title: DOG-FIA-2_S95

city: College Station

first: Ivan

last: Rusyn

state: TX

address: VIDI(Veterinary Medicine 2), 2, 402 Raymond Stotzer Pkwy

department: Interdisciplinary Faculty of Toxicology

institute: Texas A&M University

description:
Library name: DOG-FIA-2_S95
normcounts_dog.csv
qc_hash_dog.csv

data_processing:
The gene count matrices were generated from paired-end reads with 2 lanes per sample with ~150 base pairs (bp).
The reads from the 2 lanes for each sample were combined to generate a single fastq file.
Reads were quality trimmed using fastp (version 0.23.2) with the following steps: (1) automatic detection of adapter sequences for paired-end reads, (2) trimming of homopolymer stretches, and (3) removal of bases with Phred quality scores below 20 (Chen et al., 2018).
Reads were aligned to the species-specific genome assembly using STAR version 2.7.10b (Dobin et al. 2013). Aligned reads were sorted with SAMtools version 1.14 (Danecek et al. 2021), and gene-level counts were generated using HTSeq version 2.0.1 (Anders et al. 2014).
Samples with fewer than 1 million total counts or with uniquely mapped reads below 50% were excluded from further analysis. For transcript-level quality control, transcripts that were not expressed across the entire sample set for each species were removed. The Bioconductor package DESeq2 (Love et al., 2014) was used to perform within-species normalization and differential expression analysis across all experimental conditions.
Assembly: Reads from dog, monkey, human, and rat were mapped to the ROS_Cfam_1.0, Macaca_fascicularis_6.0, GRCh38, and mRatBN7 genome assemblies, respectively, using STAR. Gene annotations from Ensembl GTF files corresponding to each assembly were used for quantification.
Supplementary files format and content: normcounts_* files contain normalized count data using DESeq2 R package (Love et al., 2014) for each species
Supplementary files format and content: qc_hash* files contain meta data of the qc-passed samples for each species

organism: Canis lupus familiaris

source_name: hepatocytes

extract_protocol:
Total RNA was extracted using TRIzol reagent (Invitrogen) and the Direct-zol RNA Miniprep (Zymo, USA) following the manufacturer’s protocol with slight modifications detailed here. MPS Scaffolds from each experiment were preserved in RNAlater at -80°C until RNA extraction. On the day of extraction, samples were thawed at room temperature, and each scaffold was cut into two pieces before transferring to a 1.5 mL microcentrifuge tube. Next, 700 µL of TRIzol reagent was added to the samples, which were then incubated at room temperature for 20 minutes. During incubation, the samples were vortexed for 30 seconds every 5 minutes to ensure complete lysis. The scaffolds were removed from the microcentrifuge and the RNA was precipitated using equal volume of 100% molecular grade ethanol, then transferred to a Zymo-Spin™ IICR column. The remainder of the extraction was performed according to the standard protocol. RNA was eluted in 60 uL of DNAase/RNAse free water.
mRNA was isolated from 150ng total RNA using a Nextflex Poly-A Selection kit. Unique dual indexed cDNA libraries were prepared on a Sciclone NGSx liquid handler, using a Nextflex Rapid Directional RNA 2.0 kit miniaturized to 2/5 reaction volume. Optimal PCR cycle count was empirically determined to be 17 by cycling a pool of samples to the plateau phase on a real-time PCR machine. The concentration of libraries was determined using a Molecular Devices (San Jose, CA) Spectramax M2 fluorescent plate reader and Thermo Fisher Scientific (Waltham, MA) Picogreen Assay. Libraries were then pooled by equal mass using a Janus Mini liquid handler. The libraries were pooled by equal mass. The pool was sequenced on a Illumina (San Diego, CA) NovaSeq X+ 1.5B flowcell using the 2x150 bp recipe.

value: hepatocytes

value: fialuridine


title: HEK293T cells, Parental, rep 1

status: Public on Dec 04 2025

type: SRA

city: Albany

first: Cara

middle: T

last: Pager

state: NY

address: 1400 Washington Avenue

country: USA

institute: University at Albany, State University of New York

library_source: transcriptomic

data_processing:
After sequencing the quality of the fastq files were checked using the program FastQC (version 0.11.9).
STAR (version 2.7.10a) was used to align the fastq files to the hg38 genome with the --quantModeGeneCounts option added to count the reads per gene.
Assembly: hg38
Supplementary files format and content: Tab-delimited text files contain the raw read count abundances for each sample.

molecule: total RNA

organism: Homo sapiens

growth_protocol: HEK293Tcells (parental) were stably transfected using the piggyBAC transposon system tointegrate both ATG-(CAG)60-Myc-NLuc and ATG-(CAG)0-Myc-FLucconstructs into the HEK293T cell genome. Cells were single cellsorted and clonesthat express both integrands at high (clone 37) and low (clone 15) levels were selectedfor detailedcharacterisation by RNA sequencing. Parental, clone 15 and clone 37cell lines were cultured in DMEM supplementedwith 10% fetal bovine serum (FBS)and 1x penicillin/streptomycin in a humidified atmosphere at 37 degrees C at 5%CO2.

extract_protocol:
RNA for CAG containingreporter cell lines was extracted using Aurum total RNA mini kit (BioRad) withon-column DNase treatment, following themanufacturer’s protocol. The Qubit RNA high sensitivity assay wasused to obtain RNA concentrations(Thermo Fisher Scientific). RNA quality waschecked via capillary electrophoresis on a 5300 fragment analyzer (AgilentBioanalyzer) using the RNA AnalysisDNF-471 RNA kit (Agilent).
The NEBNext Ultra IIDirectional RNA Library Prep Kit (Illumina) with NEBNext rRNA Depletion Kit(New EnglandBiolabs) was used to prepare RNA-seq libraries, with a total of500 ng input RNA from each sample. The manufacturer’sprotocols were followed,with the following exceptions: 40× adaptor dilutions were used, and 13 cyclesof libraryamplification were performed. The resulting libraries were pooled inequimolar amounts, quantified using the NEBNextLibrary Quant Kit for Illumina(New England Biolabs), quality checked via capillary electrophoresis on the5300fragment analyzer (Agilent Bioanalzyer) using the DNF-474 HS NGS kit(Agilent). Libraries with sufficient quantitywere sequenced using paired-end, 75 base pairsequencing (parental and clone 37), or 100 base pair sequencing (clone 15) onthe Illumina NextSeq 500 sequencer at the University at AlbanyCenter for functional genomics. All datasets had total read depths greaterthan 40 million reads.

tag: cell line

tag: cell type

value: Human embryonic kidney

tag: genotype

value: Parental


title: HEK293T cells, Clone 37, rep 3

value: Clone 37


title: DOG-FIA-1_S94

description:
Library name: DOG-FIA-1_S94
normcounts_dog.csv
qc_hash_dog.csv

tag: treatment


title: Drug_2

city: Shanghai

first: Jun

last: Mi

address: 280 South Chongqing Rd.

department: Biochemistry & Molecular Cell Biology

country: China

institute: Shanghai Jiao Tong University

description: Library name: Sample 4

data_processing:
After sequencing, demultiplexing with Bcl2fastq2 was employed to generate FASTQ files for each tumor sample.
Assembly: hg38
Supplementary files format and content: Tab-delimined text files includes raw counts for each sample
Supplementary files format and content: CSV files include FPKM value for each sample

source_name: liver cancer

extract_protocol:
RNA was extracted by trizol.
For each RNA sequencing experiment, approximately 500 ng of total RNA was used for cDNA library preparation. cDNA library construction followed the Illumina TruSeq Stranded mRNA-seq Sample Preparation Guide.

tag: tissue

value: liver cancer

value: liver cell line PLC8024

value: tumor cell line

value: drug


title: HS1

city: Wuhan

first: Jinyang

last: Gu

address: 1277 Jiefang avenue

institute: Union hospital, Tongji medical college, Huazhong university of science and technology

data_processing:
CLC Genomics Workbench v 11.0.1
Sequence reads were trimmed for adapter sequence/low quality sequence using CLC Genomics Workbench (parameter-Quality limit: 0.05).
Trimmed sequence reads were mapped to GRCm39/mm39 using CLC Genomics Workbench (parameters-mismatch cost: 2 insertion cost: 3 deletion cost: 3 length fraction: 0.8 similarity fraction: 0.8)
Read count extraction and normalization were performed using CLC genomic workbench.
Assembly: mm39
Supplementary files format and content: xlsx file includes TPM for each sample.

organism: Mus musculus

source_name: Liver

extract_protocol:
RNA was extracted by Trizol method.
1) By utilizing the structural characteristic that most eukaryotic mRNAs carry a polyA tail, mRNAs with polyA tails were enriched by Oligo(dT) magnetic beads; 2) The purified mRNAs were cleaved into small fragments with fragmentation buffer at a suitable temperature; 3) First-strand cDNAs were produced by reverse transcription using a random hexamer primer; 4) Second-strand cDNAs are synthesized (strand-specific library: dUTPs were used instead of dTTPs in the second-strand synthesis to incorporate dUTPs in the second-strand cDNAs, while the high-fidelity DNA polymerase used in this method could not amplify uracil-containing DNA templates, thus realizing the strand specificity), while simultaneously performing end repair and dA-Tailing; 5) Sequencing adapter ligation was performed, followed by DNA magnetic bead purification and fragment selection after ligation was completed to yield a library with 250-350 bp insert fragments; 6) The ligated products were amplified by PCR and purified again using DNA magnetic beads, where the products were solubilized with nuclear-free water; 7) After the initial library was constructed, a Qubit fluorescence quantifier was used for concentration detection, followed by a Qsep400 high-throughput biofragment analyzer for fragment size detection; 8) The double-stranded DNA library obtained by PCR was subjected to single-stranded cyclization to obtain a single-stranded circular DNA library; 9) The final library was amplified with phi29 before uploading to make DNA nanoballs (DNB) with more than 300 copies of a molecule, and the DNBs were loaded into the sequencing chip and sequenced on the BGI sequencing platform.

