNA

source_name: colon biopsy

extract_protocol:
Colon biopsies were obtained with standard cold endoscopic biopsy forceps from two different regions of the colon (R=right/proximal and L=left/distal). Six biopsies were collected in a conical tube with Basal Media (Advanced DMEM/F12 with NEAA and Sodium Pyruvate, Thermo cat. No. 12634-010; 2mM Glutamax, Thermo cat. No. 35050061; 10 mM HEPES (Corning); Penicillin-Streptomycin-Neomycin (PSN) Antibiotic Mixture, Thermo cat. No. 15640055; Normocin 100 μg/mL, Invivogen cat. No. ant-nr-2; 1 mM N-acetylcysteine, Sigma-Aldrich, A9165) with 10 μM Y-27632 (MedChem Express) at 4°C. Biopsies were transferred into cryovials containing freezing media (90% (v/v) FCS, 10% (v/v) DMSO and 10 μM Y-27632). Colon biopsies were thawed for 2min with gentle agitation in a 37 C water bath, transferred to a gentleMACS Tube (Miltenyi Biotec), washed twice with basal media containing 10 μM Y-27632, and then incubated in 2 mL digestion buffer (basal media, 10 μM Y-27632, 600 U/mL Collagenase IV (Worthington cat. No.LS004189), 0.1 mg/mL DNAse I (Sigma-Aldrich, D4513) and digested for 20 min at 37°C in a shaking incubator set at 225 rpm. Subsequently, samples were placed in the gentleMACS Dissociator, processed with the gentleMACS program m_intestine_01, followed by 15 min incubation at 37°C in a shaking incubator set at 225 rpm. The suspension was then strained through a 100 μm strainer (Miltenyi) and centrifuged at 450g for 5 min at RT. Two additional washes were performed in Hanks’ Balanced Salt Solution (HBSS) (Corning) containing 0.1 mg/mL DNAse I (Sigma-Aldrich, D4513). Dead cells were removed with the Dead Cell Removal Kit (Miltenyi) according to the manufacturer’s instructions. Cell suspensions were counted using a TC20 Automated Cell Counter (Bio-Rad) with 0.4% Trypan Blue Solution (Thermo Fisher Scientific). Live cell enriched colon single-cell suspensions were used for tissue scRNA-seq and CITE-seq, with a final pooled viability >75%. Colon single-cell suspensions from each patient were pooled with equivalent number of live cells. Samples from unique individuals were pooled, and samples from the proximal (Right,R) or distal (Left,L) colon of the same individual were placed in separate pools, so each sample could later be uniquely identified using demuxlet.
cDNA library generation using the Chromium Single Cell 3′ v2 Reagent Kits, following the detailed protocol provided by 10x Genomics.

value: colon biopsy

tag: treatment


title: GL78_086_WN-S2-P1_A12_AGM_E10-5_SP_32-33_Gfi1_HE__S86


title: XAUT1-POOL-SCG3

description:
XAUT1-POOL-SCG3_pool1: XAUT1-HS1-CLR1 XAUT1-HS2-CLL1 XAUT1-HS3-CLR1 XAUT1-HS4-CLL1 XAUT1-HS5-CLR1 XAUT1-HS6-CLL1 XAUT1-HS7-CLR1 XAUT1-HS8-CLL1 XAUT1-HS9-CLR1 XAUT1-HS11-CLR1
biopsy_RNA.rds
biopsy_RNA.h5ad


title: Kouiki_I_140514_G05

source_name: Ibaraki, May-14,  replicate 1


title: Control090

description: 3'-biased mRNA


title: GL78_085_WN-S2-P1_J11_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S85

value: Hematopoietic cells


title: Kouiki_I_140712_G05

source_name: Ibaraki, Jul-14,  replicate 1


title: Kouiki_I_140409_G08

source_name: Ibaraki, Apl-14,  replicate 2


title: XAUT1-POOL-SCG2

description:
XAUT1-POOL-SCG2_pool1: XAUT1-HS1-CLR1 XAUT1-HS2-CLL1 XAUT1-HS3-CLR1 XAUT1-HS4-CLL1 XAUT1-HS5-CLR1 XAUT1-HS6-CLL1 XAUT1-HS7-CLR1 XAUT1-HS8-CLL1 XAUT1-HS9-CLR1 XAUT1-HS11-CLR1
biopsy_RNA.rds
biopsy_RNA.h5ad


title: Control089

description: 3'-biased mRNA


title: XAUT1-POOL-SCG1

description:
XAUT1-POOL-SCG1_pool1: XAUT1-HS1-CLR1 XAUT1-HS2-CLL1 XAUT1-HS3-CLR1 XAUT1-HS4-CLL1 XAUT1-HS5-CLR1 XAUT1-HS6-CLL1 XAUT1-HS7-CLR1 XAUT1-HS8-CLL1 XAUT1-HS9-CLR1 XAUT1-HS11-CLR1
biopsy_RNA.rds
biopsy_RNA.h5ad


title: GL78_084_WN-S2-P1_I11_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S84


title: Control088

description: 3'-biased mRNA


title: GL78_083_WN-S2-P1_H11_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S83


title: GL78_082_WN-S2-P1_G11_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S82


title: Control087

description: 3'-biased mRNA


title: Control086

description: 3'-biased mRNA


title: LN5, scTCRseq

city: Chicago

first: Xiufen

last: Chen

state: IL

address: 900 E. 57th street KCBD 6230A

institute: University of Chicago

description: 10x Genomics

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were made using the CellRanger software v3.1.0 (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger)
Supplementary files format and content: GRCh38
Supplementary files format and content: Tab-separated values files and matrix files

source_name: Lymphoid tissue

extract_protocol:
Fresh lymph node biopsies from patients with newly diagnosed (13) or relapsed/refractory (1) cHL, as well as reactive lymphoid tissues (5) from patients were utilized in this study. To prepare cHL tissues for downstream analyses, they were cut into small pieces and incubated in 1 mg/ml collagenase IV (Sigma-Aldrich) and 20 mg/ml DNAse I (Roche) at 37 °C for 30 min and then passed through a 100 micron filter to generate single cell suspensions, which were frozen for future analysis.
B-cell depletion of each sample was achieved using CD19-biotin and anti-biotin microbeads (Miltenyi) and negative selection to enrich T cell populations. A total of 8,000-10,000  cells per sample were processed using a Chromium Next GEM Single Cell 5’ Kit v1.1 to generate cDNA libraries, and sequencing was performed using the 10x Genomics Chromium Single Cell 5’ Gene Expression workflow for Single Cell RNA Sequencing. Single-cell libraries from four samples were pooled and sequenced on one NovaSEQ SP 100 base PET lane to achieve an average sequencing depth of 2,981 reads per cell.

value: Lymphoid tissue

value: Leukocyte

value: reactive lymph tissue


title: LN5, scRNAseq

description: 10x Genomics


title: Kouiki_I_140409_G07

source_name: Ibaraki, Apl-14,  replicate 1


title: GL78_081_WN-S2-P1_F11_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S81


title: LN4, scTCRseq

description: 10x Genomics


title: Control091

description: 3'-biased mRNA


title: Kouiki_I_140311_G06

source_name: Ibaraki, Mar-14,  replicate 2


title: Control085

description: 3'-biased mRNA


title: LN4, scRNAseq

description: 10x Genomics


title: GL78_079_WN-S2-P1_C11_AGM_E10-5_SP_32-33_Gfi1_HE__S79


title: LN3, scTCRseq

description: 10x Genomics


title: Control083

description: 3'-biased mRNA


title: GL78_077_WN-S2-P1_K10_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S77


title: Kouiki_I_140311_G05

source_name: Ibaraki, Mar-14,  replicate 1


title: Control082

description: 3'-biased mRNA


title: Kouiki_I_140218_G03

source_name: Ibaraki, Feb-14,  replicate 1


title: Kouiki_I_140218_G04

source_name: Ibaraki, Feb-14,  replicate 2


title: Kouiki_Y_150210_G30

source_name: Yamagata, Feb-15,  replicate 2

value: Yamagata


title: GL78_076_WN-S2-P1_J10_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S76


title: Kouiki_Y_150210_G26

source_name: Yamagata, Feb-15,  replicate 1


title: LN3, scRNAseq

description: 10x Genomics


title: LN2, scTCRseq

description: 10x Genomics


title: GL78_075_WN-S2-P1_I10_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S75


title: Control081

description: 3'-biased mRNA

source_name: Whole Blood


title: Kouiki_Y_150108_G27

source_name: Yamagata, Jan-15,  replicate 2


title: LN2, scRNAseq

description: 10x Genomics


title: Control080

description: 3'-biased mRNA


title: Control084

description: 3'-biased mRNA


title: LN1, scTCRseq

description: 10x Genomics


title: Kouiki_Y_150108_G13

source_name: Yamagata, Jan-15,  replicate 1


title: GL78_078_WN-S2-P1_B11_AGM_E10-5_SP_32-33_Gfi1_HE__S78

source_name: AGM


title: Control079

description: 3'-biased mRNA


title: GL78_073_WN-S2-P1_G10_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S73


title: Kouiki_Y_141211_G30

source_name: Yamagata, Dec-14,  replicate 2


title: Control110

description: 3'-biased mRNA


title: scRNA-seq, Neural organoid, Line FOK4, Day 90 Chr-Veh, Replicate 1

city: Munich

first: Elisabeth

last: Binder

address: Kraepelinstr. 2-10

department: Department Genes and Environment

institute: Max Planck Institute of Psychiatry

description:
Single-cell RNA sequencing (10xGenomics Chromium)
FOK4-D90-Chr-Veh-C1

data_processing:
Filtered count bc matrices were produced from fastq files using 10x Genomics Cell Ranger v3.0.2 with the transcriptome hg38_ensrel94.
Assembly: hg38
Supplementary files format and content: barcodes.tsv: CellRanger output tab-delimited text file listing the barcodes appearing in the matrix.mtx filtered barcodes x features count matrix.
Supplementary files format and content: features.tsv: CellRanger output tab-delimited text file listing the features appearing in the matrix.mtx filtered barcodes x features count matrix.
Supplementary files format and content: matrix.mtx: CellRanger output matrix market file containing the filtered barcodes x features count matrix.

source_name: Unguided neural organoid

growth_protocol: Two primary human induced pluripotent stem cell (hiPSC) lines were used in this study. The first cell line was reprogrammed using hiPSCs from skin fibroblasts (HPS0076:409b2, RIKEN BRC cell bank, female) and is referred to as “Line 409b2” here. The second cell line was reprogrammed using a plasmid-based protocol for integration-free hiPSCs from peripheral blood mononuclear cells from a female donor through the BeCOME study and is referred to as “Line FOK4”here. MTA approvals were obtained for the use of both hiPSC lines. hiPSCs were cultured in Matrigel-coated (1:100 diluted in DMEM-F12 (Gibco™, 31330-038), Corning Incorporated, 354277) Costar® 6-well cell culture plates (Corning Incorporated, 3516) in mTESR1 Basal Medium (STEMCELL Technologies, 85851) supplemented with 1× mTESR1 Supplement (STEMCELL Technologies, 85852) at 37°C with 5 % CO2. Passaging was performed with Gentle Cell Dissociation Reagent (STEMCELL Technologies, 07174). RevitaCell Supplement (1:100 diluted, Gibco™, A2644501) was added for 24 hours after passaging to promote cell survival. Human neural organoids were created as described by Lancaster et al. with some modifications. Briefly, hiPSCs were dissociated in StemPro Accutase Cell Dissociation Reagent (Life Technologies, A1110501). Single cells (n=9000) were dispensed into each well of an Ultra-low attachment 96-well plate with round bottom wells (Corning Incorporated, 7007) in human embryonic stem cell medium (hESC, DMEM/F12-GlutaMAX (Gibco™, 31331-028) with 20 % Knockout Serum Replacement (Gibco™, 10828-028), 3 % FBS (Fetal Bovine Serum, Gibco™, 16141-061), 1 % non-essential amino acids (Gibco™, 11140-035), 0.1 mM 2-mercaptoethanol (Gibco™, 31350-010)) supplemented with 4 ng/ml human recombinant FGF (Fibroblast Growth Factor, Peprotech, 100-18B) and 50 µM Rock inhibitor Y27632 (Millipore, SCM075) for 4 days and in hESC medium without bFGF and Rock inhibitor for an additional 2 days to form embryoid bodies (EBs). On day 6, the medium was changed to neural induction medium (NIM, DMEM/F12 GlutaMAX supplemented with 1:100 N2 supplement (Gibco™, 15502-048), 1 % Non-essential amino acids and 1 µg/ml Heparin (Sigma, H3149)) and cultured for an additional 6 days. On day 12, the EBs were embedded in Matrigel (Corning Incorporated, 354234) drops and transferred to 10-cm cell culture plates (TPP, 93100) in neural differentiation medium without vitamin-A (NDM-A, DMEM/F12GlutaMAX and Neurobasal (Gibco™, 21103-049) in a 1:1 ration, additionally supplemented with 1:100 N2 supplement 1:100 B27 without Vitamin A (Gibco™, 12587-010), 0.5 % non-essential amino acids, insulin 2.5 µg/ml (Gibco™, 19278), 1:100 Antibiotic-Antimycotic (Gibco™, 15240-062) and 50 µM 2-mercaptoethanol) for 4 days. On day 16, Organoids were transferred onto an orbital shaker in NDM+A medium (same composition as NDM-A with the addition of B27 with Vitamin A (Gibco™, 17504-044) in the place of B27 without Vitamin A) and were grown in these conditions at 37°C with 5 % CO2. NDM+A medium was changed twice per week until the organoids were collected for cryopreservation or single-cell dissociation or fixation in paraformaldehyde.

