title: SF11873_snRNA

description: Single Nuclei RNA-Seq

value: Primary


title: IR_Treated_48hrs

status: Public on Jul 05 2022

type: SRA

city: San Francisco

first: Aaron

last: Diaz

state: CA

department: Neurological Surgery

country: USA

institute: University of California, San Francisco

description: RNA-Seq

library_source: transcriptomic

data_processing:
We utilized CellRanger (version 3.0.2) for data pre-processing and gene expression quantification, following the guidelines from the CellRanger website (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/advanced/references#premrna).
The CellRanger ATAC software (version 1.1.0) was used for read alignment, deduplication, and identifying transposase cut sites.
The presence/absence of somatic CNVs was assessed with CONICSmat. ELSA was used for cell type classification.
Assembly: hg38
Supplementary files format and content: SnRNA-seq: feature read count matrix (.txt)  files.   Includes cell barcodes, genes and count. SnATAC-seq: peak, matrix, barcode file.

molecule: total RNA

organism: Homo sapiens

source_name: Brain

extract_protocol:
Nuclei were extracted from frozen tissues following the “Frakenstein” protocol developed by Luciano Martelotto, Ph.D., Melbourne, Centre for Cancer Research, Victorian Comprehensive Cancer Centre, and available from 10X Genomics
For snRNA-seq(frozen tissues), ~15,000 nuclei were loaded per capture. Single-nucleus capture, reverse transcription, cell lysis, and library preparation were performed per manufacturer’s protocol. Sequencing was performed on an Illumina NovaSeq using a paired-end 100bp protocol.

tag: progression

tag: diagnosis

value: Brain tumor


title: iPS-derived Endothelial Cell Clone 5A No Dox Replicate 2

city: Worcester

first: Jennifer

middle: Eunmi

last: Moon

state: MA

address: 55 Lake Ave N

institute: University of Massachusetts Medical School

description:
EC_DE_Summary.tab.txt
EC_TRANSC5AND_Rep2

data_processing:
For all samples, FastQC was used to evaluate quality of data.
Libraries were aligned to genome build GRch38 using HISAT2 (Kim et al., 2015, 2019)
Mapped reads were quantified using featureCounts (Liao et al., 2014).
Differential expression analysis was conducted by using the R package edgeR (McCarthy et al. 2012)
Genome_build: GRch38
Supplementary_files_format_and_content: Tab delimited files containg differentially expressed genes generated by edgeR within R (McCarthy et al., 2012)

molecule: polyA RNA

growth_protocol:
Differentiation of iPS cells into endothelial cells were adapted from Lian et al. (2014) and Patsch et al. (2015). Cells were treated with 8 uM CHIR99021 (Tocris Bioscience) for on day 0 and day 1, and 2 uM CHIR99021 on day 2-4. Day 5, cells were collected for CD34+ were selected using the CD34 MicroBead Kit (Miltenyi Biotec). Purified CD34 cells were cultured on collagen coated plates with EGM-2 media (Lonza) supplemented with 25 ng/ml VEGF165 (PEPROTECH).
iPS cells were grown in feeder free conditions on vitronectin coated plates and cultured in Essential E8 medium (Invitrogen). Differentiation of iPS cells into endothelial cells were adapted from Lian et al. (2014) and Patsch et al. (2015). Cells were treated with 8 uM CHIR99021 (Tocris Bioscience) for on day 0 and day 1, and 2 uM CHIR99021 on day 2-4. Day 5, cells were collected for CD34+ were selected using the CD34 MicroBead Kit (Miltenyi Biotec). Purified CD34 cells were cultured on collagen coated plates with EGM-2 media (Lonza) supplemented with 25 ng/ml VEGF165 (PEPROTECH).

extract_protocol:
RNA samples were extracted using Trizol Reagent (ThermoFisher) adapted from the manufacturer’s instructions, using 100% ethanol in place of isopropanol. Precipitated RNA was resuspended in RNase- and DNase-free water and DNA digestion was performed using DNase I (Roche) with 2U/ul of RNasin Plus (Promega) for 1 hr at room temperature. RNeasy Mini Kit was used for RNA purification and DNase I removal following the RNA Cleanup protocol from the manufacturer’s instructions.
The NEBNext Ultra™ II Directional RNA Library Prep Kit for Illumina, NEBNext Poly(A) mRNA Magnetic Isolation Module, and NEBNext Multiplex Oligos for Illumina were used to prepare sequencing libraries following the manufacturer’s instructions (New England Biolabs, Inc). All libraries were prepared with 1 ug of starting RNA.

treatment_protocol: - or + doxycycline (500 ng/ml); 6 days for iPS cells and 14 days for iPS-derived endothelial cells

tag: cell type

value: iPS cell line derived Endothelial Cells

tag: cell line type

value: Clone 5A

tag: treatment

value: control


title: Day7.60min_APRIL_D3

city: York

first: Matthew

middle: Anthony

last: Care

state: North Yorkshire

address: Wentworth Way

department: Department of Biology

country: United Kingdom

institute: The University of York

data_processing:
The fastq files were assessed for initial quality using FastQC v0.11.8, trimmed for adapter sequences using TrimGalore v0.6.0  and aligned to GRCh38.p12 (Gencode release 28) with STAR aligner (v2.6.0c) using twopassMode.
Transcript abundance was estimated using RSEM v1.3.0 and imported into R v3.5.1 using txImport v1.10.1 and then processed using DESeq2 v1.22.2, generating VST transformed data.
Assembly: GRCh38.p13/hg38 (Gencode release 35)
Supplementary files format and content: Tab separated sheets containing DESeq2 VST transformed data

source_name: Peripheral blood

growth_protocol: Total B-cells were isolated by negative selection with the Memory B-cell Isolation Kit (Miltenyi). Cells were maintained in Iscove’s Modified Dulbecco Medium (IMDM) supplemented with Glutamax and 10% heat-inactivated fetal bovine serum (HIFBS, Invitrogen); Lipid Mixture 1, chemically defined and MEM Amino Acids solution (both at 1x final concentration) were added from day 3 onwards. Day 0 to day 3 – B cells were cultured in 24 well plates at 2.5 X 10^5/ml with IL2 (20 U/ml), IL21 (50 ng/ml), F(ab’)2 goat anti-human IgM & IgG (2 µg/ml) on γ-irradiated CD40L expressing L-cells (6.25 X 10^4/well). Day 3 to day 6 – At day 3, cells were detached from the CD40L L-cell layer and reseeded at 1 X 10^5/ml in media supplemented with IL2 (20 U/ml) and IL21 (50 ng/ml). Gene expression data sets were generated from differentiating PBs (day 7). At day 6, PBs from 3 healthy donors were seeded at 1 X 10^6/ml in phenol red free IMDM supplemented with 0.5% HIFBS, IL6 (10 ng/ml), IL21 (10 ng/ml) and GSI (100nM) and incubated for 20 hours. A pre-treatment sample (0 minutes) and post treatment samples were removed at +60, and +360 minutes following treatments with Multimeric-APRIL (100ng/ml), sCD40L(100ng/ml), SDF1(10ng/ml) and IFNα (100U/ml).

extract_protocol:
Peripheral blood was obtained from healthy donors after informed consent. Mononuclear cells were isolated by Lymphoprep (Axis Shield) density gradient centrifugation. Total B-cells were isolated by negative selection with the Memory B cell Isolation Kit (Miltenyi).
RNA was extracted with TRIzol (Invitrogen) then subjected to DNAseI treatment (DNAfree, Ambion).
RNA samples were quantified with a Qubit HS RNA assay kit (Thermo Fisher Scientific) and the quality was assessed by RNA HS assay using the TapeStation system (Agilent). Sequencing libraries were generated by using a TruSeq Stranded Total RNA Human/Mouse/Rat kit (Illumina) according to the manufacturer’s protocol. Library performance was checked on the TapeStation followed by picogreen quantification. Equimolar pooling was performed across all the libraries and the final pool was sequenced on a NextSeq 500 platform (Illumina) using 76-bp single-end sequencing.

tag: time

tag: condition

value: APRIL

value: Plasmablast


title: IR_Control_48hrs

description: RNA-Seq


title: iPS-derived Endothelial Cell Clone 5 No Dox Replicate 1

description:
EC_DE_Summary.tab.txt
EC_TRANSC5ND_Rep1

value: Clone 5


title: T5224_3days_treated

description: Single Cell RNA-Seq


title: iPS-derived Endothelial Cell Clone 1 Dox Replicate 3

description:
EC_DE_Summary.tab.txt
EC_TRANSC1D_Rep3

value: Clone 1

value: doxycycline


title: T5224_3days_control

description: Single Cell RNA-Seq


title: iPS-derived Endothelial Cell Clone 1 Dox Replicate 2

description:
EC_DE_Summary.tab.txt
EC_TRANSC1D_Rep2


title: ATAC-Seq PANC1 cells, PARP1 shRNA stable KD, replicate 3

city: San Diego

first: Xia

last: Ding

address: 10724 Science Center Dr

institute: Pfizer, Inc.

library_source: genomic

data_processing:
For ATAC-seq data analysis, paired-end sequencing reads for each sample were mapped to hg38 human reference genome with Bowtie2 using the following parameters: -t -X 2000 --no-mixed --no-discordant --trim-to 3:30.
Sam files were converted into Bam with Samtools. Peak calling was performed with MACS2 using these parameters: callpeak -g hs --nolambda --nomodel --shift -100 --extsize 200 --bdg --SPMR -q 5e-2. Peaks overlap with ENCODE black list regions were removed from downstream analysis using Bedtools. Bedgraph files were converted to Bigwig using bedGraphToBigWig
To correct for ATAC-seq bias, we extracted genome wide signal at 100 bp resolution using DeepTools, and performed quantile normalization on the signal data using the R package preprocessCore (https://rdrr.io/bioc/preprocessCore/).
Differential peak calling between the control and drug treated samples was performed with DESeq2
Assembly: HG38
Supplementary files format and content: bigwig

molecule: genomic DNA

growth_protocol: Cells were maintained in RPMI1640+10%FBS

extract_protocol:
Permeabilized nuclei were obtained by resuspending cells in 250 µL Nuclear Permeabilization Buffer [(10 mM Tris-HCl (pH 7.4) (BP-152, Thermo Fisher Scientific), 10 mM NaCl (S271-3, Thermo Fisher Scientific), 3 mM MgCl2 (194698, MP Biomedicals), 0.1% Tween-20 (P7949, Sigma), 0.1% IGEPAL-CA630 (I8896, Sigma), 0.01% digitonin (G9441, Promega) in Molecular biology water (46000-CM, Corning)], and incubating for 5 min on a rotator at 4°C. Nuclei were then pelleted by centrifugation for 5 min at 500 × g at 4°C. The pellet was resuspended in 25µL ice-cold Tagmentation Buffer [33 mM Tris-acetate (pH = 7.8) (BP-152, Thermo Fisher Scientific), 66 mM K-acetate (P5708, Sigma), 11 mM Mg-acetate (M2545, Sigma), 16 % DMF (DX1730, EMD Millipore) in Molecular biology water (46000-CM, Corning)]. An aliquot was then taken and counted by hemocytometer to determine nuclei concentration. Approximately 50,000 nuclei were resuspended in 10 µL ice-cold Tagmentation Buffer, and incubated with 1µL Tagmentation enzyme (FC-121-1030; Illumina) at 37 °C for 30 min with shaking 500 rpm. The tagmentated DNA was purified using MinElute PCR purification kit (28004, Qiagen).
The libraries were amplified using NEBNext High-Fidelity 2X PCR Master Mix (M0541, NEB) with primer extension at 72°C for 5 min, denaturation at 98°C for 30s, followed by 8 cycles of denaturation at 98°C for 10 s, annealing at 63°C for 30 s and extension at 72°C for 60 s. Amplified libraries were then purified using MinElute PCR purification kit (28004, Qiagen), and two size selection steps were performed using SPRIselect bead (B23317, Beckman Coulter) at 0.55X and 1.5X bead-to-sample volume rations, respectively. All libraries were sequenced on an Illumina NovaSeq 6000 in paired-end mode.

tag: cell line

value: Pancreatic cancer cell

tag: chip antibody

value: NA (ATACseq)


title: ATAC-Seq PANC1 cells, nonsense shRNA, replicate 3


title: iPS-derived Endothelial Cell Clone 1 No Dox Replicate 3

description:
EC_DE_Summary.tab.txt
EC_TRANSC1ND_Rep3


title: ATAC-Seq PANC1 cells, nonsense shRNA, replicate 2


title: ATAC-Seq PSN1 cells, Talazoparib treated 50 nM, 72 hr, replicate 3


title: ATAC-Seq PSN1 cells, Talazoparib treated 50 nM, 72 hr, replicate 2


title: ATAC-Seq PANC1 cells, nonsense shRNA, replicate 1


title: ATAC-Seq PSN1 cells, Talazoparib treated 50 nM, 72 hr, replicate 1


title: iPS cell line Disomy (322-3) Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
DIS322-3D_Rep2

growth_protocol:
iPS cells were grown in feeder free conditions on vitronectin coated plates and cultured in Essential E8 medium (Invitrogen).
iPS cells were grown in feeder free conditions on vitronectin coated plates and cultured in Essential E8 medium (Invitrogen). Differentiation of iPS cells into endothelial cells were adapted from Lian et al. (2014) and Patsch et al. (2015). Cells were treated with 8 uM CHIR99021 (Tocris Bioscience) for on day 0 and day 1, and 2 uM CHIR99021 on day 2-4. Day 5, cells were collected for CD34+ were selected using the CD34 MicroBead Kit (Miltenyi Biotec). Purified CD34 cells were cultured on collagen coated plates with EGM-2 media (Lonza) supplemented with 25 ng/ml VEGF165 (PEPROTECH).