value: Liver

value: WT

value: HFD-Sham


title: HEK293T cells, Clone 37, rep 2


title: RDv16_vag

city: Minneapolis

first: David

last: Masopust

address: 2101 6th St. SE

institute: University of Minnesota

data_processing:
RNA sequencing (50-bp single-end) with the HiSeq 2500 Illumina was done at the University of Minnesota Genomics Center (UMGC). The sequences were processed with the Collection of Hierarchical UMII-RIS Pipelines (CHURP, v 0.2.2) developed by the Research Informatics Solutions (RIS) group at the Minnesota Supercomputing Institute. The reference implementation of the CHURP package is at: https://github.com/msi-ris/CHURP.git 64. For whole tissue cell lysates, 2x 50bp FastQ paired-end reads were trimmed using Trimmomatic (v 0.33) enabled with the optional “-q” option; 3bp sliding-window trimming from 3’ end requiring minimum Q30. For CM9-specific CD8 T cells, 2 x 50bp FastQ paired-end reads were trimmed using Trimmomatic (v 0.33) enabled with the optional “-q” option; 3bp sliding-window trimming from 3’ end requiring minimum Q30 and headcrop 3. Quality control on raw sequence data for all samples was performed with FastQC.
Read mapping was performed via Hisat2 (v2.1.0) using the Macaca mulata genome (Mmul_10) as reference.
Assembly: Mmul_10
Supplementary files format and content: csv files contain raw read counts for each sample

organism: Macaca mulatta

source_name: vagina

extract_protocol:
For bulk RNA sequencing experiments, reproductive tract tissue was isolated directly into RNAlater (Qiagen). Tissue was homogenized in Trizol (Sigma-Aldrich) using a Tissuemiser homogenizer (Fisher Scientific), and RNA was extracted using a RNeasy mini kit as per the manufacturer’s instructions (QIAGEN). RNA integrity was assessed using capillary electrophoresis with the Agilent 2100 BioAnalyzer system (Agilent Technologies).
Sequencing libraries for CM9-specific CD8 T cells were prepared using the Clontech SMARTer® Stranded Total RNA-Seq Kit v2 - Pico Input Mammalian kit. Libraries for whole tissue cell extracts were prepared using the TruSeq stranded mRNA library prep kit (Illumina).

value: vagina

value: all

tag: Sex

value: female

tag: haplotype

tag: vaccination

value: heterologous virus vectors VSVnj, rVV, rAd5 encoding SIV-gag

value: 20mg CM9 peptide intravaginal challenge

tag: timepoint


title: NS1

value: ND-Sham


title: S25

type: RNA

city: Zhuhai

first: Hongzhen

last: Tang

state: Guangdong

address: Cuijing Road, Qianshan, Xiangzhou District

institute: BioMega Spatiotemporal Pathology Research Center

hyb_protocol: Overnight (37 °C) hybridisation with 980-plex CosMx ISH probe mix

scan_protocol: Cyclic RNA imaging & Z-stack acquisition on NanoString CosMx SMI (0.34 µm pixel, 100 × objective)

data_processing: AtoMx SIP v3.0 for segmentation & molecule assignment ; downstream analysis in Python 3.8 (squidpy + scanpy 1.9.6 : PCA 50 PC, KNN, UMAP, Louvain r = 1)

molecule: polyA RNA

source_name: gastric cancer

label_protocol: not applicable — in-situ FISH, no separate molecular label

growth_protocol: primary tumour tissue

extract_protocol: Sections baked 60 °C o/n; HIER; PK; fiducials; formalin re-fix; NHS-acetate block

value: gastric cancer

tag: fov

tag: age

tag: grade

tag: stage#

tag: Stage

value: Ib

tag: ln

tag: t

tag: n

tag: m


title: DOG-D8-3_S45

description:
Library name: DOG-D8-3_S45
normcounts_dog.csv
qc_hash_dog.csv

tag: cell type

value: hepatocytes


title: Old CR 28 months old_18

city: Liverpool

first: Gulam

last: Altab

address: 6 West Derby Street

country: United Kingdom

institute: University of Liverpool

data_processing:
The raw Fastq files were trimmed for the presence of Illumina adapter sequences using Cutadapt version 1.2.1.
The reads were further trimmed using Sickle version 1.200 with a minimum window quality score of 20
The filtered reads were mapped to the rat reference genome (ftp.ensembl.org/pub/release-100/fasta/rattus_norvegicus/dna/) using bowtie 2 2.4.0. Alignment score was above 90% for all the samples.
To count the number of reads that mapped to each gene,miRDeep2 version 2.0.1.2 was used to quantify the miRNA expression
Assembly: Ensembl Rattus_norvegicus.Rnor_6.0.96.gtf
Supplementary files format and content: rawcounts_miRNA_all_samples.txt

organism: Rattus norvegicus

source_name: gastrocnemius muscle

extract_protocol:
Total RNA from gastrocnemius muscle tissues was isolated using RNeasy Plus Universal Mini Kit (Qiagen, UK) according to manufacturer instructions.
Utilizing the NEBNext small RNA library preparation Kit for Illumina, strand-specific RNASeq library preparation was performed

treatment_protocol: Between 21–28 days of age, rats were kept in barrier conditions on a 12-h light:12-h dark cycle (08:00–20:00). Regular health monitoring occurred through sentinel animal screening. Until 2 months old, rats were group-caged and fed ad libitum. At 2 months, rats were individually housed and randomly assigned to either caloric restriction (CR) or ad libitum (AL) groups. AL and CR rats were sacrificed at 6, 12, and 28 months. CR rats were fed the CRM diet at 55% of the daily intake of control rats to maintain body weight at ~55% of age-matched AL-fed animals. CR rats received pre-weighed rations daily between 10:30 and 11:00 h.

value: gastrocnemius muscle

value: 28 months

value: Caloric Restriction


title: Control_2

description: Library name: Sample 3

value: liver cancer

tag: cell line

value: liver cell line PLC8024

value: tumor cell line

value: control


title: Old CR 28 months old_17

tag: treatment


title: Old CR 28 months old_16

tag: tissue


title: DOG-D8-2_S44

description:
Library name: DOG-D8-2_S44
normcounts_dog.csv
qc_hash_dog.csv


title: Drug_1

description: Library name: Sample 2

value: drug


title: HEK293T cells, Clone 15, rep 3

value: Human embryonic kidney

tag: genotype

value: Clone 15


title: DOG-D8-1_S43

description:
Library name: DOG-D8-1_S43
normcounts_dog.csv
qc_hash_dog.csv


title: HEK293T cells, Clone 37, rep 1

value: Clone 37


title: DOG-FIA-3_S96

description:
Library name: DOG-FIA-3_S96
normcounts_dog.csv
qc_hash_dog.csv

value: fialuridine


title: Control_1

description: Library name: Sample 1


title: DOG-D4-3_S42

description:
Library name: DOG-D4-3_S42
normcounts_dog.csv
qc_hash_dog.csv


title: HEK293T cells, Clone 15, rep 2


title: HEK293T cells, Clone 15, rep 1


title: Old CR 28 months old_14


title: DOG-D4-2_S41

description:
Library name: DOG-D4-2_S41
normcounts_dog.csv
qc_hash_dog.csv


title: PC3 KO cells n=2

city: Sheffield

first: Jiepei

last: He

state: Yorkshire

address: Medical School, Beech Hill Road

department: Oncology and metabolism

institute: University of Sheffield

data_processing:
Bioinformatics analysis was performed on the online platform Galaxy Europe (http://usegalaxy.eu/storage)
Online tools trimmomatic and FastQC, were used to ensure the quality of RNA-seq data
Hisat2 was used to align all reads
Feature counts was used to calculated genes read
Limma was used to compare the differentiation of genes expression
Assembly: hg38
Supplementary files format and content: limma_data_PC3WT_P3KO_Report,gz format
Supplementary files format and content: Galaxy(limma-voom_PC3KO-PC3WT).xlsx format

source_name: Cancer cell

growth_protocol: PC3 prostate cancer cells were maintained in DMEM supplemented with 10% fetal bovine serum (FBS) and antibiotics in humidified atomsphere with 5% CO2 at 37℃

extract_protocol:
RNA was harversted using the ReliaPrep RNA Cell Miniprep System (Promega)
The RNA library construction were performed by company Source BioScience (Nottingham, UK).RNA sequencing package indudes entry QC,polyAselection and stranded cDNA library preparation using IDTkits and reagents with 150bp PE sequencing to 30 millionread pairs per sample (9 Gb per sample)

treatment_protocol: CRISPR/Cas9 knockout P2X4R in prostate cancer cell PC3 cell line,comparation of the gene expression with PC3 wild type

value: Cancer cell

value: prostate cancer cells

value: CRISPR/Cas9 knockout


title: Old CR 28 months old_13


title: Mutant CPXM2-treated 2

description: Library name: Sample8

source_name: pancreas

growth_protocol: 90% DMEM medium supplemented with 10% FBS and 50 IU of penicillin/streptomycin

value: pancreas

value: PaTu 8988t

value: pancreatic tumor cell

value: CPXM2R213A OE ECM


title: DOG-D4-1_S40

description:
Library name: DOG-D4-1_S40
normcounts_dog.csv
qc_hash_dog.csv

value: hepatocytes


title: Mutant CPXM2-treated 3

description: Library name: Sample9


title: Old CR 28 months old_15

value: gastrocnemius muscle

value: 28 months

value: Caloric Restriction


title: PC3 KO cells n=1


title: Mutant CPXM2-treated 1

description: Library name: Sample7


title: DOG-D14-3_S48

description:
Library name: DOG-D14-3_S48
normcounts_dog.csv
qc_hash_dog.csv


title: PC3 WT cells n=3

value: WT


title: Old 28 months old_11

value: Ad Libitum


title: PC3 WT cells n=2


title: DOG-D14-2_S47

description:
Library name: DOG-D14-2_S47
normcounts_dog.csv
qc_hash_dog.csv


title: Wildtype CPXM2-treated 2

description: Library name: Sample5

value: CPXM2WT OE ECM


title: Wildtype CPXM2-treated 3

description: Library name: Sample6


title: Old 28 months old_12


title: Old 28 months old_9


title: PC3 WT cells n=1


title: Old 28 months old_10


title: DOG-D14-1_S46

description:
Library name: DOG-D14-1_S46
normcounts_dog.csv
qc_hash_dog.csv


title: Bulk_Hela_StrippedDNA

city: Santa Barbara

first: Robert

middle: Anthony

last: Piscopio

state: CA

address: 2002 BioEngineering Bldg, CNTR for Bioengineering

institute: University of California, Santa Barbara

description: Method is detailed in Piscopio et al.