extract_protocol:
Single cells were dissociated using StemPro Accutase Cell Dissociation Reagent (Life Technologies), filtered through 30uM and 20uM filters (Miltenyi Biotec) and cleaned of debris using a Percoll (Sigma, P1644) gradient. Single cells were resuspended in ice-cold Phosphate-Buffered Saline (PBS) supplemented with 0.04 % Bovine Serum Albumin and prepared for single-cell separation.
Experiments were performed in a paired case-control design with 2 or 4 conditions at day 70 and day 90 respectively run in parallel. Single cells were run through the Chromium controller to form gel emulsion beads containing barcoded single cells and prepared into single-cell libraries using the Chromium Single Cell 3’ Reagent Kits v2 according to the manufacturer’s recommendations without any modifications (10x Genomics). To reach an optimal target cell number, 10,000 cells per sample were loaded onto a channel of the 10x chip. All libraries were assessed using a High Sensitivity DNA Analysis Kit for the 2100 Bioanalyzer (Agilent) and KAPA Library Quantification kit for Illumina (KAPA Biosystems). Sequencing of the 10x Genomics single-cell RNA-seq libraries was performed on an Illumina NovaSeq 6000 (Illumina, San Diego, CA) at the sequencing core facility of the Max Planck Institute for Molecular Genetics (Berlin, Germany).

treatment_protocol: Organoids were treated with glucocorticoids by dissolving Dexamethasone (Dex) in DMSO (dimethyl sulfoxide) and subsequently in the NDM+A culture medium. To achieve the final concentration of 100nM, Dex was first diluted in DMSO in a concentration of 100μM and subsequently diluted in NDM+A culture medium to a final concentration of 100nM. Vehicle control (Veh) organoids received equal amounts of DMSO. Chronic exposures (days 60-70 in culture) were performed by replacing supplemented media every two days. Some organoids were collected at day 70 following 10 days of exposure. Other organoids from the same batch were subsequently cultured in normal unsupplemented media for a 20-day wash-out period followed by an acute exposure (day 90 in culture) with 100nM Dex or DMSO for 12 hours.

value: Unguided neural organoid

tag: cell line

tag: age (days)

tag: biological replicate_number

value: 100nM Dexamethasone in DMSO for 10 days (day 60 to day 70) + DMSO for 12 hours before collection at D90


title: Control078

description: 3'-biased mRNA


title: LN1, scRNAseq

description: 10x Genomics


title: Control077

description: 3'-biased mRNA


title: GL78_071_WN-S2-P1_E10_AGM_E10-5_SP_32-33_Gfi1_HE__S71


title: Kouiki_Y_141211_G28

source_name: Yamagata, Dec-14,  replicate 1


title: HL14, scTCRseq

description: 10x Genomics

value: Hodgkin lymphoma


title: Control076

description: 3'-biased mRNA


title: GL78_070_WN-S2-P1_C10_AGM_E10-5_SP_32-33_Gfi1_HE__S70


title: HL14, scRNAseq

description: 10x Genomics


title: GL78_074_WN-S2-P1_H10_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S74


title: HL13, scTCRseq

description: 10x Genomics


title: Control075

description: 3'-biased mRNA


title: GL78_069_WN-S2-P1_B10_AGM_E10-5_SP_32-33_Gfi1_HE__S69


title: Kouiki_Y_141110_G26

source_name: Yamagata, Nov-14,  replicate 1


title: Kouiki_Y_141110_G27

source_name: Yamagata, Nov-14,  replicate 2


title: HL13, scRNAseq

description: 10x Genomics

source_name: Lymphoid tissue


title: Control074

description: 3'-biased mRNA


title: Kouiki_Y_141014_G13

source_name: Yamagata, Oct-14,  replicate 1


title: HL12, scTCRseq

description: 10x Genomics


title: Control073

description: 3'-biased mRNA


title: GL78_068_WN-S2-P1_A10_AGM_E10-5_SP_32-33_Gfi1_HE__S68


title: Kouiki_Y_141014_G28

source_name: Yamagata, Oct-14,  replicate 2


title: GL78_067_WN-S2-P1_K9_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S67


title: GL78_066_WN-S2-P1_I9_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S66


title: Kouiki_Y_140910_G26

source_name: Yamagata, Sep-14,  replicate 1


title: Control072

description: 3'-biased mRNA


title: Control071

description: 3'-biased mRNA


title: GL78_072_WN-S2-P1_F10_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S72


title: Kouiki_Y_140910_G30

source_name: Yamagata, Sep-14,  replicate 2


title: HL12, scRNAseq

description: 10x Genomics


title: GL78_065_WN-S2-P1_F9_AGM_E10-5_SP_32-33_Gfi1_HE__S65


title: Control070

description: 3'-biased mRNA


title: GL78_064_WN-S2-P1_E9_AGM_E10-5_SP_32-33_Gfi1_HE__S64


title: Kouiki_Y_140808_G28

source_name: Yamagata, Aug-14,  replicate 2


title: HL10, scTCRseq

description: 10x Genomics


title: Control069

description: 3'-biased mRNA


title: HL11, scRNAseq

description: 10x Genomics


title: Kouiki_Y_140808_G27

source_name: Yamagata, Aug-14,  replicate 1


title: Kouiki_Y_140710_G13

source_name: Yamagata, Jul-14,  replicate 1


title: GL78_063_WN-S2-P1_D9_AGM_E10-5_SP_32-33_Gfi1_HE__S63


title: Control068

description: 3'-biased mRNA

source_name: Whole Blood


title: GL78_062_WN-S2-P1_C9_AGM_E10-5_SP_32-33_Gfi1_HE__S62


title: Kouiki_Y_140710_G30

source_name: Yamagata, Jul-14,  replicate 2


title: HL10, scRNAseq

description: 10x Genomics


title: HL9, scTCRseq

description: 10x Genomics


title: Control067

description: 3'-biased mRNA


title: HL11, scTCRseq

description: 10x Genomics


title: GL78_061_WN-S2-P1_A9_AGM_E10-5_SP_32-33_Gfi1_HE__S61


title: Control066

description: 3'-biased mRNA


title: Kouiki_Y_140609_G28

source_name: Yamagata, Jun-14,  replicate 2


title: GL78_060_WN-S2-P1_I8_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S60

source_name: AGM


title: Control065

description: 3'-biased mRNA


title: Kouiki_Y_140512_G27

source_name: Yamagata, May-14,  replicate 2


title: GL78_059_WN-S2-P1_H8_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S59


title: Kouiki_Y_140609_G26

source_name: Yamagata, Jun-14,  replicate 1


title: Kouiki_Y_140512_G13

source_name: Yamagata, May-14,  replicate 1


title: GL78_058_WN-S2-P1_F8_AGM_E10-5_SP_32-33_Gfi1_HE__S58


title: Control064

description: 3'-biased mRNA


title: HL8, scRNAseq

description: 10x Genomics


title: GL78_057_WN-S2-P1_E8_AGM_E10-5_SP_32-33_Gfi1_HE__S57


title: Control063

description: 3'-biased mRNA


title: Kouiki_Y_140410_G30

source_name: Yamagata, Apl-14,  replicate 2


title: HL7, scRNAseq

description: 10x Genomics


title: Kouiki_Y_140410_G28

source_name: Yamagata, Apl-14,  replicate 1


title: Control062

description: 3'-biased mRNA


title: GL78_056_WN-S2-P1_D8_AGM_E10-5_SP_32-33_Gfi1_HE__S56


title: HL9, scRNAseq

description: 10x Genomics

source_name: Lymphoid tissue


title: HL6, scTCRseq

description: 10x Genomics


title: GL78_055_WN-S2-P1_C8_AGM_E10-5_SP_32-33_Gfi1_HE__S55


title: HL8, scTCRseq

description: 10x Genomics


title: HL6, scRNAseq

description: 10x Genomics


title: Control061

description: 3'-biased mRNA


title: GL78_054_WN-S2-P1_B8_AGM_E10-5_SP_32-33_Gfi1_HE__S54


title: HL5, scTCRseq

description: 10x Genomics


title: Control060

description: 3'-biased mRNA


title: Control059

description: 3'-biased mRNA


title: GL78_053_WN-S2-P1_A8_AGM_E10-5_SP_32-33_Gfi1_HE__S53


title: 3BKO_T73BDN_chip_rep_1_ChIPseq

city: Edinburgh

first: Duncan

last: Sproul

state: Mid Lothian

address: Crewe Road South

department: MRC Human Genetics Unit

country: United Kingdom

institute: University of Edinburgh

description: 3BKO_T73BDN_norm_mean_scaled_ChIPseq.bigWig

library_source: genomic

data_processing:
All ChIP-seq experiments were processed previously described (Masalmeh et al., 2021, PMID: 33514701). Read quality was checked using FASTQC (v0.11.4, https://www.bioinformatics.babraham.ac.uk/projects/fastqc), with low quality reads and adaptors removed using TrimGalore with default settings (v0.4.1). Reads were aligned to hg38 using bowtie 2 (v2.3.1, with settings: -N 1 -L 20 --no-unal) (Langmead and Salzberg, 2012, PMID: 22388286). For paired end data, additional settings were used during alignment to remove discordant reads: --no-mixed –nodiscordant -X 1000. ChIP-Rx-seq reads were aligned to a combination of hg38 and dm6 genomes. Multimapping reads excluded using SAMtools (v1.6, with settings: -bq 10) (Li et al., 2009) and PCR duplicates excluded using SAMBAMBA (v0.5.9) (Tarasov et al., 2015, PMID: 25697820). For T7 ChIP-Rx-seq UMIs were used for de-duplication instead of SAMBAMBA. We first extracted UMIs from the appropriate read and placed them in the read FASTQ headers using UMI tools extract (v1.0.0 and setting: --bc-pattern=NNNNNNNNNNN). Then deduplication was performed using the UMI tools dedup function (v1.0.0 and setting: --paired). Tracks for visualisation of histone ChIP-seq were generated using Deeptools (v3.2.0) (Ramirez et al., 2016, PMID: 27079975). Counts per million normalised tracks were generated using the bamCoverage function (settings: --normalizeUsing CPM) with the default bin size of 50 bp. The mean of replicate tracks and normalisation over the input was calculated using the bigwigCompare function (settings: --operation mean). For the single-ended H3K36me3 ChIP-seq, the estimated fragment length of 150 bp was used. For the other paired-end experiments the actual fragment size was used.
Assembly: hg38
Supplementary files format and content: Processed data files are bigWig files generated by normalisation over input and from the mean of the two replicates as described under data processing.

molecule: genomic DNA

source_name: HCT116 DNMT3B KO

growth_protocol: HCT116 and DNMT3B cells were gifts from B. Vogelstein (Rhee et al 2002 PMID: 11932749). Cells were cultured in McCoy’s 5A medium (Gibco) supplemented with 10% fetal calf serum (Life technologies) and penicillin-streptomycin antibiotics at 140 and 400 µg/ml respectively.