value: induced pluripotent stem cells


title: ATAC-Seq PANC1 cells, PARP1 shRNA stable KD, replicate 1


title: ATAC-Seq PSN1 cells, DMSO treated, replicate 2


title: iPS-derived Endothelial Cell Clone 1 No Dox Replicate 1

description:
EC_DE_Summary.tab.txt
EC_TRANSC1ND_Rep1


title: ATAC-Seq PSN1 cells, DMSO treated, replicate 1


title: iPS-derived Endothelial Cell Clone 1 No Dox Replicate 2

description:
EC_DE_Summary.tab.txt
EC_TRANSC1ND_Rep2


title: iPS cell line Disomy (322-3) Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
DIS322-3D_Rep1


title: CH05 10X

city: New York

first: Neville

last: Dusaj

state: NY

institute: Weill Cornell Medicine

data_processing:
10x data was processed using Cell Ranger 3.0.1 using default parameters. RNA reads were aligned to the human or mouse referece genome, or human and mouse reference concatenation for species mixing experiment. Cell barcodes were appended with tags "A" or "B" corresponding to the two runs of the experiment that were completed.
Amplicon reads was processed following steps: 1) Identification of reads with proper priming, 2) Identification of cell barcodes (CB) that match with the whitelisted CB, 3) Replacement of CB that are not perfectly matched with the whitelist CB, 4) Deduplication of reads, 5) Analyze reads with CB that are also observed in the 10x scRNA-seq data, 6) Collapse networks of closely related UMIs within CBs to correct for sequencing error, 7) Matching of CB/UMI pairs found in 10x scRNA-seq data, discarding those matching non-DNMT3A genes and those matching no gene below a supporting read threshold.
Genome_build: hg19
Supplementary_files_format_and_content: Processed 10X data is matrix table of gene counts per cell for each sample
Supplementary_files_format_and_content: Output matrices processed from amplicon data have three columns: barcodes (BC), number of wildtype calls (num.WT.call), and number of mutant calls (num.MUT.call).

source_name: Bone marrow aspirate from patient

extract_protocol:
Cryopreserved patient samples approved by the Institutional Review Board of the University of Chicago were thawed and stained using standard procedures (10 min, 4°C) with the surface antibody CD34-PE-Vio770 (clone AC136, Miltenyi Biotec) and DAPI (Sigma-Aldrich). Cells were then sorted for DAPI-negative, CD34+ and CD34-negative cells using BD Influx.
The standard 10x Genomics Chromium v.3 protocol was carried out according to manufacturer’s recommendations (10x Genomics, Pleasanton, CA) until after emulsion breakage and recovery of first strand cDNA. A gene-specific DNMT3A primer was spiked into 10x primer mix at 1% of the concentration of the cDNA amplification primers for the initial cDNA PCR step. Then, during the amplification step, the 10x cDNA library underwent an extra cycle of PCR beyond the manufacturer’s recommended number of cycles. After cleanup with SPRIselect, a small portion of the cDNA library, 3 µL (~10% of total) was aliquoted for targeted genotyping, and the remaining cDNA underwent the standard 10x protocol. The cDNA set aside for GoT was amplified for 3 to 4 additional cycles using KAPA HiFi HotStart ReadyMix (KAPABiosystems) and 10x primer mix to provide sufficient material for the enrichment step. After clean-up, locus-specific reverse primers and the generic forward SI-PCR were used to amplify the site of interest of the cDNA template using ~10 PCR cycles. The locus-specific reverse primers contain a partial Illumina read 2 handle, a stagger to increase the complexity of the library for optimal sequencing and a gene specific region to allow specific priming. The SI-PCR oligo (10x) anneals to the partial Illumina read 1 sequence at the 3’ end of the molecule, preserving the cell barcode (CB) and UMI. After the initial amplification and SPRI purification to remove unincorporated primers, a second PCR was performed with a generic forward PCR primer (P5_generic) to retain the CB and UMI together with an RPI-x primer (Illumina) to complete the P7 end of the library and add a sample index. The targeted amplicon library was subsequently sequenced separately. The cycle settings were as follows: 28 cycles for read 1, 98 or 130 cycles for read 2, and 8 cycles for sample index.
Single-cell RNA-seq (10X) with targeted amplification of a mutation of interest. Two runs of the experiment, A and B, were completed for sample CH05.

tag: tissue

value: Bone marrow


title: ATAC-Seq PSN1 cells, DMSO treated, replicate 3


title: iPS cell line Disomy (322-3) No Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
DIS322-3ND_Rep2


title: ATAC-Seq PANC1 cells, PARP1 shRNA stable KD, replicate 2


title: Human tumor 2

city: NASHVILLE

first: Marisol

last: Ramirez

state: TENNESSEE

address: 2220 Pierce Avenue, 571 Preston Research Building

department: Center for Quantitative Sciences, Department of Biostatisticss

institute: Vanderbilt University Medical Center

data_processing:
DropEst pipeline was used to process scRNA-seq data and to generate count matrices of each gene in each cell (Petukhov et al., 2018). Specifically, cell barcodes and UMIs were extracted by dropTag, reads were aligned to the human reference transcriptome hg38 using STAR (Dobin et al., 2013) and cell barcode errors were corrected and gene-by-cell count matrices and three other count matrices for exons, introns and exon/intron spanning reads were measured by dropEst. Spliced and unspliced reads were annotated and RNA expression dynamics of single cells were estimated by velocyto (Manno et al., 2018).
We initially filter out cells with < 100 genes using scanpy.pp.filter_cells with min_genes = 100. This reduces the total number of cells (in both tumors) from 8,649 to 4,863. We remove genes found in < 3 total spliced counts across all samples with scanpy.pp.filter_genes with min_counts < 3. This reduces the number of genes from 43,306 to 15,344. These filtering steps ensure we remove any cells or genes with low or no reads, to prepare for further filtering steps below.
We then use Dropkick v1.2.6 (https://github.com/KenLauLab/dropkick) to label and filter out low quality cells from each sample. Once each sample has dropkick scores and labels (using dropkick score > 0.5 to identify high quality cells), we then concatenate the datasets with a batch key for each tumor.
We normalize the data using scanpy.pp.normalize_total and used default arguments for this normalization (as described in the Scanpy API v 1.8), such that after normalization, each cell has a total count equal to the median of total counts for cells before normalization. Next, numpy.log1p is used to log transform the data (+1, which keeps zeroes from being transformed to negative infinity). Finally, the log-transformed, normalized counts are then scaled using scapy.pp.scale, which rescales the data to unit variance and mean-centers each gene. We then use scanpy.tl.pca to compute a 50-component PCA embedding of the data, using all genes, and use scvelo.pp.neighbors and scvelo.tl.umap to generate a UMAP dimensionality reduction.
We use Scrublet to determine the number of possible doublets in the data. One tumor had 6 predicted doublets (out of 7741 original cells before filtering); the other had 3 (out of 580 original cells), which were removed.  Because these samples may contain non-tumor cells, we annotated clusters by cell type based on expression of tissue compartment markers. First, we clustered cells by the Leiden algorithm and then analyzed expression of markers across these clusters. To remove immune cells, we filtered clusters by expression of PTPRC. To remove fibroblasts, we filtered cells used COL1A1 expression, and we used CLDN5 expression to remove endothelial cells. We also used EPCAM to identify epithelial cells. We found several small clusters of immune cells and a single small population of likely fibroblasts. A single cluster had a few cells with low expression of CLDN5, and higher average expression of EPCAM, so we chose not to remove this cluster. Removing the other non-cancer cells reduced the number of cells from 4,863 to 4,485.   After removing so many cells, we re-filter the genes with a low threshold (min_cells = 3) to remove any genes that were only expressed in the low-quality cells. This reduces the number of genes from 15,344 to 13,938. Therefore, we moved forward with the analysis with 1618 cells and 13,938 across two tumors.
We use Scrublet to determine the number of possible doublets in the data. One tumor had 6 predicted doublets (out of 7741 original cells before filtering); the other had 3 (out of 580 original cells), which were removed.
Because these samples may contain non-tumor cells, we annotated clusters by cell type based on expression of tissue compartment markers. First, we clustered cells by the Leiden algorithm and then analyzed expression of markers across these clusters. To remove immune cells, we filtered clusters by expression of PTPRC. To remove fibroblasts, we filtered cells used COL1A1 expression, and we used CLDN5 expression to remove endothelial cells. We also used EPCAM to identify epithelial cells. We found several small clusters of immune cells and a single small population of likely fibroblasts. A single cluster had a few cells with low expression of CLDN5, and higher average expression of EPCAM, so we chose not to remove this cluster. Removing the other non-cancer cells reduced the number of cells from 4,863 to 4,485.
After removing so many cells, we re-filter the genes with a low threshold (min_cells = 3) to remove any genes that were only expressed in the low-quality cells. This reduces the number of genes from 15,344 to 13,938. Therefore, we moved forward with the analysis with 1618 cells and 13,938 across two tumors.
Genome_build: Hg38
Supplementary_files_format_and_content: 3359-PK-1-GCCAAT-ATCAGT_S1_L001.loom, 236D_236D_V300044428.loom. Loom format containing spliced, unspliced, ambiguous and spanning matrices.

source_name: Human SCLC tumor

growth_protocol: The two human SCLC tumors were collected in collaboration with Vanderbilt University Medical Center. Tumor was immediately placed on cold RPMI on ice for dissociation for sequencing.

extract_protocol:
Tissue was washed in an RBC lysis buffer, passed through a 70 μm filter, and washed in PBS. Cells were dissociated with cold DNAse and proteases and titrated every 5-10 minutes to increase dissociation.
Tumor 1: . Library preparation for scRNA-seq was performed according to previous protocols (Banerjee et al., 2020), and cells were sequenced using BGI MGI-seq.   Tumor 2: Tumor was immediately placed in cold RPMI on ice for dissociation. Library preparation for scRNA-seq was performed as described previously (Banerjee et al., 2020). Cells were prepared for sequencing using TruDrop (Southard-Smith et al., 2020) and sequenced on Nova-seq.

treatment_protocol: Both patients received etoposide and cisplatin. Tumor 1 came from a patient who also received prophylactic cranial irradiation

value: SCLC tumor

tag: collection method

value: Upper right lobectomy

tag: stage/treatment

value: Stage 1B mixed SCLC/LCNEC, relapsed (EP standard of care)


title: CH05 Amplicon


title: 132 10C10-DMSO10 rep1

city: Nantes

first: Delphine

last: Fradin

address: 8 quai Moncousu

country: France

institute: CRCI2NA, Inserm UMR1307, CNRS UMR6075

data_processing:
Pipeline details have been published by Charpentier E et al. 2018 (10.21203/rs.3.pex-1336/v1)
Genome_build: Hg38
Supplementary_files_format_and_content: Matrix table of raw gene counts for all genes in all samples

source_name: monoclonal CD8+

growth_protocol: CD8+ T cells were grown during 24h in RPMI+8%human serum

extract_protocol:
miRNeasy kit (Qiagen)
Libraries were prepared according to NovaSeq 6000 Sequencing Sytem's instructions accompanying the NovaSeq 6000 SP Reagent Kit (Ref# 20028401). Libraries were sequenced on the NovaSeq 6000 following the manufacturer's protocols.