library_source: genomic

data_processing:
Supplementary files format and content: text file where each row is a mapped read, and column 1 indicates cell number, column 2 contains chromosome number, column 3 contains genomic location, and column 4 indicates sense and antisense strands of DNA.
scSTAMP-seq data processing and gene expression analysis. Sequencing FASTQ data files were processed using custom Perl scripts on a Linux high performance computing (HPC) cluster at the University of California, Santa Barbara. Sequencing data was post-processed and analyzed using Seurat (version 4.3.0) in R (version 4.2.2) and RStudio (version 2022.12.0+353). Representative figures were generated in RStudio and compiled in Adobe Illustrator (version 26.5).  scSTAMP-seq barcode processing and analysis. A custom Perl script was written to evaluate the spatial barcode portion of the sequencing library. For PHO spatial barcode sequences, read 1 was used to determine cell barcode/UMI. Read 2 was used to distinguish ‘inner’ and ‘outer’ sequence reads by identifying unique ‘inner’ and ‘outer’ primer sequences as well as the downstream spatial barcode/UMI sequence. MPO barcode counts were determined similarly, but an additional multiplex identifier sequence from read 2 was used to distinguish multiplex from spatial barcodes. Unique counts for each molecule were defined as reads containing a unique combination of the UMI on read 1 and on read 2.  Plate-based scSTAMP-seq data processing. For plate-based sequencing, read 2 of the paired-end read was aligned in the sense direction using BWA (version 0.7.15-r1140) to the RefSeq gene model based on the human genome release hg19, incorporating a collection of 92 ERCC spike-in molecules. In cases where a read aligned to multiple locations, it was evenly distributed across those locations. Gene isoforms were consolidated into a single gene count, and the unique molecular identifiers (UMIs) were used to remove duplicate reads and generate transcript counts at the single-molecule level for each gene within individual cells. Genes that were not detected in at least one cell were excluded from subsequent analyses.  Droplet-based scSTAMP-seq data processing. For droplet-based sequencing, read 2 was trimmed using the default settings of TrimGalore. After trimming, STARsolo (STAR aligner version 2.7.8a) was used to map the reads to hg19 using the gene annotation file from Ensembl. Transcripts with the same UMI were deduplicated and genes that were not detected in at least one cell were removed from any downstream analysis.  scSTAMP-seq gene expression analysis. The standard analysis pipeline in Seurat (version 4.3.0) was used for single-cell RNA expression normalization and analysis36. For plate-based sequencing experiments, cells containing more than 1000 genes and more than 1000 unique transcripts, as well as less than 20% ERCC spike-ins, were used for downstream analysis. For the droplet-based sequencing experiment, cells containing more than 200 genes and more than 200 unique transcripts, as well as less than 5% mitochondrial counts, were used for downstream analysis. The default NormalizeData function was used to log normalize the data. Principal components were obtained from the 2,000 most variable genes and the elbow method was used to determine the optimal number of principal components used in clustering. UMAP based clustering was performed by running the following functions, FindNeighbors, FindClusters, and RunUMAP. After clustering, cell types were assigned to groups using known expression markers, spatial barcode counts, and multiplex barcode counts, where stated.
scSTAMP-MAT-seq genome-wide DNA accessibility and 5mC analysis. The scSTAMP-MAT-seq analysis pipeline for DNA accessibility and 5mC was performed as described in the scMAT-seq methods by Chialastri et al.31. DNase I hypersensitivity sites for Hela cells were downloaded from the UCSC table browser and sites were grouped based on detection scores35. To compare across datasets, the data was normalized to counts per million and a 75 base pair moving average was plotted for region of interest. When comparing differing genomic regions within a sample, each region was further normalized by methylated cytosines that were detected when MspJI-seq was performed on bulk Hela gDNA that had been GpC methylated after stripping off chromatin. Bulk Hela gDNA isolation, chromatin stripping, and processing was performed as described previously by Chialastri et al.31. For clustering, DNA accessibility and 5mC were quantified within 5 kb bins and then converted to binary scores. Pseudobulk profiles were generated using the assigned cell type from the transcriptome. For comparison between Hela and U2OS cells, the top 2% most variable bins between groups were retained. After removal of bins with low variance, principal component analysis was used to assign clusters. Cluster identification was performed through comparison to transcriptome derived cell types, where high similarity was observed between cluster calling for all 3 measurements. A threshold minimum of 15,000 was applied to the data shown in Supplementary Figure 14a,b.
Assembly: hg19
Library strategy: scSTAMP-MAT-seq

molecule: genomic DNA

source_name: Hela

growth_protocol: Cell culture. Hela and U2OS cells were cultured and maintained on tissue cultured treated plastic flasks (Thermo Scientific, 156367) or 6-well plates (Corning, 3516) in DMEM with high glucose, GlutaMAX and sodium pyruvate (Gibco, 10569044), supplemented with 10% fetal bovine serum (FBS) (Gibco, 10569044) and 1% Pen-Strep (Gibco, 15140122). All mammalian cell cultures were maintained in incubators at 37°C and 5% CO2. Cells were routinely passaged 1:6 after 70-80% confluence was reached using 0.25% trypsin-EDTA (Gibco, 25200056). For microscopy imaging, cells were seeded onto glass 96 well glass bottom plates (Cellvis, P96-1.5H-N) or 35 mm dishes with glass coverslips (MATTEK, P35G-1.5-20-C) coated with fibronectin (Advanced Biomatrix, 5050). For the combinatorial barcoding experiment, cells were seeded on a custom protein patterned 96-well plate (CYTOO, 20-950-00) treated with fibronectin.

extract_protocol:
For FACS sorting, a single-cell suspension was made by dissociating cells using 0.25% trypsin-EDTA and inactivated using DMEM containing serum. Afterwards, cells were washed and resuspended in 1 mL of cold 1x DPBS-/- (without calcium or magnesium) (Gibco, 14040133) before being passed through a 35 µm nylon mesh cell strainer into a 5 mL round bottom polystyrene tube (Corning, 352235) that was then kept on ice. For plate-based single-cell capture, cell suspensions were FACS sorted for single cells into 384-well plates (Bio-Rad, HSP3801). For droplet-based single-cell capture, cell suspensions were processed for single cells by a 10X Genomics Chromium Controller following the workflow of the 10X Genomics Chromium Next GEM Single Cell 3’ LT Kit v3.1 (10X Genomics, 1000325).
scSTAMP-seq plate-based library preparation. 4 µL of Vapor-Lock (QIAGEN, 981611) was dispensed into each well of a 384-well plate using a 12-channel pipette. All downstream dispensing into 384-well plates was performed using a Nanodrop II liquid handling robot (BioNex Solutions). To each well, 100 nL of uniquely barcoded 7.5 ng/µL reverse transcription primers containing 6-nucleotide UMI was added. The reverse transcription primers used here have been described by Grün et al.33. Next, 100 nL of lysis buffer (0.175% IGEPAL CA-630, 1.75 mM dNTPs, 1:1,250,000 ERCC RNA spike-in mix (Ambion, 4456740), 0.19 U RNase inhibitor (Clontech, 2313A)) was added to each well. Single cells were sorted into individual wells of a 384-well plate using FACS. After sorting, plates were heated to 65°C for 3 minutes and returned to ice. Reverse transcription was conducted next by adding 150 nL of reverse transcription mix (0.7 U RNase OUT (Invitrogen, 10777-019), 2.34x first strand buffer (Invitrogen, 10777-019), 23.34 mM DTT (Invitrogen, 10777-019), 3.5 U Superscript II (Invitrogen, 18064-071)) to each well and the plates were incubated at 42°C for 1 hour and 15 minutes, 4°C for 5 minutes, 70°C for 10 minutes, and then held at 4°C. Next, second strand synthesis was carried out by adding 1.75 µL of second strand synthesis mix (1.74x second strand buffer (Invitrogen, 10812-014), 0.35 mM dNTP (Invitrogen, 10812-014), 0.14 U E. coli DNA Ligase (Invitrogen, 18052-019), 0.56 U E. coli DNA Polymerase I (Invitrogen, 18010-025), 0.03 U RNase H (Invitrogen 18021-071)) to each well and the plates were incubated at 16°C for 2 hours and then held at 4°C. 1 µL of nuclease-free water was added into each well to minimize losses during downstream pooling. The PHO, MPO, and endogenous mRNA molecules in a well now all contain the same RNA capture barcode, indicating a unique cell, which allows for pooling. Reaction wells receiving different barcodes were pooled using a multichannel pipette, and the oil phase was discarded. The small molecule spatial barcode portion was separated from endogenous transcript cDNA using a 3.2x SPRI bead cleanup with 1.8x 100% isopropanol in 2 steps. First, 0.7x AMPure XP beads (Beckman Coulter, A63881) were added and incubated at room temp for 15 minutes. At this step, it is critical to not dispose of the supernatant, as the spatial barcode library portion is found in the supernatant and the endogenous cDNA is bound to SPRI beads. The supernatant containing the spatial barcode fragments was then transferred to a fresh Eppendorf tube. The endogenous cDNA portion was prepared as described previously by Chialastri et al.31. Second, to the spatial barcode sample, an additional 2.5x beads and 1.8x 100% isopropanol (of the original pre-cleanup sample volume) were added and incubated at room temp for 15 minutes. The samples were placed on a magnetic stand for 5 minutes. The supernatant was removed and beads were washed twice with 80% ethanol. The sample was then eluted in 30 µL of nuclease-free water. The sample was then concentrated to 15 µL using an Eppendorf Vacufuge plus. Next, samples were prepared for PCR amplification by mixing 15 µL of the concentrated elute from the previous step, 25 µL of 2x PCR master mix (NEB, M0541S), 2 µL Illumina RPI primer (10 µM), 2 µL each of an ‘inner’ and ‘outer’ PCR primer (10 µM), and nuclease-free water up to 50 µL. This first PCR was performed at 94°C for 10 seconds, 60°C for 30 seconds, and 72°C for 30 seconds, for 10 cycles. A subsequent ExoSAP-IT treatment was then applied to cleanup the PCR product as follows. 2 µL of ExoSAP-IT (Applied Biosystems, 78200.200.UL) was added to 5 µL of sample from the previous PCR (for a total volume of 7 µL) and incubated at 37°C for 15 minutes. Another PCR was performed by mixing 7 µL of the cleaned-up PCR with 22.5 µL 2x PCR master mix, 2 µL Illumina RPI primer, 2 µL Illumina sample index primer, and nuclease-free water up to 45 µL. This second PCR was performed at 94°C for 10 seconds, 60°C for 30 seconds, and 72°C for 30 seconds, for 6-12 cycles.  Another ExoSAP-IT treatment was then applied to the 45 µL reaction by adding 5 µL of ExoSAP-IT to the previous PCR (for 50 µL total) and incubated at 37°C for 15 minutes. Finally, a 1.2x double-sided selection (0.90x left/0.30x right) was used to purify the spatial library, and a final ethanol wash was performed twice. The product was then eluted in 20 µL of nuclease-free water. Library fragment size distribution was determined using a 2100 Bioanalyzer. A schematic depicting each step of this process can be found in Supplementary Figure 1.  scSTAMP-seq droplet-based library preparation. Sequencing libraries were prepared using a custom protocol based on the 10X Genomics Chromium Next GEM Single Cell 3’ LT Kit v3.1 (10X Genomics, 1000325) adapted from McGinnis et al.29. The 10X workflow was followed until cDNA amplification. We designed custom ‘inner’ and ‘outer’ PHO handle primers that allow scSTAMP-seq to be adapted for the 10X workflow (Supplementary Table 1). The scSTAMP-seq 10X ‘inner’ and ‘outer’ PHO primers were mixed to a working concentration of 2.5 µM. Next, the following cDNA amplification master-mix was made: 50 µL 10X amplification master mix, 15 µL 10X cDNA primer mix, 1 µL scSTAMP-seq 10X primers. Then, cDNA amplification was performed and an SPRI size selection was performed by incubating samples for 30 minutes with 0.6x AMPure XP beads to separate spatial and endogenous cDNA fractions. Next, the small molecule spatial barcode fraction was separated from endogenous transcript cDNA using the same strategy as described in the plate-based library preparation method above. The endogenous cDNA portion was then processed to generate Illumina libraries according to the 10X workflow and the spatial portion was processed to generate Illumina libraries as described in the plate-based library preparation workflow. A schematic depicting each step of this process can be found in Supplementary Figure 10.  scSTAMP-MAT-seq multiomic library preparation. The DNA accessibility and 5mC sequencing pipeline was performed as described previously in the scMAT-seq methods by Chialastri et al.31 up to the first SPRI bead clean up. At this step, the small molecule spatial barcode portion was separated from endogenous molecules using the same strategy as described in the plate-based library preparation workflow. The transcriptomic and epigenomic libraries were processed to generate Illumina libraries as described in scMAT-seq by Chialastri et al.31 and the spatial portion was processed to generate Illumina libraries as described in the plate-based library preparation workflow above. A schematic depicting each step of this process can be found in Supplementary Fig. 12.  Library sequencing was performed by Novogene on an Illumina HiSeq 4000 platform or at the Biological Nanostructures Laboratory (BNL) in the California NanoSystems Institute (CNSI) at the University of California Santa Barbara (UCSB) on an Illumina NextSeq 500 platform.