extract_protocol:
ChIP-seq was performed as previously described (Masalmeh et al., 2021, PMID: 33514701). For T7-DNMT3B ChIP-Rx-seq experiments, 1×107 cells were harvested, washed and crosslinked with 1% methanol-free formaldehyde in PBS for 8 min at room temperature. Crosslinked cells were lysed for 10 min on ice in 50 μl of lysis buffer (50 mM Tris-HCl pH 8, 150 mM NaCl, 1 mM EDTA, 1% SDS) freshly supplemented with proteinase inhibitor (Sigma-Aldrich). IP dilution buffer (20 mM Tris-HCl pH 8, 150 mM NaCl, 1 mM EDTA, 0.1% Triton X-100) freshly supplemented with proteinase inhibitor, DTT and PMSF was added to the samples to reach a final volume of 500 μl. As prolonged sonication caused T7-DNMT3B degradation, chromatin was fragmented using Benzonase (Pchelintsev et al., 2016, PMID: 26821228): samples were sonicated on ice with Soniprep 150 twice for 30 s to break up nuclei; then 200 U of Benzonase Nuclease (Sigma) and MgCl2 (final concentration 2.5 mM) were added and samples were incubated on ice for 15 min. The reaction was blocked by adding 10 μl of 0.5 M EDTA pH 8. Following centrifugation for 30 min at 18,407 g at 4 °C, supernatants were collected and supplemented with Triton X-100 (final concentration 1%) and 5% input aliquots were retained for later use. Protein A Dynabeads (Invitrogen) previously coupled with 10 μl of T7-Tag antibody per 1×107 cells in blocking solution (1x PBS, 0.5% BSA) were added and the samples incubated overnight under rotation at 4 °C. Beads were then washed for 10 min at 4 °C with the following buffers: IP dilution buffer 1% Triton X-100 (20 mM Tris-HCl pH 8, 150 mM NaCl, 2 mM EDTA, 1% Triton X-100), buffer A (50mM HEPES pH 7.9, 500 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate, 0.1% SDS), buffer B (20 mM Tris pH 8, 1 mM EDTA, 250 mM LiCl, 0.5% NP-40, 0.5% Na-deoxycholate), TE buffer (1mM EDTA pH 8, 10 mM Tris pH 8). Chromatin was eluted by incubating the beads in extraction buffer (0.1 M NaHCO3, 1% SDS) for 15 min at 37 °C. To reverse the cross-linking Tris-HCl pH 6.8 and NaCl were added to final concentrations of 130 mM and 300 mM respectively, and immunoprecipitations were incubated at 65 °C overnight. Samples were then incubated at 37 °C for 1 h after addition of 2 μl of RNase Cocktail Enzyme Mix (Ambion). Then 40 μg of Proteinase K (Roche) were added, followed by 2 h incubation at 55 °C. Input material was similarly de-crosslinked. Samples were purified with the MinElute PCR purification kit (QIAGEN). For ChIP-Rx-seq of endogenous T7-DNMT3B and ectopic T7-DNMT3B expressed in DNMT3B KO cells, 20 μg of Spike-in chromatin (ActiveMotif 53083) was added to each sample after sonication. 2 μl of spike-in antibody per sample (ActiveMotif 61686) was also added in a ratio 1:5 versus the T7 antibody. A similar protocol was used for H3K4me3, H3K9me3, H3K27me3 and H3K36me3 ChIP-seq experiments, except: 0.5×107 cells were harvested and crosslinked with 1% methanol-free formaldehyde in PBS for 5 min at room temperature. For H3K36me3 ChIP-Rx-seq crosslinked Drosophila S2 cells were spiked into samples before sonication at a ratio of 20:1 human to Drosophila cells. Following nuclei rupture by sonication on ice with Soniprep 150, chromatin was fragmented using Bioruptor Plus sonicator (Diagenode) for 40 cycles (30 s on/30 s off on high setting at 4 °C). 2 μl /1×106 cells of the following antibodies were used for immunoprecipitations: H3K4me3 (EpiCypher 13-00041), H3K9me3 (Active Motif 39161), H3K27me3 (Cell Signaling Technology C36B11) and H3K36me3 (Abcam ab9050).
Libraries were prepared using the NEBNext Ultra II DNA Library Prep Kit for Illumina (E7645) according to the manufacturer instructions. NEBNext Multiplex Oligos for Illumina (NEB) barcode adapters were used. Specifically, Illumina Index Primers Set 1 (E7335) for endogenous T7-DNMT3B and H3K36me3, and Unique Dual Index UMI Adaptors DNA Set 1 (E7395) for the other libraries.  For histone modifications ChIP-seq, adapter-ligated DNA was size selected for an insert size of 150 bp using Agencourt AMPure XP beads. Libraries were quantified using the Qubit dsDNA HS or BR assay kit and assessed for size and quality using the Agilent Bioanalyser. Endogenous T7-DNMT3B and H3K36me3 ChIP-Rx-seq libraries were sequenced using the NextSeq 500/550 high-output version 2.5 kit (75 bp paired end reads, DNMT3B or 75 bp single end reads, H3K36me3). The other ChIP-seq libraries were sequenced using the NextSeq 2000 P3 (50 bp paired end reads). Libraries were combined into equimolar pools to run within individual flow cells. Sequencing was performed by the Edinburgh Clinical Research Facility.

value: HCT116 DNMT3B KO

value: colorectal cancer cell line

tag: chip antibody

value: T7-Tag (D9E1X, Cell Signaling)


title: Kouiki_Y_140310_G13

source_name: Yamagata, Mar-14,  replicate 1

value: shoot


title: Control058

description: 3'-biased mRNA

value: Control


title: Kouiki_Y_140213_G29

source_name: Yamagata, Feb-14,  replicate 2


title: 3BKO_T73BDN_chip_rep_2_ChIPseq


title: GL78_052_WN-S2-P1_J7_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S52

value: AGM


title: 3BKO_T73BDN_input_rep_2_ChIPseq


title: Control057

description: 3'-biased mRNA


title: HL7, scTCRseq

description: 10x Genomics


title: HL4, scTCRseq

description: 10x Genomics


title: HL5, scRNAseq

description: 10x Genomics


title: Control056

description: 3'-biased mRNA


title: HL4, scRNAseq

description: 10x Genomics


title: Kouiki_Y_140213_G27

source_name: Yamagata, Feb-14,  replicate 1


title: 3BKO_T73BDN_input_rep_1_ChIPseq


title: Kouiki_Y_140310_G26

source_name: Yamagata, Mar-14,  replicate 2


title: HL3, scTCRseq

description: 10x Genomics


title: 3BKO_T73BW263A_chip_rep_2_ChIPseq

description: 3BKO_T73BW263A_norm_mean_scaled_ChIPseq.bigWig


title: GL78_050_WN-S2-P1_G7_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S50

value: CD1/Gf1+ (Thambyrajah et al 2015)


title: Control054

description: 3'-biased mRNA


title: GL78_048_WN-S2-P1_D7_AGM_E10-5_SP_32-33_Gfi1_HE__S48

value: Hemogenic endothelium cells

value: 6-8 weeks old


title: Control055

description: 3'-biased mRNA


title: Control053

description: 3'-biased mRNA


title: HL2, scTCRseq

description: 10x Genomics


title: GL78_049_WN-S2-P1_E7_AGM_E10-5_SP_32-33_Gfi1_HE__S49


title: 3BKO_T73BW263A_chip_rep_1_ChIPseq


title: 3BKO_T73BW263A_input_rep_2_ChIPseq


title: HL3, scRNAseq

description: 10x Genomics


title: GL78_047_WN-S2-P1_B7_AGM_E10-5_SP_32-33_Gfi1_HE__S47


title: GL78_046_WN-S2-P1_A7_AGM_E10-5_SP_32-33_Gfi1_HE__S46


title: Control052

description: 3'-biased mRNA


title: HL2, scRNAseq

description: 10x Genomics


title: GL78_051_WN-S2-P1_H7_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S51

value: Hematopoietic cells


title: HL1, scTCRseq

description: 10x Genomics

value: Lymphoid tissue


title: 3BKO_T73B_chip_rep_2_ChIPseq

description: 3BKO_T73B_norm_mean_scaled_ChIPseq.bigWig


title: 3BKO_T73BW263A_input_rep_1_ChIPseq


title: Control051

description: 3'-biased mRNA


title: 3BKO_T73B_chip_rep_1_ChIPseq


title: GL78_044_WN-S2-P1_G6_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S44


title: HL1, scRNAseq

description: 10x Genomics

value: Leukocyte


title: Control050

description: 3'-biased mRNA


title: 3BKO_T73B_input_rep_2_ChIPseq


title: Control049

description: 3'-biased mRNA


title: DNMT3B_KO_rep_3_RNA_seq

data_processing:
RNA-seq reads were aligned to the genome using the nf-core/rnaseq (v3.12.0, doi: https://doi.org/10.5281/zenodo.1400710) workflow from the nf-core collection of workflows (Ewels et al., 2020, PMID: 32055031). Briefly, the pipeline was executed with Nextflow v23.04.2 (Di Tommaso et al., 2017, PMID: 28398311) and aligned reads to the hg38 genome using the STAR aligner (Dobin et al., 2013). Reads aligning completely within ENSEMBL (version 103/GENCODE 37) transcripts were then counted using Salmon (Patro et al., 2017, PMID: 23104886).
Assembly: hg38
Supplementary files format and content: Length scaled merged gene counts (columns = ENSEMBL Gene ID, Gene Symbol, scaled gene count)

source_name: HCT116 with knocked out DNMT3B

extract_protocol:
RNA extraction was performed using RNeasy Plus Mini Kit (QIAGEN) following the manufacturer’s instructions. Quantity and quality of RNA samples were assessed using the Nanodrop spectrophotometer (Nanodrop ND-1000, Thermo Scientific) as well as the Qubit fluorometer (Invitrogen) with the Qubit High sensitivity RNA reagents. In addition, size distribution of RNA fragments and the RNA integrity number (RIN) value were determined by using Agilent 2100 BioAnalyzer with the RNA nano chip.
RNA samples for whole genome sequencing were send to Edinburgh Genomics for Library preparation and Sequence data generation. Library preparation was performed using the TruSeq stranded mRNA-seq library kit (Illumina). The libraries were then sequenced on NovaSeq with 50 paired end (PE) reads and aiming for at least 375M reads per sample.

tag: cell line_background

tag: genotype

value: DNMT3B KO


title: Control048

description: 3'-biased mRNA

tag: disease

tag: age


title: GL78_043_WN-S2-P1_F6_AGM_E10-5_SP_32-33_Gfi1_HE__S43

tag: tissue


title: DNMT3B_KO_rep_2_RNA_seq


title: DNMT3B_KO_rep_1_RNA_seq


title: HCT116_rep_3_RNA_seq

source_name: HCT116 cells

value: WT


title: 3BKO_T73B_input_rep_1_ChIPseq


title: GL78_042_WN-S2-P1_E6_AGM_E10-5_SP_32-33_Gfi1_HE__S42


title: Control047

description: 3'-biased mRNA

source_name: Whole Blood

tag: sequencing plate number (used for batch correction)


title: Control046

description: 3'-biased mRNA


title: 3BKO_mock_chip_rep_1_ChIPseq

description: 3BKO_mock_norm_mean_scaled_ChIPseq.bigWig


title: 3BKO_mock_chip_rep_2_ChIPseq


title: HCT116_rep_2_RNA_seq


title: Control045

description: 3'-biased mRNA


title: 3BKO_mock_input_rep_2_ChIPseq


title: Control044

description: 3'-biased mRNA


title: GL78_040_WN-S2-P1_C6_AGM_E10-5_SP_32-33_Gfi1_HE__S40


title: GL78_038_WN-S2-P1_A6_AGM_E10-5_SP_32-33_Gfi1_HE__S38


title: 3BKO_mock_input_rep_1_ChIPseq


title: GL78_045_WN-S2-P1_K6_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S45


title: GL78_037_WN-S2-P1_K5_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S37


title: GL78_039_WN-S2-P1_B6_AGM_E10-5_SP_32-33_Gfi1_HE__S39


title: Nuclei from s_0165 sample

city: Leuven

first: Araks

last: Martirosyan

address: Herestraat 49

department: Center for Brain & Disease Research

country: Belgium

institute: VIB-KU Leuven

description: 10X Genomics

data_processing:
Gene counts were obtained by aligning reads to the human hg38 genome (GRCh38/Ensemble 93 pre-built by 10x) using CellRanger software (v.3.0.2) (10x Genomics).
To account for unspliced nuclear transcripts, reads mapping to pre-mRNA were counted (as recommended by CellRanger v.3.0.2).
The quantification of pre-mRNA was done using the CellRanger count pipeline on each of the 29 individual libraries, providing double fastq files as an input for the libraries that were re-sequenced. Default CellRanger parameters have been used throughout the pipeline.
At a first filtering step, thresholds to exclude degrading nuclei with low RNA-content (< 500 transcripts) and doublets having high RNA-content (> 6000 transcripts) were imposed. MALAT1 was excluded from the transcript list since it was highly enriched and could drive the clustering.
Next, to make sure only singlets were left in the database, the Scrublet algorithm [61] with default parameters was run on each donor’s dataset, using default parameters, to further identify doublets. In total 2,496 doublets were identified (Supplementary Table 2) and removed from the database at this point.
finally, nuclei having more than 2% mitochondrial content were removed, leaving 83,484 nuclei expressing 30,194 unique transcripts for downstream analysis.
Assembly: hg38 genome (GRCh38/Ensemble 93 pre-built by 10x)
Supplementary files format and content: Tab-separated values files and matrix files

source_name: Brain

extract_protocol:
To identify the substantia nigra in Control and PD tissues, cryostat sections of 5μm thickness were cut, haematoxylin and eosin (H&E) stained and viewed under the microscope by a neuropathologist (DRT) to identify region populated with melanin containing neurons. Based on this observation, the substantia nigra pars compacta region was manually annotated with a waterproof pen. H&E-guided 2 mm biopsy punches were collected from the SNpc of selected samples and nuclei were isolated to prepare the sequencing libraries.
Libraries were prepared using the Chromium Single Cell 3 Reagent Kits v.3 according to the manufacturer’s protocol (10x Genomics).