treatment_protocol: DCMU or vehicle (DMSO) was added in the media during 24h prior to RNA extraction

tag: cell population

value: monoclonal CD8+

tag: exposition

value: DMSO 10uM

tag: replicate


title: Human tumor 1

value: bronchoscopy with transbronchial needle aspiration of a left hilar mass

value: Relapsed


title: iPS cell line Disomy (322-2) Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
DIS322-2D_Rep1


title: 165 CD8pol-DCMU10 rep2

source_name: PBMC

value: polyclonal CD8+

value: DRN 10uM


title: 157 CD8pol-DCMU250 rep1

value: DRN 250uM


title: iPS cell line Disomy (322-3) No Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
DIS322-3ND_Rep1

value: control


title: ADMA treated HUVEC

type: RNA

city: Beijing

first: Donglinag

last: Zhang

state: Beijing

address: Xinjiekou East Street No.31, Xicheng District, Beijing, China

country: China

institute: Beijing Jishuitan hospital

description: ADAM treated HUEVC

hyb_protocol: standard Affymetrix protocol

scan_protocol: standard Affymetrix protocol

data_processing: RMA

label: ADMA

source_name: ADAM treated HUVEC

label_protocol: standard Affymetrix protocol

extract_protocol: none provided by the submitter

value: HUVEC


title: iPS cell line Disomy (322-2) No Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
DIS322-2ND_Rep2


title: Control

description: Normal culture HUEVC

label: NC

source_name: Normal culture HUVEC


title: iPS-derived Endothelial Cell Clone 1 Dox Replicate 1

description:
EC_DE_Summary.tab.txt
EC_TRANSC1D_Rep1

value: iPS cell line derived Endothelial Cells

value: Clone 1

value: doxycycline


title: iPS cell line Disomy (322-2) Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
DIS322-2D_Rep2

value: induced pluripotent stem cells


title: 174 CD8pol-DMSO10 rep4


title: iPS cell line Disomy (322-2) No Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
DIS322-2ND_Rep1


title: 134 10C10-DCMU100 rep1

value: DRN 100uM


title: 149 10C10-DMSO100 rep4

value: DMSO 100uM


title: 156 CD8pol-DMSO10 rep1


title: 158 CD8pol-DCMU100 rep1


title: iPS cell line Trisomy (32436B) Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
TRI32436BD_Rep2

value: Trisomy (32436B)


title: iPS cell line Disomy (32316) No Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
DIS32316ND_Rep1


title: iPS cell line Trisomy (32436B) Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
TRI32436BD_Rep1


title: CD8+ T cells exposed to DCMU at 100µM during 24h (rep 3)

description: miRNA data from CD8+ T cells

hyb_protocol: Labeled samples were hybridized to the arrays at 48°C for 20h and then washed and stained with a Streptavidin-PE solution  After overnight hybridization, chips are washed on the fluidic station FS450 following specific protocols (Affymetrix) and scanned using the GCS3000 7G

data_processing: The scanned images are then analyzed with Expression Console software (Affymetrix) to obtain raw data (cel files) and metrics for Quality Controls.  RMA normalization is performed using R with Version 17 of Entrezgene CDF brain array.

label: biotin

source_name: CD8+ T cells purified from healthy PBMC

label_protocol: 4 ng of total RNA was labeled using the FlashTag Biotin RNA Labeling Kit (Thermofisher).Briefly,  RNA was poly-A tailed and a proprietary biotin-labeled dendramer molecule was joined to the 3′ end using DNA ligase

extract_protocol: Trizol extraction of total RNA was performed according to the manufacturer's instructions.

tag: tumor stage

value: healthy

value: DCMU at 100uM


title: CD8+ T cells exposed to DCMU at 250µM during 24h (rep 3)

description: miRNA data from CD8+ T cells

scan_protocol: After overnight hybridization, chips are washed on the fluidic station FS450 following specific protocols (Affymetrix) and scanned using the GCS3000 7G

value: DCMU at 250uM


title: CD8+ T cells exposed to DCMU at 10µM during 24h (rep 3)

description: miRNA data from CD8+ T cells

value: DCMU at 10uM


title: iPS cell line Disomy (32316) Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
DIS32316D_Rep1

source_name: iPS cell line Disomy (32316) No Dox Replicate 1


title: 173 CD8pol-DMSO100 rep4


title: 147 10C10-DCMU10 rep3

value: monoclonal CD8+


title: iPS cell line Disomy (32316) No Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
DIS32316ND_Rep2


title: 169 CD8pol-DCMU250 rep3

value: polyclonal CD8+


title: 137 10C10-DMSO100 rep2


title: 141 10C10-DCMU10 rep2

value: DRN 10uM


title: iPS cell line Trisomy (32436B) No Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
TRI32436BND_Rep1


title: CD8+ T cells exposed to DCMU at 10µM during 24h (rep 2)

description: miRNA data from CD8+ T cells


title: iPS cell line Trisomy (31627H) Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
TRI31627HD_Rep2

value: Trisomy (31627H)


title: iPS cell line Trisomy (31627H) Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
TRI31627HD_Rep1


title: 133 10C10-DCMU250 rep1

value: DRN 250uM


title: 145 10C10-DCMU250 rep3


title: iPS cell line Trisomy (32436B) No Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
TRI32436BND_Rep2

value: control


title: iPS cell line Disomy (32316) Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
DIS32316D_Rep2

source_name: iPS cell line Disomy (32316) No Dox Replicate 2


title: iPS cell line Trisomy (31627H) No Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
TRI31627HND_Rep2


title: CD8+ T cells exposed to DCMU at 250µM during 24h (rep 2)

description: miRNA data from CD8+ T cells


title: 142 10C10-DMSO250 rep3

value: DMSO 250uM


title: 175 CD8pol-DCMU250 rep4


title: iPS cell line Clone 7 Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
TRANSC7D_Rep2

source_name: iPS cell line Clone 7 Dox Replicate 2

value: Clone 7


title: iPS cell line Trisomy (31627H) No Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
TRI31627HND_Rep1


title: iPS cell line Clone 7 Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
TRANSC7D_Rep1

source_name: iPS cell line Clone 7 Dox Replicate 1

value: doxycycline


title: 138 10C10-DMSO10 rep2

value: DMSO 10uM


title: 161 CD8pol-DMSO100 rep2


title: 170 CD8pol-DCMU100 rep3

value: DRN 100uM


title: CD8+ T cells exposed to DMSO at 10µM during 24h (rep 3)

description: miRNA data from CD8+ T cells

value: DMSO at 10uM


title: iPS cell line Clone 7 No Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
TRANSC7ND_Rep1

value: induced pluripotent stem cells


title: CD8+ T cells exposed to DMSO at 100µM during 24h (rep 2)

description: miRNA data from CD8+ T cells

value: healthy

value: DMSO at 100uM


title: 130 10C10-DMSO250 rep1


title: 140 10C10-DCMU100 rep2


title: iPS cell line Clone 7 No Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
TRANSC7ND_Rep2


title: iPS cell line Clone 5A Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
TRANSC5AD_Rep1

source_name: iPS cell line Clone 5A Dox Replicate 1

value: Clone 5A


title: CD8+ T cells exposed to DMSO at 250µM during 24h (rep 2)

description: miRNA data from CD8+ T cells

value: DMSO at 250uM


title: 152 10C10-DCMU100 rep4

value: monoclonal CD8+


title: iPS cell line Clone 5A Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
TRANSC5AD_Rep2

source_name: iPS cell line Clone 5A Dox Replicate 2


title: iPS cell line Clone 5A No Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
TRANSC5AND_Rep2


title: CD8+ T cells exposed to DCMU at 100µM during 24h (rep 2)

description: miRNA data from CD8+ T cells

value: DCMU at 100uM


title: CD8+ T cells exposed to DMSO at 10µM during 24h (rep 2)

description: miRNA data from CD8+ T cells


title: CD8+ T cells exposed to DMSO at 250µM during 24h (rep 3)

description: miRNA data from CD8+ T cells


title: CD8+ T cells exposed to DMSO at 100µM during 24h (rep 3)

description: miRNA data from CD8+ T cells


title: 153 10C10-DCMU10 rep4

value: DRN 10uM


title: CD8+ T cells exposed to DCMU at 10µM during 24h (rep 1)

description: miRNA data from CD8+ T cells

value: DCMU at 10uM


title: CD8+ T cells exposed to DCMU at 250µM during 24h (rep 1)

description: miRNA data from CD8+ T cells

value: DCMU at 250uM


title: CD8+ T cells exposed to DMSO at 250µM during 24h (rep 1)

description: miRNA data from CD8+ T cells


title: CD8+ T cells exposed to DMSO at 100µM during 24h (rep 1)

description: miRNA data from CD8+ T cells


title: CD8+ T cells exposed to DCMU at 100µM during 24h (rep 1)

description: miRNA data from CD8+ T cells


title: CD8+ T cells exposed to DMSO at 10µM during 24h (rep 1)

description: miRNA data from CD8+ T cells


title: 131 10C10-DMSO100 rep1

value: DMSO 100uM


title: 167 CD8pol-DMSO100 rep3

value: polyclonal CD8+


title: iPS cell line Clone 5 Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
TRANSC5D_Rep2

source_name: iPS cell line Clone 5 Dox Replicate 2

value: Clone 5


title: iPS cell line Clone 5 No Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
TRANSC5ND_Rep1

value: control


title: iPS cell line Clone 5 Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
TRANSC5D_Rep1

source_name: iPS cell line Clone 5 Dox Replicate 1

value: doxycycline


title: iPS cell line Clone 5A No Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
TRANSC5AND_Rep1


title: iPS cell line Clone 1 Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
TRANSC1D_Rep2

source_name: iPS cell line Clone 1 Dox Replicate 2

value: Clone 1


title: 146 10C10-DCMU100 rep3

value: DRN 100uM


title: iPS cell line Clone 1 Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
TRANSC1D_Rep1

source_name: iPS cell line Clone 1 Dox Replicate 1

value: induced pluripotent stem cells


title: 144 10C10-DMSO10 rep3

value: DMSO 10uM


title: Diet_AspClo_63 CD-HFD Asp/Clo

city: Barcelona

first: Sara

last: Torrecilla

state: Catalonia

address: Rossello Street, 149

country: Spain

institute: IDIBAPS

description: Gene expression from Non-tumor liver tissue from diet-induced model (CD-HFD) of NASH-HCC prophylactically treated with Aspirin and Clopidogrel

hyb_protocol: Standard Affymetrix protocol

scan_protocol: Standard Affymetrix protocol

data_processing: Raw expression data from microarrays were normalized using the ExpressionFileCreator [RMA with quantile normalization method].

organism: Mus musculus

label_protocol: Standard Affymetrix protocol

growth_protocol: As previously published (Malehmir M et al, Nat Med, 2019)

extract_protocol: Total RNA was extracted from 20 mg fresh-frozen non-tumoral liver tissue with Trizol reagent (Invitrogen) and purified with RNeasy columns (Qiagen, Valencia, CA). RNA sample concentration and quality were assessed by NanoDrop ND-1000 spectrometer (NanoDrop, Wilmington, DE) and bioanalyzer (Agilent, Palo Alto, CA), respectively.

tag: liver status

value: Chronic damage

value: liver

value: Asp/Clo


title: Diet_AspClo_62 CD-HFD Asp/Clo

source_name: Non-tumor liver tissue from diet-induced model (CD-HFD) of NASH-HCC prophylactically treated with Aspirin and Clopidogrel

treatment_protocol: As previously published (Malehmir M et al, Nat Med, 2019)


title: iPS cell line Clone 1 No Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
TRANSC1ND_Rep2


title: Diet_AspClo_60 CD-HFD Asp/Clo


title: 135 10C10-DCMU10 rep1

value: monoclonal CD8+


title: 172 CD8pol-DMSO250 rep4

value: DMSO 250uM


title: iPS cell line Clone 5 No Dox Replicate 2

description:
IPSCD6_DE_Summary.tab.txt
TRANSC5ND_Rep2


title: Diet_Sulin_21 CD-HFD Sulindac

description: Gene expression from Non-tumor liver tissue from diet-induced model (CD-HFD) of NASH-HCC prophylactically treated with Sulindac

value: Sulindac


title: Diet_AspClo_58 CD-HFD Asp/Clo


title: 177 CD8pol-DCMU10 rep4

value: DRN 10uM


title: Diet_AspClo_64 CD-HFD Asp/Clo


title: Diet_Sulin_17 CD-HFD Sulindac

source_name: Non-tumor liver tissue from diet-induced model (CD-HFD) of NASH-HCC prophylactically treated with Sulindac


title: Diet_Sulin_15 CD-HFD Sulindac


title: Diet_Sulin_29 CD-HFD Sulindac


title: 139 10C10-DCMU250 rep2

source_name: monoclonal CD8+

value: DRN 250uM


title: 143 10C10-DMSO100 rep3

value: DMSO 100uM


title: iPS cell line Clone 1 No Dox Replicate 1

description:
IPSCD6_DE_Summary.tab.txt
TRANSC1ND_Rep1


title: Diet_AspClo_61 CD-HFD Asp/Clo


title: 154 CD8pol-DMSO250 rep1

source_name: PBMC

value: polyclonal CD8+


title: Diet_Sulin_26 CD-HFD Sulindac


title: ATAC-seq AMs in vitro - P4 20h rep2

city: Palo Alto

first: Lawrence

middle: Stephen

last: Prince

address: 1701 Page Mill Rd

department: Pediatrics

institute: Stanford University

data_processing:
RNAseq reads were trimmed using Trimmomatic 0.36 using the following parameters: LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 MINLEN:27
Trimmed reads were aligned using STAR v2.5.3a using default parameters
Reads were quantified using RSEM v1.3.0 using default parameters
ATAC samples were mapped to the mm10 genome using Bowtie2
IDR peak files were generated for each condition with IDR 2.0
Genome_build: mm10
Supplementary_files_format_and_content: Raw counts matrices for each ENSEMBL gene and sample. .idr peaks files

source_name: Alveolar macrophages

extract_protocol:
Alveolar macrophages (25,000) were lysed in 25 ml lysis buffer (10 mM Tris-HCl pH7.5, 10mM NaCl, 3mM MgCl2, 0.1% IGEPAL CA-630), followed by addition of 1.25 ml Tagment DNA enzyme I (Illumina 15027865). DNA was extracted with the Zymogen ChIP DNA purification kit (Zymo).
DNA was amplified using the Nextera Primer Ad1, Nextera Barcode Primers Ad2.n, SYBR Green, and NEBNext High-Fidelity 2X PCR Master Mix for 5-15 cycles. PCR reactions were size selected for 160-280 bp by gel extraction (10% TBE gels, Life Technologies EC62752BOX) and then 75 bp single-end sequenced for 51 cycles on the Illumina HiSeq 4000 following the manufacturer's protocols.

