title: Hgfac WT intact tissue_2

description: 10x Genomics


title: In Ctr_1

status: Public on Oct 26 2023

type: SRA

city: Heidelberg

first: Thomas

last: Schmidt

address: Grabengasse 1

country: Germany

institute: Heidelberg University

library_source: genomic

data_processing:
The raw ChIP-seq reads were trimmed using fastp (doi 10.1093/bioinformatics/bty560) version 0.23.2, followed by alignment to the GENCODE human reference genome hg38 using the STAR aligner version 2.7.8a (doi 10.1093/bioinformatics/bts635). Subsequently, Samtools version 1.14 (doi 10.1093/gigascience/giab008) was used to sort and merge the aligned reads by experimental condition and methylation marker. The merged alignments were used as input for genome coverage analysis, experimental quality controland peak calling.
Peak calling was performed, using MACS2 version 2.2.7.1 (doi 10.1186/gb-2008-9-9-r137) in broad peak mode, for each condition and histone marker. Consensus peaks were then constructed using bedtools version 2.30.0 (doi 10.1002/0471250953.bi1112s47) by intersecting the called peaks for the respective histone marker from the control and SAM conditions (minimum overlap 50%). Featurecounts version 2.0.1 (doi 10.1093/bioinformatics/btt656) was used to count the number of reads aligning to the consensus peaks. Next, we conducted differential peak calling using DESeq2 version 1.34.0 (doi 10.1186/s13059-014-0550-8). For ChIP-seq experimental quality control, genome coverage analysis was performed to enable visual inspection of called peaks by means of bamCoverage module of deeptools version 3.5.1 (doi 10.1093/nar/gkw257). The R-package DESeq2 version1.36.0 was used to create PCA plots based on the read counts of all consensus peaks found for each sample group (control vs SAM), across the investigated methylation markers. The R-package Pheatmap version 1.0.12 was used to generate clustered Heatmaps for each investigated methylation marker, to visualize the read counts of differentially methylated regions, shared between control and SAM samples (consensus peaks).
Supplementary files format and content: "metadata.xlsx" contains metadata sheet, bigWig files contain genome-wide signal data

molecule: genomic DNA

organism: Homo sapiens

growth_protocol: PC-3 cells were seeded in cell culture flasks in RPMI 1640 with 10% fetal calf serum and 1,2% penicillin/streptomycin.

extract_protocol: Nuclei of PC-3 cells were disrupted and sonicated to yield fragment size ranging from 200-500bp. Lysates were incubated with H3K4me3 and H3K27me3 antibody (Rabbit polyclonal) and bound DNA after washes was extracted using Qiagen PCR cleanup kit.

treatment_protocol: They were treated either with vehicle ((0.005M H2SO4) or 200 µMol of SAM. Medium was changed every other day.

tag: cell type

value: prostate cancer cells


title: In_SAM_1


title: THP-1 cells, Exp2

city: Illkirch-Graffenstaden

first: Zhirong

last: ZHANG

address: 1 Rue Laurent fries

department: Stem celll and development

country: France

institute: IGBMC

data_processing:
Image analysis and base calling were performed using RTA version 2.7.7 and bcl2fastq version 2.20.0.422.
Sequencing reads were trimmed, aligned and counted using PinAPL-Py and guide RNAs (Brunello library). The Matching Threshold for bowtie2 alignment was defined as 35.
Data were statistically analyzed using Model-based Analysis of Genome-wide CRISPR/Cas9 Knockout (MAGeCK v0.5.9.3). The method ranks sgRNAs based on p-values calculated from the negative binomial model, and uses a modified robust ranking aggregation (RRA) algorithm to identify positively or negatively selected genes. Read counts were normalized by total number of reads. Enrichment testing of paired samples was performed using the test function in MAGeCK with --paired option. Volcano plot was generated using ggplot package in R and significance was defined as FDR < 0.05.
Assembly: hg38
Supplementary files format and content: tab-delimited text file for sgRNA raw counts

source_name: THP-1 cells

growth_protocol: THP-1 cells were grown in RPMI 1640 supplement with 10 mM HEPES, 10% fetal calf serum (FCS), 2.5 g/l glucose, 1 mM sodium pyruvate, gentamycin and 50 µM β-mercaptoethanol at 37°C with 5% CO2.

extract_protocol:
Genomic DNA from both control- and treated groups was isolated using the phenol/chloroform extraction method.
Fragments containing sgRNA were amplified from genomic DNA by PCR referring to the protocol from Broad Institute GPP Web Protal (https://portals.broadinstitute.org/gpp/public/resources/protocols).

treatment_protocol: THP-1 cells stably expressing Cas9 protein were infected with the pooled Brunello lentivirus library at ROI of 0.2. After puromycin selection for 3 days, cells were further expanded. To ensure a representation of minimal 500 cells per sgRNA, at least 4.0×107 cells were used in each group. An adequate number of cells were primed with LPS in suspension for 3 hours and treated with vehicle (control group) or R837 plus Yoda1 (treated group). When pyroptotic cell death reached ~85% in the treated group, R837 and Yoda1 were washed out and the survivals were expanded. When cell number reached 4.0 × 107 cells, these cells were repeatedly treated with R837 plus Yoda1 to reach reached ~85% of pyroptotic cell death. R837 and Yoda1 were washed out and the survivals were further expanded to reach 4.0 × 107 cells.

tag: cell line

value: THP-1 cells

value: Monocytes

tag: genotype

value: wild type

tag: treatment


title: THP-1 cells, Exp1


title: Control; Timepoint-1; 22-P-1013 [22_P_1013_1_t]

city: Santiago de Compostela

first: Alberto

last: Gómez-Carballa

state: A coruña

address: Choupana sn

department: Pediatrics

country: Spain

institute: GENVIP research group

description:
raw_counts.txt
Norm_corrected_exp_ACD.txt
Norm_corrected_exp_CONTROLS.txt

library_source: transcriptomic

data_processing:
Quality control (QC) of the RNA-Seq data obtained was performed using the FastQC and MultiQC.
After the QC step, problematic 3' end and adapters were removed with the Trimmomatic package
Reads mapped against the human reference genome using ultrafast universal RNA-Seq aligner STAR
Secondary and low-quality alignments were removed using SAMtools
DESeq2 R package was used to normalize counts and carry out the differential expression (DE) analysis after removing lowly expressed genes
Assembly: (GCh38,v.104)
Supplementary files format and content: Tab-delimited raw counts file
Supplementary files format and content: Tab-delimited Normalized and corrected data (paired-samples design accounting for patient-to-patient differences with respect to TP1) for ACD samples
Supplementary files format and content: Tab-delimited Normalized and corrected data (paired-samples design accounting for patient-to-patient differences with respect to TP1) for control samples

molecule: polyA RNA

source_name: Capillary blood

extract_protocol:
We collected blood samples using a procedure of capillary puncture before (Time Point 1) and after (Time Point 2) exposure to music, including 16 ACD patients, and 14 healthy donors. Sample was collected in a tube containing the proper volume of PAXgene additive. RNA was isolated using the PAXgene blood miRNA extraction kit (Qiagen). On-column DNase I treatment was carried out during the extraction process. RNA concentration step and an additional DNase treatment were undertaken using an RNA clean & concentrator kit (Zymo Research) prior to sequencing
Strand specific library preparation was completed using the SMARTer Ultra low input RNA library (Poly-A). Sequencing was performed using a NovaSeq 6000 System at 100 paired end configuration to finally obtain 70M reads per sample

tag: tissue

value: Capillary blood

tag: gender

value: Female

tag: age

tag: timepoint

tag: group

value: Control

tag: subject id


title: Cochlear organoids in stiff gels 3

city: Shanghai

first: Mingyu

last: Xia

state: Shanghai

address: No. 83, Fenyang Road, Xuhui District

department: ENT Institute and Otorhinolaryngology Department of the Affiliated Eye and ENT Hospital, Sta

country: China

institute: Fudan University

data_processing:
RNA-seq reads were aligned to mm10 using TopHat 2.1.1
Cuffdiff2 2.2.1 was used to estimate fragments per kilobase of transcript per million mapped reads (FPKM) values and to analyze differentially expressed transcripts.
Transcripts and genes were annotated according to the RefGene database (NCBI).
Sequencing data were analyzed using the DESeq2 package (http://bioconductor.org/packages/release/bioc/html/DESeq2.html).
Assembly: mm10
Supplementary files format and content: The processed file including the baseMean and expression of the all genes from six samples in the two groups. FoldChange, log2FoldChange, p-value and q-value were also shown, the comparision regards to the mean  expression of the indicated genes  from organoids cells in stiff gels compared to in soft gels.

molecule: total RNA

organism: Mus musculus

source_name: inner ear

growth_protocol: The expansion medium was based on Dulbecco's Modified Eagle Medium/Nutrient Mixture F-12 (Thermo Fisher Scientific) supplemented with 1%N2 (GIBCO), 2% B27 (GIBCO) and 100μg/ml penicillin/streptomycin, 50 ng/mL EGF (Proteintech), 50 ng/mL bFGF (Proteintech), 50 ng/mL IGF (Proteintech), 3 μM CHIR99021 (Sigma), 500 μM valproic acid (Sigma), 100 mg /mL 2-phospho-L-ascorbic acid (Sigma), and 2μM 616452 (Sigma). After 8-10 days of expansion, the combination of growth factors and small molecules was changed into a differentiation medium containing 3 μM CHIR99021 (Sigma) and 5μM LY411575 (Sigma).

extract_protocol:
Organoids cultured in soft and stiff  hydrogels were collected. Total mRNA was extracted using TRIzol (Life Technologies) followed by purification by an RNeasy Mini Kit (QIAGEN).
RNA-seq libraries were prepared using the TruSeq RNA Library Prep Kit v2 (Illumina), then samples were sequenced by an Illumina HiSeq 2500 sequencer.

treatment_protocol: At the differentiation stage, organoids were allocated into two different stiffness hydrogels.

value: inner ear

value: cochlear progenitors-derived organoid cells

value: WT

value: Cultured in stiff gels


title: Small intestinal organoids_Young-rep1

first: Erica

last: Valentini

address: Im Neuenheimer Feld 580

institute: DKFZ

data_processing:
nf-core/rnaseq  v1.4.2
Nextflow  v20.04.1
FastQC  v0.11.8
Cutadapt v2.5
Trim Galore! v0.6.4
SortMeRNA v2.1b
STAR vSTAR_2.6.1d
HISAT2 v2.1.0
Picard MarkDuplicates v2.21.1
Samtools v1.9
featureCounts v1.6.4
StringTie v2.0
Preseq v2.0.3
deepTools v3.3.1
RSeQC v3.0.1
dupRadar v1.14.0
Qualimap v.2.2.2-dev
MultiQC v1.7
Genome_build: GRCm38
bam files generated using STAR aligner
Supplementary_files_format_and_content: count text files gernerated using R/DESeq2 v1.28.1

source_name: Small intestinal organoids

growth_protocol: small intestinal organoids were culture for 4-5 weeks (passage 3 & 4) and passaged every week

extract_protocol:
RNA was isolated from organoids via QIAGEN RNAeasy kit, including on column DNase digest
Illumina sequencing libraries were prepared using the TruSeq Stranded mRNA Library Prep Kit (Illumina) according to the manufacturer's protocol. Briefly, poly(A)+ RNA was purified from 500ng of total RNA using oligo(dT) beads, fragmented to a median insert length of 155 bp and converted to cDNA. The ds cDNA fragments were then end-repaired, adenylated on the 3′ end, adapter ligated and amplified with 15 cycles of PCR.

treatment_protocol: no treatment

value: small intestinal epithelial cells

tag: strain

value: Young

tag: organoid culture passage


title: Cochlear organoids in stiff gels 2


title: RNA-Seq STAT3-KD rep3

city: San Antonio

first: Tim

middle: Hui-Ming

last: Huang

state: TX

address: 7703 Floyd Curl Dr

country: USA

institute: University of Texas Health at San Antonio

data_processing:
Basecalls were performed using Illumina Casava software.
Reads were aligned to hg19 genome assembly using STAR 2.5.2b with default parameters. Refseq transcripts were supplied in GTF format file.
featureCounts v1.5.1 was used to count the uniquely mapped reads in union mode.
Differentially expressed genes were detected by edgeR.
Assembly: hg19
Supplementary files format and content: tab-delimited text files include reads counts for each Sample

source_name: U937-derived macrophages

growth_protocol: The Vehicle control and STAT3 KD U937 monocyte were cultured with RPMI 1640 supplemented with 10% heat shocked FBS and were differentiated into macrophages by 100 ng/mL phorbol 12-myristate 13-acetate.

extract_protocol:
Total RNA was extracted using the PureLink RNA Mini Kit (Thermo Fisher Scientific) according to the manufacturer's instructions.
1 ug of total RNA was used to construct the libraries using NEB stranded mRNA-seq library preparation kit. Samples were sequenced using Illumina NovaSeq S4 platform.