value: Brain

tag: clinical diagnosis

tag: Sex

value: male

tag: pmi hours

tag: rin measure


title: Control043

description: 3'-biased mRNA


title: Nuclei from s_0159 sample

description: 10X Genomics

value: female


title: Control042

description: 3'-biased mRNA


title: HCT116_T73B_chip_rep_2_ChIPseq

tag: cell type


title: HCT116_rep_1_RNA_seq


title: GL78_041_WN-S2-P1_D6_AGM_E10-5_SP_32-33_Gfi1_HE__S41

tag: strain

tag: developmental stage

tag: population

tag: batch


title: HCT116_T73B_input_rep_2_ChIPseq

tag: cell line


title: Nuclei from s_0158 sample

description: 10X Genomics


title: GL78_035_WN-S2-P1_H5_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S35


title: Control040

description: 3'-biased mRNA


title: GL78_034_WN-S2-P1_G5_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S34


title: Nuclei from s_0154 sample

description: 10X Genomics


title: HCT116_T73B_input_rep_1_ChIPseq

tag: chip antibody


title: HCT116_mock_chip_rep_2_ChIPseq


title: Nuclei from s_0153 sample

description: 10X Genomics


title: GL78_033_WN-S2-P1_F5_AGM_E10-5_SP_32-33_Gfi1_HE__S33


title: HCT116_T73B_chip_rep_1_ChIPseq


title: Control038

description: 3'-biased mRNA


title: GL78_032_WN-S2-P1_B5_AGM_E10-5_SP_32-33_Gfi1_HE__S32


title: Control037

description: 3'-biased mRNA


title: HCT116_mock_input_rep_2_ChIPseq


title: Nuclei from s_0151 sample

description: 10X Genomics


title: GL78_031_WN-S2-P1_K4_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S31


title: GL78_030_WN-S2-P1_J4_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S30


title: 3BKO_k36me3_chip_rep_2_ChIPseq

tag: cell line_background

tag: treatment

value: H3K36me3 (Abcam ab9050)


title: Control041

description: 3'-biased mRNA

tag: disease


title: Nuclei from s_0147 sample

description: 10X Genomics

tag: age


title: HCT116_mock_input_rep_1_ChIPseq

value: colorectal cancer cell line


title: Control035

description: 3'-biased mRNA


title: Nuclei from s_0142 sample

description: 10X Genomics

tag: tissue


title: GL78_036_WN-S2-P1_I5_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S36


title: GL78_029_WN-S2-P1_I4_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S29


title: Nuclei from s_0152 sample

description: 10X Genomics


title: Control034

description: 3'-biased mRNA

tag: sequencing plate number (used for batch correction)


title: GL78_028_WN-S2-P1_H4_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S28


title: Nuclei from s_0131 sample

description: 10X Genomics

tag: clinical diagnosis


title: Nuclei from s_0130 sample

description: 10X Genomics

tag: Sex


title: Control039

description: 3'-biased mRNA


title: Control033

description: 3'-biased mRNA


title: 3BKO_k36me3_input_rep_2_ChIPseq

tag: genotype

value: none (input)


title: GL78_027_WN-S2-P1_G4_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S27


title: Control032

description: 3'-biased mRNA


title: GL78_026_WN-S2-P1_F4_AGM_E10-5_SP_32-33_Gfi1_HE__S26


title: HCT116_mock_chip_rep_1_ChIPseq

value: T7-Tag (D9E1X, Cell Signaling)


title: 3BKO_k36me3_chip_rep_1_ChIPseq


title: Nuclei from s_0128 sample

description: 10X Genomics

value: Control

tag: pmi hours

tag: rin measure


title: Control031

description: 3'-biased mRNA


title: GL78_025_WN-S2-P1_E4_AGM_E10-5_SP_32-33_Gfi1_HE__S25

tag: cell type

value: AGM

tag: strain


title: Control030

description: 3'-biased mRNA


title: Control036

description: 3'-biased mRNA


title: Nuclei from s_0120 sample

description: 10X Genomics

value: Parkinson's

tag: lewy bodies_presence_in_midbrain

tag: cerad score_for_neuritic_plaques

tag: braak stage_for_neurofibrillary_tangles


title: GL78_023_WN-S2-P1_B4_AGM_E10-5_SP_32-33_Gfi1_HE__S23

value: Hemogenic endothelium cells

value: CD1/Gf1+ (Thambyrajah et al 2015)

tag: developmental stage

tag: population

tag: batch


title: GL78_024_WN-S2-P1_D4_AGM_E10-5_SP_32-33_Gfi1_HE__S24

value: 6-8 weeks old


title: Control029

description: 3'-biased mRNA

tag: disease


title: DNMT3B_KO_DNMT3B_deltaN_rep_2_exp_4_WGBS

data_processing:
Whole Genome Bisulfite Sequencing (WGBS):
Sequencing quality was assessed with FASTQC (v0.11.4). Low quality reads and remaining adaptors were removed using TrimGalore (v0.4.1, Settings: --adapter AGATCGGAAGAGC --adapter2 AAATCAAAAAAAC). The paired end reads were then aligned to the hg38 genome using Bismark (v 0.18.1 with Bowtie2 v2.3.1 and settings: -N 0 -L 20, trimming settings --clip_r1 10 --clip_r2 13 --three_prime_clip_r1 1 --three_prime_clip_r2 1) (Langmead and Salzberg, 2012, PMID: 22388286, Krueger and Andrews, 2011, PMID: 21493656 ) before PCR duplicates were identified and removed using Bismark’s deduplicate_bismark command. Aligned BAM files were processed to report coverage and number of methylated reads for each CpG observed. Forward and reverse strands were combined using Bismark’s methylation extractor and bismark2bedgraph modules with custom Python and AWK scripts. Processed WGBS files were assessed for conversion efficiency based on the proportion of methylated reads mapping to the phage-λ genome spike-in (>99.5% in all cases). BigWigs for visualisation of WGBS data were generated using the coverage in 2.5 kb-sized windows across the genome. These were defined using BEDtools (v2.27.1) (Quinlan and Hall, 2010, PMID: 20110278) and coverage and converted to bigWigs using UCSC tools bedGraphToBigWig (v326). Windows with a total coverage <10 were excluded before conversion to bigWigs.
Assembly: hg38
Supplementary files format and content: Processed coverage values for each CpG observed in sample (.cov files). CpG position and total coverage (T) and methylated coverage (M) combining both strands. Columns = chr, start, end, T, M. Positions are 0-based, ie BED format.
Supplementary files format and content: bigWig files for each sample also supplied. Mean methylation in 2.5Kb windows across genome. Only windows with coverage >= 10 in that sample are in each file.

source_name: HCT116 with knocked out DNMT3B expressing DNMT3B delta N

extract_protocol:
To extract genomic DNA cells were resuspended in genomic lysis buffer (300 mM NaCl, 1% SDS, 20 mM EDTA) and incubated with Proteinase K (Roche) at 55 C overnight. RNA was removed by incubation with RNase A/T1 Cocktail (Ambion) at 37°C for 1 h, in between two phenol-chloroform extraction steps. DNA was quantified by Nanodrop 8000 spectrophotometer and purity assessed by electrophoresis.
100 or 200 ng of purified DNA samples were sheared using the Covaris E220 Evolution Focused Ultrasonicator to create fragments of approximately 350 bp. 0.5 ng of unmethylated phage-λ DNA (NEB) was spiked into each DNA sample prior to shearing to allow assessment of the efficiency of the bisulfite-conversion reaction. 100 ng of each sheared DNA sample was then processed using the EZ DNA Methylation Gold Kit or EZ DNA Methylation-Lightning Kit (Zymo Research) according to the manufacturer’s protocol to create bisulfite-converted single-stranded (BC-ss) DNA. For WGBS experiment 1, the TruSeq DNA Methylation Kit (Illumina Inc.) was used.  Bisfulfite converted single-stranded DNA was randomly primed with a polymerase able to read uracil nucleotides to synthesise DNA strands containing a sequence specific tag. 3’ ends of the newly synthesized DNA strands were then selectively tagged with a second specific sequence tag to produce di-tagged DNA molecules with known sequence tags at 5’ and 3’ ends. Di-tagged molecules were then enriched by PCR with unique indexed PCR primers to provide dsDNA libraries with Illumina sequencing adapters that could be multiplexed and sequenced on a single flow cell. For all other WGBS experiments the Accel-NGS Methyl-Seq DNA Library Kit (Swift BioSciences) was used. For the Swift kit, bisfulfite converted single-stranded DNA was first denatured before simultaneous tailing and ligation of truncated adapters to 3’ ends with Adaptase. An extension step then incorporates the truncated adapter 1 by a primer extension reaction, before a ligation step adds truncated adapter 2 to the bottom strand only. 9 cycles of PCR were used to increase yield and incorporate full-length adapters for unique dual-indexed sequencing. Finally, Agencourt AMPure XP beads (Beckman Coulter) were used to remove oligonucleotides and small fragments and change enzymatic buffer composition. Libraries were quantified using the Qubit dsDNA HS assay kit, assessed for size and quality using the Agilent Bioanalyser with the DNA HS Kit and combined in a single equimolar pool. Sequencing was performed on a P2 flow cell on the Illumina NextSeq 2000 platform using NextSeq 1000/2000 P2 Reagents v3 kit (200 Cycles), or on the NextSeq 550 platform using NextSeq 500/550 High-Output v2 kit (150 cycles). PhiX Control v3 Library was spiked in at a concentration of ~5% to increase library diversity for sequencing. Library preparation and sequencing was performed by the Edinburgh Clinical Research Facility.

treatment_protocol: To express DNMT3B isoforms, HCT116 and DNMT3B KO cells were transfected with FuGENE HD transfection reagent (Promega). For stable integrants, DNMT3 expression constructs were co-transfected with a plasmid expressing piggyBac transposase. After 48 h cells stably expressing DNMT3s were selected with 1 μg/ml puromycin and expanded in the presence of puromycin for 3-4 weeks before being harvested for analysis. Knock-in mutations were introduced by CRISPR editing.

tag: cell line_background

tag: treatment

value: Expressing DNMT3B deltaN


title: Nuclei from s_0118 sample

description: 10X Genomics

tag: tissue

tag: age


title: Control027

description: 3'-biased mRNA

tag: sequencing plate number (used for batch correction)


title: GL78_020_WN-S2-P1_F3_AGM_E10-5_SP_32-33_Gfi1_HE__S20

source_name: AGM


title: Nuclei from s_0116 sample

description: 10X Genomics

tag: clinical diagnosis

tag: Sex


title: Control026

description: 3'-biased mRNA


title: Nuclei from s_0129 sample

description: 10X Genomics

tag: pmi hours

tag: rin measure


title: GL78_021_WN-S2-P1_G3_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S21

tag: cell type

value: Hematopoietic cells

tag: strain


title: DNMT3B_KO_DNMT3B_deltaN_rep_1_exp_4_WGBS

tag: genotype

value: DNMT3B KO


title: Nuclei from s_0111 sample

description: 10X Genomics

tag: lewy bodies_presence_in_midbrain

tag: cerad score_for_neuritic_plaques

tag: braak stage_for_neurofibrillary_tangles


title: Control025

description: 3'-biased mRNA


title: DNMT3B_KO_DNMT3B_rep_1_exp_4_WGBS

source_name: HCT116 with knocked out DNMT3B expressing DNMT3B

value: Expressing DNMT3B


title: GL78_018_WN-S2-P1_D3_AGM_E10-5_SP_32-33_Gfi1_HE__S18

tag: developmental stage

tag: population

tag: batch


title: Control024

description: 3'-biased mRNA

tag: disease


title: DNMT3B_KO_exp_4_WGBS

tag: cell line_background

tag: treatment


title: Nuclei from s_0127 sample

description: 10X Genomics

tag: tissue

value: Brain

tag: age


title: Nuclei from s_0110 sample

description: 10X Genomics

tag: clinical diagnosis

tag: Sex

value: male


title: Control023

description: 3'-biased mRNA

tag: sequencing plate number (used for batch correction)