treatment_protocol: Neonatal (postnatal day 4) and adult (8 to 10 weeks of age) mice were challenged with 5 ml or 50 ml of LPS diluted in sterile, endotoxin-free phosphate-buffered saline (PBS) respectively, at 0.1 mg/g body weight. The control mice received sterile PBS. All mice were euthanized 2h after intranasal delivery of LPS or PBS.

tag: strain

value: 20h, in vitro culture

tag: age

value: postnatal day 4


title: ATAC-seq AMs in vitro - P4 0h rep2

value: 0h, in vitro culture


title: 166 CD8pol-DMSO250 rep3


title: Diet_Sulin_23 CD-HFD Sulindac


title: 163 CD8pol-DCMU250 rep2


title: ATAC-seq AMs in vitro - 8W 20h rep1

treatment_protocol: Alveolar macrophages were sorted from neonatal (postnatal day 4) and adult (8 to 10 weeks of age) mice and plated in 96-well plates at 60,000 to 100,000 cells/well. Culture media consisted of RPMI-1640 supplemented with 10% FBS, 100 U/mL penicillin/streptomycin, and 4 ng/mL recombinant murine M-CSF (R&D, 416-ML010). Cells were harvested at 0 h and 20 h of tissue culture.

value: 8 weeks


title: Diet_Sulin_25 CD-HFD Sulindac


title: ATAC-seq AMs in vitro - P4 0h rep1


title: ATAC-seq AMs in vitro - 8W 20h rep2


title: ATAC-seq AMs in vitro - P4 20h rep1


title: 151 10C10-DCMU250 rep4


title: 162 CD8pol-DMSO10 rep2


title: 176 CD8pol-DCMU100 rep4

value: DRN 100uM


title: ATAC-seq AMs in vitro - 8W 0h rep1


title: ATAC-seq AMs in vitro - 8W 0h rep2


title: ATAC-seq AMs in vivo - P4 LPS rep1

value: 2h, intranasal LPS instillation


title: Diet_Sulin_18 CD-HFD Sulindac


title: Diet_Sulin_20 CD-HFD Sulindac


title: Diet_Sulin_16 CD-HFD Sulindac


title: Diet_Veh_high_1831 CD-HFD Vehicle With HCC and fibrosis

description: Gene expression from Non-tumor liver tissue from diet-induced model (CD-HFD) of NASH-HCC treated with vehicle, that developed HCC and fibrosis

value: Vehicle


title: 164 CD8pol-DCMU100 rep2


title: ATAC-seq AMs in vivo - P4 LPS rep2


title: 171 CD8pol-DCMU10 rep3


title: 168 CD8pol-DMSO10 rep3

value: DMSO 10uM


title: 159 CD8pol-DMSO10 rep1


title: 136 10C10-DMSO250 rep2


title: Diet_Veh_high_2088 CD-HFD Vehicle With HCC and fibrosis

source_name: Non-tumor liver tissue from diet-induced model (CD-HFD) of NASH-HCC treated with vehicle, that developed HCC and fibrosis

value: Chronic damage

value: liver


title: ATAC-seq AMs in vivo - P4 PBS rep1

value: 2h, intranasal PBS instillation


title: ATAC-seq AMs in vivo - P4 PBS rep2


title: Diet_Veh_high_1770 CD-HFD Vehicle With HCC and fibrosis


title: ATAC-seq AMs in vivo - 8W LPS rep2


title: 148 10C10-DMSO250 rep4

value: monoclonal CD8+

value: DMSO 250uM


title: Diet_Veh_low_2091 CD-HFD Vehicle Without HCC

description: Gene expression from Non-tumor liver tissue from diet-induced model (CD-HFD) of NASH-HCC treated with vehicle, that did not develop HCC and fibrosis


title: 150 10C10-DMSO10 rep4


title: Diet_Veh_low_794 CD-HFD Vehicle Without HCC

source_name: Non-tumor liver tissue from diet-induced model (CD-HFD) of NASH-HCC treated with vehicle, that did not develop HCC and fibrosis


title: ATAC-seq AMs in vivo - 8W PBS rep2


title: ATAC-seq AMs in vivo - 8W LPS rep1


title: Diet_Veh_high_798 CD-HFD Vehicle With HCC and fibrosis


title: ATAC-seq AMs in vivo - 8W PBS rep1


title: Diet_Veh_low_1819 CD-HFD Vehicle Without HCC


title: Diet_Veh_low_1810 CD-HFD Vehicle Without HCC


title: Diet_Veh_low_1825 CD-HFD Vehicle Without HCC


title: RNA-seq AMs in vitro - P4 20h rep2

extract_protocol:
RNA of alveolar macrophages were harvested using trizol reagent. TruSeq Stranded mRNA Sample Prep Kit with TruSeq Unique Dual Indexes (Illumina, San Diego, CA) was used with 60 ng of total RNA for generating sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols on the Illumina HiSeq 4000.


title: Hlth_1807 Control chow-diet

description: Gene expression from Healthy liver tissue

source_name: Healthy liver tissue

value: Healthy


title: RNA-seq AMs in vitro - P4 20h rep3


title: 160 CD8pol-DMSO250 rep2


title: SiO2_HSA_2

first: Jinlyu

last: Sun

address: No.1 Shuaifuyuan, Wangfujing, Dongcheng District

department: Department of Allergy

institute: Peking Union Medical College Hospital

data_processing:
Reads obtained from the sequencing machines includes raw reads containing adapters or low quality bases which will affect the following assembly and analysis. Thus, to get high quality clean reads, reads were further filtered by Cutadapt (https://cutadapt.readthedocs.io/en/stable/,version:cutadapt-1.9)
We aligned reads of all samples to the research species reference genome using HISAT2 (https://daehwankimlab.github.io/hisat2/,version:hisat2-2.0.4) package, which initially remove a portion of the reads based on quality information accompanying each read and then maps the reads to the reference genome.
The mapped reads of each sample were assembled using StringTie (http://ccb.jhu.edu/software/stringtie/,version:stringtie-1.3.4d) with default parameters. Then, all transcriptomes from all samples were merged to reconstruct a comprehensive transcriptome using gffcompare software(http://ccb.jhu.edu/software/stringtie/gffcompare.shtml,version:gffcompare-0.9.8). After the final transcriptome was generated, StringTie and ballgown (http://www.bioconductor.org/packages/release/bioc/html/ballgown.html) were used to estimate the expression levels of all transcripts and perform expression abundance for mRNAs by calculating FPKM (fragment per kilobase of transcript per million mapped reads) value.
Genes differential expression analysis was performed by DESeq2 software between two different groups (and by edgeR between two samples). The genes with the parameter of false discovery rate (FDR) below 0.05 and absolute fold change ≥ 2 were considered differentially expressed genes. Differentially expressed genes were then subjected to enrichment analysis of GO functions and KEGG pathways.and absolute fold change ≥ 2 were considered differentially expressed genes. Differentially expressed genes were then subjected to enrichment analysis of GO functions and KEGG pathways.
Supplementary files format and content: FPKM

source_name: Mouse bone marrow-derived mast cells

growth_protocol: Mouse bone marrow-derived mast cells (BMMCs) were isolated from BALB/c mouse femurs and cultured in complete RPMI-1640 with 100 U/mL penicillin, 100 U/mL streptomycin, 2 mM L-glutamine, 0.1 mM nonessential amino acids, 10% foetal bovine serum, 10 ng/mL IL-3, and 10 ng/mL SCF.

extract_protocol:
Total RNA was extracted using Trizol reagent following the manufacturer's procedure. The total RNA quantity and purity were analysis of Bioanalyzer 2100 and RNA 6000 Nano LabChip Kit, high-quality RNA samples with RIN number > 7.0 were used to construct sequencing library.
After total RNA was extracted, mRNA was purified from total RNA (5ug) using Dynabeads Oligo (dT) with two rounds of purification. Following purification, the mRNA was fragmented into short fragments using divalent cations under elevated temperature (Magnesium RNA Fragmentation Module under 94℃ 5-7min). Then the cleaved RNA fragments were reverse-transcribed to create the cDNA by SuperScript™ II Reverse Transcriptase, which were next used to synthesise U-labeled second-stranded DNAs with E. coli DNA polymerase I, RNase H and dUTP Solution. An A-base was then added to the blunt ends of each strand, preparing them for ligation to the indexed adapters. Each adapter contained a T-base overhang for ligating the adapter to the A-tailed fragmented DNA. Dual-index adapters were ligated to the fragments, and size selection was performed with AMPureXP beads. After the heat-labile UDG enzyme treatment of the U-labeled second-stranded DNAs, the ligated products were amplified with PCR by the following conditions: initial denaturation at 95℃ for 3 min; 8 cycles of denaturation at 98℃ for 15 sec, annealing at 60℃ for 15 sec, and extension at 72℃ for 30 sec; and then final extension at 72℃ for 5 min. The average insert size for the final cDNA librarys were 300±50 bp. At last, we performed the 2×150bp paired-end sequencing (PE150) on an Illumina Novaseq™ 6000 (LC-Bio Technology CO., Ltd., Hangzhou, China) following the vendor's recommended protocol.

value: BALB/c

tag: cell type

value: Mouse bone marrow-derived mast cells

value: nano-SiO2 exposure and DNP-HSA stimulation


title: RNA-seq AMs in vitro - P4 20h rep1

value: 20h, in vitro culture


title: GBM SF6996_snATAC

description: Single Nuceli ATAC-Seq

data_processing:
We utilized CellRanger (version 3.0.2) for data pre-processing and gene expression quantification, following the guidelines from the CellRanger website (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/advanced/references#premrna).
The CellRanger ATAC software (version 1.1.0) was used for read alignment, deduplication, and identifying transposase cut sites.
The presence/absence of somatic CNVs was assessed with CONICSmat. ELSA was used for cell type classification.
Genome_build: hg38
Supplementary_files_format_and_content: SnRNA-seq: feature read count matrix (.txt)  files.   Includes cell barcodes, genes and count. SnATAC-seq: peak, matrix, barcode file.

source_name: Brain

tag: progression

value: Primary

tag: diagnosis

value: GBM

tag: gender

value: Female

tag: pair#


title: GBM SF11857_snATAC

description: Single Nuceli ATAC-Seq

value: Recurrent

value: Male


title: RNA-seq AMs in vitro - P4 0h rep3

tag: condition

value: 0h, in vitro culture

value: postnatal day 4


title: RNA-seq AMs in vitro - P4 0h rep2


title: GBM SF9798_snATAC

description: Single Nuceli ATAC-Seq


title: Hlth_1965 Control chow-diet

tag: treatment

value: Vehicle


title: SiO2_HSA


title: 155 CD8pol-DMSO100 rep1

value: polyclonal CD8+

value: DMSO 100uM


title: SiO2_2

value: nano-SiO2 exposure


title: Hlth_1798 Control chow-diet

tag: tissue

value: liver


title: Hlth_1804 Control chow-diet

value: Healthy


title: RNA-seq AMs in vitro - P4 0h rep1


title: GBM SF11331_snATAC

description: Single Nuceli ATAC-Seq


title: GBM SF9510_snATAC

description: Single Nuceli ATAC-Seq


title: Hlth_1968 Control chow-diet


title: SiO2_HSA_1

value: BALB/c


title: RNA-seq AMs in vitro - 8W 20h rep3

value: 8 weeks


title: GBM SF7062_snATAC

description: Single Nuceli ATAC-Seq


title: GAM1.3

city: Villaviciosa

first: Natalia

last: Molinero Garcia

state: Asturias

address: IPLA-CSIC, Paseo Rio Linares S/N

department: DEPARTMENT OF MICROBIOLOGY AND BIOCHEMISTRY OF DAIRY PRODUCTS

institute: Dairy Products Institute of Asturias (IPLA-CSIC)

description:
Pure culture of R.gauvreauii IPLA60001 in M2 medium supplemented to D-glucose in 8hours Replicate 3
countscomparacion1Name.txt and countscomparacion2Name.txt
diffexpr-results-mrs_vs_pH1.csv and diffexpr-results-mrs_vs_pH2.csv

data_processing:
Alignment with their reference genomes using Bowtie2 v.2.2.6 (.sam files)
SAMtools v.1.4 was used to generate the ".bam" files and "sorted.bam" files
Read counts were generated using FeatureCounts
Differential gene expression analysis was performed using the Bioconductor package DESeq2
Genome_build: Accessions number: for R.gauvreauii IPLA60001 is SRR10273262 and for R. bilirensis IPLA60002 is SRR9209608

organism: Ruminococcus gauvreauii

source_name: Culture of Ruminococcus gauvreauii IPLA60001

growth_protocol: To study the interaction between Rc. biliarensis IPLA60002 and R. gauvreauii IPLA60001 we made co-cultures combinations in M2 broth medium supplemented with 30%CBRF (clarified bovine rumen fluid) and 0.2% Starch from Rice (Sigma-Aldrich) as only carbon source. The co-culture conditions were as follows: 10ml of initial exponential phase co-cultures of R. biliarensis IPLA60002 and R. gauvreauii IPLA60001 (6 h), and 10ml co-cultures for middle exponential phase (8 h). The strains were grown in monoculture to have control conditions: R. gauvreauii was growth in M2 medium supplemented with 30% CBRF and 0,2% D-glucose, and R. biliarensis was growth in M2 medium supplemented with 30%CBRF and 0.2% starch from rice. Three technical replicates were performed for each condition. The tubes were centrifuged 10 min and the pellet was immediately resuspended in 1ml of RNAlaterTM (Qiagen) and frozen at -80ᵒC.