value: U937-derived macrophages

value: STAT3 knockdown


title: THP-1 cells, Ctrl1

value: DMSO


title: In Ctr_2


title: Hgfac WT intact tissue_1

city: Cambridge

first: Ramnik

last: Xavier

state: MA

address: 415 Main Street

institute: Broad Institute of MIT and Harvard

description: 10x Genomics

library_source: transcriptomic single cell

data_processing:
Single-cell RNA-seq data was processed using Cell Ranger (version 5.0.1) via the cumulus workflow
Assembly: mm10
Supplementary files format and content: Cell Ranger output files

source_name: distal colon

extract_protocol:
Day2 colonic wounds or intact tissue samples from Hgfac WT and Hgfac KO mice were used. Endoscopic wounds were created by the biopsy forceps on day0 and excised by a 2 mm skin biopsy punch tool under the dissecting microscope on day2. Three wounds per mouse were pooled as a sample. Intact tissues excised by a 2 mm skin biopsy punch tool were also collected and used as an unwounded sample. Epithelial and IEL fractions were isolated from wound biopsies or intact colons via mechanical disruption. Briefly, tissues were disrupted via centrifugal force generated by a small magnet in a flask at 1,100 rpm for 45 min at 37°C in 4 ml of RPMI 1640 medium supplemented with 2 mM EDTA. The supernatant was used for epithelial and IEL fractions and the remaining tissues were used for the lamina proprial fraction. For epithelial and IEL fractions, the supernatant was collected and was centrifuged at 16,000 rpm for 8 min. The pellet was suspended with Advanced DMEM/F12 containing 10% FBS medium once and was centrifuged at 16,000 rpm for 8 min. After removing the supernatant, the pellets were suspended with 1 ml of TripLE at 37°C for 10 min. After washing with Advanced DMEM/F12 containing 10% FBS, the pellet was suspended with FACS buffer and kept on ice during lamina proprial isolation. For the lamina proprial (LP) fraction, the tissues were digested in media containing 20% FBS in RPMI 1640 with 0.5 mg/ml of Collagenase VIII (Sigma) for 60 min. The supernatant was collected, filtered, and centrifuged at 16,000 rpm for 8 min. After washing with FACS buffer (2% FBS in PBS) once, cells were used for the analysis. Finally, both epithelial and IEL fraction and LP fraction were combined and used for scRNA-seq.
Library was performed according to the manufacturer's instructions (single cell3' v3 protocol, 10x Genomics)

value: distal colon

value: Epithelial/fibroblast/immune cells

value: Hgfac WT

value: no wound


title: RNA-Seq STAT3-KD rep2


title: Hgfac WT day2 wound_1

description: 10x Genomics

value: wound


title: genomic DNA from fibroblasts from healthy control 2 (replicate 1)

type: genomic

city: Bologna

first: Maria Giulia

last: Bacalini

address: Via Altura 3

department: Brain Aging Laboratory

country: Italy

institute: IRCCS Istituto delle Scienze Neurologiche di Bologna

hyb_protocol: Bisulphite converted DNA was amplified, fragmented and hybridised to Infinium BeadChip using standard Illumina protocol

scan_protocol: HiScan scanner (Illumina)

data_processing: Signal intensities files were etracted using the minfi Bioconductor package. The preprocessFunnorm function implemented in minfi was applied to normalize raw data.

label: Cy3 and Cy5

source_name: Control 2 (replicate 1)

label_protocol: Standard Illumina Protocol

growth_protocol: HGPS fibroblasts were from the BioLaM biobank (Rizzoli Orthopedic Institute Ethical Committee approval no.0018250–2016) located in the CNR Institute of Molecular Genetics Unit of Bologna. Cells were cultured in Dulbecco's modified eagle medium (DMEM) supplemented with 20% foetal bovine serum, 1% Penicillin-streptomycin-amphotericin B  in 5% CO2 at 37 °C.

extract_protocol: Total genomic DNA (gDNA) from was isolated using the AllPrep DNA/RNA/Protein Mini kit (Qiagen). For microarray-based analysis of genome wide DNA methylation levels, 1µg of DNA was bisulfite-treated using the EZ DNA Methylation™ kit (Zymo Research), according to manufacturer instructions. Genone-wide DNA methylation analysis was performed using the Infinium MethylationEPIC BeadChip (Illumina) following manufacturer's instructions.

value: fibroblasts

tag: donor

value: Control 2

tag: disease

value: Healthy

tag: age at skin biopsy (yrs)


title: Hgfac KO day2 wound_2

description: 10x Genomics

value: Hgfac KO


title: OSCC_3 [miRNA]

city: Chengdu

first: Shouyi

last: Tang

state: Sichuan

address: Chengdu, Sichuan, P. R. China.

institute: Sichuan  University

data_processing:
Raw reads generated during high-throughput sequencing were fastq format sequences. In order to get high-quality reads that could be used for later analysis, raw reads needed to be further quality filtered. Trimmomatic[1] software was first used for adpter removing, And then low-quality bases and N-bases or low-quality reads were filtered out. Finally, we got high-quality clean reads. Using hisat2 to align clean reads to the reference genome of the experimental specie, the sample should be assessed by genomic and gene alignment.
Using hisat2 to align clean reads to the reference genome of the experimental specie, the sample should be assessed by genomic and gene alignment.
The result of alignment with the reference genome was stored in a binary file, called a bam file. Using the Stringtie software to assemble the reads, the new transcript was spliced. Then the candidate lncRNA transcripts were selected by comparing the gene annotation information of the reference sequence produced by Cuffcompare[4] software. Finally, transcripts with coding potential were screened out by CPC, CNCI, Pfam and PLEK to obtain lncRNA predicted sequences.
Aligning the sequencing reads of each sample with the sequence of mRNA transcript 4 sequences, known lncRNA sequences and lncRNA prediction sequences by bowtie2, and using eXpress to make gene quantitative analysis, the FPKM value and counts value(the number of reads for each gene in each sample) were obtained.
In order to generate SAM file, we used BWA software to align the sequencing reads of each sample with reference genome. Then CIRI software was used to scan for PCC signals (paired chiastic clipping signals), and circRNA sequences were predicted based on junction reads and GT-AG cleavage signals. Using the RPM algorithm to quantify the circRNAs, and normalizing the number of junction reads counts and foldchange by DESeq. Enrichment of differentially expressed circRNAs was analysised through the annotation information of circRNA source transcripts
Assembly: hg38
Supplementary files format and content: excel spreadsheet file includes raw counts for each sample
Supplementary files format and content: excel spreadsheet file includes FPKM(mRNA and lncRNA), RPM(circRNA), and TPM(miRNA) for each sample

source_name: mucosa

growth_protocol: Not applicable

extract_protocol:
Total RNA was extracted using the mirVana miRNA Isolation Kit (Ambion) following the manufacturer’s protocol. RNA integrity was evaluated using the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA). The samples with RNA Integrity Number (RIN) ≥7 were subjected to the subsequent analysis.
The libraries were constructed using TruSeq Stranded Total RNA with Ribo-Zero Gold according to the manufacturer’s instructions.

treatment_protocol: Not applicable

value: mucosa

value: OSCC


title: genomic DNA from fibroblasts from healthy control 2 (replicate 2)

source_name: Control 2 (replicate 2)


title: RNA-Seq STAT3-KD rep1


title: Cochlear organoids in stiff gels 1

value: inner ear

value: cochlear progenitors-derived organoid cells

value: WT

value: Cultured in stiff gels


title: genomic DNA from fibroblasts from HGPS patient (replicate 2)

source_name: HGPS (replicate 2)

value: HGPS


title: Hgfac KO day2 wound_1

description: 10x Genomics


title: Cochlear organoids in soft gels 2

value: Cultured in soft gels


title: Cochlear organoids in soft gels 3


title: genomic DNA from fibroblasts from HGPS patient (replicate 3)

source_name: HGPS (replicate 3)


title: OSCC_1 [miRNA]


title: Cochlear organoids in soft gels 1


title: Hgfac KO intact tissue_2

description: 10x Genomics

value: distal colon


title: hSIO - ZNF800-KO [scRNA-seq]

city: Utrecht

first: Jeff

last: DeMartino

address: Heidelberglaan 25

country: Netherlands

institute: Princess Maxima Center

description:
10x Genomics
LX198

data_processing:
Illumina output demultiplexed (bcl files) converted to fastq files using mkfastq (CellRanger v7.0.1)
Alignment, filtering, barcode and UMI counting done using cellranger count (CellRanger v7.0.1)
Assembly: Genome Reference Consortium Human Build 38 + transgene constructs (MUC2-mNeonGreen; DEFA5-IRES-dsRed; CHGA-IRES-iRFP670)
Supplementary files format and content: Per sample filtered_feature_bc_matrix output from CellRanger (v7.0.1); includes gene-barcode matrices, genes, barcodes

source_name: human small intenstinal organoids

extract_protocol:
Organoids were removed from theCultrexBasement Membrane Extract (BME, Growth Factor Reduced, Type 2, R&D Systems), digested withTryplE(Thermo Fisher) for 10~15 mins into single cells, washed with ice-coldAdDMEM/F12 (GIBCO) and filtered into FACS tubes through cell strainer (Falcon) with Rho kinase inhibitor Y-27632 dihydrochloride (10 µM,AbmoleBioscience). Cells were co-stained with DAPI (Sigma) and DRAQ5 (Biostatus) and subjected for FACS sorting of live cells. FACS was performed on BD Influx™ cell sorter (BD Bioscience). 4,000 live cells per condition were subjected to droplet-based scRNA-seq using the 10x Genomics platform.
Libraries were prepared using the 10x Genomics Chromium 3’ Gene Expression solution v3.1

value: human small intenstinal organoids

value: ZNF800-/-; MUC2-mNeonGreen; DEFA5-IRES-dsRed; CHGA-IRES-iRFP670


title: leukoplakia_withdysplasia_1 [miRNA]


title: Hgfac KO intact tissue_1

description: 10x Genomics

value: Epithelial/fibroblast/immune cells

value: no wound


title: genomic DNA from fibroblasts from healthy control 1 (replicate 2)

source_name: Control 1 (replicate 2)

value: fibroblasts

value: Control 1


title: S4

city: Boston

first: Yanhui

last: Hu

address: 77 Avenue Louis Pasteur

department: Genetics

institute: Harvard Medical School

data_processing:
The quality of the reads was evaluated with FastQC (Andrews, 2010).
The clean RNA-seq reads were mapped onto the reference genome Drosophila melanogaster BDGP6 using STAR
Assembly: BDGP6
Supplementary files format and content: raw count matrix, csv format

organism: Drosophila melanogaster

source_name: adult intestines

extract_protocol:
Total RNA was extracted from dissected adult intestines (after removal of the Malpighian tubules and crop) and RNA was extracted using Direct-zol RNA MicroPrep (Zymo Research)
RNA libraries were prepared for sequencing using standard Illumina protocols

value: adult intestines

value: mutant, CG5789 RNAi knockdown


title: hSIO - WT [scRNA-seq]

description:
10x Genomics
LX197


title: OLK_withoutdysplasia_3 [miRNA]

value: mucosa


title: RNA-Seq Vehicle control rep2

value: U937-derived macrophages

value: Vehicle control


title: genomic DNA from fibroblasts from HGPS patient (replicate 1)

source_name: HGPS (replicate 1)


title: RNA-Seq Vehicle control rep1


title: leukoplakia_withdysplasia_3 [miRNA]


title: leukoplakia_withdysplasia_2 [miRNA]


title: hBEC, signaling perturbations, donor no. 429581 [ali12]

first: Kalki

last: Kukreja

address: 200 Longwood Avenue

institute: Harvard University

description: Signaling perturbations performed on air-liquid interface (ALI) hBEC organotypic cultures

data_processing:
Raw FASTQ were prepared from Illumina bcl files in a format compatible with the inDrops.py pipeline. We ran the experiment with individual donor ( donor ID: 429581) using nextseq. Each nextseq run results in 16 FASTQ files (4 lanes x 4 reads per lane). We ran the nextseq run twice and hence have 16x2 = 32 raw files for this experiment. These contain 16 samples corresponding to 16 different conditions that are multiplexed for sequencing. For the second experiment, we ran 17 total samples with pooled donors (donor IDs: 221175 and 323353 ) on novaeseq platform. Each novaseq run results in 8 FASTQ files (2 lanes x 4 reads). The read files from an inDrops run have the following content:
*_R1_001.fastq.gz : contains the 3' UTR cDNA read
*_R2_001.fastq.gz : contains the first half of the cell barcode
*_R3_001.fastq.gz : contains the library index for demultiplexing libraries
*_R4_001.fastq.gz : contains the second half of the barcode and the UMI.
All subsequent processing steps from FASTQ files to count matrixes were performed using the custom pipeline for inDrops data analysis (github.com/indrops), using the parameter values specified in the .yaml files provided in the supplementary material. The yaml files also provide library indices that can be used for demultiplexing.
When cells were pooled prior to droplet capture, the cell barcodes were then assigned to each donor respectively by their SNP profiles using the scSplit (https://github.com/jon-xu/scSplit).
To obtain the final counts matrix used for data analysis, total count and a mitochondrial count filters were applied as described in McCauley,Kukreja et. al. 2022, biorXiv
Assembly: GRCh38/91
Supplementary files format and content: Raw counts (tsv.gz): library identity, barcode, gene names, and raw counts for all cells
Supplementary files format and content: Library barcodes and metadata (metadata.csv.gz): library identity, barcode, and associated metadata for all cells across in the sample

source_name: Airway epthelium

growth_protocol: Primary human bronchial epithelial cells (HBECs) from normal donors were obtained from Lonza (Cat. # CC-2540) and were expanded for two passages with Growth Medium (BEGM supplemented with one SingleQuots kit, Lonza Cat. # CC-3171) in uncoated T75 flasks (Corning Cat. # 430641U). After expansion, cells were seeded on uncoated 12-well TransWell (Corning Cat. # 3460) plates at a density of 1e6 cells per plate and expanded for 7 days in Differentiation Medium (50:50 BEGM (Lonza Cat. # CC-3171)/high-glucose DMEM (ThermoFisher Cat. # 11965175)) supplemented with one SingleQuots kit with no T3 and final retinoic acid concentration of 50 nM (Lonza # CC-4175). After 7 days, media was removed from the apical surface and cells were differentiated at air-liquid interface for an additional 30-45 days with media replaced every 48-72 hours.