title: GL78_019_WN-S2-P1_E3_AGM_E10-5_SP_32-33_Gfi1_HE__S19

tag: cell type


title: Nuclei from s_0109 sample

description: 10X Genomics

tag: pmi hours

tag: rin measure


title: 3BKO_k36me3_input_rep_1_ChIPseq

tag: genotype

tag: chip antibody


title: Control028

description: 3'-biased mRNA


title: GL78_017_WN-S2-P1_C3_AGM_E10-5_SP_32-33_Gfi1_HE__S17

tag: strain


title: HCT116_exp_4_WGBS

value: WT


title: GL78_022_WN-S2-P1_H3_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S22

tag: developmental stage


title: GL78_016_WN-S2-P1_B3_AGM_E10-5_SP_32-33_Gfi1_HE__S16

tag: population


title: HCT116_DNMT3B_control_clone_2_exp_3_WGBS

source_name: HCT116 control clone cells unedited by CRISPR


title: Nuclei from s_0119 sample

description: 10X Genomics

tag: lewy bodies_presence_in_midbrain

tag: cerad score_for_neuritic_plaques

tag: braak stage_for_neurofibrillary_tangles


title: Control021

description: 3'-biased mRNA

tag: disease


title: Nuclei from s_0107 sample

description: 10X Genomics

tag: tissue


title: Nuclei from s_0105 sample

description: 10X Genomics

tag: age


title: Control020

description: 3'-biased mRNA


title: DNMT3B_KO_DNMT3B_rep_2_exp_4_WGBS

tag: cell line_background

tag: treatment


title: Nuclei from s_0104 sample

description: 10X Genomics

tag: clinical diagnosis

tag: Sex


title: GL78_015_WN-S2-P1_A3_AGM_E10-5_SP_32-33_Gfi1_HE__S15

tag: cell type

tag: batch


title: HCT116_DNMT3B_control_clone_1_exp_3_WGBS

tag: genotype


title: GL78_014_WN-S2-P1_J2_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S14


title: Control019

description: 3'-biased mRNA

tag: sequencing plate number (used for batch correction)


title: GL78_013_WN-S2-P1_I2_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S13

tag: strain


title: Nuclei from s_0103 sample

description: 10X Genomics

tag: pmi hours

tag: rin measure


title: HCT116_DNMT3B_W263A_clone_3_exp_3_WGBS

source_name: HCT116 cells with DNMT3B W263A mutation knocked in by CRISPR

value: DNMT3B W263A


title: HCT116_DNMT3B_W263A_clone_2_exp_3_WGBS

value: colorectal cancer cell line


title: Control018

description: 3'-biased mRNA


title: Nuclei from s_0102 sample

description: 10X Genomics

value: female

tag: lewy bodies_presence_in_midbrain


title: GL78_012_WN-S2-P1_G2_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S12

tag: developmental stage

tag: population


title: Control017

description: 3'-biased mRNA

tag: disease


title: GL78_011_WN-S2-P1_F2_AGM_E10-5_SP_32-33_Gfi1_HE__S11

tag: tissue


title: Nuclei from s_0100 sample

description: 10X Genomics

tag: age

tag: cerad score_for_neuritic_plaques

tag: braak stage_for_neurofibrillary_tangles


title: HCT116_exp_3_WGBS

tag: cell line_background

tag: treatment


title: Control022

description: 3'-biased mRNA


title: HCT116_DNMT3B_W263A_clone_1_exp_3_WGBS

tag: cell type


title: Control016

description: 3'-biased mRNA


title: Nuclei from s_0098 sample

description: 10X Genomics

tag: clinical diagnosis

tag: Sex


title: Nuclei from s_0096 sample

description: 10X Genomics


title: Control015

description: 3'-biased mRNA

tag: sequencing plate number (used for batch correction)


title: GL78_010_WN-S2-P1_E2_AGM_E10-5_SP_32-33_Gfi1_HE__S10

tag: strain

tag: batch


title: DNMT3B_KO_DNMT3Acd_exp_2_WGBS

source_name: HCT116 with knocked out DNMT3B expressing DNMT3A catalytic dead

tag: genotype

value: Expressing DNMT3A catalytic dead


title: H2195_9d

city: Houston

first: Carl

last: Gay

state: TX

address: 1515 Holcombe Blvd

department: Thoracic-Head & Neck Med Onc

institute: The University of Texas MD Anderson Cancer Center

description: NuGEN Ovation RRBS Methyl-Seq

data_processing:
Seqeuncing reads were trimmed for adapters using TrimGalore-0.6.5
Trimmed reads were mapped to reference genome using bwa
Mapped reads were deduped using NuGEN nudup tool package
Methylation beta values were computed using Bismark-0.22.3
Assembly: hg38
Supplementary files format and content: gzipped tab delimited text file for CpG locations and beta values

growth_protocol: RPMI-1640 with 10% fetal calf serum

extract_protocol:
Qiagen AllPrep DNA/RNA Mini Kit
Briefly, approximately 100ng of genomic DNA was digested using MspI, and Illumina-compatible cytosine-methylated adaptor were ligated to the enzyme-digested DNA. RRBS libraries were then visualized using Bioanalyzer High Sensitivity DNA chips (Agilent, Santa Clara, CA), and those passing QC were subsequently sequenced as 100bp paired-end reads on an Illumina NovaSeq instrument.
NuGEN Ovation RRBS Methyl-Seq

treatment_protocol: drugs were diluted in DMSO and directly added to cell culture medium. Medium and drug was replaced every other day during treatment period.

tag: cell line

value: Small Cell Lung Cancer

value: 2uM cisplatin

tag: time

value: 9 days


title: GL78_009_WN-S2-P1_C2_AGM_E10-5_SP_32-33_Gfi1_HE__S9

value: Hemogenic endothelium cells

value: AGM

tag: developmental stage

tag: population


title: Control014

description: 3'-biased mRNA

tag: disease

value: Control


title: GL78_008_WN-S2-P1_A2_AGM_E10-5_SP_32-33_Gfi1_HE__S8

tag: tissue

value: CD1/Gf1+ (Thambyrajah et al 2015)

tag: age

value: 6-8 weeks old


title: Control013

description: 3'-biased mRNA


title: H2195_5d

description: NuGEN Ovation RRBS Methyl-Seq

value: 5 days


title: DNMT3B_KO_DNMT3B_exp_2_WGBS

value: DNMT3B KO

value: Expressing DNMT3B


title: H2195_2d

description: NuGEN Ovation RRBS Methyl-Seq

value: 2 days


title: DNMT3B_KO_DNMT3B_W263A_exp_2_WGBS

source_name: HCT116 with knocked out DNMT3B expressing DNMT3B W263A

value: Expressing DNMT3B W263A


title: Control012

description: 3'-biased mRNA


title: GL78_006_WN-S2-P1_I1_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S6

value: Hematopoietic cells


title: H2195_0d

description: NuGEN Ovation RRBS Methyl-Seq

value: WT

value: 0 day


title: Control010

description: 3'-biased mRNA


title: GL78_005_WN-S2-P1_H1_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S5


title: DNMT3B_KO_exp_2_WGBS

value: colorectal cancer cell line


title: Control011

description: 3'-biased mRNA


title: H1876_9d

description: NuGEN Ovation RRBS Methyl-Seq

value: Small Cell Lung Cancer


title: HCT116_exp_2_WGBS


title: Control009

description: 3'-biased mRNA


title: GL78_004_WN-S2-P1_G1_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S4


title: DNMT3B_KO_DNMT3A_exp_2_WGBS

source_name: HCT116 with knocked out DNMT3B expressing DNMT3A

value: Expressing DNMT3A


title: DNMT3B_KO_DNMT3Bcd_exp_1_WGBS

source_name: HCT116 with knocked out DNMT3B expressing DNMT3B catalytic dead

value: Expressing DNMT3B catalytic dead


title: H1876_5d

description: NuGEN Ovation RRBS Methyl-Seq

value: 2uM cisplatin


title: DNMT3B_KO_DNMT3B_exp_1_WGBS


title: DNMT3B_KO_exp_1_WGBS


title: GL78_002_WN-S2-P1_E1_AGM_E10-5_SP_32-33_Gfi1_HE__S2

value: Hemogenic endothelium cells

value: AGM


title: Control008

description: 3'-biased mRNA

value: Control


title: GL69_1475_WN-S2-1_K16_Roshana_Gfi1_IAHC_S384_L001

value: CD1/Gf1+ (Thambyrajah et al 2015)

value: 6-8 weeks old


title: GL78_007_WN-S2-P1_J1_AGM_E10-5_SP_32-33_Gfi1_pos_IAHC__S7


title: SHP_77

description: NuGEN Ovation RRBS Methyl-Seq

value: untreated


title: H1876_2d

description: NuGEN Ovation RRBS Methyl-Seq


title: Control006

description: 3'-biased mRNA


title: H1876_0d

description: NuGEN Ovation RRBS Methyl-Seq


title: GL69_1474_WN-S2-1_K15_Roshana_Gfi1_IAHC_S383_L001


title: mMEL-kit (K641E) C3-T

city: Boston

first: Hensin

last: Tsao

state: MA

address: 70 Blossom St, Edwards 202

institute: Massachusetts General Hospital

data_processing:
fastqc to QC fastq file.
StringTie was used to assemble RNA-Seq alignments into potential transcripts.
Using STAR aligner, read counts were converted to FPKM (Fragments per kilobase of transcript per million read pairs). FPKMs are calculated from the number of reads mapped to each particular gene sequence taking into account the gene length and the sequencing depth.
Assembly: mm10
Supplementary files format and content: Tab-delimited text files include FPKM and TPM values.

source_name: MelanA

growth_protocol: Cells were grown in RPMI 1640 with 10% FBS.

extract_protocol:
Total cell RNA was extracted using TRIzol Reagent (Invitrogen; Thermo Fisher Scientific, Waltham, MA) and RNeasy Mini kit (Qiagen, Hilden, Germany). RNA quality was examined by Qubit 3 (Thermo Fisher Scientific) and Fragment Analysis (Whitehead Institute Genome Technology Core, Massachusetts Institute of Technology, Cambridge, MA).
Sequencing libraries were prepared following the NEBNext Ultra II RNA Library Prep kit (New England Biolabs, MA) protocol and sequenced at Admera Health (South Plainfield, NJ) using the Illumina platform.

value: MelanA

value: melanoma

value: Kit K641E


title: HCT116_exp_1_WGBS


title: Control004

description: 3'-biased mRNA


title: GL69_1472_WN-S2-1_K11_Roshana_Gfi1_IAHC_S381_L001

value: Hematopoietic cells


title: mMEL-kit (K641E) C3#2


title: mMEL-kit (K641E) C1-T


title: NJH29

description: NuGEN Ovation RRBS Methyl-Seq

value: WT


title: Control003

description: 3'-biased mRNA


title: GL69_1471_WN-S2-1_K08_Roshana_Gfi1_IAHC_S380_L001


title: H889

description: NuGEN Ovation RRBS Methyl-Seq

value: 0 day


title: mMEL-kit (K641E) C3#1


title: H847

description: NuGEN Ovation RRBS Methyl-Seq


title: Control002

description: 3'-biased mRNA


title: mMEL-kit (K641E) C1#2


title: GL69_1470_WN-S2-1_K07_Roshana_Gfi1_IAHC_S379_L001


title: GL78_003_WN-S2-P1_F1_AGM_E10-5_SP_32-33_Gfi1_HE__S3


title: GL69_1469_WN-S2-1_K02_Roshana_Gfi1_IAHC_S378_L001


title: H82

description: NuGEN Ovation RRBS Methyl-Seq


title: H841

description: NuGEN Ovation RRBS Methyl-Seq


title: mMEL-kit (K641E) C1#1


title: GL69_1468_WN-S2-1_K01_Roshana_Gfi1_IAHC_S377_L001

description:
SMART-Seq2
Not included in final analysis


title: H774

description: NuGEN Ovation RRBS Methyl-Seq


title: Control005

description: 3'-biased mRNA


title: GL69_1467_WN-S2-1_J16_Roshana_Gfi1_IAHC_S376_L001


title: mMEL-kit (K641E) N1#1


title: GL69_1473_WN-S2-1_K13_Roshana_Gfi1_IAHC_S382_L001


title: H865

description: NuGEN Ovation RRBS Methyl-Seq


title: H740

description: NuGEN Ovation RRBS Methyl-Seq


title: H748

description: NuGEN Ovation RRBS Methyl-Seq


title: GL69_1466_WN-S2-1_J14_Roshana_Gfi1_IAHC_S375_L001


title: shCTRL_RNAseq_timepoint44 [SRR26830817]

city: Blacksburg

first: Benjamin

middle: Alex

last: Unruh

email: baunruh@vt.edu, skojima@vt.edu

state: Virginia

address: 1015 Life Science Cir

department: Biological Sciences

institute: Virginia Tech

description:
SRR26830817
Strand specific, paired-end sequencing, rRNA depleted

data_processing:
Azenta used their in-house quality control bioinformatical pipeline to return FASTQ files with a quality score of > Q30 on average. We further used TrimGalore (Martin et al. 2010) to remove adapter sequences and remove reads of low quality (< Q20), which we found to be about  0.2% per file. Reads were mapped with STARv2.7.7a (Dobin et al. 2013) using options --outFilterScoreMinOverLread 0.3 and --sjdbOverhang 100. Reads mapped to the NR003278.3 (Mus musculus 18S rRNA) and NR003279.1 (Mus musculus 28S rRNA) were eliminated first. Remaining reads were then mapped to the mouse mm10 genome (GENECODE: GRCm38.p6.genome.fa) and read counts against gene (transcript per million or TPM), exon, and intron were independently quantified with HOMER (V4.11.1) (Heinz et al. 2010) using gencode.vM25.annotation.gtf and option condenseGenes. Transcripts with TPM < 0.25 (as an average of all time-points and conditions) were removed for all downstream analysis. Raw reads were  (v4.11) to quantify the reads mapped to exon or intron separately with the –exon or –intron option for both RNA-seq and 4sU-seq
Reported RPKMs for exons and introns were manually calculated for each timepoint using the total reads mapping to both introns and exons.
Assembly: mm10
Supplementary files format and content: comma-delimited text file(CSV) include RPKM values for exons and introns for each sample.