treatment_protocol: PHO labeling of cell membranes. First, cell culture media was removed, and cells were washed three times with cold DPBS+/+ (with calcium and magnesium) (Gibco, 14040133). Hybridized PHO/CMO-anchor at 500 nM was then incubated with live-cell samples over ice for 15 minutes. Next, cells were washed again three times with cold DPBS+/+. After washing, CMO co-anchors were incubated with live-cell samples over ice for 15 minutes. Cells were then washed again three times with cold DPBS+/+. This subsequent two-step anchor/co-anchor incubation is optimal for oligo embedding rate and retention time, as described previously by Weber et al.28. Cells were then imaged and illuminated with UV light. After illumination, cells were either processed for single cell sorting, or the labeling procedure was repeated for experiments that utilized iterative rounds of unique spatial barcode labeling. For experiments that incorporated MPOs, both PHOs and MPOs were incubated simultaneously.  Microscopy and photomask illumination. All microscopy was performed using a Nikon Ti-2 inverted confocal microscope equipped with a Yokogawa CSU-W1 SoRa spinning disk and an Andor iXon Life 888 EMCCD camera. Images were captured with a Nikon CFI S Plan Fluor ELWD 20x objective lens and a Nikon CFI Plan Apo Lambda 10x objective lens. For fluorescence confocal imaging, 488 nm, 561 nm, and 640 nm lasers at 100% power were delivered through liquid light guides for FAM, Cy3, and Cy5 fluorophores, respectively. Digital light processing (DLP) patterned light projection was done using a Mightex Polygon400-G digital micromirror device (DMD) coupled with an Excelitas X-Cite 360-380 nm UVA liquid light guide-coupled LED, or a 365 nm 50W LED flood light (Everbeam, B08635F9CX) with an acrylic light diffusion film and custom 3D printed plate stand for consistent and uniform illumination.

value: Hela


title: DOG-D0-2_S38

description:
Library name: DOG-D0-2_S38
normcounts_dog.csv
qc_hash_dog.csv


title: scSTAMP-seq_ErrorCorrection_Plate2_Lib4_Spatial

description: Method is detailed in Piscopio et al.

library_source: other

data_processing:
Supplementary files format and content: text file for PHO barcodes where each row contains counts for inner and outer spatial barcodes.
scSTAMP-seq data processing and gene expression analysis. Sequencing FASTQ data files were processed using custom Perl scripts on a Linux high performance computing (HPC) cluster at the University of California, Santa Barbara. Sequencing data was post-processed and analyzed using Seurat (version 4.3.0) in R (version 4.2.2) and RStudio (version 2022.12.0+353). Representative figures were generated in RStudio and compiled in Adobe Illustrator (version 26.5).  scSTAMP-seq barcode processing and analysis. A custom Perl script was written to evaluate the spatial barcode portion of the sequencing library. For PHO spatial barcode sequences, read 1 was used to determine cell barcode/UMI. Read 2 was used to distinguish ‘inner’ and ‘outer’ sequence reads by identifying unique ‘inner’ and ‘outer’ primer sequences as well as the downstream spatial barcode/UMI sequence. MPO barcode counts were determined similarly, but an additional multiplex identifier sequence from read 2 was used to distinguish multiplex from spatial barcodes. Unique counts for each molecule were defined as reads containing a unique combination of the UMI on read 1 and on read 2.  Plate-based scSTAMP-seq data processing. For plate-based sequencing, read 2 of the paired-end read was aligned in the sense direction using BWA (version 0.7.15-r1140) to the RefSeq gene model based on the human genome release hg19, incorporating a collection of 92 ERCC spike-in molecules. In cases where a read aligned to multiple locations, it was evenly distributed across those locations. Gene isoforms were consolidated into a single gene count, and the unique molecular identifiers (UMIs) were used to remove duplicate reads and generate transcript counts at the single-molecule level for each gene within individual cells. Genes that were not detected in at least one cell were excluded from subsequent analyses.  Droplet-based scSTAMP-seq data processing. For droplet-based sequencing, read 2 was trimmed using the default settings of TrimGalore. After trimming, STARsolo (STAR aligner version 2.7.8a) was used to map the reads to hg19 using the gene annotation file from Ensembl. Transcripts with the same UMI were deduplicated and genes that were not detected in at least one cell were removed from any downstream analysis.  scSTAMP-seq gene expression analysis. The standard analysis pipeline in Seurat (version 4.3.0) was used for single-cell RNA expression normalization and analysis36. For plate-based sequencing experiments, cells containing more than 1000 genes and more than 1000 unique transcripts, as well as less than 20% ERCC spike-ins, were used for downstream analysis. For the droplet-based sequencing experiment, cells containing more than 200 genes and more than 200 unique transcripts, as well as less than 5% mitochondrial counts, were used for downstream analysis. The default NormalizeData function was used to log normalize the data. Principal components were obtained from the 2,000 most variable genes and the elbow method was used to determine the optimal number of principal components used in clustering. UMAP based clustering was performed by running the following functions, FindNeighbors, FindClusters, and RunUMAP. After clustering, cell types were assigned to groups using known expression markers, spatial barcode counts, and multiplex barcode counts, where stated.
scSTAMP-MAT-seq genome-wide DNA accessibility and 5mC analysis. The scSTAMP-MAT-seq analysis pipeline for DNA accessibility and 5mC was performed as described in the scMAT-seq methods by Chialastri et al.31. DNase I hypersensitivity sites for Hela cells were downloaded from the UCSC table browser and sites were grouped based on detection scores35. To compare across datasets, the data was normalized to counts per million and a 75 base pair moving average was plotted for region of interest. When comparing differing genomic regions within a sample, each region was further normalized by methylated cytosines that were detected when MspJI-seq was performed on bulk Hela gDNA that had been GpC methylated after stripping off chromatin. Bulk Hela gDNA isolation, chromatin stripping, and processing was performed as described previously by Chialastri et al.31. For clustering, DNA accessibility and 5mC were quantified within 5 kb bins and then converted to binary scores. Pseudobulk profiles were generated using the assigned cell type from the transcriptome. For comparison between Hela and U2OS cells, the top 2% most variable bins between groups were retained. After removal of bins with low variance, principal component analysis was used to assign clusters. Cluster identification was performed through comparison to transcriptome derived cell types, where high similarity was observed between cluster calling for all 3 measurements. A threshold minimum of 15,000 was applied to the data shown in Supplementary Figure 14a,b.
Assembly: hg19
Library strategy: scSTAMP-seq

molecule: other

value: Hela cell samples were labeled with PHO1, PHO2, PHO4, PHO5, PHO7, and PHO8. Hela cell samples then received 0, 1.5, 4.5, 13.5, 18, or 25 seconds of UV DLP light illumination.