extract_protocol:
HBECs were collected 14 days post-injury by treating with 0.05% Trypsin-EDTA (prepared by dilution of 0.25% Trypsin- EDTA, ThermoFisher Cat. # 25200056, with calcium/magnesium-free PBS), for 20 minutes at 37 °C. Trypsin dissociation was halted with Trypsin Neutralizer Solution (ThermoFisher Cat. # R002100) at 1:1 dilution and cells were filtered through a 40 μm strainer and pelleted by centrifugation at 300x g for 5 minutes. Cells were resuspended in phosphate buffered saline (ThermoFisher Cat. # 10010072) with 0.1% bovine serum albumin (ThermoFisher Cat. # B14) and counted on a Countess Cell Counter (ThermoFisher) to determine cell number and viability. For cell counting, 10 μL of cell suspension was mixed with 10 μL of Trypan Blue Solution (ThermoFisher Cat. # 15250061) and 10 μL of that 1:1 mix was loaded into Countess chip and read with default program. Viable cell numbers were used to determine cell concentration. Optiprep Density Gradient Medium (Sigma Cat. # D1556) was added to achieve a final concentration of 15% Optiprep and 120-150,000 cells/ml. Cells were captured by inDrops as detailed in Zilionis, R. et al. Single-cell barcoding and sequencing using droplet microfluidics. Nat. Protoc. 12, 44–73 (2017).
inDrops v3 scRNA-seq; Library construction as detailed in Zilionis, R. et al. Single-cell barcoding and sequencing using droplet microfluidics. Nat. Protoc. 12, 44–73 (2017).

treatment_protocol: After air-liquid interface differentiation, cells were washed 3X with warm 1XS PBS and treated with MEM (ThermoFisher Cat. # 10370088, 500 μL apical, 1 mL basal) for 1 hour at 37°C to remove differentiated luminal cells. Following incubation, luminal cells were removed by pipetting gently up and down 3-5X before removing apical media and washing with 500 μL MEM in apical chamber. Cytokines were diluted in Differentiation Medium at concentrations indicated. Cells were cultured for an additional 14 days, replacing media every 48-72 hours.

value: Airway epthelium

tag: donor ids

value: signaling molecules as specficied


title: 135722_INTER

first: Konstantin

last: Okonechnikov

state: Germany

address: Im Neuenheimer Feld 280

department: Pediatric Neurooncology

institute: German Cancer Research Center

description: Bulk RNA-seq

data_processing:
Reads aligned with STAR to hg19, gene expression counts computed with FeatureCount using gencode v.19 annoitation
Assembly: hg19
Supplementary files format and content: Gene expression matrix (raw)

source_name: Tumor

extract_protocol:
RNA were extracted from microdissected formalin-fixed and paraffin-embedded (FFPE) tissue samples using the automated Maxwell system with the Maxwell 16 FFPE Plus LEV DNA Purification Kit or Maxwell 16 LEV RNA FFPE Kit (Promega, Madison, WI, USA), according to the manufacturer’s instructions.
RNA libraries were generated with TruSeq RNA Access (Illumina) with the use of manufacturer-supplied protocols.

value: Tumor

tag: tumor class

value: Medulloblastoma

tag: tumor subclass

value: MB SHH

tag: tissue id

tag: location

value: Internodular


title: OSCC_2 [miRNA]

value: OSCC


title: RNA-Seq Vehicle control rep3

tag: cell type

tag: genotype


title: Hgfac KO intact tissue_3

description: 10x Genomics

tag: tissue

value: distal colon

value: Hgfac KO

tag: treatment


title: SNU601-vec-3

city: Cheonan

first: Keunsoo

last: Kang

state: Chungnam

address: Dandae-ro 119

department: Microbiology

country: South Korea

institute: Dankook University

data_processing:
Sequenced reads were subjected to trimming based on sequencing quality using Trim Galore (https://github.com/FelixKrueger/TrimGalore).
STAR (Dobin et al. Bioinformatics 2013) was utilized to align these cleaned reads to the reference human genome (hg38).
The abundance of all known genes was then estimated using StringTie (Pertea et al. Nat Biotechnol. 2015) according to the transcripts per kilobase million (TPM) method.
Differentially expressed genes (DEGs) were identified using DESeq2 (Love et al. Genome Biol. 2014) with default parameters
Assembly: hg38
Supplementary files format and content: tab-delimited text files include TPM values for each Sample
Supplementary files format and content: tab-delimited text files include raw read count values for each Sample

growth_protocol: SNU601 human gastric cancer cells were cultured in RPMI1640 with 10% fetal bovine serum at 37 °C in the presence of 5% CO2.

extract_protocol:
Total RNA was isolated in triplicate from SNU601-vec, SNU601-miR199a, and SNU601-miR199b cells using the AccuPrep Universal RNA Extraction Kit (Bioneer) and sent to Macrogen (Seoul, Korea) for RNA sequencing.
After checking the quantity and integrity of total RNA, an RNA library was constructed from 1 µg of total RNA using an Illumina TruSeq Stranded mRNA Sample Prep Kit (Illumina, San Diego, CA).

treatment_protocol: miR-199a_ or miR-199b_pLVX-Puro was introduced into SNU601 cells by lentiviral infection. Cells were infected with lentiviruses produced from 293T cells co-transfected with the lentiviral vectors, pMD2.G, and psPAX2 for 2–3 days, and then selected with puromycin (2 μg/mL) for more than 2 weeks.

tag: cell line

value: gastric cance cells

value: control


title: 135724_INTER

description: Bulk RNA-seq


title: OLK_withoutdysplasia_1 [miRNA]

tag: group


title: 135716_INTER

description: Bulk RNA-seq


title: NHDF_SBNO2KO_siSBNO1_Total3

city: Bethesda

first: Markus

last: Hafner

state: MD

address: 50 South Drive

department: NIAMS

institute: NIH

description: TotalNC_SBNO2KOplusSBNO1KD_gene_exp.diff

data_processing:
Base calling and demultiplexing was performed using bcl2fastq/2.20.0
For RNA-Seq fastq files were aligned to human genome (hg38) using STAR aligner star/2.7.2b.
Differential expression was performed using cuffdiff (cufflinks/2.2.1)
PAR-CLIP reads where analyzed using PARpipes including: removal of PCR duplicates, adapter trimming (cutadapt 2.4 with Python 3.7.2 ),  alignment to hg38 using bowtie/1.2.2 binding sites, cluster identification paralyzer
Genome_build: hg38
Supplementary_files_format_and_content: PAR-CLIP: comma separated values (clusters.csv) RNA-Seq: tab separated values .diff files

source_name: Human normal dermal fibroblast

growth_protocol: HEK293 and human normal dermal fibroblasts were cultured in DMEM supplied with 10% FBS and penicillinG/Streptomycin

extract_protocol:
For PAR-CLIP, Cells were cross-linked with 5000 uJ/cm2 of UV light (312nm) and fractionated nuclei were lysed and ribonucleocomplex were immunoprecipitated with anti-FLAGAb. Fluorescently labeled adaptors were ligated to the RNA and separated by SDS-PAGE. cDNA were synthesized by reverse transcript and subjected to library prep.
For PAR-CLIP, the cDNA was amplified by PCR and libraries were size selected by Pippinprep. For RNA seq, rRNA depletion was performed with the NEBNext® rRNA Depletion Kit (Human/Mouse/Rat) (New England BioLabs) and sequencing library construction was performed with NEBNext® Ultra™ II Directional RNA Library Prep with Sample Purification Beads (New England BioLabs) according to the manufacturer’s instructions. Each sample was multiplexed with NEBNext® Multiplex Oligos for Illumina® (96 Index Primers) (New England BioLabs). For poly A plus RNA-Seq from human fibroblasts, RNA  was extracted using RNeasy Plus Mini Kit (Qiagen).  Purified RNA was poly  (A) enriched and sequenced by Novogene Co.Ltd (California, USA).
PAR-CLIP-seq, RNA-seq

treatment_protocol: For PAR-CLIP, FH-SBNO2 were induced by 1 μg/ml tetracycline for 24 hours before cross-linking in the 70% confluent dishes. 100 mM of 4 thiouridine (4-SU) was added to the culture medium 12 hours before cross-linking.  For Poly A selected RNA-sequencing of SBNO2 KD fibroblasts, 0.2 million of NHFD from three different healthy neonatal donors (Lonza) were transfected with siSBNO2 (Thermo, HSS177015) as described above. After 72 hours, the cells were left without stimulation or stimulated with 10 ng/ml recombinant human TNF-a (PeproTech) for 1 hour or 6 hours.

tag: gene manipulation

value: SBNO2gRNA#2#3, siSBNO1#1-#3

tag: stimulation

value: No stimulation

tag: subcellular fractionation

value: Total

value: normal dermal fibroblast


title: S2

value: adult intestines

value: mutant, CG5789 RNAi knockdown


title: S3

value: wild type


title: OLK_withoutdysplasia_2 [miRNA]

value: mucosa


title: 135704_INTER

description: Bulk RNA-seq

value: Tumor


title: SNU601-vec-2


title: S1


title: SNU601-199B-2

value: miR-199b overexpression


title: normal_3 [miRNA]

value: normal


title: 135728_INTER

description: Bulk RNA-seq

value: Medulloblastoma

value: MB SHH

value: Internodular


title: 135710_INTER

description: Bulk RNA-seq


title: 135712_INTER

description: Bulk RNA-seq


title: SNU601-199B-3


title: NHDF_SBNO2KO_siSBNO1_Total1


title: NHDF_SBNO2KO_siSBNO1_Chrm3

value: Chromatin


title: NHDF_SBNO2KO_siSBNO1_Total2


title: OSCC_3

value: OSCC


title: OSCC_2


title: NHDF_SBNO2KO_siSBNO1_Chrm2


title: 135714_INTER

description: Bulk RNA-seq


title: 135708_INTER

description: Bulk RNA-seq


title: normal_1 [miRNA]


title: SNU601-vec-1

value: gastric cance cells

value: control


title: SNU601-199A-3

value: miR-199a overexpression


title: SNU601-199A-2


title: OSCC_1


title: normal_2 [miRNA]


title: 135720_INTER

description: Bulk RNA-seq


title: 136706_INTER

description: Bulk RNA-seq


title: NHDF_SBNO2KO_siSBNO1_Chrm1

value: SBNO2gRNA#2#3, siSBNO1#1-#3

value: No stimulation


title: SNU601-199A-1


title: leukoplakia_withdysplasia_3


title: SNU601-199B-1


title: NHDF_SBNO2KO_siNC_Total3

value: SBNO2gRNA#2#3, siRNA NC

value: Total

value: normal dermal fibroblast


title: leukoplakia_withdysplasia_2

value: mucosa


title: NHDF_SBNO2KO_siNC_Total2


title: NHDF_SBNO2KO_siNC_Total1


title: 135726_INTER

description: Bulk RNA-seq

value: Tumor


title: FV3

city: Shenzhen

first: Rufan

address: No.66,Gongchang Road,Guangming District

institute: Sun Yat-sen University

data_processing:
reads connector and quality filtering software:  First round: fastp, version 0.19.5, parameters -q 20, -f 1, --length_required 18  Second round: cutadapt, version 1.18, parameter -m 20-j 50
Rfam comparison:bowtie, version 1.2.2, parameter -n 1-e 80-l 18-a-m 3
Genome comparison:bowtie, version 1.2.2, parameters -n 1-e 80-l 18-a-m 3 --best
Small RNA identification：miRDeep2, version 2.0.1.2, mapper parameter -c - i-j-l 18
DESeq2 (with replicated samples) or EdgeR (without replicated samples) were used for differential expression analysis
Assembly: hg38
Supplementary files format and content: Tab-delimited text files include TPM values for each Sample

source_name: fusion vesicles

extract_protocol:
RNA was extracted using an RNA miRNeasy Micro Kit (Qiagen), and the quantity and purity were quantified by Quantus Fluorometer (Promega, USA) and Qsep100 (BIOptic, China)
Library construction was performed using Multiplex Small RNA Library Prep Set for Illumina (NEB, USA) following manufacturer's protocols.