extract_protocol:
Total RNA was extracted from the pellets that were resuspended in 200 μl of lysozyme (concentración) and mutanolysin (concentración) solution and incubated at 37°C for 1 h. Then, 800 μl of QIAzol (Qiagen) were added and placed in a tube containing 0.8 g of glass beads (106 μm diameter; Sigma). The cells were lysed by shaking the mix on a BioSpec homogenizer (BioSpec Products, Inc., USA) for 2 min followed by 2 min in ice. This step was repeated three times. Samples were further treated with chloroform and then centrifuged at 12,000 rpm for 15 min at 4°C. The upper phase containing the RNA was recovered. Samples were further purified using the RNeasy Mini Kit (Qiagen) following the manufacturer’s protocol. Quality and integrity of the RNA was checked by an Agilent 2200 Tape Station Nucleic Acid System (Agilent Technologies, Palo Alto, Calif., USA). The amount of RNA was quantified in a Qubit™ 4 fluorometer with Qubit RNA assay kits (Thermo Fisher Scientific, Inc., Waltham, MA, USA). One to 2.5 μg of total RNA was depleted using Ribo-Zero rRNA removal kit (Illumina) to remove rRNA according to the supplier’s instructions. The yield of rRNA depletion was checked in the Agilent Tape Station and rRNA-depleted RNA samples were treated with the TruSeq Stranded mRNA sample preparation kit (Illumina). Size evaluation and quantification of RNA samples was performed as above described.
Whole transcriptome library was sequenced by means of a NextSeq 550 Sequencing System (Illumina) using a High Output kit v2.5 (75 cycles) flow cell

tag: treaatment


title: RNA-seq AMs in vitro - 8W 20h rep2


title: GBM SF4297_snATAC

description: Single Nuceli ATAC-Seq


title: GAM1.2

description:
Pure culture of R.gauvreauii IPLA60001 in M2 medium supplemented to D-glucose in 8hours Replicate 2
countscomparacion1Name.txt and countscomparacion2Name.txt
diffexpr-results-mrs_vs_pH1.csv and diffexpr-results-mrs_vs_pH2.csv


title: GAM1.1

description:
Pure culture of R.gauvreauii IPLA60001 in M2 medium supplemented to D-glucose in 8hours Replicate 1
countscomparacion1Name.txt and countscomparacion2Name.txt
diffexpr-results-mrs_vs_pH1.csv and diffexpr-results-mrs_vs_pH2.csv


title: HSA_2

value: Mouse bone marrow-derived mast cells

value: DNP-HSA stimulation


title: SiO2


title: RNA-seq AMs in vitro - 8W 20h rep1

value: 20h, in vitro culture


title: RNA-seq AMs in vitro - 8W 0h rep3


title: HSA_1


title: GBM SF10592_snATAC

description: Single Nuceli ATAC-Seq

value: Primary

value: GBM


title: GBM SF6621_snATAC

description: Single Nuceli ATAC-Seq

value: Recurrent


title: HSA


title: SiO2_1


title: RNA-seq AMs in vitro - 8W 0h rep1


title: Blank_2

value: blank control


title: GBM SF9494_snATAC

description: Single Nuceli ATAC-Seq

value: Male


title: TS_NonLrcRep2

city: ITHACA

first: Sangeeta

last: Ghuwalewala

state: New York

address: 107 Biotechology Building 526 Campus Road

department: Molecular Biology and Genetics

institute: Cornell University

library_source: transcriptomic single cell

data_processing:
The raw data files were demultiplexed to generate the sample-specific FASTQ files, which were aligned to the mouse reference genome (mm10-3.0.0) using the 10X Genomics Cell Ranger pipeline (v3.1.0)
The raw scRNA-seq data was processed using Cell Ranger from the 10X platform to generate an expression matrix that was further analyzed in R using the Seurat package version 3.1
Principle Component Analysis (PCA) was performed on the gene expression matrix using the least number of principal components (PCs) that could be used to explain the majority of the variance in the data
The PCA embeddings were used by Harmony to integrated the datasets. Unsupervised shared nearest neighbor (SNN) clustering was done on the Harmony integrated Seurat object
Assembly: mm10-3.0.0
Supplementary files format and content: feature.tsv, barcode.tsv and matrix.mtx files for each sample

source_name: Tail skin IFE

extract_protocol:
Interfollicular epidermis was FACS sorted using alpha6-integrin+ and Sca1+ staining and doxycycline inducible K5tta promoter driven H2BGFP
Single-cell 3′ cDNA libraries generation using Chromium Single Cell 3′ gel bead and library Kit v3, following the manufacturer’s protocols (10x Genomics)

treatment_protocol: Mice were fed with doxy chow for 2 or 3 weeks before harvest.

tag: tissue/cell type

value: Tail skin IFE

value: Post natal day 51

tag: genotype

value: wild type (K5-tTA x pTRE-H2BGFP)


title: TS_MidLrcRep2

description: Interfollicular epidermis was FACS sorted using alpha6-integrin+ and Sca1+ staining and doxycycline inducible K5tta promoter driven H2BGFP


title: GBM SF4449_snRNA

description: Single Nuceli RNA-Seq

value: Female


title: TS_LrcRep2


title: Blank_1


title: RNA-seq AMs in vivo - P4 LPS rep4

value: 2h, intranasal LPS instillation

value: postnatal day 4


title: GBM SF9871_snRNA

description: Single Nuceli RNA-Seq


title: Blank


title: TS_NonLrcRep1


title: RNA-seq AMs in vivo - P4 LPS rep3


title: GBM SF12754_snRNA

description: Single Nuceli RNA-Seq


title: RNA-seq AMs in vivo - P4 PBS rep4

value: 2h, intranasal PBS instillation


title: GBM SF11981_snRNA

description: Single Nuceli RNA-Seq


title: TS_MidLrcRep1


title: TS_LrcRep1


title: GBM SF4324_snRNA

description: Single Nuceli RNA-Seq


title: RNA-seq AMs in vivo - 8W LPS rep4


title: RNA-seq AMs in vitro - 8W 0h rep2

value: 0h, in vitro culture


title: RNA-seq AMs in vivo - P4 PBS rep2


title: BS_NonLrcRep2

source_name: Back skin IFE

value: Back skin IFE

value: Post natal day 47


title: RNA-seq AMs in vivo - P4 PBS rep3


title: GBM SF12333_snRNA

description: Single Nuceli RNA-Seq


title: GBM SF12704_snRNA

description: Single Nuceli RNA-Seq


title: RNA-seq AMs in vivo - 8W PBS rep4

value: 8 weeks


title: GBM SF12751_snRNA

description: Single Nuceli RNA-Seq


title: BS_LrcRep2


title: RNA-seq AMs in vivo - 8W PBS rep3


title: Mouse_IR_Treated

description: RNA-Seq

value: Brain tumor


title: RNA-seq AMs in vivo - 8W LPS rep3


title: BS_NonLrcRep1


title: BS_MidLrcRep1


title: GBM SF12165_snRNA

description: Single Nuceli RNA-Seq


title: RNA-seq AMs in vivo - 8W PBS rep2


title: Mouse_Sham

description: RNA-Seq

value: Primary


title: BS_LrcRep1


title: GBM SF6118_snRNA

description: Single Nuceli RNA-Seq

value: GBM


title: KU_rep4

city: Trieste

first: Silvano

last: Piazza

address: localita' Padriciano 99

country: Italy

institute: ICGEB

description: Hippocampus from C57BL/6 KP treated with rna

data_processing:
Raw sequences files’ quality was checked via FastQC (v0.11.9, http://www.bioinformatics.babraham.ac.uk/projects/fastqc/)
Reads alignment to the mouse reference genome using Salmon tool (v1.4.0)
Salmon outputs were analysed using R’s package DESeq2 (v1.32.0)
Assembly: Mus musculus GRCm39 DNA primary assembly, transcript version GRCm39.103

source_name: Hippocampus

extract_protocol:
Hippocampal RNAs from mice intranasally treated with the ATM kinase blocker KU55933 (n=4) or vehicle (4% DMSO in saline; n = 4) were purified using standard column purification according to the manufacturer's protocol (RNAeasy Mini Kit, QIAGEN). All procedures were conducted in RNAase-free conditions. RNA concentration was evaluated using Qubit RNA BR Assay Kit (Life Technologies). RNA purity was assessed by determining UV 260/280 and 260/230 absorbance ratios using a Nanodrop® ND‐1000 spectrophotometer (Thermo Fisher Scientific). RNA quality was evaluated by measuring the RNA integrity number (RIN) using an Agilent RNA 6000 Nano Kit with an Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA) according to the manufacturer’s instructions.
Libraries were prepared with the TruSeq® Stranded mRNA Sample Preparation kit (Illumina, San Diego, Ca, USA) according to manufacturer’s instructions. For each sample of hypothalamus and PFC, 1000 ng and 500 ng were used as input quantity, respectively.

treatment_protocol: U55933 or vehicle (DMSO) was administered to P40 wt mice by intranasal route at a dosage of 7.5 mg/kg. The mice had been previously anesthetized, and the total volume was administered 3 μL at a time, alternating the 2 nostrils.

value: C57BL/6 KP

value: Hippocampus

value: WT

value: KU55933 was administered to P40 wt mice by intranasal route at a dosage of 7.5 mg/kg.


title: BS_MidLrcRep2


title: RNA-seq AMs in vivo - P4 LPS rep2


title: RNA-seq AMs in vivo - 8W PBS rep1


title: KU_rep3


title: Dlx1.F.Tail.P6.R2

data_processing:
Libraries were sequenced on an Illumina instrument (Hiseq4000). 2x150 bp HiSeq reads were generated with a depth of 20M.
Trimmed sequence reads were mapped to GrCM38/mm10 using CLC genomic benchwork (parameters- mismath cost: 2 insertion cost: 3 deletion cost: 3 length fraction: 0.8 similarity fraction: 0.8)
reads were trimmed for low quality and adaptor sequences with TrimGalore v0.6.0, a wrapper for cutadapt and fastQC ;unwanted reads were removed with STAR v 2.7.0e
SARTools and DESeq2 v1.26.0 were used to generate normalized counts and statistical analysis of differential gene expression
PCA was performed on the raw counts for the different tdTomato + sorted populations normalized in both back and tail skin with their respective BL (integrin/+Sca1+) sorted control and the scores were represented in a three-dimensional scatter plot.
Assembly: mm10/mouse
Supplementary files format and content: tab-delimited text files include RPKM values for each Sample

source_name: Skin Tissue

extract_protocol:
RNA was harvested using TRIzol (Thermo Fisher) according to the manufacture's protocol. Poly A+RNA was isolated with the NEBNext Poly(A) mRNA magnetic isolation module (New England Biolabs).
Since our RNA was low input (<20 nt total RNA), truSeq-barcoded RNAseq libraries were generated with the Ultra II RNA Library Prep Kit (non-directional) (New England Biolabs).

treatment_protocol: Mice at PD30-34 were injected with tamoxifen 100ug/ml for five consecutive days and harvested after 2 weeks at PD49 (telogen)

value: Skin Tissue

value: Tail Skin Dlx1+ IFE

value: WT(Dlx1CreER x RosatdTomato)

value: 2-week post tamoxifen injection

tag: time


title: GBM SF12243_snRNA

description: Single Nuceli RNA-Seq

extract_protocol:
Nuclei were extracted from frozen tissues following the “Frakenstein” protocol developed by Luciano Martelotto, Ph.D., Melbourne, Centre for Cancer Research, Victorian Comprehensive Cancer Centre, and available from 10X Genomics
For snRNA-seq(frozen tissues), ~15,000 nuclei were loaded per capture. Single-nucleus capture, reverse transcription, cell lysis, and library preparation were performed per manufacturer’s protocol. Sequencing was performed on an Illumina NovaSeq using a paired-end 100bp protocol.

value: Recurrent

value: Female


title: RNA-seq AMs in vivo - 8W LPS rep2

value: 2h, intranasal LPS instillation


title: KU_rep1


title: RNA-seq AMs in vivo - P4 PBS rep1

value: 2h, intranasal PBS instillation

value: postnatal day 4


title: GBM SF9962_snRNA

description: Single Nuceli RNA-Seq

value: Male


title: GBM SF12407_snRNA

description: Single Nuceli RNA-Seq


title: Dlx1.F.Tail.P5.R2

value: Tail Skin Total IFE


title: KU_rep2


title: GBM SF10565_snRNA

description: Single Nuceli RNA-Seq


title: Aspm.F.Tail.P5.R2

value: WT(AspmCreER x RosatdTomato)


title: Dlx1.F.Tail.P5.R1


title: Aspm.F.Tail.P6.R2

value: Tail Skin Aspm+ IFE


title: RNA-seq AMs in vivo - P4 LPS rep1


title: DMSO_rep4

value: C57BL/6 KP

value: DMSO was administered to P40 wt mice by intranasal route at a dosage of 7.5 mg/kg.