source_name: NIH 3T3

growth_protocol: NIH3T3 cells were cultured in Dulbecco’s Modified Eagle Medium (DMEM) (Gibco: Cat #11965118) supplemented with 10% fetal bovine serum (Corning, Cat#MT35010CV) at 37C with 5% CO2.

extract_protocol:
RNAs were extracted with TRIzol (Invitrogen, Cat# 15596018) according to manufacturer’s instructions, and then treated with DNase I (ThermoFisher, AM2239) at 37°C for 1 hr. After re-purifying RNAs using TRIzol, 1 µg of yeast spike-in RNA (gift from Dr. Silke Hauf: Virginia Tech) labeled with 4-thiouracil (Neta Scientific, CMX-21484-1G) for 1 hr, and 300 fmol 4sU-labeled DNA/RNA hybrid spike-in control (ATTTAGGTGACACTATAGGATCCTCTAGAGTCGACCTTCTCCCTATAGTGAGTC GTATTAGCA[4sU]CAG) were added to each RNA sample to normalize the RNA pull-down efficiency (see below). 4sU-labeled RNA was then biotinylated with 1 mg/ml EZ-link HPDP-Biotin (ThermoFisher, cat: 21341) for 90 min at room temperature as previously described (34, 69, 70). 30 ug RNA (approximately 50% for shCtrl and 60-70% for shBmal1 samples) were subsequently subjected to streptavidin pull-down, while the remainder was set aside as ‘total RNA’.
Approximately 100-200 ng of total and newly synthesized RNAs from each time point were subjected to transcriptomic analysis (Azenta Biosciences). RNA quality of each sample was verified by the RNA integrity number (RIN) > 9.5 measured by Tapestation 4200 (Agilent). Additional DNase I treatment was performed before proceeding to the library preparation per recommendation from Azenta. Paired-end, strand-specific, and rRNA depleted sequencing library was then prepared using QIAGEN FastSelect rRNA HMR Kit (Qiagen, cat#334378) and NEB Directional Ultra II RNA library prep kit (NEB, E7760) by following the manufacturer’s recommendations (NEB, cat# E7765). Qubit 2.0 Fluorometer (ThermoFisher Scientific) and quantitative PCR (KAPA Biosystems) method were used to quantify each library before multiplexing and the distribution of reads lengths were measured by the Agilent Tapestation 4200 (Agilent Technologies). Libraries were then sequenced by Illumina Hi-seq.

treatment_protocol: Cells were synchronized with 50% horse serum media for 2 hours.

value: NIH 3T3

value: mouse fibroblast

value: shCTRL

tag: time point


title: GL69_1464_WN-S2-1_J09_Roshana_Gfi1_IAHC_S373_L001


title: shCTRL_RNAseq_timepoint46 [SRR26830816]

description:
SRR26830816
Strand specific, paired-end sequencing, rRNA depleted


title: mVEC #2

value: vector


title: H735

description: NuGEN Ovation RRBS Methyl-Seq

value: Small Cell Lung Cancer


title: mMEL-kit (K641E) PT#1


title: shCTRL_RNAseq_timepoint42 [SRR26830772]

description:
SRR26830772
Strand specific, paired-end sequencing, rRNA depleted


title: shCTRL_RNAseq_timepoint40 [SRR26830773]

description:
SRR26830773
Strand specific, paired-end sequencing, rRNA depleted


title: shCTRL_RNAseq_timepoint38 [SRR26830774]

description:
SRR26830774
Strand specific, paired-end sequencing, rRNA depleted


title: GL69_1465_WN-S2-1_J12_Roshana_Gfi1_IAHC_S374_L001


title: Control007

description: 3'-biased mRNA


title: Control001

description: 3'-biased mRNA


title: H719

description: NuGEN Ovation RRBS Methyl-Seq


title: mMEL-kit (K641E) N1#2


title: GL69_1463_WN-S2-1_J08_Roshana_Gfi1_IAHC_S372_L001


title: shCTRL_RNAseq_timepoint36 [SRR26830775]

description:
SRR26830775
Strand specific, paired-end sequencing, rRNA depleted


title: mVEC #1


title: shCTRL_RNAseq_timepoint34 [SRR26830776]

description:
SRR26830776
Strand specific, paired-end sequencing, rRNA depleted


title: GL69_1462_WN-S2-1_J06_Roshana_Gfi1_IAHC_S371_L001


title: RNA_T33

city: jinan

first: congcong

last: liu

address: wenhua road

country: China

institute: Shandong university

data_processing:
We used Bowtie2 to align Clean reads to the reference gene sequence and then used RSEM to calculate gene expression levels for individual samples.
The DEseq2 method is based on the principle of negative binomial distribution and this project is based on the method described in Michael I, et al for differential gene DEG.
Assembly: mm10
Supplementary files format and content: Text file with raw gene counts for every gene and every sample

source_name: Liver

extract_protocol:
1 μg of total RNA was subjected to library synthesis using Trizol (Invitrogen, Carlsbad, CA,USA)
RNA libraries were prepared for sequencing using standard Illumina protocols.

treatment_protocol: Fasting 6h

value: Liver

value: liver


title: H69

description: NuGEN Ovation RRBS Methyl-Seq


title: H526

description: NuGEN Ovation RRBS Methyl-Seq


title: shCTRL_RNAseq_timepoint32 [SRR26830785]

description:
SRR26830785
Strand specific, paired-end sequencing, rRNA depleted


title: mMEL-kit (K641E) PT#2


title: GL69_1461_WN-S2-1_J05_Roshana_Gfi1_IAHC_S370_L001

value: AGM


title: RNA_T25


title: shCTRL_RNAseq_timepoint30 [SRR26830796]

description:
SRR26830796
Strand specific, paired-end sequencing, rRNA depleted


title: GL69_1460_WN-S2-1_J03_Roshana_Gfi1_IAHC_S369_L001

value: CD1/Gf1+ (Thambyrajah et al 2015)


title: H524

description: NuGEN Ovation RRBS Methyl-Seq


title: RNA_C28

value: Control


title: GL69_1458_WN-S2-1_I12_Roshana_Gfi1_IAHC_S367_L001

value: 6-8 weeks old


title: shCTRL_RNAseq_timepoint26 [SRR26830818]

description:
SRR26830818
Strand specific, paired-end sequencing, rRNA depleted


title: H510A

description: NuGEN Ovation RRBS Methyl-Seq

value: untreated


title: H446

description: NuGEN Ovation RRBS Methyl-Seq


title: RNA_C26


title: GL69_1457_WN-S2-1_I07_Roshana_Gfi1_IAHC_S366_L001


title: shCTRL_RNAseq_timepoint24 [SRR26830819]

description:
SRR26830819
Strand specific, paired-end sequencing, rRNA depleted


title: H378

description: NuGEN Ovation RRBS Methyl-Seq


title: RNA_C22


title: shCTRL_4sUseq_timepoint46 [SRR26830790]

description:
SRR26830790
Strand specific, paired-end sequencing, rRNA depleted

library_source: other

data_processing:
library strategy: 4sUseq
Azenta used their in-house quality control bioinformatical pipeline to return FASTQ files with a quality score of > Q30 on average. We further used TrimGalore (Martin et al. 2010) to remove adapter sequences and remove reads of low quality (< Q20), which we found to be about  0.2% per file. Reads were mapped with STARv2.7.7a (Dobin et al. 2013) using options --outFilterScoreMinOverLread 0.3 and --sjdbOverhang 100. Reads mapped to the NR003278.3 (Mus musculus 18S rRNA) and NR003279.1 (Mus musculus 28S rRNA) were eliminated first. Remaining reads were then mapped to the mouse mm10 genome (GENECODE: GRCm38.p6.genome.fa) and read counts against gene (transcript per million or TPM), exon, and intron were independently quantified with HOMER (V4.11.1) (Heinz et al. 2010) using gencode.vM25.annotation.gtf and option condenseGenes. Transcripts with TPM < 0.25 (as an average of all time-points and conditions) were removed for all downstream analysis. Raw reads were  (v4.11) to quantify the reads mapped to exon or intron separately with the –exon or –intron option for both RNA-seq and 4sU-seq
Reported RPKMs for exons and introns were manually calculated for each timepoint using the total reads mapping to both introns and exons.
Assembly: mm10
Supplementary files format and content: comma-delimited text file(CSV) include RPKM values for exons and introns for each sample.

molecule: other


title: GL69_1459_WN-S2-1_I18_Roshana_Gfi1_IAHC_S368_L001


title: GL69_1455_WN-S2-1_H21_Roshana_Gfi1_IAHC_S364_L001


title: GL69_1456_WN-S2-1_I06_Roshana_Gfi1_IAHC_S365_L001


title: H2330

description: NuGEN Ovation RRBS Methyl-Seq


title: H250

description: NuGEN Ovation RRBS Methyl-Seq


title: HiC_cKO6: Satb2 flx/flx::Nes-Cre DIV14 primary cortical cultures, 1h bic, rep6

city: Innsbruck

first: Nico

last: Wahl

address: Innrain 66

department: Institute for Neuroscience

country: Austria

institute: Medical University Innsbruck

data_processing:
Paired-end sequencing reads were mapped to the mouse reference genome assembly (mm10), artefacts were filtered, and libraries were ICED normalized using  HiC-Pro (v.2.11.4) at diffrent resolutions.
Differential compartment analysis was carried out by using dcHiC. Input file was created following dcHiC guidelines using HiC-Pro matrices at 100 kb.
TADs were computed at 25 kb resolution using SpectralTAD. Differential TAD boundaries between cKO and Flx Hi-C contact matrices were identified by TADCompare.
Chromatin loops were called by Mustache (v1.2.0) at 5 kb resolution using pooled replicate Hi-C matrices. The following parameters for differential loop detection were used: P value (-p) of 0.05 and sparsity (-st) of 0.8 thresholds together with KR normalization.
Frequently interacting regions (FIREs) were called using FIREcaller. Briefly, ‘sparseToDense.py’ HiC-Pro utility was used to create quadratic matrices at 10 kb resolution.
Assembly: mm10
Supplementary files format and content: bedgraph / genomic coordinates and EV1 values of called compartments in Flx and cKO samples
Supplementary files format and content: txt / differential TAD boundaries between cKO and Flx samples
Supplementary files format and content: bedpe / genomic coordinates of chromatin loops in Flx and cKO samples called 5kb resolution
Supplementary files format and content: txt / FIRE analysis in Flx and cKO samples

source_name: Cortex

growth_protocol: Corical tissue was dissected at P0-P1, dissociated with Papain and titurated. Cells were plated in serum-containing medium (MEM, 1% horse serum, 0.1% Pen/Strep, 0.1 % sodium piruvate). After 2h, MEM was excahnged with serum-free feeding medium (Neurobasal-A Medium, containing B-27, 0.1% Pen/Strep and 2 mM Glutamine.  Cells were treated with 5µM cytosine arabinoside on DIV3 to inhibit glial cell proliferation.  At DIV4, one third of the medium was replaced by fresh serum-free feeding medium. Neurons were cultured for 14 days in 5% CO2 at 37°C.