title: Control 3

description: Library name: Sample3

value: Ctrl ECM


title: Control 2

description: Library name: Sample2


title: Old 28 months old_8


title: Old 28 months old_7


title: DOG-D0-1_S37

description:
Library name: DOG-D0-1_S37
normcounts_dog.csv
qc_hash_dog.csv


title: Control 1

description: Library name: Sample1


title: Young 6 months old_6

value: 6 months


title: scSTAMP-seq_ErrorCorrection_Plate2_Lib3_Spatial

description: Method is detailed in Piscopio et al.


title: DOG-CPZ-3_S90

description:
Library name: DOG-CPZ-3_S90
normcounts_dog.csv
qc_hash_dog.csv

value: chlorpromazine


title: scSTAMP-seq_ErrorCorrection_Plate2_Lib2_Spatial

description: Method is detailed in Piscopio et al.


title: PDAC sample4

description: Library name: Sample 6

source_name: tumor tissue of pancreas

value: tumor tissue of pancreas

value: pancreatic cell


title: DOG-CPZ-2_S89

description:
Library name: DOG-CPZ-2_S89
normcounts_dog.csv
qc_hash_dog.csv


title: Young 6 months old_4


title: DOG-D0-3_S39

description:
Library name: DOG-D0-3_S39
normcounts_dog.csv
qc_hash_dog.csv


title: DOG-CPZ-1_S88

description:
Library name: DOG-CPZ-1_S88
normcounts_dog.csv
qc_hash_dog.csv


title: PDAC sample3

description: Library name: Sample 5


title: Wildtype CPXM2-treated 1

description: Library name: Sample4

value: pancreas

value: PaTu 8988t


title: scSTAMP-seq_ErrorCorrection_Plate1_Lib4_Spatial

description: Method is detailed in Piscopio et al.


title: PDAC sample1

description: Library name: Sample 3


title: Young 6 months old_2


title: DOG-CON-3_S87

description:
Library name: DOG-CON-3_S87
normcounts_dog.csv
qc_hash_dog.csv

value: DMSO control


title: Young 6 months old_3


title: scSTAMP-seq_ErrorCorrection_Plate1_Lib3_Spatial

description: Method is detailed in Piscopio et al.


title: Paratumor 2

description: Library name: Sample 2

source_name: paratumor tissue of pancreas

value: paratumor tissue of pancreas


title: scSTAMP-seq_ErrorCorrection_Plate1_Lib2_Spatial

description: Method is detailed in Piscopio et al.


title: DOG-CON-2_S86

description:
Library name: DOG-CON-2_S86
normcounts_dog.csv
qc_hash_dog.csv

value: hepatocytes


title: scSTAMP-seq_ErrorCorrection_Plate1_Lib1_Spatial

description: Method is detailed in Piscopio et al.


title: Paratumor 1

description: Library name: Sample 1


title: DOG-CON-1_S85

description:
Library name: DOG-CON-1_S85
normcounts_dog.csv
qc_hash_dog.csv


title: Young 6 months _1

value: gastrocnemius muscle


title: DOG-BOS-3_S93

description:
Library name: DOG-BOS-3_S93
normcounts_dog.csv
qc_hash_dog.csv

value: bosentan


title: scSTAMP-seq_ProofOfConcept_Plate2_Lib2_Spatial

description: Method is detailed in Piscopio et al.

value: Hela cells were labeled with PHO1. Hela cell samples then received 0, 1, or 10 minutes of UV flood light illumination.


title: DOG-BOS-2_S92

description:
Library name: DOG-BOS-2_S92
normcounts_dog.csv
qc_hash_dog.csv


title: Young 6 months old_5

value: Ad Libitum


title: perihematomal tissues, ICH mouse model, 24h, rep1

city: Wuhan City

first: Zhuojin

last: Wu

address: 1095 Jiefang Avenue

institute: Tongji Hospital affiliated to Tongji Medical College of Huazhong University of Science & Technology

description: 10x genomics

library_source: transcriptomic single cell

data_processing:
Cell Ranger (v5.0.1)
Assembly: mm10
Supplementary files format and content: Tab-separated values files and matrix files

molecule: nuclear RNA

source_name: perihematomal tissues

extract_protocol:
Single-nucleus isolation was performed using the Chromium Nuclei Isolation Kit 16 rxns (10x Genomics) according to the manufacturer's instructions. Then, 9 μl aliquot of the nuclear suspension was mixed with 1 μl of trypan blue, and the concentration of nuclei was determined under a microscope using a hemocytometer, with the count averaged over three replicates. The concentration of the nuclei suspension was adjusted to approximately 1,000 nuclei/μl by adding Wash and Resuspension Buffer.
library was performed accordong to the manufacter's instructions (Chromium Next GEM Single Cell 3ʹ Reagent Kits v.3.1, 10x Genomics).

value: perihematomal tissues


title: perihematomal tissues, ICH mouse model, 72h, rep1

description: 10x genomics


title: scSTAMP-seq_ErrorCorrection_Plate2_Lib1_Spatial

description: Method is detailed in Piscopio et al.

value: Hela


title: 5-day-old chicks, H9N2 infection, rep 3

city: Fuzhou

first: Song

last: Wang

state: Fujian

address: Cangshan District

institute: Fujian Agriculture and Forestry University

description:
Library name: B3.R1
B3

data_processing:
Residual rRNA sequences were removed using sortmerna, and clean reads were obtained after quality checking them by fastqc v0.11.5.
Sequence comparison of CleanReads with the specified reference genome is performed using hisat2.2.1.0 to obtain position information on the reference genome or gene, as well as sequence feature information specific to the sequenced sample.
The htseq-count was used to obtain the number of reads aligned to protein-coding genes in each sample, and cufflinks were used to calculate the FPKM value of protein-coding gene expression.
Gene Ontology (GO) enrichment analysis was performed on the differentially expressed genes, with all the protein-coding genes as the background list and the list of differentially protein-coding genes as the candidate list filtered out from the background list, and the p-value representing whether the GO functional set was significantly enriched in the list of differentially protein-coding genes was calculated using the hypergeometric distribution test, and then the p-value was subjected to Benjamini and The p-value was then subjected to Benjamini and Hochberg's multiple testing, and genes with P-value <0.05  were categorized as differentially expressed genes.
KEGG is the main public database on Pathways. Pathway analyses of differentially protein-coding genes were performed using the KEGG database (in conjunction with the KEGG annotation results), and the significance of differential gene enrichment in each Pathway entry was calculated using the hypergeometric distribution test to assess the function of lncRNA-adjacent genes. enriched GO functions of gene set. KEGG is a database resource for understanding high-level functions and utilities of the biological system, such as the cell, the organism and the ecosystem, from molecularlevel information, especially large-scale molecular datasets generated by genome sequencing and other high-through put experimental technologies (http://www.genome.jp/kegg/). We used KOBAS to test the statistical enrichment of differential expression genes in KEGG pathways.
Assembly: GRCg6a
Supplementary files format and content: tab-delimited text files include genes readcount and transcripts readcount for each Sample.

organism: Gallus gallus

source_name: lung

extract_protocol:
RNA was harvested using Trizol reagent(Invitrogen, CA, USA).
Total RNA was extracted using the Trizol reagent. RNA purity and quantification were evaluated using the NanoDrop 2000 spectrophotometer (Thermo Scientific, USA). RNA integrity was assessed using the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA). Ribo-off rRNA Depletion Kit（vazyme, Nanjing, China）was used to remove ribosomal RNA, then the libraries were constructed using VAHTS Universal V6 RNA-seq Library Prep Kit according to the manufacturer’s instructions.

treatment_protocol: The 5-day-old chicks were infected with H9N2 virus for 48 h.

value: lung

value: H9N2 virus infection


title: 5-day-old chicks, H9N2 infection, rep 2

description:
Library name: B2.R1
B2


title: scSTAMP-seq_ProofOfConcept_Plate2_Lib1_Spatial

description: Method is detailed in Piscopio et al.


title: PDAC sample2

description: Library name: Sample 4


title: DOG-BOS-1_S91

description:
Library name: DOG-BOS-1_S91
normcounts_dog.csv
qc_hash_dog.csv


title: perihematomal tissues, ICH mouse model, 6h, rep1

description: 10x genomics


title: 5-day-old chicks, H9N2 infection, rep 1

description:
Library name: B1.R1
B1


title: scSTAMP-seq_ProofOfConcept_Plate2_Lib1_Trancriptome

description: Method is detailed in Piscopio et al.