title: DMSO_rep3

value: Hippocampus

value: WT


title: GBM SF11857_snRNA

description: Single Nuceli RNA-Seq

value: GBM


title: Dlx1.F.Tail.P6.R1

value: Skin Tissue

value: Tail Skin Dlx1+ IFE


title: GBM SF11488_snRNA

description: Single Nuceli RNA-Seq


title: GBM SF12408_snRNA

description: Single Nuceli RNA-Seq


title: Aspm.F.Tail.P5.R1

value: 2-week post tamoxifen injection


title: DMSO_rep1


title: DMSO_rep2


title: Slc1a3.F.Tail.P5.R2

value: WT(Slc1a3CreER x RosatdTomato)


title: GBM SF9798_snRNA

description: Single Nuceli RNA-Seq

value: Recurrent


title: Stat1_KO_LK

city: Cambridge

first: Hugo

middle: P.

last: Bastos

address: Jeffrey Cheah Biomedical Centre, Puddicombe Way

department: Haematology

institute: Wellcome - MRC Cambridge Stem Cell Institute

data_processing:
Library strategy: scRNA-Seq
Reads processed and aligned to mm10 using CellRanger v2.1.1
Genome_build: mm10 (GRCm38)
Supplementary_files_format_and_content: Processed files are 10x-Genomics-formatted hdf5 files containing the UMI counts for each processed library

source_name: bone marrow

extract_protocol:
Bone marrow cells were harvested from STAT1KO and WT control mice, lineage negative cKit positive LK cells (lin-cKit+) were sorted to PBS/2% FBS
10X genomics Chromium, v2 chemistry

value: LK

value: KO

value: bone marrow


title: Aspm.F.Tail.P6.R1


title: WT_LK


title: RNA-seq AMs in vivo - 8W LPS rep1

value: 2h, intranasal LPS instillation

value: 8 weeks


title: GBM SF9510_snRNA

description: Single Nuceli RNA-Seq

value: Male


title: Ad-10XSRSF1-U1_Ad

city: La Jolla

first: Kaushik

last: Saha

address: 9500 Gilman Drive

department: Chemistry and Biochemistry

institute: University of California San Diego

description: MiSeq Nano

data_processing:
Basecalls performed with Realtime Analysis (RTA) software launched automatically by MiSeq Control Software
All steps of the processing were carried out by SHAPEMapper 2.0 pipeline

organism: Human adenovirus 2

source_name: In vitro transcribed RNA

growth_protocol: In vitro transcribed RNA

extract_protocol:
Purification by Monarch RNA cleanup kit
Two-step PCR

treatment_protocol: In the Fastq file names, N indicates treatment with NMIA, D DMSO, DC denatured control

value: In vitro transcribed RNA

tag: cell line

value: Wild-type


title: Ad-10XSRSF1-65-35-U1_Ad

description: MiSeq Nano


title: Ad-DC

description: MiSeq Nano


title: SCVS3

city: Guangzhou

first: Huang

last: Shuying

address: 中国广州市中山二路74号中山大学广州校区北校园

institute: Sun Yat-Sen University

data_processing:
1 μg total RNA was used for following library preparation. The poly(A) mRNA isolation was performed  using Oligo(dT) beads. The mRNA fragmentation was performed using divalent cations and high  temperature. Priming was performed using Random Primers. First strand cDNA and the second-strand  cDNA were synthesized. The purified double-stranded cDNA was then treated to repair both ends and  add a dA-tailing in one reaction, followed by a T-A ligation to add adaptors to both ends. Size selection of  Adaptor-ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by  PCR using P5 and P7 primers and the PCR products were validated.  fragments thereof, and quality of bases lower than 20, pass filter data of fastq format were processed by  Cutadapt (V1.9.1, phred cutoff: 20, error rate: 0.1, adapter overlap: 1bp, min. length: 75, proportion of N:  0.1) to be high quality clean data.
Then libraries with different indexs were multiplexed and loaded on an Illumina HiSeq/ Illumina  Novaseq/ MGI2000 instrument for sequencing using a 2x150 paired-end (PE) configuration according to  manufacturer’s instructions.  fragments thereof, and quality of bases lower than 20, pass filter data of fastq format were processed by  Cutadapt (V1.9.1, phred cutoff: 20, error rate: 0.1, adapter overlap: 1bp, min. length: 75, proportion of N:  0.1) to be high quality clean data.
Quality Control:In order to remove technical sequences, including adapters, polymerase chain reaction (PCR) primers, or  fragments thereof, and quality of bases lower than 20, pass filter data of fastq format were processed by  Cutadapt (V1.9.1, phred cutoff: 20, error rate: 0.1, adapter overlap: 1bp, min. length: 75, proportion of N:  0.1) to be high quality clean data.
Alignment:Firstly, reference genome sequences and gene model annotation files of relative species were downloaded  from genome website GRCm38.100   Mus musculus. Secondly, Hisat2 (v2.0.1) was used to index  reference genome sequence. Finally, clean data were aligned to reference genome via software Hisat2  (v2.0.1).
Expression analysis:In the beginning transcripts in fasta format are converted from known gff annotation file and indexed  properly. Then, with the file as a reference gene file, HTSeq (v0.6.1) estimated gene and isoform  expression levels from the pair-end clean data.
Differential expression analysis:Differential expression analysis used the DESeq2 Bioconductor package, a model based on the negative  binomial distribution. the estimates of dispersion and logarithmic fold changes incorporate data-driven  prior distributions, Padj of genes were setted <0.05 to detect differential expressed ones.
Assembly: GRCm38.100   Mus musculus
Supplementary files format and content: tab-delimited text files include counts for each Sample ...

source_name: meningeal lymphatic endothelial cells

extract_protocol:
Meningeal lymphatic endothelial cells were collected following previous reported protocol with modifications. Mice transcardially perfused with ice-cold PBS for 1 min under anesthesia. Individual meninx was quickly dissected from the skullcap in DMEM with 2% FBS, and digested in preheated DMEM with 2% FBS, 1 mg/mL collagenase VIII, 0.5 mg/mL DNase I, 45 μM actinomycin D for 12 min at 37°C. At the end of the digestion, 1 mL DMEM with 10% FBS were added to the solution to terminate digestion. Individual samples consisted of cell suspensions pooled from 3-4 meninges were filtered through a 70μm nylon-mesh filter and washed with ice-cold fluorescence-activated cell sorting (FACS) buffer (PBS free of calcium and magnesium, 1 mM EDTA and 1% BSA, pH 7.4). The cells were pelleted by centrifugation at 400 g, 4°C for 5min, resuspended in 400 μL FACS buffer with anti-CD45–BB515 (BD Biosciences, clone 30-F11, 1:200), anti-CD31-Alexa Fluor 647 (BD Biosciences, clone 390, 1:200), anti-Podoplanin-PE (eBioscience, clone 8.1.1, 1:200) and DAPI (Sigma, 0.2 μg/mL), and incubated for 15min at 4°C. Cells were then washed and resuspended in ice-cold FACS buffer. Cells were sorted by MoFlo Astrios EQs (Beckman Coulter, Indianapolis, IN, US) and analyzed by the FlowJo software (Tree Star, Ashland, OR, USA). Singlets were gated using the height, area and the pulse width of the forward-scatter cells, and cells negative for DAPI were selected as viable cells. The LECs were gated as CD45-CD31+podoplanin+ and sorted into a PCR tube containing 1 μL lysis buffer. Total RNA was isolated using RNAeasy Micro Kit following the manufacturer’s protocol (Qiagen, Hilden, Germany)
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: Sub-chronic variable stress

value: meningeal lymphatic endothelial cells

value: 3 month old

value: Sub-chronic variable stress


title: K14.BS.F.P5.R4

value: Back Skin Total IFE

value: WT(K14CreER x RosatdTomato)


title: SCVS5

address: 中国广州市中山二路74号中山大学广州校区北校园


title: GBM SF6621_snRNA

first: Aaron

last: Diaz

institute: University of California, San Francisco

description: Single Nuceli RNA-Seq

value: GBM


title: Ad-5XSRSF1-65-35-U1

description: MiSeq Nano


title: Slc1a3.F.Tail.P5.R1

value: Skin Tissue

value: Tail Skin Total IFE

value: 2-week post tamoxifen injection


title: GBM SF7388_snRNA

description: Single Nuceli RNA-Seq

value: Recurrent


title: Slc1a3.F.Tail.P6.R2

value: Tail Skin Slc1a3+ IFE

value: WT(Slc1a3CreER x RosatdTomato)


title: GBM SF7025_snRNA

description: Single Nuceli RNA-Seq


title: SCVS2

address: 中国广州市中山二路74号中山大学广州校区北校园


title: Slc1a3.F.Tail.P6.R1


title: GBM SF7062_snRNA

description: Single Nuceli RNA-Seq

value: Female


title: K14.F.BS.P5.R1


title: SCVS1

address: 中国广州市中山二路74号中山大学广州校区北校园


title: AdML_5X SRSF1_DMSO

description:
Reverse transcribed with Superscrit II and analyzed by SHAPEMapper 2.0 pipeline
Ad-S1_Ad.shape

data_processing:
Library strategy: Selective 2′ hydroxyl acylation followed by primer extension by mutational profiling (SHAPE-MaP)
Basecalls performed with Realtime Analysis (RTA) software launched automatically by MiSeq Control Software
All steps of the processing were carried out by SHAPEMapper 2.0 pipeline
Supplementary_files_format_and_content: .SHAPE

source_name: In vitro transcription

extract_protocol:
Proteinase K digestion (where protein is added) followed by RNA purification by Monarch RNA cleanup kit (New England Biolabs)
cDNA was PCR amplified using primers containing partial Illumina adapter for 20 cycles and then the library was further amplified for 8 cycles with dual index primers

treatment_protocol: RNAs were treated with 4 mM N-Methylisatoic anhydride (NMIA) for 15 min at 30 °C and 15 in at room temperature or the vehicle (DMSO) under the same conditions

value: wild type

value: 5X SRSF1-RE and 3.33% DMSO for 15 min at 30 °C and then 15 min at room temperature


title: K14.BS.F.P6.R4

value: Back Skin K14+ IFE


title: Naive3

address: 中国广州市中山二路74号中山大学广州校区北校园

value: Naive


title: Naive5

address: 中国广州市中山二路74号中山大学广州校区北校园

value: meningeal lymphatic endothelial cells


title: K14.F.BS.P6.R1


title: GBM SF3243_snRNA

description: Single Nuceli RNA-Seq

value: Male


title: AdML_5X SRSF1_NMIA

value: 5X SRSF1-RE and 4 mM NMIA for 15 min at 30 °C and then 15 min at room temperature


title: GBM SF6186_snRNA

description: Single Nuceli RNA-Seq


title: Naive2

address: 中国广州市中山二路74号中山大学广州校区北校园

value: 3 month old


title: Naive1

address: 中国广州市中山二路74号中山大学广州校区北校园


title: GBM SF6098_snRNA

description: Single Nuceli RNA-Seq


title: RNP_AdML + SRSF1 + SDA

description: Reverse transcribed with Superscript II and analyzed by SHAPEMapper 2.0 pipeline

data_processing:
library strategy: RNA-protein interactions by mutational profiling (RNP-MaP)
Basecalls performed with Realtime Analysis (RTA) software launched automatically by MiSeq Control Software
Per-nucleotide mutation frequencies (number of mutation events/effective read depth) for both crosslinked (SDA + UV-treated) and uncrosslinked (UV-treated) samples were calculated from output ShapeMapper 2 profiles.
RNP-MaP ‘Reactivity’ was computed as the ratio of nucleotide crosslinked mutation frequency to un-crosslinked mutation frequency (SDA + UV rate/UV-only rate).
To be designated as RNP-MaP sites, nucleotide positions had to pass three quality filters: 1) sites were required to have at least 50 more mutation events in the SDA + UV-treated sample than the UV-treated sample; 2) site reactivities had to exceed the nucleotide-dependent empirical thresholds described in the next section; and 3) nucleotide reactivities were required to achieve a Z′ factor greater than 0.
Details can be found in Weidmann CA et al. (2021). Nat Biotechnol, 39, 347-356
Genome_build: NOT PROVIDED
Supplementary_files_format_and_content: NOT PROVIDED

source_name: In vitro transcribed from a plasmid

growth_protocol: Not Applicable

extract_protocol:
Proteinase K digestion followed by RNA purification by Monarch RNA cleanup kit (New England Biolabs)
cDNA was PCR amplified using primers containing partial Illumina adapter for 20 cycles and then the library was further amplified for 8 cycles with dual index primers

treatment_protocol: RNAs were treated with 4 mM SDA (NHS-diazirine, succinimidyl 4,4′-azipentanoate) for 2 hours at 4 °C with rocking or the vehicle (DMSO) under the same conditions followed by exposure to 3 J/cm2 of 365 nm UV light