extract_protocol:
For in situ Hi-C, Arima-HiC kit was used, according to the manufacturer’s protocol. Briefly, 2.5x106 primary cortical neurons were fixed directly on the dish by addition of formaldehyde to a final concentration of 1 % for 10 min on RT. After washing twice with 1X PBS cells were detached using a cell lifter and collected in 1X PBS containing protease inhibitors. Cells were pelleted at 500 g for 10 min at 4 °C and pellets were directly snap-frozen in liquid nitrogen. After nuclei isolation and chromatin digestion, biotin was added and filled ends were ligated. After reverse crosslinking and purification, DNA was shared using a M220 Covaris sonicator at duty factor 12 %, PIP 75, CPB 200 for 60 s to achieve input fragment size of 400 bp. Additionally, DNA was size-selected using AMPure XP beads, according to Arima HiC kit manual.
Hi-C libraries were prepared using Swift Biosciences Accel-NGS 2S Plus DNA Library Kit according to Arima-HiC Kit User Guide (November 2018)

treatment_protocol: Cottical neurons were treated with 20 µM NBQX for 16 h on DIV13. After silencing, neurons were stimulated with 50 µM biccuculine for 1h on DIV14 and then collected.

value: Cortex

value: DIV14 primary cortical cultures

value: Satb2 flx/flx::Nes-Cre

value: 1h bicucculine


title: shCTRL_4sUseq_timepoint44 [SRR26830791]

description:
SRR26830791
Strand specific, paired-end sequencing, rRNA depleted


title: H345

description: NuGEN Ovation RRBS Methyl-Seq

value: WT

value: 0 day


title: HiC_cKO5: Satb2 flx/flx::Nes-Cre DIV14 primary cortical cultures, 1h bic, rep5


title: shCTRL_4sUseq_timepoint42 [SRR26830792]

description:
SRR26830792
Strand specific, paired-end sequencing, rRNA depleted

value: NIH 3T3

value: mouse fibroblast


title: HiC_Flx6: Satb2 flx/flx DIV14 primary cortical cultures, 1h bic, rep6

value: Satb2 flx/flx


title: H2227

description: NuGEN Ovation RRBS Methyl-Seq

value: Small Cell Lung Cancer


title: RNA_T24

value: Liver


title: shCTRL_4sUseq_timepoint40 [SRR26830793]

description:
SRR26830793
Strand specific, paired-end sequencing, rRNA depleted

value: shCTRL


title: shCTRL_RNAseq_timepoint28 [SRR26830807]

description:
SRR26830807
Strand specific, paired-end sequencing, rRNA depleted


title: GL69_1453_WN-S2-1_H06_Roshana_Gfi1_IAHC_S362_L001

value: Hematopoietic cells

value: AGM


title: H2198

description: NuGEN Ovation RRBS Methyl-Seq


title: shCTRL_4sUseq_timepoint38 [SRR26830794]

description:
SRR26830794
Strand specific, paired-end sequencing, rRNA depleted


title: HiC_Flx5: Satb2 flx/flx DIV14 primary cortical cultures, 1h bic, rep5


title: GL69_1451_WN-S2-1_G23_Roshana_Gfi1_IAHC_S360_L001

value: CD1/Gf1+ (Thambyrajah et al 2015)


title: H2196

description: NuGEN Ovation RRBS Methyl-Seq


title: shCTRL_4sUseq_timepoint36 [SRR26830795]

description:
SRR26830795
Strand specific, paired-end sequencing, rRNA depleted


title: GL69_1454_WN-S2-1_H09_Roshana_Gfi1_IAHC_S363_L001

value: 6-8 weeks old


title: H2195

description: NuGEN Ovation RRBS Methyl-Seq

value: untreated


title: GL69_1450_WN-S2-1_G14_Roshana_Gfi1_IAHC_S359_L001


title: H2171

description: NuGEN Ovation RRBS Methyl-Seq


title: shCTRL_4sUseq_timepoint32 [SRR26830798]

description:
SRR26830798
Strand specific, paired-end sequencing, rRNA depleted


title: GL69_1449_WN-S2-1_G09_Roshana_Gfi1_IAHC_S358_L001


title: GL69_1448_WN-S2-1_G08_Roshana_Gfi1_IAHC_S357_L001


title: Donor_1285_FVIII_POOL

city: Gif-sur-Yvette

first: Valeria

last: Porcheddu

address: Université de Paris-Saclay, CEA, INRAE, Département Médicaments et Technologies pour la Santé, SIMoS

country: France

institute: CEA Paris-Saclay

data_processing:
All data were aligned using the MiXCR v4.2.0 tool (https://github.com/milaboratory/mixcr).
The high-quality reads were further assembled into clonotypes, correcting for PCR and sequencing errors.
Shared clonotypes between the unstimulated and antigen-stimulated samples were removed.
The downstream analysis was performed with MIXCR, “Immunarch” R package, and in-house R script (https://github.com/porchedduvl/CD4TFVIIIepitopes.git).
Assembly: MIXCR
Supplementary files format and content: tab-delimited txt file including read raw counts and, VJ usage, TCR sequence

source_name: TCR CDR3β sequencing

growth_protocol: CD4 T cells were isolated from autologous PBMCs by positive selection using magnetic labelling with anti-CD4 mAbs conjugated to magnetic microbeads followed by magnetic cell sorting, as recommended by the manufacturer (Miltenyi Biotec, Bergisch Gladbach, Germany). Immature DCs were loaded overnight at 37˚C with either 0.2 µM of rFVIII, 0.2 µM of Ovalbumin, or 0.25 µM of KLH in 96-wells U-bottom plates, while mature DCs were used for FVIII peptides (10 ug/ml). CD4 T-cell lines were generated in vitro in IMDM (Invitrogen) supplemented with 10% human serum, 0.24 mM of glutamine, 0.55 mM of asparagine, 1.5 mM of arginine (all amino acids from Sigma-Aldrich),  50 U/mL of penicillin/50 μg/mL of streptomycin (Invitrogen), 20 I.U/mL human IL-2 and 17 I.U/ml of human IL-7 (premium grade; Miltenyi Biotec) by 3 once-per-week rounds of stimulation of CD4 T cells (200 000 per well) with autologous DCs (20.000 per well).

extract_protocol:
RNA was extracted from antigen-specific T cells using the RNeasy Plus Mini Kit (Qiagen) and sample RNA was reverse transcribed using a TCRB chain constant region reverse primer with Superscript IV First-Strand Synthesis System (Invitrogen).
cDNA was column purified with the Oligo Clean and Concentrator Kit (Zymo Research). cDNA was used as a template for PCR with KAPA2G Fast Multiplex Mix (Roche). Forward TCRB FR3 primers were used with a single nested TCRB constant region reverse primer. 20 cycles for NGS library preparation for the PCR1 and 20 cycles of PCR2 were performed on the PCR1 product to incorporate sample barcodes and Illumina sequencing adaptors using Herculase II Fusion DNA Polymerase (Agilent). Specific principles and methodology have been previously reported. PCR2 product underwent a run of agarose gel electrophoresis. Bands at 300pb were purified using a GeneJET purification kit (Thermo Fisher) and quantified using the Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific).
RNA-seq on an Illumina NextSeq 500 with 15% of PhiX control v3 (Illumina)

treatment_protocol: Following 24h stimulation, cells were harvested in Cell Staining Buffer (phosphate-buffered saline/EDTA 2mM /bovine serum albumin 0.5%), then stained for CD4 (Vio-Blue, clone REA623, Miltenyi Biotec) and activation-induced markers CD134 (OX-40) (PE anti-human, clone Ber-ACT35, Biolegend), CD137 (4-1BB) (APC anti-human, clone 4B4-1, Bio legend). The CD4/CD134/CD137 positive population for each condition was sorted by flow cytometry (FACSARIA III; Becton Dickinson, San Jose, CA).

value: Buffy coat

value: lymphocytes T

value: CD4 T

tag: subject status

value: healthy donors


title: shCTRL_4sUseq_timepoint34 [SRR26830797]

description:
SRR26830797
Strand specific, paired-end sequencing, rRNA depleted


title: shCTRL_4sUseq_timepoint30 [SRR26830799]

description:
SRR26830799
Strand specific, paired-end sequencing, rRNA depleted


title: Donor_1327_FVIII


title: Donor_1255_FVIII_POOL


title: Donor_1327_FVIII_POOL


title: H2141

description: NuGEN Ovation RRBS Methyl-Seq

value: WT

tag: treatment


title: Donor_1285_FVIII

tag: cell type


title: GL69_1447_WN-S2-1_F23_Roshana_Gfi1_IAHC_S356_L001


title: shCTRL_4sUseq_timepoint28 [SRR26830800]

description:
SRR26830800
Strand specific, paired-end sequencing, rRNA depleted


title: H2108

description: NuGEN Ovation RRBS Methyl-Seq

value: 0 day


title: Donor_1255_FVIII


title: shCTRL_4sUseq_timepoint26 [SRR26830801]

description:
SRR26830801
Strand specific, paired-end sequencing, rRNA depleted

value: NIH 3T3


title: GL69_1446_WN-S2-1_F17_Roshana_Gfi1_IAHC_S355_L001


title: GL69_1452_WN-S2-1_H02_Roshana_Gfi1_IAHC_S361_L001


title: H2107

description: NuGEN Ovation RRBS Methyl-Seq


title: Donor_1245_FVIII_POOL


title: shCTRL_4sUseq_timepoint24 [SRR26830802]

description:
SRR26830802
Strand specific, paired-end sequencing, rRNA depleted

value: mouse fibroblast


title: Input

data_processing:
Raw reads were filtered to obtain high-quality clean reads by removing sequencing adapters, short reads (length <35 bp) and low-quality reads using trim-galore (v0.6.4)
Then FastQC (v0.11.9) and Multiqc (v1.8) is used to ensure high reads quality
The clean reads were mapped to the mouse genome (assembly mouse genome GRCm38) using the Burrow-Wheeler Aligner (BWA v0.7.17) software
Peak detection was performed using the MACS2 (v2.2.6) peak finding algorithm with 0.05 set as the q-value cutoff
Annotation of peak sites to gene features was performed using the homer annotatePeaks.pl (v4.10)
Assembly: mm10
Supplementary files format and content: bigWig

source_name: mouse primary hepatocytes

extract_protocol:
Cells were fixed with 1% formaldehyde for 10 minutes. After quenching and cell lysis, chromatin was sonicated using sonication.
The DNA libraries were amplified for 15 cycles and sequenced using Illumina NovaSeq 6000 with paired-end 150 as the sequencing mode

value: mouse primary hepatocytes

tag: chip antibody

value: none (input)


title: Donor_1245_FVIII


title: H2081

description: NuGEN Ovation RRBS Methyl-Seq

value: Small Cell Lung Cancer


title: Rep2

value: TOX3 (Sigma,SAB1303181)


title: Donor_1238_FVIII_POOL


title: shBmal1_RNAseq_timepoint44 [SRR26830804]

description:
SRR26830804
Strand specific, paired-end sequencing, rRNA depleted


title: Rep1


title: H211

description: NuGEN Ovation RRBS Methyl-Seq


title: shBmal1_RNAseq_timepoint42 [SRR26830805]

description:
SRR26830805
Strand specific, paired-end sequencing, rRNA depleted


title: H2029

description: NuGEN Ovation RRBS Methyl-Seq


title: Donor_1238_FVIII


title: shBmal1_RNAseq_timepoint40 [SRR26830806]

description:
SRR26830806
Strand specific, paired-end sequencing, rRNA depleted


title: Donor_1180_OVA


title: H196

description: NuGEN Ovation RRBS Methyl-Seq


title: shBmal1_RNAseq_timepoint38 [SRR26830808]

description:
SRR26830808
Strand specific, paired-end sequencing, rRNA depleted


title: Donor_1180_FVIII


title: H1963

description: NuGEN Ovation RRBS Methyl-Seq


title: E11_limb_RNA_WT_rep2

city: Beijing

first: Chunsheng

last: Han

address: No.5, Yard 1, Beichen West Road, Chaoyang District, Beijing

institute: Institute of Zoology, Chinese Academy of Sciences

data_processing:
The sequencing reads were mapped to the mouse genome (UCSC mm10) with TopHat (v2.0.6).
The mRNA expression levels were represented by fragments per kilobase of transcript sequence per millions (FPKM) calculated by Cufflinks (v2.0.2).
The differentially expressed genes were identified with the Cuffdiff package.
Assembly: mouse genome (UCSC mm10)
Supplementary files format and content: xlsx

source_name: Hindlimb buds

extract_protocol:
Hindlimb buds were extracted from E11.5 embryos using forceps under a stereoscope. Total RNAs were isolated using TRIzol (15596-026; Invitrogen).
High-throughput sequencing libraries were constructed using the NEB-Next Ultra RNA Library Prep Kit for Illumina (E7760, NEB).