value: fusion vesicles


title: FV2


title: 141798_NOD

description: Bulk RNA-seq

value: Nodular


title: FV1


title: leukoplakia_withdysplasia_1


title: 141800_NOD

description: Bulk RNA-seq

value: Medulloblastoma


title: NV1

source_name: cell membrane-based vesicles

value: cell membrane-based vesicles


title: 135718_INTER

description: Bulk RNA-seq

value: MB SHH

value: Internodular


title: OLK_withoutdysplasia_3


title: NV3


title: 141804_NOD

description: Bulk RNA-seq


title: normal_2

value: normal


title: NV2


title: 145482_NOD

description: Bulk RNA-seq


title: 141792_NOD

description: Bulk RNA-seq


title: ccRCC tumor - patient 3

city: Paris

first: Lucile

last: Massenet-Regad

address: 1 avenue Claude Vellefaux

institute: INSERM

data_processing:
FASTQ files were aligned on GRCh38-2020-A human reference genome using the CellRanger pipeline (version 4.0.0). Sequencing saturation was greater than 90% for all samples based on to Cellranger output files.
Processing of seurat object containing all samples : True cells were first distinguished from empty droplets using « emptyDrops » from the DropletUtils package (1.12.3). Data were preprocessed to keep cells expressing at least 200 genes, 1000 UMIs, and less than 20% of mitochondrial genes. Cells identified as doublets by scDblFinder (1.6.0) or scds (1.8.0) packages were removed from the analysis. Data were normalised for library size and log2-transformed. Dimensionality was reduced by principal component analysis (PCA) on the space of the 2,000 most variable genes, based on the variance-stabilizing transformation method. The six samples were integrated using Harmony (0.1.0) based on the 50 first principal components of their respective PCA. After sample integration, Louvain graph-based clustering was conducted to identify cell clusters (resolution = 1). Cells identities are stored in Celltype_Harmony metadata variable. Celltype_Harmony2 contains cell identities after tumoral cell from tumoral sample sub-clustering at 0.1 resolution.
Assembly: GRCh38
Supplementary files format and content: Output files from CellRanger (raw matrices : barcodes, features and matrix) for each scRNA-seq sample.
Supplementary files format and content: Seurat object of all merged samples

source_name: kidney ccRCC tumor

extract_protocol:
Tissue samples were mechanically and enzymatically digested in two rounds of 15 min incubation with agitation at 37°C with 2 mg/mL collagenase I (Sigma C0130), 2 mg/mL hyaluronidase (Sigma H3506), 0.1 mg/mL Liberase (Roche 05401020001), and 25 µg/mL DNAse I (Roche 1010415900). Cell suspension was filtered on 100µm filter and washed. Red blood cells were removed by magnetic isolation and cell suspension was splitted into two parts. One part was further enriched in CD45- cells by removing CD45+ cells using human CD45 microbeads kit. Cell suspensions were stained for viability and surface specific markers to be sorted on a BD FACS Aria III. Cells debris and doublets were excluded from FSC/SSC gatings. Dead cells were excluded based on viability marker gating. Following viable cell populations were isolated and sorted: 1) From a first enriched CD45- cell suspension fraction, we sorted CD45-CD3- cells; 2) From an other cell suspension fraction, we sorted CD45+CD3+cells, CD45+CD3-CD56+ cells, CD45+CD3-CD45-CD14-CD19+ cells, CD45+CD3-CD45-CD19-CD14+ cells, CD45+CD3-CD45-CD19-CD14-CD4+ cells, CD45+CD3-CD45-CD19-CD14- cells, and CD45+CD3-CD45-CD19-CD14-CD11c-CD4- cells. The detailed gating strategy for cell sorting is provided in Supp. Figure 1. Cells were sorted in Eppendorf tubes containing RPMI medium supplemented with 10% FCS (Hyclone), 100 U/ml Penicillin/Streptomycin, 1% MEM Non-Essential Amino Acids , and 1% Pyruvate.
Single-cell libraries were generated using the Chromium 10X Next GEM Single-cell 3’ Kit (v3.1) according to the manufacturer instructions (10X Genomics). Quality of the libraries was assessed by High Sensitivity D1000 ScreenTape assay (Agilent). Next-generation sequencing was performed on an Illumina NovaSeq 6000, with 200 000 reads/cell sequencing depth.

value: kidney ccRCC tumor

tag: patient

value: ccRCC patient 3

tag: tumor grade

tag: tumor stage

tag: Sex

value: Female

tag: age


title: OLK_withoutdysplasia_2


title: normal_3


title: NHDF_SBNO2KO_siNC_Chrm1

value: Chromatin


title: NHDF_NCgRNA_siNC_Total1

description: TotalNC_SBNO2KO_gene_exp.diff,TotalNC_SBNO2KOplusSBNO1KD_gene_exp.diff

value: NCgRNA, siRNA NC

value: No stimulation


title: NHDF_SBNO2KO_siNC_Chrm2

value: SBNO2gRNA#2#3, siRNA NC


title: OLK_withoutdysplasia_1


title: NHDF_NCgRNA_siNC_Total3

value: Total

value: normal dermal fibroblast


title: adjacent kidney tissue - patient 3

source_name: healthy kidney

value: healthy kidney


title: ccRCC tumor - patient 2

value: ccRCC patient 2

value: Male


title: NHDF_NCgRNA_siNC_Total2


title: ATAC-Seq - SUV39H1-edited 1928z Day50-3

city: New York

first: Michel

last: Sadelain

address: 1275 York Avenue, Box 182

institute: Memorial Sloan Kettering Cancer Cener

data_processing:
Reads were trimmed for both quality (<=15) and Illumina adaptor sequences using Trim Galore v0.4.5 (https://github.com/FelixKrueger/TrimGalore) then aligned to human assembly hg38 using bowtie2 with the default parameters. Duplicated reads that have the same start site and orientation were removed using the Picard tool MarkDuplicates (http://broadinstitute.github.io/picard). Accessible regions were called using MACS2 v2.1.2 against standard input as the control (fold change > 2 and p value < 0.001). Regions overlapping with genomic ‘blacklisted’ regions were removed. The peaks from all samples were then merged within 500 bp to create a full peak atlas. Raw read counts were tabulated over this peak atlas using featureCounts . All genome browser tracks were normalized to a sequencing depth of ten million mapped reads. Peaks were annotated using linear genomic distance, with a gene assigned to a peak if it was within 50 kb up- or down- stream of the gene start or end, respectively. Raw read counts in the peak atlas were normalized in DESeq2 prior differential and motif analyses. Significant differential regions were generally accepted with fold change greater than 1.5 and adjusted p value less than 0.1.
Assembly: Hg38
Supplementary files format and content: Processed data files contain log2foldchange (SUVKO-1928z/WT-1928z) counts along with associated statistics. The files are in .txt format

source_name: Primary immune cells

extract_protocol:
50,000 fresh T cells were washed in cold PBS and lysed. The transposition reaction containing TDE1 Tagment DNA Enzyme (Illumina catalogue # 20034198) was incubated at 37°C for 30 minutes. The DNA was cleaned with the MinElute PCR Purification Kit (QIAGEN catalogue # 28004) and material was amplified for 5 cycles using NEBNext High-Fidelity 2X PCR Master Mix (New England Biolabs catalogue # M0541L). After evaluation by real-time PCR, 8 additional PCR cycles were done. The final product was cleaned by aMPure XP beads (Beckman Coulter catalogue # A63882) at a 1X ratio, and size selection was performed at a 0.5X ratio.
Libraries were sequenced on a HiSeq4000 in a PE50 run, using the HiSeq 3000/4000 SBS Kit (Illumina).  An average of 108 million paired reads were generated per sample.

value: Primary immune cells

value: Primary T cells

value: CAR T cells isolated from mouse bone marrow at day 50 (post 3 NALM6 rechallenges)


title: NHDF_NCgRNA_siNC_Chrm2

description: ChrmNC_SBNO2KO_gene_exp.diff,ChrmNC_SBNO2KOplusSBNO1KD_gene_exp.diff


title: 141794_NOD

description: Bulk RNA-seq

value: Tumor

tag: tumor class

value: Medulloblastoma

tag: tumor subclass

value: MB SHH

value: Nodular


title: normal_1

value: mucosa

value: normal


title: adjacent kidney tissue - patient 1

value: ccRCC patient 1


title: 141812_NOD

description: Bulk RNA-seq

tag: tissue id

tag: location


title: NHDF_SBNO2KO_siNC_Chrm3

value: No stimulation

value: Chromatin

tag: cell type


title: adjacent kidney tissue - patient 2

tag: tissue


title: ccRCC tumor - patient 1

value: kidney ccRCC tumor


title: 141814_NOD

description: Bulk RNA-seq


title: NHDF_NCgRNA_siNC_Chrm3

value: NCgRNA, siRNA NC

value: normal dermal fibroblast


title: 145484_NOD

description: Bulk RNA-seq


title: NHDF_NCgRNA_siNC_Chrm1


title: ATAC-Seq - SUV39H1-edited 1928z Day50-4

tag: genotype

tag: treatment


title: ATAC-Seq - SUV39H1-edited 1928z Day50-1

tag: cell line


title: 141808_NOD

description: Bulk RNA-seq


title: 141796_NOD

description: Bulk RNA-seq


title: scB15 (P12),scRNAseq

city: Kalyani

first: Arindam

last: Maitra

state: West Bengal

address: PO: NSS

country: India

institute: National institute of Biomedical Genomics

description: 10x Genomics

data_processing:
The demultiplexing, barcoded processing and gene counting were made using the Cell Ranger software v4.0.0  (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger)
Assembly: GRCh38
Supplementary files format and content: Tab-separated values files and matrix files

source_name: OSCC-GB primary tumor tissue

extract_protocol:
Resected OSCC-GB tissue samples were collected in pre-chilled MACS tissue storage solution (Milteny Biotec) and transported on ice immediately after surgical procurement. The Freshly collected tissue was then cut into small pieces ≤ 1-2 mm with sterile scissors in a dish containing 1 mL of enzyme mix (enzymes H, R, and A from Miltenyi Biotech in RPMI1640 medium). The minced tissue suspension and the remaining enzyme mix were transferred to a gentleMACS C tube and then placed onto the gentleMACS Octo dissociator (Miltenyi Biotech). The GentleMACS program 37C_h_TDK_3 was run for dissociation after attaching the heating elements. Following completion of the program, the homogenate was briefly centrifuged and filtered through a 70-μm cell strainer (BD Bioscience) into a 50-mL centrifuge tube (Axygen). The strainer was then rinsed with 15 mL of RPMI1640 medium (Gibco) and the cell suspension was then centrifuged at 300g for 7 min. The supernatant was aspirated and the pellet was resuspended in RPMI1640 medium.  Red blood cells in the cell pellet were lysed by incubating the pellet in RBC lysis buffer (Miltenyi Biotec) for 10 minutes at 4°C. Then, 10 ml of chilled RPMI1640 medium was added and the tube was centrifuged at 300g for 10 min at 4°C. The supernatant was then aspirated and the cell pellet was resuspended in RPMI1640 medium.  The number of cells and fractions of live cells in suspension was counted using trypan blue and Live/Dead Viability/Cytotoxicity Kit (Thermo Fisher Scientific) in Countess II FL Automated Cell Counter (Thermo Fisher Scientific).
The scRNA-seq library was generated using single cell 3’ v3 protocol, 10x Genomics according to the manufacturer’s instructions. Briefly, isolated single cells were resuspended in the master mix and loaded together with partitioning oil and gel beads into the chip to generate the gel bead-in-emulsion (GEM). The poly-A RNA from the cell lysate contained in every single GEM was retrotranscripted to cDNA, which contains an Ilumina R1 primer sequence, Unique Molecular Identifier (UMI), and the 10x Barcode. The pooled barcoded cDNA was then cleaned up with Silane DynaBeads, amplified by PCR and the appropriated-sized fragments were selected with SPRIselect reagent for subsequent library construction. During the library construction Ilumina R2 primer sequence, paired-end constructs with P5 and P7 sequences and a sample index were added.