data_processing:
Supplementary files format and content: text file where column 1 contains gene names and columns 2-97 contain gene counts for 96 single cells.
scSTAMP-seq data processing and gene expression analysis. Sequencing FASTQ data files were processed using custom Perl scripts on a Linux high performance computing (HPC) cluster at the University of California, Santa Barbara. Sequencing data was post-processed and analyzed using Seurat (version 4.3.0) in R (version 4.2.2) and RStudio (version 2022.12.0+353). Representative figures were generated in RStudio and compiled in Adobe Illustrator (version 26.5).  scSTAMP-seq barcode processing and analysis. A custom Perl script was written to evaluate the spatial barcode portion of the sequencing library. For PHO spatial barcode sequences, read 1 was used to determine cell barcode/UMI. Read 2 was used to distinguish ‘inner’ and ‘outer’ sequence reads by identifying unique ‘inner’ and ‘outer’ primer sequences as well as the downstream spatial barcode/UMI sequence. MPO barcode counts were determined similarly, but an additional multiplex identifier sequence from read 2 was used to distinguish multiplex from spatial barcodes. Unique counts for each molecule were defined as reads containing a unique combination of the UMI on read 1 and on read 2.  Plate-based scSTAMP-seq data processing. For plate-based sequencing, read 2 of the paired-end read was aligned in the sense direction using BWA (version 0.7.15-r1140) to the RefSeq gene model based on the human genome release hg19, incorporating a collection of 92 ERCC spike-in molecules. In cases where a read aligned to multiple locations, it was evenly distributed across those locations. Gene isoforms were consolidated into a single gene count, and the unique molecular identifiers (UMIs) were used to remove duplicate reads and generate transcript counts at the single-molecule level for each gene within individual cells. Genes that were not detected in at least one cell were excluded from subsequent analyses.  Droplet-based scSTAMP-seq data processing. For droplet-based sequencing, read 2 was trimmed using the default settings of TrimGalore. After trimming, STARsolo (STAR aligner version 2.7.8a) was used to map the reads to hg19 using the gene annotation file from Ensembl. Transcripts with the same UMI were deduplicated and genes that were not detected in at least one cell were removed from any downstream analysis.  scSTAMP-seq gene expression analysis. The standard analysis pipeline in Seurat (version 4.3.0) was used for single-cell RNA expression normalization and analysis36. For plate-based sequencing experiments, cells containing more than 1000 genes and more than 1000 unique transcripts, as well as less than 20% ERCC spike-ins, were used for downstream analysis. For the droplet-based sequencing experiment, cells containing more than 200 genes and more than 200 unique transcripts, as well as less than 5% mitochondrial counts, were used for downstream analysis. The default NormalizeData function was used to log normalize the data. Principal components were obtained from the 2,000 most variable genes and the elbow method was used to determine the optimal number of principal components used in clustering. UMAP based clustering was performed by running the following functions, FindNeighbors, FindClusters, and RunUMAP. After clustering, cell types were assigned to groups using known expression markers, spatial barcode counts, and multiplex barcode counts, where stated.
scSTAMP-MAT-seq genome-wide DNA accessibility and 5mC analysis. The scSTAMP-MAT-seq analysis pipeline for DNA accessibility and 5mC was performed as described in the scMAT-seq methods by Chialastri et al.31. DNase I hypersensitivity sites for Hela cells were downloaded from the UCSC table browser and sites were grouped based on detection scores35. To compare across datasets, the data was normalized to counts per million and a 75 base pair moving average was plotted for region of interest. When comparing differing genomic regions within a sample, each region was further normalized by methylated cytosines that were detected when MspJI-seq was performed on bulk Hela gDNA that had been GpC methylated after stripping off chromatin. Bulk Hela gDNA isolation, chromatin stripping, and processing was performed as described previously by Chialastri et al.31. For clustering, DNA accessibility and 5mC were quantified within 5 kb bins and then converted to binary scores. Pseudobulk profiles were generated using the assigned cell type from the transcriptome. For comparison between Hela and U2OS cells, the top 2% most variable bins between groups were retained. After removal of bins with low variance, principal component analysis was used to assign clusters. Cluster identification was performed through comparison to transcriptome derived cell types, where high similarity was observed between cluster calling for all 3 measurements. A threshold minimum of 15,000 was applied to the data shown in Supplementary Figure 14a,b.
Assembly: hg19
Library strategy: scSTAMP-seq


title: perihematomal tissues, ICH mouse model, sham, rep1

description: 10x genomics


title: UM-UC-3 cells, shCDH11#2, Xenograft, 1-5

city: Toon

first: Osuke

last: Arai

state: Japan

address: Ehime

department: Urology

country: Japan

institute: Ehime Univercity

description:
Library name: UM-UC-3 CDH11KD#2 Xenograft rep1-5
Analysis methods: MaterialMethod_template_BRBN10.docx

data_processing:
Fastq files were trimmed using CLC Genomics Workbench (Filgen) or trim galore (xenograft model).
Assembly: hg38
Supplementary files format and content: Comma-separated values file includes raw counts for each Sample

organism:
Homo sapiens
Mus

source_name: cell line xenograft tissue

extract_protocol:
Ctrl and CDH11 KD cell lines were precultured in 12-well plate (2 × 105 cells/well) for 24 hours. Total RNA samples were extracted using RNeasy spin column kits and DNase I. Ctrl and CDH11 KD cell lines were suspended in Matrigel solution (Corning; 1 × 107 cells/ml). The cell suspension (100 μl) was inoculated subcutaneously into 5-week-old nude mice. Tumor tissues were sampled at 4 weeks after transplantation. Tissue pieces sampled from the Xenograft model were lysed using ISOGEN. Total RNA was extracted using RNeasy spin column kits and DNase I.
RNA integrity number was verified using an Agilent 2100 Bioanalyzer.RNA-seq was performed on an Illumina NextSeq 500 with a read configuration of 75 bp for single reads; 3 million reads were generated per sample. In xenograft model, RNA-seq was performed on the NovaSeq6000, with a read configuration of 75 bp for single reads; 3 million reads were generated per sample.

value: cell line xenograft tissue

value: CDH11 knockdown


title: scSTAMP-seq_ProofOfConcept_Plate1_Lib2_Spatial

description: Method is detailed in Piscopio et al.


title: 5-day-old chicks, Mock, rep 3

description:
Library name: A3.R1
A3

value: Mock treatment


title: UM-UC-3 cells, shCDH11#1, Xenograft, 1-5

description:
Library name: UM-UC-3 CDH11KD#1 Xenograft rep1-5
Analysis methods: MaterialMethod_template_BRBN10.docx


title: PBMC_Aged

first: Hailiang

last: Liu

address: NO 1239, siping road

institute: Tongji university

data_processing:
The analysis of raw sequencing data involves the utilization of the Cell Ranger Single Cell Software Suite (version 3.1.0) from the 10x Genomics website for the generation of a gene expression matrix. Subsequent downstream analysis is conducted using the R package Seurat (version 3.1.0). Standardization of the gene expression dataset includes the exclusion of cells with fewer than 200 genes or exceeding 90% of the highest gene count. Furthermore, cells are sorted based on mitochondrial read percentages in descending order, with the top 15% being excluded from the analysis. Gene expression values are normalized based on library size, log-transformed, and only genes with significant variance are selected for analysis. Principal component analysis is used for dimensionality reduction, followed by clustering using graph-based methods in the PCA space. UMAP is used to visualize these clusters in two dimensions. The FindConservedMarkers function within the Seurat software is utilized to identify marker genes for individual clusters based on specified criteria (logfc.threshold > 0.25 and minPct > 0.25). These clusters are then annotated with known cell types sourced from a cell marker database. Additionally, the same Seurat function is employed to detect genes that exhibit differential expression in the samples, following specific criteria (logfc.threshold > 0.25, minPct > 0.25, and Padj ≤ 0.05). Subsequently, pseudotime trajectory analysis is conducted using the R package Monocle2, while iTALK is utilized for the analysis of cell communication networks.
Assembly: mm10
Supplementary files format and content: Tab-separated values files and matrix files

source_name: Peripheral blood mononuclear cells

extract_protocol:
Peripheral blood mononuclear cells (PBMCs) were isolated from C57BL/6 mice by collecting 1-2 ml of peripheral blood in EDTA-coated tubes, diluting it in an equal volume of RPMI 1640 medium, and layering it over lymphocyte separation medium in centrifuge tubes. After centrifugation at 400 g for 25 minutes, the intermediate white layer containing PBMCs was collected, centrifuged at 500 g for 10 minutes, and half of the supernatant was discarded. The cells were subsequently rinsed with 2 ml of RPMI 1640 medium and subjected to centrifugation at 500 g for 5 minutes. The resulting supernatant was removed, and the cell pellet was reconstituted in 500 µL of fetal bovine serum (FBS) (Gibco). Cell viability was assessed by staining with 0.4% trypan blue (Merck Millipore) and examining under a microscope, with only cells demonstrating over 80% viability being utilized for library preparation.
The single-cell libraries were prepared utilizing the Chromium™ Controller and the Chromium™ Single Cell 3ʹ Reagent Kits v3 (10x Genomics, Pleasanton, CA). Single cells, reagents, and gel beads containing 10X Barcodes were enclosed within Gel Bead-In-EMulsions (GEMs), facilitating single-cell lysis and reverse transcription. Subsequent to GEM disruption, cDNA underwent amplification through PCR, with subsequent size selection (200-300 bp). The fragments were subjected to end-repair, A-tailing at the 3' ends, and adapter ligation. The adapter-ligated fragments underwent purification utilizing double-sided Solid Phase Reversible Immobilization (SPRI) beads. Following sample index PCR, a subsequent round of SPRI purification was carried out. The fragment distribution of the prepared library was assessed using an Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA), and its concentration was quantified via real-time quantitative PCR employing TaqMan probes. Subsequent sequencing was conducted on an Illumina platform.

value: Peripheral blood mononuclear cells

value: aged

value: PBS for 1 month


title: scSTAMP-seq_ProofOfConcept_Plate1_Lib1_Trancriptome

description: Method is detailed in Piscopio et al.


title: PBMC_Young

value: young


title: 5-day-old chicks, Mock, rep 1

description:
Library name: A1.R1
A1


title: Eu-MycSTAT6-D419N-N2

status: Public on Dec 03 2025

city: Montréal

first: Madelyn

middle: J

last: Abraham

state: QC

address: 3755 Chemin de la Cote Sainte Catherine, Lab E504

country: Canada

institute: Lady Davis Institute for Medical Research

description: Library name: A13-575

data_processing:
Trimming: raw FASTQ sequences were trimmed and filtered with Trimmomatic v0.32. Parameters: cut adapter and other Illumina-specific sequences; reject low quality bases (quality < 30), short reads (n < 30); remove the first and last three bases if their qualities are below 30; remove the first three bases of each read; perform sliding window trimming, cutting once the average quality of a window of 4 bases falls below 30. Quality control of raw and processed reads was assessed with FASTQC v.0.11.2.
Alignment and read count: Alignment was performed using STAR v.2.3.0e with mm10 as reference genome. Read counting was done using featureCounts v.1.4.4 and the mm10 UCSC gene annotation.
For clustering and PCA, all exonic reads mapped to the same gene (official gene symbol) were used to summarize gene expression, discarding information on alternative transcripts, thus avoiding overrepresentation of certain genomic regions in the unsupervised clustering.
Differential expression analysis was performed at the gene level (exonic reads mapping to each UCSC gene ID were computed).
Assembly: mm10
Supplementary files format and content: csv files include readcounts for each sample

source_name: Tumour

growth_protocol: Eu-Myc murine lymphoma cells were maintained in DMEM/IMDM supplemented with 10% fetal bovine serum (FBS), antibiotics, and BME in humidified atmosphere with 5% CO2 at 37C

extract_protocol:
Tumour cells were isolated from lymph nodes using CD19 microbeads, then bulk RNA was extracted from tumour cells
Libraries were prepared by Genome Quebec, prior to sequencing on an Illumina HiSeq PE 100bp

treatment_protocol: 1x10^6 cells were transplanted into C57BL6 mice via tail vein injection. After 11 days of growth, mice were sacrificed and lymph node tumours were harvested.