treatment_protocol: Trizol for RNA-seq

tag: background

value: ICR


title: E11_limb_RNA_WT_rep3


title: E11_limb_RNA_WT_rep1


title: H1930

description: NuGEN Ovation RRBS Methyl-Seq


title: shBmal1_RNAseq_timepoint36 [SRR26830809]

description:
SRR26830809
Strand specific, paired-end sequencing, rRNA depleted


title: shBmal1_RNAseq_timepoint34 [SRR26830810]

description:
SRR26830810
Strand specific, paired-end sequencing, rRNA depleted


title: H209

description: NuGEN Ovation RRBS Methyl-Seq


title: shBmal1_RNAseq_timepoint46 [SRR26830803]

description:
SRR26830803
Strand specific, paired-end sequencing, rRNA depleted


title: Donor_1195_OVA


title: H187

description: NuGEN Ovation RRBS Methyl-Seq


title: Donor_1207_FVIII


title: H1876

description: NuGEN Ovation RRBS Methyl-Seq


title: E11_limb_RNA_KO_rep3

value: KO


title: Donor_1207_OVA


title: H1836

description: NuGEN Ovation RRBS Methyl-Seq


title: E11_limb_RNA_KO_rep1


title: E11_limb_RNA_KO_rep4


title: H1688

description: NuGEN Ovation RRBS Methyl-Seq


title: shBmal1_RNAseq_timepoint32 [SRR26830811]

description:
SRR26830811
Strand specific, paired-end sequencing, rRNA depleted


title: E11_limb_RNA_KO_rep2


title: Donor_1195_FVIII


title: shBmal1_RNAseq_timepoint30 [SRR26830812]

description:
SRR26830812
Strand specific, paired-end sequencing, rRNA depleted


title: Donor_1179_OVA


title: WT_Input

data_processing:
The raw reads of ChIP-seq was converted into a fastq file, and the sequencing quality was evaluated by FastQC, the reads were trimmed to remove the adapter sequence.
The trimmed ChIP-seq reads were mapped to UCSC mm10 genome by using Bowtie2 (v2.5.0) with default parameters. For PBX4, peak calling was performed using Pepr (v1.1.10) with default parameters and corresponding input as background.
The annotation of genome location and repeat type is generated by HOMER (v4.11), and the heatmap is generated by the command line version of deepTools (v3.5.0).
HOMER (v4.11) was used to identify the rich motif in PBX4 peak under the default setting.
Assembly: mouse genome (UCSC mm10)
Supplementary files format and content: bigWig files

source_name: testis

extract_protocol:
The CHIP and input DNA samples were purified with phenol/chloroform
DNA was used to construct a library with the NEBNext Ultra II DNA Library Prep Kit for Illumina (E7645 and E7335, NEB); High-throughput sequencing libraries were constructed using the NEB-Next Ultra RNA Library Prep Kit for Illumina(E7760, NEB)

treatment_protocol: Lysis for CHIP-seq


title: shBmal1_RNAseq_timepoint26 [SRR26830814]

description:
SRR26830814
Strand specific, paired-end sequencing, rRNA depleted


title: Donor_1179_FVIII


title: WT_IP

value: α-FLAG (F1804, SIGMA)


title: miRNAs-NPW8

city: Mexico City

first: Mariana

last: Flores Torres

address: Periferico Sur 4809

country: Mexico

institute: Instituto Nacional de Medicina Genomica

data_processing:
Data quality control analysis was performed before and after adaptor removal using FastQC.
3'-adaptors were trimmed with Cudadapt (Phred score >30, min lenght 18).
Remaining sequences were aligned to the human genome (Hg38p12) with  BWA, miRNA profile was determined using miRDeep2 and miRBase v.22 as miRNA reference.
Reads were tallied to generate total counts for each miRNA.
Genome_build: Hg38p12
Supplementary_files_format_and_content: csv file reports raw counts

source_name: Plasma

extract_protocol:
Total RNA was extracted from pregnant women plasma samples (2 mL) at 1st, 2nd and 3rd trimester (same women for each trimester) using TRI Reagent Solution (Ambion), in a 3:1 sample volume ratio. Samples were mixed and incubated for 5 minutes at room temperature and centrifuged at 12,000 x g for 10 minutes at 4 °C. The remaining aqueous phase was removed and mixed with chloroform (0.2 mL per mL of TRI Reagent), the new aqueous phase was transferred to fresh tubes and isopropanol (0.5 mL per mL of TRI Reagent) was added, mixed, and incubated overnight at -75 °C. After incubation samples were centrifuged at 12,000 x g for 15 minutes at 4 °C. RNA pellet was resuspended in 75% ethanol and centrifuged at 7,500 x g at 4 °C for 5 minutes. RNA pellets were resuspended in 10 uL of RNase-free water and stored at -75 °C.
Library construction for small RNA sequencing analysis was performed with the NextFLEX small RNA-Seq kit v3 (Bioo Scientific), bands in an acrylamide gel, ranging from 50 to 150 bp were processed. Libraries were sequenced for 150 cycles, single-end, on an Illumina NextSeq 500.

value: Non pregnant

value: Plasma


title: miRNAs-NPW9


title: Input-rep4

value: Pbx4-FLAG KI


title: miRNAs-NPW5


title: miRNAs-NPW4


title: Input-rep3


title: H1694

description: NuGEN Ovation RRBS Methyl-Seq

value: untreated


title: H1618

description: NuGEN Ovation RRBS Methyl-Seq


title: shBmal1_RNAseq_timepoint24 [SRR26830815]

description:
SRR26830815
Strand specific, paired-end sequencing, rRNA depleted


title: Input-rep2


title: shBmal1_4sUseq_timepoint46 [SRR26830777]

description:
SRR26830777
Strand specific, paired-end sequencing, rRNA depleted


title: H1436

description: NuGEN Ovation RRBS Methyl-Seq


title: shBmal1_4sUseq_timepoint44 [SRR26830778]

description:
SRR26830778
Strand specific, paired-end sequencing, rRNA depleted


title: shBmal1_RNAseq_timepoint28 [SRR26830813]

description:
SRR26830813
Strand specific, paired-end sequencing, rRNA depleted


title: Input-rep5


title: shBmal1_4sUseq_timepoint42 [SRR26830779]

description:
SRR26830779
Strand specific, paired-end sequencing, rRNA depleted


title: H1417

description: NuGEN Ovation RRBS Methyl-Seq


title: miRNAs-NPW19


title: miRNAs-NPW18


title: H1341

description: NuGEN Ovation RRBS Methyl-Seq


title: H1522

description: NuGEN Ovation RRBS Methyl-Seq


title: H146

description: NuGEN Ovation RRBS Methyl-Seq


title: Pbx4-rep4


title: Input-rep1


title: shBmal1_4sUseq_timepoint40 [SRR26830780]

description:
SRR26830780
Strand specific, paired-end sequencing, rRNA depleted


title: miRNAs-NPW17


title: H1672

description: NuGEN Ovation RRBS Methyl-Seq


title: H128

description: NuGEN Ovation RRBS Methyl-Seq


title: miRNAs-NPW20


title: Pbx4-rep5


title: Pbx4-rep1


title: shBmal1_4sUseq_timepoint36 [SRR26830782]

description:
SRR26830782
Strand specific, paired-end sequencing, rRNA depleted


title: shBmal1_4sUseq_timepoint30 [SRR26830786]

description:
SRR26830786
Strand specific, paired-end sequencing, rRNA depleted


title: H1238

description: NuGEN Ovation RRBS Methyl-Seq


title: Pbx4-rep3


title: exRNaseA digested, rep2

city: New Haven

first: Jun

last: Lu

state: CT

address: 10 Amistad St.

department: Yale Stem Cell Center

institute: Yale University

data_processing:
RNA-seq reads were aligned to the mouse reference genome (mm10) using STAR v2.4.2a and annodated with Genecode M21
Normalized gene counts was calculated using  RSEM v1.3.3
Assembly: mm10
Supplementary files format and content: tab-delimited text files include gene expected count, TPM and FPKM for each gene and each sample

source_name: Bone marrow neutrophils

growth_protocol: Bone marrow neutrophils were freshly isolated from wildtype mice.

extract_protocol:
RNA was harvested using TRIzol.
RNA libraries for RNA-seq were prepared using the polyA selection protocol from Illumina.

treatment_protocol: Cells were suspected in HBSS buffer, and treated with RNaseA for 10 minutes at 37C.

value: Bone marrow neutrophils

value: Extracellular RNase A


title: shBmal1_4sUseq_timepoint34 [SRR26830783]

description:
SRR26830783
Strand specific, paired-end sequencing, rRNA depleted


title: shBmal1_4sUseq_timepoint32 [SRR26830784]

description:
SRR26830784
Strand specific, paired-end sequencing, rRNA depleted


title: shBmal1_4sUseq_timepoint26 [SRR26830788]

description:
SRR26830788
Strand specific, paired-end sequencing, rRNA depleted


title: DMS79

description: NuGEN Ovation RRBS Methyl-Seq

value: WT


title: exRNaseA digested, rep1


title: Control RNA, rep2

value: Mock


title: Pbx4-rep2


title: miRNAs-NPW10


title: miRNAs-M32_3T

value: 3rd trimester


title: DMS153

description: NuGEN Ovation RRBS Methyl-Seq

value: 0 day


title: shBmal1_4sUseq_timepoint38 [SRR26830781]

description:
SRR26830781
Strand specific, paired-end sequencing, rRNA depleted

value: NIH 3T3

value: mouse fibroblast


title: shBmal1_4sUseq_timepoint28 [SRR26830787]

description:
SRR26830787
Strand specific, paired-end sequencing, rRNA depleted


title: miRNAs-M30_2T

value: 2nd trimester


title: H1048

description: NuGEN Ovation RRBS Methyl-Seq

value: Small Cell Lung Cancer


title: miRNAs-M32_1T

value: 1st trimester


title: miRNAs-M3_1T


title: shBmal1_4sUseq_timepoint24 [SRR26830789]

description:
SRR26830789
Strand specific, paired-end sequencing, rRNA depleted


title: DC2.4 cells, EXOmock, Preparation 1

city: Chapel Hill

first: Ying

last: Wang

state: NC

address: 125 Mason Farm Road, 3118G Marsico Hall, Marsico Hall/3118G/3nd

institute: University of North Carolina at Chapel Hill

data_processing:
After removing reads with adapter or of low quality, clean reads were aligned to the mouse GRCm39 reference genome using hisat2 software, and only the uniquely mapped reads were retained for downstream analysis. The gene expression level was quantified using FeatureCounts function from Subread version 2.0.1.
Assembly: GRCm39
Supplementary files format and content: tab-delimited text file includes raw counts for each Sample

growth_protocol: DC2.4 cells were cultured with RPMI1640 supplemented with 10% FBS, 1× non-essential amino acids (Gibco), 1× HEPES buffer (Gibco), and 0.0054× β-mercaptoethanol (Gibco). 4T1 wild-type cells were cultured in RPMI1640 supplemented with 10% FBS. All the adherent cell lines were cultured at 37 °C and 5% CO2 in a humidified atmosphere.

extract_protocol:
Total RNA was extracted from DC2.4 cells and 4T1 cells after different treatments by using RNeasy Plus Mini Kit (Qiagen) according to manufacturer’s instructions. 1.0 µg of total RNA was used for the construction of sequencing libraries.
RNA library was generated by using NEBNext® UltraTM RNA Library Prep Kit for Illumina® (NEB) following manufacturer’s recommendations

treatment_protocol: DC2.4 cells and 4T1 tumor cells were treated with mRNAs (uniSTING mRNA or EGFP mRNA) either mixed with TransIT®-mRNA transfection agent (Mirus Bio) or encapsulated into SS-OP LNPs in DMEM culture medium supplemented with 10% FBS, according to the manufacturer’s instructions or conditions optimized in the Liu lab. For the investigation of the gene regulation in DC2.4 cells by miRNAs, the extracellular vesicles isolated from uniSTING (EXOuniSTING)- or Mock (EXOmock)-treated 4T1 tumor cells were incubated with DC2.4 cells for 24 hours before total RNA extraction.

value: Immortalized murine dendritic cells

value: Extracellular vesicles isolated from Mock treated 4T1 tumor cells


title: DC2.4 cells, EXOuniSTING, Preparation 2

value: Extracellular vesicles isolated from uniSTING treated 4T1 tumor cells


title: CORL88

description: NuGEN Ovation RRBS Methyl-Seq


title: Control RNA, rep1


title: miRNAs-M30_AB

value: After birth

value: Plasma


title: 4T1 tumor cells, uniSTING mRNA, Preparation 2

value: Breast cancer cells

value: uniSTING mRNA