value: OSCC-GB primary tumor tissue

value: F


title: NHDF_SBNO2KD_Total2

tag: gene manipulation

value: siSBNO2 #1-#3

value: Total


title: ATAC-Seq - WT 1928z_Day50-2

value: Primary immune cells

value: Primary T cells

value: WT

value: CAR T cells isolated from mouse bone marrow at day 50 (post 3 NALM6 rechallenges)


title: scB14 (P11),scRNAseq

description: 10x Genomics


title: ATAC-Seq - WT 1928z_Day50-1


title: 141810_NOD

description: Bulk RNA-seq

value: Tumor

value: Medulloblastoma

value: MB SHH

value: Nodular


title: NHDF_SBNO2KD_Total1

tag: stimulation

value: No stimulation


title: ATAC-Seq - SUV39H1-edited 1928z Day50-2


title: NHDF_NC_Total3

value: si Negative Control

tag: subcellular fractionation


title: scB12 (P9),scRNAseq

description: 10x Genomics

value: M


title: scB13 (P10),scRNAseq

description: 10x Genomics


title: NHDF_NC_Total2


title: 141806_NOD

description: Bulk RNA-seq


title: scB10 (P8),scRNAseq

description: 10x Genomics


title: SUV39H1-edited 1928z Day50-5

data_processing:
The output FASTQ data were mapped to the target genome using the STAR RNA-seq aligner, resolving reads across splice junctions. The first mapping pass uses a list of known annotated junctions from Ensemble. Junctions found during the first pass are then added to the known junctions and a second mapping pass is done using the RemoveNoncanonical flag. After mapping, the output SAM files (SAMtools, version 0.1.19) were processed using PICARD tools (version 1.124; https://broadinstitute.github.io/picard/) to add read groups and AddOrReplaceReadGroups, which in addition sorts the file and converts it to the compressed BAM format. The expression count matrix was then computed from the mapped reads using HTSeq (version 0.5.3; https://htseq.readthedocs.io/en/release_0.10.0/) and one of several possible gene model databases. The R (version 3.2.0) package DESeq (https://www.huber.embl.de/users/anders/DESeq/) was used to both normalize the full dataset and analyze differential expression between sample groups.
Assembly: Hg38
Supplementary files format and content: Processed data files contain log2foldchange (SUVKO-1928z/WT-1928z) counts along with associated statistics. The files are in .txt format

extract_protocol:
Cell pellets were resuspended in RLT buffer and nucleic acids were extracted using the AllPrep DNA/RNA Mini Kit (QIAGEN catalogue # 80204) according to the manufacturer’s instructions. RNA was eluted in nuclease-free water and DNA in 0.5X Buffer EB. Phase separation in cells lysed in TRIzol Reagent (ThermoFisher catalogue # 15596018) was induced with chloroform. RNA was precipitated with isopropanol and linear acrylamide and washed with 75% ethanol. The samples were resuspended in RNase-free water.
RNA libraries for RNA-seq were prepared using SMARTER mRNA-Seq Library Prep Kit following manufacturer's protocols.


title: SUV39H1-edited 1928z Day50-4


title: NHDF_SBNO2KD_Total3


title: NHDF_NC_Total1


title: MB295_SS2

description: Smart-seq v2.5

data_processing:
10X data: reads aligned, assigned to cells and gene expression counts computed with CellRanger v3
SS2 data: reads aligned with STAR, gene expression counts per cell computed with FeatureCount
Filtering of low quality cells performed with Seurat
Assembly: hg19
Supplementary files format and content: Gene expression matrix (raw)

molecule: nuclear RNA

extract_protocol:
The tissue was put into 5ml of lysis buffer containing DTT and Triton-X (0.1 %), and subsequently lysed using a glass douncer and pestle. The nuclei were filtered two times (100µM and 40µM filters) and centrifuged for 5min at 1600rpm and 4°C, followed by two washing steps
The 10X Genomics 3’-Single Cell RNA-sequencing V2 protocol was applied to all samples according to the manufacturer’s instructions. Whole transcriptome snRNA-seq was performed following an adapted SMARTseq protocol (SMARTseq V2.5) for nuclei.

value: MB SHH INF

tag: m stage

tag: gender


title: scB9 (P7),scRNAseq

description: 10x Genomics


title: NHDF_SBNO2KD_Chrm3

value: Chromatin

value: normal dermal fibroblast


title: SUV39H1-edited 1928z Day50-2


title: MB266_SS2

description: Smart-seq v2.5


title: scB7 (P5),scRNAseq

description: 10x Genomics

value: OSCC-GB primary tumor tissue


title: MB263_SS2

description: Smart-seq v2.5

value: F


title: WT 1928z_Day50-5


title: NHDF_SBNO2KD_Chrm2

value: siSBNO2 #1-#3


title: NHDF_NC_Chrm3


title: NHDF_SBNO2KD_Chrm1


title: MBEN_AK3_SS2

description: Smart-seq v2.5


title: MB299_SS2

description: Smart-seq v2.5


title: scB5 (P4),scRNAseq

description: 10x Genomics


title: SUV39H1-edited 1928z Day50-3

value: Primary immune cells


title: Hgfac KO day2 wound_3

description: 10x Genomics

source_name: distal colon

value: distal colon

value: Epithelial/fibroblast/immune cells

value: Hgfac KO

value: wound


title: WT 1928z_Day50-4

value: Primary T cells

value: WT

value: CAR T cells isolated from mouse bone marrow at day 50 (post 3 NALM6 rechallenges)


title: MBEN_AK2_10X

description: 10X kit v2

value: Tumor

value: Medulloblastoma


title: WT 1928z_Day50-3


title: NHDF_NC_Chrm1

value: si Negative Control

value: No stimulation


title: OSCC (P1),scRNAseq

description: 10x Genomics

value: M


title: scB8 (P6),scRNAseq

description: 10x Genomics


title: WT 1928z_Day50-2


title: MBEN_AK1_10X

description: 10X kit v2

value: MB SHH INF


title: WT 1928z_Day50-1


title: PolyA_SBNO2KD NHDF_TNF6h_h6

value: 10ng /ml TNF 6h

value: Total

value: normal dermal fibroblast


title: genomic DNA from fibroblasts from healthy control 1 (replicate 1)

source_name: Control 1 (replicate 1)

value: fibroblasts

tag: donor

value: Control 1

tag: disease

value: Healthy

tag: age at skin biopsy (yrs)


title: MBEN_AK3_10X

description: 10X kit v2

tag: tumor class


title: SUV39H1-edited 1928z Day50-1

value: Primary immune cells


title: IL13+Labetuzumab treated ALI #4

city: Macclesfield

first: Carlos

middle: Lopez

last: Garcia

state: Cheshire

address: Alderley Park, Congleton Rd, Alderley Edge, West Didsbury

department: TLCB

country: United Kingdom

institute: CRUK-MI

description:
Group_3
CLG01_163

data_processing:
Adapter trimming performed using trimgalore (version 0.6.5)
Alignment performed using STAR aligner (version 2.5.1b) to GRCh38
BAM alignments were quantified in R (version 3.6.1) using featureCounts from the Rsubread library (version 2.0.1)
Assembly: GRCh38
Supplementary files format and content: raw counts (.csv)

source_name: Primary human bronchial epithelial cells Lonza batch #646466

growth_protocol: Cells were expanded following the protocol described by Hynds et al., (2016) (https://pubmed.ncbi.nlm.nih.gov/27539459/). To carry out ALI cultures, 40,000 HBECs in 100 µl bronchial epithelial growth medium (BEGM, Promocell) were seeded into Transwell PET cell culture inserts (12 well, 6.5mm, 0.4 µm pore, Corning). 500 µl of BEGM were added to the well (basolateral compartment) and both basolateral and apical media were replaced every two days. Air-lift was carried out when 100% confluency  (typically 7 days) was reached by removing the apical media and replacing the basolateral BEGM with differentiation medium containing 50% BEGM (without T3 supplement) and 50% DMEM. ALI cultures were grown for 3 weeks with basolateral medium changes every two days.

extract_protocol:
Rneasy MinELute Minikit
rRNA depletion using QIAGEN FastSelect rRNA H/M/R kit. RNA sequencing library preparation using NEBNext Ultra RNA Library Prep Kit

treatment_protocol: IL-13 (10ng/mL)was carried out during the third week of ALI development. It was added in the basolateral media of the ALIs and replaced every two days. Labetuzumab (2 µg/mL) was added in the apical compartment of the ALIs in 20 µl of PBS also during the third week of ALI development.

tag: age

tag: Sex

value: male

value: wild-type


title: IMB88_SS2

city: Heidelberg

description: Smart-seq v2.5

tag: tumor subclass


title: scB2 (P3),scRNAseq

description: 10x Genomics

value: OSCC-GB primary tumor tissue

value: F


title: Plasma PD4

city: Nanjing

first: Han

last: Xiaojuan

state: Jiangsu

address: Zhongshan Road 321

institute: Nanjing Drum Tower Hospital,  The Affiliated Hospital of Nanjing University Medical School

data_processing:
Paired-end reads were harvested from Illumina HiSeq 4000 sequencer, and were quality controlled by Q30. After 3’ adaptor-trimming and low quality reads removing by cutadapt software (v1.9.3). The high quality trimmed reads were used to analyse circRNAs, LncRNAs and mRNAs
The high quality reads were aligned to the human reference genome (UCSC hg19) with hisat2 software (v2.0.4). Then, guided by the Ensembl gtf gene annotation file, cuffdiff software (v2.2.1, part of cufflinks) was used to get the FPKM as the expression profiles of mRNA, and fold change and p-value were calculated based on FPKM, differentially expressed mRNA were identified.
And GO and Pathway analysis were performed on these target genes. GO and Pathway enrichment analysis were also performed based on the differentially expressed mRNAs.
Genome_build: Hg19
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

source_name: plasma

extract_protocol:
Plasma were collected, flash frozen on dry ice. Total RNA was isolated using trizol LS Reagent.
Total RNA was used for removing the rRNAs using Ribo-Zero rRNA Removal Kits (Illumina, USA) following the manufacturer's instructions. RNA libraries were constructed by using rRNA-depleted RNAs with TruSeq Stranded Total RNA Library Prep Kit (Illumina, USA) according to the manufacturer’s instructions. Libraries were controlled for quality and quantified using  the BioAnalyzer 2100 system (Agilent Technologies, USA).
10 pM libraries were denatured as single-stranded DNA molecules, captured on Illumina flow cells, amplified in situ as clusters and finally sequenced for 150 cycles on Illumina HiSeq Sequencer according to the manufacturer’s instructions.

tag: disease state

value: Parkinson

value: plasma

value: 72 year old

value: untreated


title: MB299_10X

description: 10X kit v2

tag: tissue

value: Tumor

value: Medulloblastoma

value: M


title: NHDF_NC_Chrm2

value: si Negative Control

value: No stimulation

value: Chromatin

tag: cell type


title: IL13+Labetuzumab treated ALI #3

description:
Group_3
CLG01_162

value: Primary human bronchial epithelial cells Lonza batch #646466

tag: genotype

tag: treatment


title: Plasma PD3

value: 59 year old


title: MB295_10X

description: 10X kit v2

value: MB SHH INF


title: IL13+Labetuzumab treated ALI #2

description:
Group_3
CLG01_161

value: male


title: Plasma PD2

value: 68 year old


title: PolyA_SBNO2KD NHDF_TNF6h_h1

tag: gene manipulation

value: 10ng /ml TNF 6h

value: Total

value: normal dermal fibroblast


title: IL13+Labetuzumab treated ALI #1

description:
Group_3
CLG01_160

value: wild-type


title: Plasma PD1

value: Parkinson

value: plasma

value: 65 year old


title: MB287_10X

description: 10X kit v2

value: Tumor

value: Medulloblastoma

value: F


title: MB266_10X

description: 10X kit v2

value: M


title: PolyA_NC NHDF_TNF6h_h5

value: si Negative Control

tag: stimulation


title: IL-13-treated ALI #4

description:
Group_2
CLG01_157

value: Primary human bronchial epithelial cells Lonza batch #646466


title: PolyA_SBNO2KD NHDF_TNF6h_h5

tag: subcellular fractionation


title: PolyA_SBNO2KD NHDF_TNF1h_h6

value: 10ng /ml TNF 1h


title: Plasma NC3

value: control

value: 64 year old

value: untreated


title: IL-13-treated ALI #2

description:
Group_2
CLG01_153

value: male


title: MB263_10X

description: 10X kit v2

value: MB SHH INF

tag: m stage


title: Plasma NC4

value: 67 year old


title: IL-13-treated ALI #3

description:
Group_2
CLG01_155

value: wild-type


title: MB88_10X

description: 10X kit v2

tag: gender


title: IL-13-treated ALI #1

description:
Group_2
CLG01_152


title: PolyA_NC NHDF_TNF6h_h1

value: 10ng /ml TNF 6h

value: Total

value: normal dermal fibroblast


title: PolyA_SBNO2KD NHDF_TNF1h_h5


title: Plasma NC2

value: plasma

value: 52 year old


title: SHAPE-MaP of NAI-treated NRAS-Luciferase, Replicate #2

city: Groningen

first: Danny

last: Incarnato

state: Netherlands

address: Nijenborgh 7

department: Molecular Genetics

institute: University of Groningen

description:
NRAS-Luciferase, NAI (rep. #2)
In vitro transcribed RNA
NRAS_Luciferase.wig