value: SRSF1 and 4 mM SDA at 4 °C for 2 hours with rocking


title: GBM SF3448_snRNA

description: Single Nuceli RNA-Seq

value: GBM


title: Naive4

address: 中国广州市中山二路74号中山大学广州校区北校园


title: SCVS4

address: 中国广州市中山二路74号中山大学广州校区北校园

value: Sub-chronic variable stress


title: RNP_AdML + SRSF1 + DMSO

value: SRSF1 and 4% DMSO at 4 °C for 2 hours with rocking


title: GBM SF3073_snRNA

description: Single Nuceli RNA-Seq


title: Aspm.BS.F.P5.R4

value: Skin Tissue

value: Back Skin Total IFE

value: WT(AspmCreER x RosatdTomato)

value: 2-week post tamoxifen injection


title: Aspm.F.BS.P6.R2

value: Back Skin Aspm+ IFE


title: Aspm.BS.F.P6.R4


title: RE 20-2 Liver

city: irvine

first: Siwei

last: Chen

address: 66304 Verano Road South

department: Computer Science

institute: University of California, Irvine

data_processing:
Alignment to reference assembly mm10 and known splice junctions extracted from the RefSeq database performed using Tuxedo pipeline tophat
Read counting for each gene and normalization to FPKM values  using Tuxedo pipeline
Genome_build: mm10
Supplementary_files_format_and_content: Gene expression results FPKM values) of the standard RNAseq analysis are reported in the genes_fpkm.csv file

source_name: liver

extract_protocol: Standard Illumina RNA-seq protocol

tag: genotype/variation

value: RE

tag: time point


title: RNP_AdML + U1 snRNP + SRSF1 + DMSO

value: wild type

value: U1 snRNP + SRSF1 and 4% DMSO at 4 °C for 2 hours with rocking


title: RNP_AdML + U1 snRNP + SRSF1 + SDA

value: U1 snRNP + SRSF1 and 4 mM SDA at 4 °C for 2 hours with rocking


title: Aspm.F.BS.P5.R2


title: RE 20-3 Liver


title: GBM SF1343_snRNA

city: San Francisco

description: Single Nuceli RNA-Seq

value: Recurrent


title: GBM SF2628_snRNA

description: Single Nuceli RNA-Seq

value: Female


title: Aspm.F.BS.P5.R1


title: RNP_AdML + U1 snRNP + SDA

value: U1 snRNP and 4 mM SDA at 4 °C for 2 hours with rocking


title: RE 20-1 Liver


title: RNP_AdML + U1 snRNP + DMSO

value: U1 snRNP and 4% DMSO at 4 °C for 2 hours with rocking


title: GBM SF2979_snRNA

description: Single Nuceli RNA-Seq


title: GBM SF12460_snRNA

description: Single Nuceli RNA-Seq


title: KO 20-3 Liver

value: KO


title: Slc1a3.BS.F.P5.R4

value: WT(Slc1a3CreER x RosatdTomato)


title: Aspm.F.BS.P6.R1


title: AdML_2X SRSF1_NMIA

description: Reverse transcribed with Superscrit II and analyzed by SHAPEMapper 2.0 pipeline

data_processing:
library strategy: Selective 2′ hydroxyl acylation followed by primer extension by mutational profiling (SHAPE-MaP)
Basecalls performed with Realtime Analysis (RTA) software launched automatically by MiSeq Control Software
All steps of the processing were carried out by SHAPEMapper 2.0 pipeline
Genome_build: NOT PROVIDED
Supplementary_files_format_and_content: NOT PROVIDED

value: 2X SRSF1-RE and 4 mM NMIA for 15 min at 30 °C and then 15 min at room temperature


title: AdML_DC_NMIA

treatment_protocol: RNA was treated with 4 mM N-Methylisatoic anhydride (NMIA) under denaturing condition

value: denatured control with 4 mM NMIA


title: GBM SF12382_snRNA

description: Single Nuceli RNA-Seq

value: GBM

value: Male


title: WT 20-3 Liver

value: WT


title: GBM SF12008_snRNA

description: Single Nuceli RNA-Seq


title: GBM SF12427_snRNA

description: Single Nuceli RNA-Seq


title: AdML_5X SRSF1+U1 snRNP_NMIA

value: 5X SRSF1-RE+U1 snRNP and 4 mM NMIA for 15 min at 30 °C and then 15 min at room temperature


title: AdML_5X SRSF1+U1 snRNP_DMSO

value: 5X SRSF1-RE+U1 snRNP and 3.33% DMSO (vehicle) for 15 min at 30 °C and then 15 min at room temperature


title: Ad-5XSRSF1-U1_Ad

description: MiSeq Nano

value: In vitro transcribed RNA

value: Wild-type


title: GBM SF10592_snRNA

description: Single Nuceli RNA-Seq

value: Primary


title: WT 20-2 Liver


title: KO 20-1 Liver


title: AdML_U1 snRNP_NMIA

value: wild type

value: U1 snRNP and 4 mM NMIA for 15 min at 30 °C and then 15 min at room temperature


title: Slc1a3.F.BS.P5.R2

value: Skin Tissue

value: Back Skin Total IFE

value: 2-week post tamoxifen injection


title: AdML_2X SRSF1_DMSO

value: 2X SRSF1-RE and 3.33% DMSO (vehicle) for 15 min at 30 °C and then 15 min at room temperature


title: Slc1a3.F.BS.P6.R2

value: Back Skin Slc1a3+ IFE

value: WT(Slc1a3CreER x RosatdTomato)


title: GBM SF11720_snRNA

description: Single Nuceli RNA-Seq

value: Recurrent


title: Slc1a3.F.BS.P6.R1


title: KO 20-2 Liver

value: KO


title: GBM SF11248_snRNA

description: Single Nuceli RNA-Seq

value: GBM

value: Female


title: Slc1a3.F.BS.P5.R1


title: GBM SF10514_snRNA

description: Single Nuceli RNA-Seq


title: AdML_U1 snRNP_DMSO

value: U1 snRNP and 3.33% DMSO (vehicle) for 15 min at 30 °C and then 15 min at room temperature


title: AdML_NMIA

value: 4 mM NMIA for 15 min at 30 °C and then 15 min at room temperature


title: Slc1a3.BS.F.P6.R4


title: GBM SF10484_snRNA

description: Single Nuceli RNA-Seq

value: Male


title: RE 16-1 Liver

value: RE


title: Aire-GFP+ILC3, stimulated Aire KO, biol rep2

city: Rehovot

first: Jakub

last: Abramson

address: 234 Herzl st.

department: Department of Immunology

country: Israel

institute: Weizmann Institute of Science

data_processing:
Data was processed using UTAP pipeline (Kohen et al., 2019). Bcl files were converted into fastq by bcl2fastq. Demultiplexing is done using the sample barcode found on R2, the UMI found on R2 is inserted in the read name (in R1 fastq).  R1 Fastq data submitted.
Reads were trimmed using cutadapt and mapped to genome (mm10) STAR v2.4.2a (default parameters).
The pipeline quantifies the 3’ of Gencode annotated genes (The 3’ region contains 1,000 bases upstream of the 3’ end and 100 bases downstream). Counting proceeded over genes annotated in RefSeq release using htseq-count (union mode). Genes having minimum 5 UMI-corrected reads in at least one sample, were considered.
Differential expression analysis was performed using DESeq2 with the betaPrior set to False. Raw P values were adjusted for multiple testing using the procedure of Benjamini and Hochberg.
Assembly: mm10
Supplementary files format and content: tab delimited files include raw counts values for each sample, normalized abundance measurements and DESeq2 analysis

source_name: lymph nodes

extract_protocol:
Lymph nodes were collected and subjected to several rounds of enzymatic digestion by Dispase I. Single cell suspension was depleted of lineage positive cells using LS-column based MACS enrichment by cocktail of biotinylated antibodies (TCR-β, CD3, CD19, B220, CD11b, F4/80, Gr1, CD11c) and anti-biotin microbeads. T-cells were isolated by meshing the skin-draining lymph nodes and spleens through 40 μm nylon mesh. Single cell suspension was subjected to flow cytonetry based sorting.
Single cell suspensions were directly FACS sorted to Lysis/Binding buffer (Invitrogen) and frozen on dry ice. RNA was isolated using Dynabeads (Invitrogen) according to manufactures protocol. The MARS-seq protocol described elsewhere was followed to generate the sequencing libraries (Jaitin D.A. et al., Science, 2014)
RNA sequencing 3' end with bulk MARS-Seq

value: lymph nodes

tag: strain

value: Aire KO

value: Candida albicans


title: RE 16-2 Liver


title: Aire-GFP+ILC3, stimulated Aire KO, biol rep1


title: KO 16-3 Liver


title: IL, N8mg/L, Control, 9

city: Johnson City

first: Lev

middle: Y

last: Yampolsky

state: Tennessee

address: 811 Lazywood Dr

department: Biological sciences

institute: East Tennessee State University

data_processing:
Basecalling: ONT guppy_basecaller
Demultiplexing, trimming: ONT guppy_barcoder
Alignment to reference transcriptome: ONT guppy_aligner to longest transcript of DM3 reference (BioProject ID: PRJNA624896)
Merging and indexing BAM files, exporting read counts: samtools
Filtering: retain transcripts with >=72 reads total
Assembly: N/A
Supplementary files format and content: Hyp84AC.readCountsRPKM.txt (read counts and RPKMs in tab-separated text file)

organism: Daphnia magna

source_name: Whole body homogenates, 2 individuals pooled

growth_protocol: 20 oC, density 1 ind.20 mL, food 100,000 Scenedesmus cells/mL/day; 25 days; either at normoxia (8 mg O2) or mild intermittent hypoxia (4 mg/L O2 twice daily)

extract_protocol:
Qiagen RNAeasy kit (Cat ID: 74134) with DNAse treatment; Qubit quantification
ONT PCR-cDNA Barcoding kit protocol (SQK-PCB109)

treatment_protocol: either no treatment (Control), or 12 h at acute severa hypoxia (<1 g/L O2)

value: Whole body

tag: clone

tag: chronic hypoxia

value: normoxia 8mgO2/L

tag: acute hypoxia

value: Control

tag: Sex

value: female


title: GBM SF10441_snRNA

description: Single Nuceli RNA-Seq

tag: progression

tag: diagnosis

tag: age

tag: gender


title: IL, N8mg/L, Control, 8

tag: tissue


title: Aire-GFP+ILC3, stimulated Aire KO, biol rep3

tag: cell type

tag: treatment


title: KO 16-1 Liver


title: IL, N8mg/L, Control, 7


title: IL, N8mg/L, AcuteExp1mg/L, 3


title: KO 16-2 Liver


title: GBM SF9715_snRNA

description: Single Nuceli RNA-Seq

tag: pair#


title: Rorc-GFP+ OTII T cells, KO stimulated, biol rep4

value: Rorc-GFP+ OTII

tag: genotype

value: Rorc-cre+ Aire-flox


title: WT 20-1 Liver

value: WT


title: Aire-GFP+ILC3, stimulated Aire WT, biol rep3

value: Aire WT


title: GBM SF10433_snRNA

description: Single Nuceli RNA-Seq

value: Recurrent

value: GBM


title: Rorc-GFP+ OTII T cells, KO stimulated, biol rep3


title: IL, N8mg/L, AcuteExp1mg/L, 2


title: Aire-GFP+ILC3, stimulated Aire WT, biol rep2


title: GBM SF10108_snRNA

description: Single Nuceli RNA-Seq


title: AdML_DMSO_vehicle

value: wild type

value: 3.33% DMSO (vehicle) for 15 min at 30 °C and then 15 min at room temperature