value: Tumour

value: Eu-Myc

value: B Cell Lymphoma

tag: time point

value: Day 11 post-transplantation

tag: batch


title: Eu-MycSTAT6-D419N-N4

description: Library name: A13-572


title: Eu-MycSTAT6-D419N-N1

description: Library name: A13-569


title: Eu-MycSTAT6-WT-N3

description: Library name: A10-324


title: Eu-MycSTAT6-D419N-N3

description: Library name: A13-573


title: Eu-MycSTAT6-WT-N2

description: Library name: A16-394


title: 956158_HDM19wk_anti-IL33+anti-IL4Ra_5

city: Tarrytown

first: Kaitlyn

last: Gayvert

address: 777 Old Saw Mill River Rd

department: Molecular Profiling & Data Science

institute: Regeneron Pharmaceuticals

description:
Library name: 956158
3794.8755.956158.S33.W101102

data_processing:
FASTQ files were aligned and quantified using Omicsoft ArrayStudio
Assembly: mm10
Supplementary files format and content: tab-delimited text files include read counts for  each Sample

extract_protocol:
Tissues/Cells were homogenized in TRIzol and chloroform was used for phase separation. The aqueous phase, containing total RNA, was purified using MagMAX™-96 for Microarrays Total RNA Isolation Kit (Ambion by Life Technologies) according to manufacturer’s specifications. Genomic DNA was removed using RNase-Free DNase Set (Qiagen) on the Kingfisher Flex machine (Thermofisher).
The strand-specific RNA-seq libraries were prepared from 500 ng RNA using KAPA stranded mRNA-Seq Kit (KAPA Biosystems). The libraries were amplified by twelve-cycle PCR. Sequencing was performed on Illumina HiSeq®2000 (Illumina) by multiplexed single-read run with 33 cycles.

tag: hdm

value: HDM 19 weeks


title: Eu-MycSTAT6-WT-N1

description: Library name: A16-393


title: 956155_HDM19wk_anti-IL33+anti-IL4Ra_2

description:
Library name: 956155
3794.8755.956155.S33.W101102


title: 956156_HDM19wk_anti-IL33+anti-IL4Ra_3

description:
Library name: 956156
3794.8755.956156.S33.W101102


title: PBMC_Aged+AA

value: AA for 1 month


title: 956123_HDM19wk_anti-IL4Ra+isotype_8

description:
Library name: 956123
3794.8755.956123.S33.W101102


title: 956161_HDM19wk_anti-IL33+anti-IL4Ra_8

description:
Library name: 956161
3794.8755.956161.S33.W101102


title: scSTAMP-seq_ProofOfConcept_Plate1_Lib1_Spatial

description: Method is detailed in Piscopio et al.

value: Hela cells were labeled with PHO1. Hela cell samples then received 0, 1, or 10 minutes of UV flood light illumination.


title: UM-UC-3 cells, shCtrl, Xenograft, 1-5

description:
Library name: UM-UC-3 Ctrl Xenograft rep1-5
Analysis methods: MaterialMethod_template_BRBN10.docx

value: WT


title: 5-day-old chicks, Mock, rep 2

description:
Library name: A2.R1
A2

value: lung


title: 956120_HDM19wk_anti-IL4Ra+isotype_5

description:
Library name: 956120
3794.8755.956120.S33.W101102


title: 956122_HDM19wk_anti-IL4Ra+isotype_7

description:
Library name: 956122
3794.8755.956122.S33.W101102


title: Siliques_abc1k2 mutant_rep2

city: Verona

first: Elisa

last: Gecchele

address: viale del lavoro

country: Italy

institute: DiamanteSB srl

description: Gene expression in pools of siliques of abc1k2 plants sampled at all developmental stages

hyb_protocol: Aliquots of Cy3-labeled cRNA (1.65 µg) was fragmented following the manufacturers instructions. On completion of the fragmentation reaction, 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized on one sub-array of the Arabidopsis Gene Expression Microarray, 4 x 44K (G2519F-021169, Arabidopsis V4) according to manufacturer’s manual for 17 h at 65 °C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then dried immediately by brief centrifugation, according to manufacturer’s manual (One-Color Microarray-Based Gene Expression Analysis-Low Input Quick Amp Labelling-Protocol).

scan_protocol: Slides were scanned immediately after washing on the Agilent Microarray Scanner System (G2565CA) using one color scan setting for 1x44k array slides, according to the instructions of the Agilent technical manual.

data_processing: The scanned images were analyzed with Feature Extraction Software 10.5.1.1 (Agilent) using default parameters (protocol GE1_105_Dec08 and Grid: 021169_D_F_20080807) to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.

organism: Arabidopsis thaliana

source_name: Siliques, abc1k2 mutant plant, replicate 2

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

growth_protocol: Arabidopsis thaliana (L.) plants, variety Columbia-0 (Col-0), were cultivated in vivo in standard soil under growth chamber conditions (16/8 h light/dark photoperiod, 23°C). For in vitro cultivation, seeds were surface sterilised with 70% ethanol for 1 min followed by incubation for 15 min in 10% sodium hypochlorite and 0.03% Triton X-100. Seeds were rinsed four times with sterile water and sown on solid MS medium (Murashige and Skoog, 1962) supplemented with or without (as described in the text where appropriate) 30 g L-1 sucrose. Upon vernalization for 2 days at 4°C, seeds were kept in vitro with a 16/8 h light/dark photoperiod at 23°C and 150 µE m-2 sec-1.

extract_protocol: RNA was extracted following the lithium chloride protocol optimised for sugar-rich samples (Vennapusa et al., 2020) and was quantified using NanoDropTM 1000 (Thermo Scientific, Waltham, MA, USA). The RNA quality was evaluated with an Agilent 2100 Bioanalyzer.

treatment_protocol: Immature siliques were removed from the plants and dipped in liquid nitrogen to promote opening. Valves were removed and seeds were kept in liquid nitrogen for grinding.

tag: developmental stage

value: All

value: Siliques


title: Siliques_wild type_rep1

description: Gene expression in pools of siliques of wild-type plants sampled at all developmental stages