data_processing:
Library strategy: SHAPE-MaP
[RNA-seq]
Upon generation of FASTQ files by CCR-SF, the sequencing analysis was performed using the Center for Cancer Research Collaborative Bioinformatics Resource (CCBR) RNA-Seq pipeline (https://github.com/CCBR/Pipeliner). The CCBR pipeline was then deployed on the National Institutes of Health (NIH) HPC Biowulf cluster (https://hpc.nih.gov). In brief, the CCBR pipeline trims the sequencing adapters using the Cutadapt tool before alignment with the reference genome (Human - hg38) using STAR’s two-pass alignment system. The mapping statistics were calculated using Picard software. The average mapping rate of all samples was 95% and unique alignment was above 86%. There were 3.49 to 7.70% unmapped reads, and the samples have 0.20% ribosomal bases. Percent coding bases were between 56-61%, percent UTR bases were 32-36%, and mRNA bases were between 90-94% for all the samples. Library complexity was measured in terms of unique fragments in the mapped reads using Picard’s MarkDuplicate utility. The samples have 83-87% non-duplicate reads. Then, gene expression quantification analysis was performed using the RSEM tool yielding raw and normalized counts. Upon generation of gene counts by RSEM, a CPM (counts per million) filter (CPM < 0.5) was used to remove lowly expressed genes. Then, DESeq2 package was used to perform comparative analysis and derive fold change in expression and false discovery rate (FDR) adjusted p-values needed for generation of volcano plots. In addition to the output from the CCBR pipeline, we applied an additional Transcripts Per Kilobase Million (TPM) filter to only keep genes that have TPM > 3 in more than two samples when comparing an experimental set to DMSO controls. Only genes passing this TPM filter were shown in the volcano plots and used for gene ontology (GO) and pathway analysis.
[SHAPE-MaP]
All the relevant analysis steps have been conducted using the RNA Framework (https://github.com/dincarnato/RNAFramework). SHAPE raw reactivities were calculated via the rf-norm module, by using the previously published method4, followed by box-plot normalization.
Genome_build: hg38 (GRCh38)
Supplementary_files_format_and_content: RNA-seq: bigWig files
Supplementary_files_format_and_content: SHAPE-MaP: NRAS_Luciferase.wig

source_name: NRAS-Luciferase

extract_protocol:
Then all the transcripts were purified using Monarch RNA Cleanup Kit (NEB). The concentration was determined using a Nanodrop. Integrity and size of each transcript were confirmed using a 1% agarose gel. In vitro RNA folding and probing were carried out as previously described. Briefly, 2 ug of NRAS-G4-FL RNA in 89ul nuclease free H2O was heated at 95°C for 2 min, then immediately transferred to ice for 1 min. Then 10 ul of ice-cold RNA Folding Buffer 10X (500 mM HEPES pH 7.5; 1 M KCl) were then added, the solution mixed, and incubated at 37°C for 15 min. Then, 1 ul of 1 M MgCl2, pre-warmed at 37°C, was added, the solution mixed, and further incubated for 15 min. Probing was performed by adding 11 ul of NAI (1M stock), incubating at 37°C with moderate shaking for 10 min. For the control reaction, 11ul of neat DMSO was added. Reaction was quenched by adding 111ul of 1M DTT. RNA was recovered by purification on a Monarch RNA 10ug column (New England Biolabs).
Both DMSO- and NAI-treated samples, were fragmented in a buffer containing 4mM final MgCl2, by incubating at 94°C for 8 min. The fragmented RNA was then subjected to random-primed reverse transcription, as per standard SHAPE-MaP conditions. Briefly, fragmented RNA was mixed with 1ul of 10uM random decamers, and 1ul 10mM dNTPs, then incubated at 70°C for 5 min, and immediately transferred to ice for 1 min. Then, 4 ul of 5X RT Buffer (250mM Tris pH 8.0; 375mM KCl), 2 ul of DTT (0.1M), 1 ul of SuperaseIN RNase inhibitor (Ambion), 1 ul of SuperScript II RT (ThermoFisher Scientific), and 1 ul 120mM MnCl2 were added, the reactions mixed, and incubated at 25°C for 10 min, followed by 2 hours at 42°C. After reaction cleanup, the buffer was replaced with the standard SuperScript II RT First Strand Buffer with MgCl2, and the RNA-cDNA hybrids were used as input for the NEBNext® Ultra™ II Non-Directional RNA Second Strand Synthesis Module (New England Biolabs). After having converted the RNA into dsDNA, the remainder of the library preparation was then carried out using the NEBNext® Ultra™ II DNA Library Prep Kit for Illumina® (New England Biolabs), as per manufacturer instructions.

treatment_protocol: The plasmids pSKC11 and pSKC12, which encode the transcripts NRAS-G4-FL and NRAS-G4-Del-FL, respectively were received from Balasubramanian group. Then the plasmids were linearized at the 3' end using EcoRI, and linearized plasmids were used as a template for in vitro transcription. The 5'-capped transcripts were synthesized in vitro using mMessage mMachine T7 (ThermoFisher Scientific), followed by incubated with DNase for 15minutes at 37 °C to remove the template DNA.

tag: sample

value: NAI


title: PolyA_SBNO2KD NHDF_TNF1h_h1


title: PolyA_NC NHDF_TNF6h_h6


title: SHAPE-MaP of NAI-treated NRAS-Luciferase, Replicate #1

description:
NRAS-Luciferase, NAI (rep. #1)
In vitro transcribed RNA
NRAS_Luciferase.wig


title: Control  ALI (no IL-13, no labetuzumab) #4

description:
Group_1
CLG01_133


title: Control ALI (no IL-13, no labetuzumab) #2

description:
Group_1
CLG01_131


title: SHAPE-MaP of DMSO-treated NRAS-Luciferase, Replicate #2

description:
NRAS-Luciferase, DMSO (rep. #2)
In vitro transcribed RNA
NRAS_Luciferase.wig

value: DMSO


title: control+M.marinum 2

city: Camperdown

first: RENHUA

last: SONG

state: NSW

address: Charles Perkins Centre - D17, Level 4 West. The University of Sydney Johns Hopkins Drive

country: Australia

institute: Centenary Institute

data_processing:
Raw reads were checked for quality using FASTQC (v 0.11.8).
Low-quality reads and adapter sequences were removed to obtain clean reads using TRIMMOMATIC (v 0.39).
Clean reads were then mapped to Ensembl zebrafish genome (Danio_rerio.GRCz11.108) using STAR aligner.
The aligned bam files were quantified with featureCounts (v 2.0.2) [5] to generate read counts of each gene for all samples.
Differential expression analysis was performed using the DESeq2.
Assembly: GRCz11
Supplementary files format and content: Tab-separated raw count file
Supplementary files format and content: Bigwig files

organism: Danio rerio

source_name: Whole embryo

growth_protocol: Zebrafish embryos raised in E3 + PTU at 28 degrees

extract_protocol:
Trizol, phenol chloroform extraction, Qiagen RNAeasy
TruSeq RNA Library Prep Kit v2
RNA-seq (poly-A enriched)

treatment_protocol: Single cell embryos injected with scrambled or tsc1a gRNA/Cas9, infected at 2 dpf with M. marinum

value: Whole embryo

value: Scram gRNA/Cas9 injected and M. marinum infected


title: PolyA_NC NHDF_TNF1h_h5

value: si Negative Control

value: 10ng /ml TNF 1h


title: tsc1a KD+M.marinum 2

value: tsc1a KD+M.marinum

value: tsc1a gRNA/Cas9 injected and M. marinum infected


title: tsc1a KD+M.marinum 1


title: Control ALI (no IL-13, no labetuzumab) #3

description:
Group_1
CLG01_132

value: Primary human bronchial epithelial cells Lonza batch #646466

value: male


title: PolyA_NC NHDF_TNF1h_h6


title: control+M.marinum 1


title: Plasma NC1

value: control

value: 56 year old

value: untreated


title: Control ALI (no IL-13, no labetuzumab) #1

description:
Group_1
CLG01_128

value: wild-type


title: scB1 (P2),scRNAseq

description: 10x Genomics

value: OSCC-GB primary tumor tissue

value: M


title: PolyA_NC NHDF_TNF1h_h1

value: Total

value: normal dermal fibroblast


title: PolyA_SBNO2KD NHDF_Nostimulation_h6

value: No stimulation


title: tsc1a KD2

value: Whole embryo

value: tsc1a KD

value: tsc1a gRNA/Cas9 injected


title: RNA of scRNA-seq of a technical replicate of 3 single rosette organoids at 5-month old each with HTO tag A, B or C

city: Ann Arbor

first: Qianyi

last: Ma

state: MI

address: 1141 E Catherine St

department: Human Genetics

institute: University of Michigan

description: 5mo SOSRS RNA-2

data_processing:
Raw paired-end 10x scRNA-seq data were processed and aligned using CellRanger v4.0.0.
Raw RNA UMI count matrix were filtered to keep cells with >500 detected genes and <10% mitochondrial transcripts.
Raw paired-end HTO data were processed using CITE-seq-Count v1.4.4.
Cell barcodes present in both raw RNA UMI count matrix and raw HTO count matrix were kept for joint analysis.
Genome_build: GRCh38
Supplementary_files_format_and_content: raw RNA UMI count matrix; raw HTO counts matrix

source_name: Brain organoid cells

growth_protocol: iPSC lines were passaged using Accutase (Innovative cell) and replated onto Geltrex-coated (1:50 dilution in DMEM/F12) 12-well plates at 3-4 x 105 cells/well  in mTeSR1 with 10 µM rho-kinase inhibitor (Y-27632; Tocris, 1254). Medium without the inhibitor was replaced daily until the cells reach 80-100% confluency. The medium was then changed to 3N (50:50 DMEM/F12:neurobasal with N2 and B27 supplements)36 without vitamin A with 2 µM DMH1 (Tocris, 4126), 2 µM XAV939 (Cayman Chemical, 13596) , and 10 µM SB431542 (Cayman Chemical, 13031) (2 mL of media per well). Daily 75% media changes with 1 µM cyclopamine (Cayman Chemical, 11321) added began on day 1. On day 4, the monolayer was cut into squares using the StemPro EZ passage tool, and 1 mL of conditioned media was reserved. The squares were incubated for 1 minute with L7 hPSC passaging solution (Lonza). After aspirating the L7 solution, the squares were sprayed off the bottom of the culture plate with the 1 mL of preconditioned culture media with a P1000 micropipette. An additional 2 mL of fresh culture medium with the 4 inhibitors was added for a final volume of 3 mL. Approximately 200 µL of resuspended squares were then transferred into each well of a 96-well plate preincubated at 37 C for 20 minutes with 35 µL of 100% Geltrex solution/well. Up to 16 wells can be made using one monolayer. Higher densities will result in fusion between nearby SOSRS forming multi-rosette doublets or “strings”. After 48 hours, daily 50% media changes were performed. On day 6 (2 days after cutting), the four inhibitors were no longer added but replaced with 3 µM CHIR99021. After 3-4 additional days of culture. The organoids were individually removed using the STRIPPER Micropipetter (Cooper Surgical) with 275 µM tips and plated individually in the wells of a low-adherence U-bottom 96-well plate with 200 µL of 3N medium without vitamin A with 3 µM CHIR99021, BDNF (20 ng/mL), and NT3 (20 ng/mL). The ideal diameter for picking the SOSRS is ~ 250 µM. Starting 2 days later, half-media changes of 3N with vitamin A, BDNF (20 ng/mL), and NT3 (20 ng/mL) were repeated every other day. After 35 days of differentiation, 3N media with vitamin A (but without BDNF and NT3) was used for half-media changes every other day. At this time, SOSRS were transferred from the low-adherence 96-well plates to low-adherence 24-well plates for a necessary increase in media volume due to expanded SOSRS size.