title: RE 16-3 Liver


title: Aire-GFP+ILC3, stimulated Aire WT, biol rep1


title: IL, H4mg/L, Control, 12

value: hypoxia 4mgO2/L


title: RE 12-2 Liver

value: RE


title: WT 16-3 Liver


title: IL, H4mg/L, Control, 11


title: GBM SF9494_snRNA

description: Single Nuceli RNA-Seq

value: Primary

value: Male


title: WT 16-2 Liver


title: RE 12-1 Liver


title: GBM SF9372_snRNA

description: Single Nuceli RNA-Seq

value: Female


title: IL, N8mg/L, AcuteExp1mg/L, 1

value: Whole body


title: GBM SF5581_snRNA

description: Single Nuceli RNA-Seq


title: KO 12-3 Liver

value: KO


title: KO 12-2 Liver


title: GBM SF4849_snRNA

description: Single Nuceli RNA-Seq


title: OTII T cells proliferating, KO stimulated, biol rep1

value: lymph nodes

value: proliferating OTII

value: Candida albicans


title: GBM SF9358_snRNA

description: Single Nuceli RNA-Seq


title: GBM SF8963_snRNA

description: Single Nuceli RNA-Seq


title: Rorc-GFP+ OTII T cells, KO stimulated, biol rep2

value: Rorc-GFP+ OTII

value: Rorc-cre+ Aire-flox


title: RE 12-3 Liver


title: OTII T cells proliferating, KO stimulated, biol rep4


title: Rorc-GFP+ OTII T cells, KO stimulated, biol rep1


title: GBM SF4810_snRNA

description: Single Nuceli RNA-Seq


title: IL, H4mg/L, AcuteExp1mg/L, 5

value: female


title: IL, H4mg/L, Control, 10

value: Control


title: GBM SF7307_snRNA

description: Single Nuceli RNA-Seq

value: GBM


title: GBM SF3996_snRNA

description: Single Nuceli RNA-Seq


title: WT 16-1 Liver

value: WT


title: IL, H4mg/L, AcuteExp1mg/L, 4


title: IL, H4mg/L, AcuteExp1mg/L, 6


title: GBM SF6809_snRNA

description: Single Nuceli RNA-Seq


title: HU, N8mg/L, Control, 4

value: normoxia 8mgO2/L


title: OTII T cells proliferating, KO stimulated, biol rep3


title: OTII T cells proliferating, KO stimulated, biol rep2


title: KO 12-1 Liver


title: GBM SF3076_snRNA

description: Single Nuceli RNA-Seq


title: OTII T cells nonproliferating, KO stimulated, biol rep2

value: nonproliferating OTII


title: OTII T cells nonproliferating, KO stimulated, biol rep3


title: WT 12-3 Liver


title: GBM SF2777_snRNA

description: Single Nuceli RNA-Seq


title: HU, N8mg/L, AcuteExp1mg/L, 10


title: GBM SF4209_snRNA

description: Single Nuceli RNA-Seq


title: WT 12-2 Liver


title: HU, N8mg/L, Control, 6


title: HU, N8mg/L, Control, 5


title: GBM SF2501_snRNA

description: Single Nuceli RNA-Seq


title: RE 8-1 Liver

value: RE


title: OTII T cells proliferating, WT stimulated, biol rep2

value: Rorc-cre- Aire-flox


title: OTII T cells nonproliferating, KO stimulated, biol rep1


title: HU, H4mg/L, Control, 8

value: hypoxia 4mgO2/L


title: Rorc-GFP+ OTII T cells, WT stimulated, biol rep3


title: WT 12- Liver


title: GBM SF3391_snRNA

description: Single Nuceli RNA-Seq

value: Primary

value: Female


title: RE 8-3 Liver


title: HU, N8mg/L, AcuteExp1mg/L, 12

value: Whole body


title: GBM SF12616_snRNA

description: Single Nuceli RNA-Seq


title: KO 8-1 Liver

value: KO


title: GBM SF12115_snRNA

description: Single Nuceli RNA-Seq

value: Male


title: Rorc-GFP+ OTII T cells, WT stimulated, biol rep2

value: lymph nodes

value: Candida albicans


title: OTII T cells nonproliferating, WT stimulated, biol rep3


title: WT 8-3 Liver


title: OTII T cells nonproliferating, WT stimulated, biol rep2


title: GBM SF1199_snRNA

description: Single Nuceli RNA-Seq


title: RE 8-2 Liver


title: GBM SF2990_snRNA

description: Single Nuceli RNA-Seq


title: WT 8-2 Liver


title: HU, N8mg/L, AcuteExp1mg/L, 11


title: WT 8-1 Liver


title: Rorc-GFP+ OTII T cells, WT stimulated, biol rep1

value: Rorc-GFP+ OTII


title: HU, H4mg/L, Control, 9

value: female


title: KO 8-3 Liver


title: OTII T cells proliferating, WT stimulated, biol rep3

value: proliferating OTII


title: MHCII+ILC3, stimulated, biol rep3


title: KO 8-2 Liver


title: GBM SF12707_snRNA

description: Single Nuceli RNA-Seq

value: GBM


title: OTII T cells proliferating, WT stimulated, biol rep1


title: GBM SF11916_snRNA

description: Single Nuceli RNA-Seq


title: HU, H4mg/L, AcuteExp1mg/L, 3


title: MHCII+ILC3, stimulated, biol rep2


title: HU, H4mg/L, Control, 7

value: Control


title: MHCII+ILC3, stimulated, biol rep1


title: GBM SF11815_snRNA

description: Single Nuceli RNA-Seq


title: GB, N8mg/L, Control, 8

value: normoxia 8mgO2/L


title: MHCII+ILC3, nonstimulated, biol rep3

value: nonstimulated


title: HU, H4mg/L, AcuteExp1mg/L, 2


title: OTII T cells nonproliferating, WT stimulated, biol rep1

value: nonproliferating OTII


title: MHCII+ILC3, nonstimulated, biol rep2


title: GB, N8mg/L, AcuteExp1mg/L, 2


title: GBM SF11344_snRNA

description: Single Nuceli RNA-Seq


title: GBM SF11587_snRNA

description: Single Nuceli RNA-Seq


title: HU, H4mg/L, AcuteExp1mg/L, 1


title: KO 4-1 Liver


title: GBM SF11977_snRNA

description: Single Nuceli RNA-Seq


title: RE 4-3 Liver

value: RE


title: WT 4-3 Liver

value: WT


title: GBM SF11082_snRNA

description: Single Nuceli RNA-Seq

value: Primary


title: GB, N8mg/L, Control, 9


title: GBM SF11780_snRNA

description: Single Nuceli RNA-Seq

value: Female


title: WT 4-2 Liver


title: GBM SF10857_snRNA

description: Single Nuceli RNA-Seq


title: RE 4-2 Liver


title: GBM SF10432_snRNA

description: Single Nuceli RNA-Seq


title: ILC3, stimulated, biol rep1


title: GB, N8mg/L, Control, 7

value: Whole body


title: KO 4-3 Liver

value: KO


title: GB, H4mg/L, Control, 10

value: hypoxia 4mgO2/L


title: GB, N8mg/L, AcuteExp1mg/L, 1


title: RE 0-3 Liver


title: KO 4-2 Liver


title: MHCII+ILC3, nonstimulated, biol rep1

value: lymph nodes


title: GB, H4mg/L, AcuteExp1mg/L, 5


title: RE 0-2 Liver


title: ILC3, nonstimulated, biol rep1


title: ILC3, stimulated, biol rep3

value: Candida albicans


title: GB, H4mg/L, AcuteExp1mg/L, 4


title: RE 0-1 Liver


title: FI, N8mg/L, Control, 9


title: GB, H4mg/L, Control, 12


title: RE 4-1 Liver


title: ILC3, stimulated, biol rep2


title: WT 4-1 Liver


title: CH02_1 scRREMseq

first: Neville

last: Dusaj

institute: Weill Cornell Medicine

description: genomic DNA digested with MspI and treated with TET2/APOBEC

data_processing:
Library strategy: enzymatic methyl-seq (reduced representation)
scRNA data was aligned with STAR (v2.5.2) using twopassMode, gene level counts were generated using featureCounts (subread v1.5.2) with default parameters. The number of mutant and WT reads per cell was counted using a pileup at the mutation site (pySAM v0.8.2.1). Only reads with MAPQ > 40 and a specified minimum base quality at the mutation site were considered.
For scDNAme, each pool of 96 cells was first demultiplexed by Illumina i7 barcodes, resulting in four pools of 24 cells. Each pool of 24 cells was further demultiplexed by unique cell barcodes. Reads were assigned to a given cell if they matched 80% of the template adapters. Adapters and adapter reverse complements (6 bp) were trimmed from the raw sequence reads. After adapter removal, reads were trimmed by 5 bp from their 3’ end. We aligned trimmed reads to the hg38 human genome assembly using Bismark117 (v.0.14.5; parameters: -multicore 4 -X 1000 --un –ambiguous) running on bowtie2-2.2.8 aligner118. Bismark methylation extractor (--bedgraph --comprehensive) was used to determine the methylation state of each individual CpG. For downstream analyses, a site was considered methylated or unmethylated only if there was 90% agreement of the methylation state for all reads mapped to the site. Cells with coverage of at least 25,000 unique CpGs and bisulfite conversion rates of at least 99% were retained for downstream analyses.
Genome_build: hg38
Supplementary_files_format_and_content: Text files include  counts overlapping with annotated genes for all cells. Excel and comma-separated values files contain the filename conversion key to match scRNA with scDNAme cells, the sequencing summary statistics for each cell, and genotype call.
Supplementary_files_format_and_content: all_CpG_meth.csv: CpG by cell matrix with methylation values for all detected CpGs
Supplementary_files_format_and_content: scRNA_filename_key_to_scDNAme.csv: Filename conversion key to match scRNA with scDNAme cells
Supplementary_files_format_and_content: scDNAme_cell_metadata.xlsx: Summary statistics and metadata of all cells
Supplementary_files_format_and_content: scRNA_cell_metadata.xlsx: Summary statistics and metadata of all cells
Supplementary_files_format_and_content: gene_exp_mtx.txt: Gene count matrix from single-cell RNA-seq

source_name: GCSF mobilized stem cells from patient

extract_protocol:
Cryopreserved patient samples approved by the Institutional Review Board of Dana Farber Cancer Institute were thawed and stained using standard procedures (10 min, 4°C) with the surface antibody CD34-PE-Vio770 (clone AC136, Miltenyi Biotec) and DAPI (Sigma-Aldrich). Cells were then sorted for DAPI-negative, CD34+ and CD34-negative cells using BD Influx.
Methylome and whole-transcriptome library preparation and sequencing were performed as previously described. Briefly, genomic DNA (gDNA) and mRNA have been separated manually. A modified oligo-dT primer (5′-biotin-triethyleneglycol-AAGCAGTGGTATCAACGCAGAGTACT30VN-3′, where V is either A, C or G, and N is any base; IDT) was conjugated to streptavidin-coupled magnetic beads (Dynabeads, Life Technologies) according to the manufacturer’s instructions. To capture polyadenylated mRNA, we added the conjugated beads (10 μl) directly to the cell lysate and incubated them for 20 min at room temperature with mixing to prevent the beads from settling. The mRNA was then collected to the side of the well using a magnet, and the supernatant, containing the gDNA, was transferred to a fresh plate. Single-cell complementary DNA was amplified from the tubes containing the captured mRNA according to a variation of the Smart-Seq2 protocol using molecular crowding to increase sensitivity. After amplification and purification using 0.8X SPRI beads, 0.5ng cDNA was used for Nextera Tagmentation and library construction. Library quality and quantity were assessed using Agilent Bioanalyzer 2100 and Qubit, respectively. Genomic DNA present in the pooled supernatant and wash buffer from the mRNA isolation step was concentrated on 0.8X SPRI beads and eluted directly into the reaction mixtures for Msp1(+/- HaeIII) (Fermentas) enzymatic reaction (10mL final reaction) for 90 min at 37°C. Heat-inactivation was performed for 10 min at 70°C. Digested DNA was filled-in and A-tailed at the 3’ sticky ends in 8.5 mL final volume of 1X CutSmart with 2.5 units of Klenow fragment (Exo-, Fermentas). Reaction was supplemented with 1 mM dATP and 0.1 mM dCTP and 0.1 mM dGTP (NEB) and performed as follows in a thermocycler: 30°C for 25 min, 37°C for 25 min and heat-inactivation at 70°C for 10 min. Custom barcoded methylated adaptors (0.1 mM) were then ligated overnight at 16°C with the dA-tailed DNA fragments in the presence of 800 units of T4 DNA ligase (NEB) and 1 mM ATP (Roche) in a final volume of 11.5 mL of 1X CutSmart buffer. T4 DNA ligase heat-inactivation was performed at 70°C for 15 min the next day. Genomic DNA from 24 individual cells were pooled together according to their barcodes, giving, for a 96-well plate, 4 pools of 24 cells. Pooled genomic DNA was cleaned-up and concentrated using 1.8X SPRI beads (Agencourt AMPure XP - Beckman Coulter). Each pool was then sodium bisulfite converted (Fast Epitect Bisulfite, Qiagen) following manufacture recommendations or converted using EM-seq kit (New Englands Biolabs) following manufacture recommendations. Converted DNA was then amplified using primers containing Illumina i7 and i5 index. Following Illumina pooling guidelines, a different i7 index was used for every 24-cell pool, allowing multiplexing of 96 cells for sequencing on one Illumina HiSeq lane. Library enrichment was done using KAPA HiFi Uracil+ master mix (Kapa Biosystems) and the following PCR condition was used: 98°C for 45 secs; 6 cycles of: 98°C for 20 secs, 58°C for 30 secs, 72°C for 1 min; followed by 12 cycles of: 98°C for 20 secs, 65°C for 30 secs, 72°C for 1 min. PCR was terminated by an incubation at 72°C for 5 min. Enriched libraries were cleaned-up and concentrated using 1.3X SPRI beads. DNA fragments between 200 bp and 1 Kb were size-selected and recovered after resolving on a 3% NuSieve 3:1 agarose gel. Libraries molarity concentration calculation was obtained by measuring concentration of double stranded DNA (Qubit) and quantifying the average library size (bp) using an Agilent Bioanalyzer. Every 24-cells pool was mixed with the others pool in an equimolar ratio. All cells from a 96-well plate were sequenced as paired-end on HiSeq 2500 with 10% PhiX spike-in. Negative controls (empty wells with no cells) were used to control for non-specific amplification of the libraries. Single cell bisulfite-seq or enzymatic methyl-seq (reduced representation) and single-cell RNA-seq (full-length transcriptome profiling)
Single cell bisulfite-seq or enzymatic methyl-seq (reduced representation) and single-cell RNA-seq (full-length transcriptome profiling)

value: Blood

tag: disease state

value: Clonal hematopoiesis


title: GB, H4mg/L, Control, 11

value: female