extract_protocol:
One-, three-, or five-month SOSRS were dissociated using the Worthington papain dissociation kit. Cells from each organoid were labelled with sample-specific hashtag-labelled antibodies (HTO-A, Biolegend) to allow multi-sample pooled analyses to reduce batch effects, with subsequent de-multiplexing of scRNA-seq data based on the hashtags.
Single-cell suspensions were processed using the 10X Genomics Chromium platform, and scRNA-seq libraries were sequenced by Illumina NovaSeq. Each paired-end read contains a cell barcode and a UMI in Read 1, and the transcript-specific read in Read 2 with a read length of 151 nt.  Separately, each cell generates an HTO count vector for the hashtags used, allowing the cells to be assigned to organoid samples they come from.

value: Induced pluripotent stem cells

tag: cell line

tag: derived cell type

value: Brain organoid

value: 5 months


title: RNA-seq of SK-MEL-2, compound 18-treated, Replicate #2

description: SK-MEL-2, compound 18 (rep. #2)

data_processing:
[RNA-seq]
Upon generation of FASTQ files by CCR-SF, the sequencing analysis was performed using the Center for Cancer Research Collaborative Bioinformatics Resource (CCBR) RNA-Seq pipeline (https://github.com/CCBR/Pipeliner). The CCBR pipeline was then deployed on the National Institutes of Health (NIH) HPC Biowulf cluster (https://hpc.nih.gov). In brief, the CCBR pipeline trims the sequencing adapters using the Cutadapt tool before alignment with the reference genome (Human - hg38) using STAR’s two-pass alignment system. The mapping statistics were calculated using Picard software. The average mapping rate of all samples was 95% and unique alignment was above 86%. There were 3.49 to 7.70% unmapped reads, and the samples have 0.20% ribosomal bases. Percent coding bases were between 56-61%, percent UTR bases were 32-36%, and mRNA bases were between 90-94% for all the samples. Library complexity was measured in terms of unique fragments in the mapped reads using Picard’s MarkDuplicate utility. The samples have 83-87% non-duplicate reads. Then, gene expression quantification analysis was performed using the RSEM tool yielding raw and normalized counts. Upon generation of gene counts by RSEM, a CPM (counts per million) filter (CPM < 0.5) was used to remove lowly expressed genes. Then, DESeq2 package was used to perform comparative analysis and derive fold change in expression and false discovery rate (FDR) adjusted p-values needed for generation of volcano plots. In addition to the output from the CCBR pipeline, we applied an additional Transcripts Per Kilobase Million (TPM) filter to only keep genes that have TPM > 3 in more than two samples when comparing an experimental set to DMSO controls. Only genes passing this TPM filter were shown in the volcano plots and used for gene ontology (GO) and pathway analysis.
[SHAPE-MaP]
All the relevant analysis steps have been conducted using the RNA Framework (https://github.com/dincarnato/RNAFramework). SHAPE raw reactivities were calculated via the rf-norm module, by using the previously published method4, followed by box-plot normalization.
Genome_build: hg38 (GRCh38)
Supplementary_files_format_and_content: RNA-seq: bigWig files
Supplementary_files_format_and_content: SHAPE-MaP: NRAS_Luciferase.wig

source_name: SK-MEL-2 human melanoma cells

growth_protocol: MCF-7 and SKE-MEL-2 cells were seeded in 6-well culture plates at a density of 1 × 10^6 cells/mL for MCF-7 cells and 2.5 × 10^6 cells/mL for SK-MEL-2 cells.

extract_protocol:
Total RNAs were isolated using E.Z.N.A.® MicroElute Total RNA Kit (Omega), and DNA was removed by on-membrane DNase I (Omega) digestion according to the manufacturer's protocol. Then, isolated RNA integrity and quality were determined by Bioanalyzer (Agilent 2100) by using the Agilent RNA 6000 Nano kit.
500 ng of total RNA was used as the input for mRNA capture with oligo-dT coated magnetic beads. The library was prepared using Illumina TruSeq Stranded mRNA Library kit according to manufactured protocol and final purified products were then quantitated by qPCR before cluster generation and sequencing. Then, samples were sequenced 2x 76 cycles run on NextSeq500 sequencer. The samples have 21 to 56 million pass filter reads with more than 94% of bases above the quality score of Q30.

treatment_protocol: 24-hour post-seeding, 1 uM compound 18 or 1% DMSO (control) were added to the cells and were further incubated for 48 hours.

value: melanoma cells

value: Compound 18


title: HTO of scRNA-seq of a technical replicate of 3 single rosette organoids at 5-month old each with HTO tag A, B or C

description: 5mo SOSRS HTO-A,B,C-2


title: RNA-seq of SK-MEL-2, compound 18-treated, Replicate #1

description: SK-MEL-2, compound 18 (rep. #1)


title: SHAPE-MaP of DMSO-treated NRAS-Luciferase, Replicate #1

description:
NRAS-Luciferase, DMSO (rep. #1)
In vitro transcribed RNA
NRAS_Luciferase.wig

value: DMSO


title: tsc1a KD1


title: PolyA_SBNO2KD NHDF_Nostimulation_h1


title: HTO of scRNA-seq of 3 single rosette organoids at 5-month old each with HTO tag A, B or C

description: 5mo SOSRS HTO-A,B,C-1


title: control2

value: WT

value: Scram gRNA/Cas9 injected


title: control1


title: RNA-seq of SK-MEL-2, compound 18-treated, Replicate #3

description: SK-MEL-2, compound 18 (rep. #3)


title: PolyA_NC NHDF_Nostimulation_h5

value: si Negative Control


title: RNA-seq of SK-MEL-2, untreated, Replicate #3

description: SK-MEL-2, untreated (rep. #3)


title: RNA-seq of SK-MEL-2, untreated, Replicate #2

description: SK-MEL-2, untreated (rep. #2)


title: PolyA_NC NHDF_Nostimulation_h6


title: HTO of scRNA-seq of additional 3 single rosette organoids at 3-month old each with HTO tag D, E or G

description: 3mo SOSRS HTO-D,E,F

value: 3 months


title: PolyA_NC NHDF_Nostimulation_h1


title: RNA-seq of SK-MEL-2, untreated, Replicate #1

description: SK-MEL-2, untreated (rep. #1)


title: PolyA_SBNO2KD NHDF_Nostimulation_h5


title: RNA of scRNA-seq of additional 3 single rosette organoids at 3-month old each with HTO tag D, E or G

description: 3mo SOSRS RNA-D,E,F


title: HEK293 FLAG-HA-SBNO2-WT Clone #5 rep4

description: cDNA bound to FH-SBNO2

source_name: Human embryonic kidney (HEK) 293

value: FH-SBNO2 WT induced by tetracycline

value: 10 ng/ml INF a 2h

tag: par-clip antibody

value: a-FLAG (M2)

value: Nuclear


title: RNA of scRNA-seq of 3 single rosette organoids at 5-month old each with HTO tag A, B or C

description: 5mo SOSRS RNA-1

tag: age


title: HEK293 FLAG-HA-SBNO2-K284A rep5

description: cDNA bound to FH-SBNO2

value: FH-SBNO2 K284A induced by tetracycline


title: HTO of scRNA-seq of 3 single rosette organoids at 3-month old each with HTO tag A, B or C

description: 3mo SOSRS HTO-A,B,C


title: RNA of scRNA-seq of 3 single rosette organoids at 3-month old each with HTO tag A, B or C

description: 3mo SOSRS RNA-A,B,C


title: RNA-seq of MCF-7, compound 18-treated, Replicate #1

description: MCF-7, compound 18 (rep. #1)

source_name: MCF-7 human breast cancer cells

value: breast cancer cells


title: RNA-seq of MCF-7, compound 18-treated, Replicate #2

description: MCF-7, compound 18 (rep. #2)


title: HEK293 FLAG-HA-SBNO2- K284A rep3

description: cDNA bound to FH-SBNO2

value: No stimulation


title: Std1-MO, replicate 1

city: Ames

first: Raquel

last: Espin-Palazon

state: Iowa

address: 2437 Pammel Drive

institute: Iowa State University

data_processing:
Samples demultiplexed with bcl2fastq v2.20 Conversion Software (Illumina, San Diego, CA).
Adapters removed with AdapterRemoval v2.3.3 with read 1 adapter AGATCGGAAGAGCACACGTCTGAACTCCAGTCA, read 2 adapter AGATCGGAAGAGCGTCGTGTAGGGAAAGAGTGT and default settings.
Remaining technical sequences removed with Flexbar v3.5.0 command line options "--reads {fastq_read1} --reads2 {fastq_read2} --max-uncalled 3 --output-reads {out_fastq_read1} --output-reads2 {out_fastq_read2} --adapters {adapters_file} --adapter-trim-end LEFT --adapter-revcomp ON --adapter-revcomp-end RIGHT --adapter-min-overlap 5 --htrim-left GT --htrim-right CA --htrim-min-length 1 --htrim-adapter --htrim-error-rate 0.01 --min-read-length 15", where the adapters file provides the Template Switching Oligo (TSO) GCTAATCATTGCAAGCAGTGGTATCAACGCAGAGTACAT and reverse transcription primer AAGCAGTGGTATCAACGCAGAGTAC.
Gene expression quantified against the GRCz11 (RefSeq assembly GCF_000002035.6) RNA sequences, annotation release 106, using Salmon v1.9.0 with "--libType A --gcBias".
R library tximport v1.28.0 was used to convert Salmon isoform counts to gene counts.
DESeq2 v1.40.1 was used to find differentially expressed genes, with integer-rounded counts at least 10 across all samples, between pairs of libraries prepared on the same day.
Assembly: GRCz11
Supplementary files format and content: Nod1-Std1.csv: Comma-separated values file includes the output of both DESeq2 and Salmon, with columns: Ensembl gene ID, baseline mean expression, log2 fold change, standard error of log2 fold change, test statistic, p-value, adjusted p-value, then (count, transcripts per million [tpm] from Salmon, normalized counts from DESeq2) repeated for each replicate, with all rows ordered by p-value.

source_name: kdrl+ hemogenic endothelial cells

growth_protocol: mated, staged, raised and processed as described (Westerfield, 2000) in a circulating aquarium system maintained at 28C in accordance with ISU IACUC guidelines.

extract_protocol:
approximately 150 24hpf zebrafish embryos were dechorionated with pronase, anesthetized in 1% tricane, gently shaken at 28C for 5 mins with 0.05mg/ml liberase TM (Roche) in PBS solution with Ca2 + and Mg2+. The resulting cell suspension was filtered and stained with Sytox Red to exclude dead cells. kdrl+ cells were sorted with BD FACS Aria III.
RNA was isolated and purified with RNeasy Micro Kit (Qiagen). Total RNA was assessed for quality and quantity using an Agilent 2100 Bioanalyzer with RNA 6000 Pico Kit. Samples were then used to generate RNA sequencing libraries using NEBNext Single Cell/Low Input RNA Library Prep Kit (NEB #E6420S/L) for Illumina.

treatment_protocol: morpholinos were resuspended in nuclease-free water at 2mM, diluted with nuclease-free water and 1ul phenol red solution to reach a working concentration of 1mM. 1nl was injected in the yolk ball of one-cell-stage embryos using PLI-90A pico-injector warner instruments.

value: kdrl+ hemogenic endothelial cells

tag: strain

value: WT AB* x Tg(kdrl:HsHRAS-mCherry)s896

tag: Stage

value: 22 hours post fertilization (hpf)

value: Std-MO morpholino (CCTCTTACCTCAGTTACAATTTATA) injected at one-cell stage


title: RNA-seq of MCF-7, untreated, Replicate #3

description: MCF-7, untreated (rep. #3)


title: RNA-seq of MCF-7, compound 18-treated, Replicate #3

description: MCF-7, compound 18 (rep. #3)

value: Compound 18


title: HEK293 FLAG-HA-SBNO2-WT clone #2 rep2

description: cDNA bound to FH-SBNO2


title: RNA-seq of MCF-7, untreated, Replicate #2

description: MCF-7, untreated (rep. #2)

value: DMSO


title: HEK293 FLAG-HA-SBNO2-WT clone #5 rep1

description: cDNA bound to FH-SBNO2


title: RNA of scRNA-seq of 4 single rosette organoids at 1-month old each with HTO tag A, B, C or D

description: 1mo SOSRS RNA

value: Induced pluripotent stem cells

value: Brain organoid

value: 1 month


title: RNA-seq of MCF-7, untreated, Replicate #1

description: MCF-7, untreated (rep. #1)


title: Nod1-MO2, replicate 2

treatment_protocol: MOs were resuspended in nuclease-free water at 2mM, diluted with nuclease-free water and 1ul phenol red solution to reach a working concentration of 1mM. 1nl was injected in the yolk ball of one-cell-stage embryos using PLI-90A pico-injector warner instruments.

value: Nod1-MO2 morpholino (ACCAAATAAACATTACCTGGTCTGT) injected at one-cell stage


title: Std1-MO, replicate 3


title: Nod1-MO2, replicate 3


title: Nod1-MO2, replicate 1


title: G3BP_Enrich_Mock_4

status: Public on Oct 25 2023

city: Cleveland

first: Soon Yeul

last: Yi

state: OH

address: CWRU Wood Research Building W127A, 2109 Adelbert Rd,

department: Biochemistry

institute: Case Western Reserve University

data_processing:
*library strategy: CLIP-seq
Raw fastq files were first treated with fastX collapser to remove duplicated reads. Then, using CTK stripBarcode.pl the 7nt unique molecular identifier sequence at the 5'end was removed. Next, fastX trimmer and clipper was used to remove the barcode, spacer (GGG), and L32 sequence. Processed reads were then mapped to GRCh38.p14 genome using Bowtie2 (seed length 15nt, allow no mismatch). Mapped output (.sam) files were further processed using samtools (view, sort, index) and bedtools (bamtobed). Lastly, PCR duplicates were collapsed using CTK tag2collapse.pl.
Processed final bed files from each library were all combined into a single bed file and HOMER program was used to call peaks using makeTagDirectory and findPeaks (options: -o auto -style factor -L 2 -localSize 10000 -strand separate -minDist 50 -size 20 -fragLength 25). Finally, read counts per peak were normalized by the respective mapped depth of each library to generate the final normalized peak matrix.
Assembly: GRCh38.p14
Supplementary files format and content: tab-delimited text file containing peak coordinates, mapped-depth normalized read counts per peaks per libraries, peak annotation, and replicate counts.

growth_protocol: Huh-7.5 cell lines were grown and maintained at 37 °C and 5% CO2 in DMEM supplemented with 0.1 mM nonessential amino acids and 5% FBS.

extract_protocol:
Cell pellets were lysed with PBS lysis buffer (1x PBS, 0.1% SDS, 0.5% NaDoc, 0.5% NP-40, 1 protease inhibitor tablet (Roche)) for 10 minutes on ice. Lysate was then treated with RQ1 DNase (Promega) for 5 minutes at 37'C with shaking at 1100RPM.  The lysates were then briefly treated with RNase I at 1:100 or 1:5000 dilutions at the same condition as DNase treatment. Lysates were clarified by centrifuging on tabletop 4'C centrifuge for 10 minutes. HuR IP was performed overnight at 4'C with nutation, with an additional streptavidin capture step for coCLIP samples. Immunoprecipitated samples were labeled with radioactivity (gamma-32P-ATP (Revity)) and ran on SDS-PAGE. The gels were transferred to a nitrocellulose membrane, and bands 10-30kDa above the HuR (37kDa) were excised. The excised membranes were treated with proteinase K (Roche) to release the protein-bound RNAs from the membrane.
CLIP footprints were reverse-transcribed using the BrdU incorporation and bead-capture strategy described previously (PMID: 24613350).

treatment_protocol:
Cellular stress was induced by treating cells with 0.5mM sodium arsenite for 45 minutes. At 30 minutes prior to harvest, cells were prepared for proximity labeling  by incubating samples with 0.5mM biotin phenol for 30 minutes. Subsequently, the APEX reaction was initiated with 1mM hydrogen peroxide for 1 minute. After labeling, the reaction was stopped using ice cold quencher solution (10mM sodium ascorbate, 5mM Trolox in PBS). Quencher solution was replaced by ice-cold PBS for crosslinking using 254nm UV light, on ice, for a dose of 400mJ/cm^2 and a dose of 200mJ/cm^2. Samples were scraped into tubes, centrifuged and cell pellets stored at -80C until use.
Cellular fractions were acquired using a fractionation protocol described previously (PMID:26240403).

value: Hepatocellular Carcinoma

value: Mock

tag: apex-reaction

value: Yes

tag: hur antibody

value: HuR 3A2 (Santa Cruz, cat. #sc-5261)


title: G3BP_Enrich_Mock_3

tag: cell type


title: G3BP_Enrich_Arsenite_4

tag: genotype

value: Sodium Arsenite


title: G3BP_Input_Arsenite_4

tag: treatment


title: HTO of scRNA-seq of 4 single rosette organoids at 1-month old each with HTO tag A, B, C or D

description: 1mo SOSRS HTO-A,B,C,D


title: Std1-MO, replicate 2


title: G3BP_Enrich_Arsenite_5


title: NES_Enrich_Mock_1

value: APEX-NES


title: NES_Enrich_Arsenite_2


title: NES_Input_Arsenite_1


title: NES_Enrich_Mock_2


title: G3BP_Input_Mock_3


title: NLS_Input_Mock_1

value: APEX-NLS


title: NES_Input_Mock_1


title: NLS_Enrich_Arsenite_1


title: NLS_Enrich_Arsenite_2


title: NLS_Input_Arsenite_1


title: G3BP_Enrich_Arsenite_3


title: G3BP_Enrich_Arsenite_2


title: G3BP_Enrich_Arsenite_1


title: NES_Enrich_Arsenite_1


title: NLS_Enrich_Mock_2


title: NLS_Enrich_Mock_1


title: G3BP_Enrich_Mock_2


title: G3BP_Enrich_Mock_1


title: G3BP_Input_Arsenite_1


title: G3BP_Input_Arsenite_3


title: Nuc_Fraction_Arsenite_3

value: WT

value: No


title: G3BP_Input_Arsenite_2


title: G3BP_Input_Mock_2


title: G3BP_Input_Mock_1


title: Nuc_Fraction_Mock_1


title: Cyto_Fraction_Arsenite_3

molecule: cytoplasmic RNA


title: Nuc_Fraction_Arsenite_1


title: Nuc_Fraction_Mock_3


title: Nuc_Fraction_Arsenite_2


title: Cyto_Fraction_Mock_3


title: Cyto_Fraction_Mock_2


title: Silica + ATL-III sample 3

city: Changsha

first: Hanqin

last: Wang

address: Tongzipo road No.371

institute: Hunan Normal University

data_processing:
Illumina HiSeq/ Illumina Novaseq/ MGI2000 instrument
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.
Firstly, reference genome sequences and gene model annotation files of relative species were downloaded from genome website, such as UCSC, NCBI, ENSEMBL. 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).
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.
Assembly: mm10
Supplementary files format and content: tab-delimited text file includes raw counts for each Sample
Supplementary files format and content: tab-delimited text files include RPKM values for each Sample

source_name: lung tissue

extract_protocol:
Total RNA was extracted from the lung tissue using TRIzol (Genview) according to the manufacturer’s protocol.
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.

treatment_protocol: Lung tissue samples were collected and immediately snap-frozen in liquid nitrogen to preserve RNA integrity.

tag: Sex

value: male

tag: tissue

value: lung tissue

tag: time

value: Day 28

value: Silica + ATL-III


title: Silica sample 2

value: Silica


title: Silica + ATL-III sample 1


title: Control sample 3

value: Control


title: Control sample 1


title: Cyto_Fraction_Mock_1


title: Cyto_Fraction_Arsenite_1


title: Nuc_Fraction_Mock_2


title: Silica + ATL-III sample 2


title: Silica sample 1


title: Silica sample 3


title: Control sample 2


title: CD4_Michigan

first: Stephen

middle: R

last: Brooks

address: 50 SOUTH DR, RM 1140

department: Biodata Mining and Discovery Section

institute: NIAMS/NIH

data_processing:
Raw sequencing data were processed with CASAVA 1.8.2 to generate FastQ files. Reads of  were mapped to the mouse genome mm9 using TopHat 2.0.8.  FPKM was calculated using Cufflinks 2.1.1
Intron-less bigWig files were generated using USCS genome browser’s program bedGraphToBigWig, from bedGraph files made with the BEDtools package (Quinlan and Hall, Bioinformatics:26(6):841-2, 2010).
Assembly: mm9
Supplementary files format and content: bw files
Supplementary files format and content: Excel file containing FPKM values

source_name: Spleen and Lymph nodes

growth_protocol: ex vivo cells

extract_protocol:
RNA-seq was performed as described previously with slight modification (Hirahara et al., Immunity 2015). Total RNA was prepared from approximately 30-50,000 cells by using TRIzol following manufacture’s protocol (Thermo Fisher Scientific Inc).
Total RNA was subsequently processed to mRNA-seq library using TruSeq SR mRNA sample prep kit (FC-122-1001, Illumina). The purified libraries were multiplexed and captured on an Illumina flow cell for cluster generation. Libraries were sequenced for 50 single read cycles on HiSeq 2000 following the manufacturer's protocols.

value: Spleen and Lymph nodes

value: primary cells

value: CD4 T

value: {delta}CARD NOD1


title: CD4_WT

value: wild type


title: BM_Millenium

source_name: Bone marrow

value: Bone marrow

value: Lineage negative bone marrow cells


title: KO_MI rep2

city: Beijing

first: Yan

last: Liu

address: 2 Anzhen Road, Chaoyang District

institute: Beijing Institute of Heart Lung and Blood Vessel Diseases, Beijing Anzhen Hospital, Capital Medical University

data_processing:
Raw reads are filtered into clean reads. Filtering steps are as follows: 1) Remove reads with adaptors; 2) Remove reads in which unknown bases are more than 10%; 3) Remove low quality reads (the percentage of low quality bases is over 50% in a read, we define the low quality base to be the base whose  sequencing quality  is no more than 5).
Bowtie2 was used to map clean reads to reference gene and BWA was used to reference genome.
FPKM (Fragments Per Kilobase Millon Mapped Reads) was then calculated to estimate the expression level of genes
Assembly: mm9
Supplementary files format and content: Tab-delimited text files includes FPKM values for each sample

source_name: Heart

extract_protocol:
Total RNA was extracted from the tissues using Trizol (Invitrogen, Carlsbad, CA, USA) according to the manufacturer’s instructions. 1 ug of total RNA was used for the construction of sequencing libraries.
RNA libraries for RNA-seq were generated using Illumina's TruSeq RNA Library Prep Kit (Illumina, San Diego, CA) following the manufacturer’s protocols. Briefly, mRNA is enriched by using the oligo (dT) magnetic beads. Mixed with the fragmentation buffer, the mRNA is fragmented into short fragments. Then the first strand of cDNA is synthesized by using random hexamer-primer. Buffer, dNTPs, RNase H and DNA polymerase I are added to synthesize the second strand. The double strand cDNA is purified with magnetic beads. End reparation and 3’-end single nucleotide A (adenine) addition is then performed. Finally, sequencing adaptors are ligated to the fragments. The products were then purified and amplified using PCR.

value: Heart

value: Card9 KO

value: MI


title: wildtype heart tissue, biological replicate 1

type: RNA

city: Baltimore

first: Daniela

last: Cihakova M.D. Ph.D. D(ABMLI)

address: 720 Rutland Avenue

department: Pathology-Immunopathology

institute: The Johns Hopkins University School of Medicine

description: wildtype

hyb_protocol: 17hrs at 45° C with rotation (60rpm) as described by Affymetrix in their GeneChip Expression Analysis Technical Manual

scan_protocol: Using the Affymetrix GeneChip Scanner 3000 7G and default parameters described by the manufacturer in their GeneChip Expression Analysis Technical Manual

data_processing: Affymetrix CEL files were extracted with the Partek GS 6.6 platform and their data RMA and quantile-normalized to create log2 transcript signal values, which were summarized by Partek from exon to genes and used in subsequent ANOVA analysis.    Transcript annotation was subsequently updated.

label: biotin

source_name: wildtype heart tissue

label_protocol: The Affymetrix Ambion Expression WT kit was used as described in the manufacturer's manual.

growth_protocol: mouse left ventricular heart tissue dissected from 10-15 weeks old mice without perfusion

extract_protocol: Tissue was frozen at -80C. RNA was isolated with Trizol.

treatment_protocol: not applicable

value: 11.7 weeks

value: heart, left ventricle

value: wildtype


title: BM_WT


title: IL-5 transgenic heart tissue, biological replicate 2

description: IL-5 transgenic (NJ.1638)

source_name: IL-5 transgenic heart tissue

value: 13.7 weeks

value: male

value: IL-5 transgenic (NJ.1638)


title: ddblGATA1 IL-5 transgenic heart tissue, biological replicate 1

description: ddblGATA1 IL-5 transgenic (NJ.1638)

source_name: ddblGATA1 IL-5 transgenic heart tissue

value: 12.3 weeks

value: ddblGATA1 IL-5 transgenic (NJ.1638)


title: IL-5 transgenic heart tissue, biological replicate 3

description: IL-5 transgenic (NJ.1638)


title: ddblGATA1 IL-5 transgenic heart tissue, biological replicate 2

description: ddblGATA1 IL-5 transgenic (NJ.1638)

value: 10.6 weeks


title: wildtype heart tissue, biological replicate 3

description: wildtype

value: 10.9 weeks


title: IL-5 transgenic heart tissue, biological replicate 1

description: IL-5 transgenic (NJ.1638)

value: 15.6 weeks