source_name: Siliques, wild-type plant, replicate 1


title: 956129_HDM19wk_anti-IL4Ra+isotype_2

description:
Library name: 956129
3794.8755.956129.S33.W101102


title: Siliques_abc1k2 mutant_rep3

source_name: Siliques, abc1k2 mutant plant, replicate 3


title: 956127_HDM19wk_anti-IL33+isotype_7

description:
Library name: 956127
3794.8755.956127.S33.W101102


title: 956119_HDM19wk_anti-IL4Ra+isotype_4

description:
Library name: 956119
3794.8755.956119.S33.W101102


title: Siliques_abc1k2 mutant_rep1

source_name: Siliques, abc1k2 mutant plant, replicate 1


title: Siliques_wild type_rep2

source_name: Siliques, wild-type plant, replicate 2


title: 956126_HDM19wk_anti-IL33+isotype_6

description:
Library name: 956126
3794.8755.956126.S33.W101102


title: Siliques_wild type_rep3

source_name: Siliques, wild-type plant, replicate 3


title: 956125_HDM19wk_anti-IL33+isotype_5

description:
Library name: 956125
3794.8755.956125.S33.W101102


title: 956153_HDM19wk_anti-IL33+isotype_3

description:
Library name: 956153
3794.8755.956153.S33.W101102


title: 956154_HDM19wk_anti-IL33+isotype_4

description:
Library name: 956154
3794.8755.956154.S33.W101102


title: 956152_HDM19wk_anti-IL33+isotype_2

description:
Library name: 956152
3794.8755.956152.S33.W101102


title: 956124_HDM19wk_anti-IL33+anti-IL4Ra_1

description:
Library name: 956124
3794.8755.956124.S33.W101102


title: 956121_HDM19wk_anti-IL4Ra+isotype_6

description:
Library name: 956121
3794.8755.956121.S33.W101102


title: 956149_HDM19wk_isotype_8

description:
Library name: 956149
3794.8755.956149.S33.W101102

value: isotype


title: 956142_HDM19wk_isotype_1

description:
Library name: 956142
3794.8755.956142.S33.W101102


title: 956150_HDM19wk_isotype_9

description:
Library name: 956150
3794.8755.956150.S33.W101102


title: 956141_HDM19wk_None_9

description:
Library name: 956141
3794.8755.956141.S33.W101102


title: 956147_HDM19wk_isotype_6

description:
Library name: 956147
3794.8755.956147.S33.W101102


title: 956145_HDM19wk_isotype_4

description:
Library name: 956145
3794.8755.956145.S33.W101102


title: 956140_HDM19wk_None_8

description:
Library name: 956140
3794.8755.956140.S33.W101102


title: 956139_HDM19wk_None_7

description:
Library name: 956139
3794.8755.956139.S33.W101102


title: 956143_HDM19wk_isotype_2

description:
Library name: 956143
3794.8755.956143.S33.W101102


title: 956130_HDM19wk_anti-IL4Ra+isotype_3

description:
Library name: 956130
3794.8755.956130.S33.W101102


title: 956138_HDM19wk_None_6

description:
Library name: 956138
3794.8755.956138.S33.W101102


title: 956137_HDM19wk_None_5

description:
Library name: 956137
3794.8755.956137.S33.W101102


title: 956134_HDM19wk_None_2

description:
Library name: 956134
3794.8755.956134.S33.W101102


title: 956132_HDM11wk_None_9

description:
Library name: 956132
3794.8755.956132.S33.W101102

value: HDM11 weeks


title: 956113_HDM11wk_None_2

description:
Library name: 956113
3794.8755.956113.S33.W101102


title: 956112_HDM11wk_None_1

description:
Library name: 956112
3794.8755.956112.S33.W101102


title: 956111_Saline_None_5

description:
Library name: 956111
3794.8755.956111.S33.W101102

value: Saline


title: 956110_Saline_None_4

description:
Library name: 956110
3794.8755.956110.S33.W101102


title: 956108_Saline_None_2

description:
Library name: 956108
3794.8755.956108.S33.W101102


title: 956115_HDM11wk_None_4

description:
Library name: 956115
3794.8755.956115.S33.W101102


title: 956144_HDM19wk_isotype_3

description:
Library name: 956144
3794.8755.956144.S33.W101102


title: 956114_HDM11wk_None_3

description:
Library name: 956114
3794.8755.956114.S33.W101102


title: 7_mut48A

city: Málaga

first: Ana

last: Álvarez Mena

address: IHSM, Avenida Louis Pasteur 49

country: Spain

institute: University of Málaga

description:
matrix_48h
hunter_annot48hours
Muestra21_S21

data_processing:
Bowtie2 was used for mapping and quantification and qualimap for quality control.
Sam2counts to calculate the read count for each gene.
Degenes Hunter for the analysis of differentially expressed genes.
Assembly: GCA_000007825.1
Supplementary files format and content: tab-delimited excel files include Log2FC of the comparison of wild-type and bc1280 mutant strains, with p-value and padj values
Supplementary files format and content: abundance measurements
Supplementary files format and content: Differentially expressed genes

organism: Bacillus cereus

source_name: Bacillus cereus ATCC14579

extract_protocol:
Planktonic cells of B. cereus wild-type and bc1280 mutant strain were grown in 24-well plates at 28ºC without agitation during 24 and 48 hours. For the RNA extraction, the protocol described by Caro-Astorga et al., 2020 was followed.
RNA libraries were prepared for sequencing using standard Illumina protocols.

tag: strain

value: Bacillus cereus ATCC14579

value: bc1280 mutant

value: 48 hours


title: 6_E3

city: Godollo

first: Eva

last: Varallyay

address: 4 Szent-Györgyi Albert Street

department: Institute of Plant Protection

country: Hungary

institute: Hungarian University of Agriculture and Life Sciences

description:
Library name: 182_E3
small RNA

data_processing:
The quality check of the sequenced small RNA libraries was done by FastQC program, embedded in CLC
Trimming of raw reads was performed using the CLC
Removing redundant reads using CLC
Assembly: small RNA reads, no genome has been assembled
Supplementary files format and content: trimmed redundant list, each read with its number of repeats

organism: Malus domestica

source_name: leaf

extract_protocol:
CTAB method (Gambino, 2008: https://doi.org/10.1002/pca.1078 )
modified Illumina Trueseq small RNA kit protocol: https://doi.org/10.1007/978-1-4939-7683-6_9

value: leaf

tag: cultivar

value: Remo

tag: time

tag: grow type

value: production orchard

tag: infection

value: not infected


title: 8_mut48B

description:
matrix_48h
hunter_annot48hours
Muestra22_S22


title: 5_mut24A

description:
matrix_24h
hunter_annot24hours
Muestra19_S19

value: 24 hours


title: 3_AP3

description:
Library name: 179_AP3
small RNA

value: AP phytoplasma


title: 2_AP2

description:
Library name: 178_AP2
small RNA

value: Reglindis


title: 6_mut24B

description:
matrix_24h
hunter_annot24hours
Muestra20_S20


title: 2_WT24B

description:
matrix_24h
hunter_annot24hours
Muestra16_S16

value: Wild-type


title: 4_WT48B

description:
matrix_48h
hunter_annot48hours
Muestra18_S18


title: 956131_HDM11wk_None_8

description:
Library name: 956131
3794.8755.956131.S33.W101102

value: lung


title: 1_AP1

description:
Library name: 177_AP1
small RNA


title: WTA-ST_FKDL220004448-1a_1_SampleTag04-11

city: Guangzhou

first: Binfeng

last: Chen

address: 58 Zhongshan Second Road

institute: Department of Rheumatology, The First Affiliated Hospital, Sun Yat-sen University

description: BD Rhapsody system

data_processing:
The BD Resolve analysis pipeline was used to process sequencing data (fastq files).
Assembly: mm10
Supplementary files format and content: matrix files including raw counts for each sample

source_name: colon

extract_protocol:
The entire process of the Rhapsody experiment for scRNA-seq was conducted following the manufacturer's (BD Biosciences) instruction. Sorted colonic cells were captured. Cell-lysis buffer was added so that poly-adenylated RNA molecules hybridized to the beads. 8 barcoded samples were pooled together after labeling.
The whole transcriptome libraries were prepared following the BD Resolve single-cell whole-transcriptome amplification algorithm. The libraries were then subjected to an indexed paired-end sequencing run on an Illumina HiSeq 4000 system (Illumina, San Diego, CA, USA) with a 20% PhiX spike-in. Sequencing was conducted according to the manufacturer’s instructions (Illumina).

value: colon

value: CD45+ cell

value: immune cell

value: multiplexed sample:wild-type or VISTA-KO

value: dextran sulfate sodium


title: 956133_HDM19wk_None_1

description:
Library name: 956133
3794.8755.956133.S33.W101102

value: HDM 19 weeks


title: 3_WT48A

description:
matrix_48h
hunter_annot48hours
Muestra17_S17

value: Bacillus cereus ATCC14579


title: GD2572 CUT&RUN seq RPMI 8226 CEBPB KO DMSO 2h H3K27ac

city: Morgantown

first: Lei

state: West Virginia

address: 64 Medical Center Drive

department: Department of Microbiology, Immunology and Cell Biology

institute: west virginia university

description: Library name: Sample 12

data_processing:
Reads were aligned to human genome (hg38) with bowtie2 version 2.4.2 (PMID: 22388286),Reads mapped to multiple genomic positions were removed. For reads mapped to the same genomic coordinates, only one copy was kept.
Peaks were called using MACS v3.0 with the significance cut-off q-value <= 0.001
Assembly: hg38
Supplementary files format and content: txt file; columns from left to right: chromosome, start, end, name, int(-10*log10qvalue), reserved column, fold-change at peak summit, -log10pvalue at peak summit, -log10qvalue at peak summit, relative summit position to peak start
Library strategy: CUT&Run

source_name: RPMI 8226

extract_protocol:
Performed using the CUT&RUN Assay Kit from Cell Signaling Technology (#86652) following the manufacturer’s protocol. 100,000 RPMI 8226 cells were used along with acetyl-histone H3 (Lys27) (D5E4) #23349.
Libraries were made using NEB ultra II DNA library protocol

treatment_protocol: 1 × 10⁶ RPMI 8226 cells were plated in 6-well dishes and treated with or without 10 μM dexamethasone for 2h.

value: RPMI 8226

value: CEBPB KO

value: DMSO

tag: replicate


title: GD2571 CUT&RUN seq RPMI 8226 CEBPB KO DMSO 2h H3K27ac

description: Library name: Sample 11


title: 1_WT24A

description:
matrix_24h
hunter_annot24hours
Muestra15_S15


title: EPA cells 6h blue light, rep3

city: Eggenstein-Leopoldshafen

first: Alessandra

last: Boiti

address: Hermann-von-Helmholtz-Platz 1

country: Germany

institute: Karlsruhe Institute of Technology

description:
Library name: Sample 24
column name in de novo analysis files: BLCF_6h_rep3

data_processing:
For standard pipeline: Raw reads quality check; mapping to refecence genome; quantitation by sample; transcript exclusion TPM>5 only; quantitation by timepoint; DEG analysis with p<.001 and |logFC|>=1; functional analysis (GO enrichment analysis 0h vs all timepoints)
For de novo pipeline: Raw reads quality check; trinity assembly with Trinity software; quantitation by sample; transcript exclusion TPM>5 only; annotation against databases (blastx, uniprot, pfam); quantitation by timepoint; DEG analysis with p<.001 and |logFC|>=1; functional analysis (GO enrichment analysis 0h vs all timepoints)
Assembly: GRCz11 for PAC-2 cells, de novo Trinity software assembly for EPA
Supplementary files format and content: Tab-delimited text file contains TPM values for each sample
Supplementary files format and content: BLZF_readcount_genename.xls contains raw counts for each sample
Supplementary files format and content: BLZF_fpkm_genename.xls file contains FPKM values for each sample

organism: Garra andruzzii

source_name: EPA

growth_protocol: Cells are cultured in Leibovitz L-15 medium with 15% FBS (PAC-2) or 20% FBS (EPA), 100U/ml penicillin, 100ug/ml streptomycin, and 50ug/ml gentamicin

extract_protocol:
total RNA is harvested using Trizol reagent according to manufacturer’s instruction.
Messenger RNA was purified from total RNA using poly-T oligo-attached magnetic beads. After fragmentation, the first strand cDNA was synthesized using random hexamer primers, followed by the second strand cDNA synthesis using either dUTP for directional library or dTTP for non-directional library
The library was checked with Qubit and real-time PCR for quantification and bioanalyzer for size distribution detection. Quantified libraries will be pooled and sequenced on Illumina platforms, according to effective library concentration and data amount.
The clustering of the index-coded samples was performed according to the manufacturer’s instructions. After cluster generation, the library preparations were sequenced on an Illumina platform and paired-end reads were generated.

treatment_protocol: Cells are seeded with transparent medium (no phenol red), kept in darkness for 2 days, then exposed to blue light (468nm) for up to 6h and samples are taken before exposure (0h), and after 1h, 3h, and 6h.

value: EPA

value: fibroblasts

value: blue light exposure

tag: type


title: 956109_Saline_None_3

description:
Library name: 956109
3794.8755.956109.S33.W101102

value: Saline


title: EPA cells 6h blue light, rep2

description:
Library name: Sample 23
column name in de novo analysis files: BLCF_6h_rep2


title: GD2569 CUT&RUN seq RPMI 8226 CEBPB KO Dex 2h H3K27ac

description: Library name: Sample 9

value: 10 μM dexamethasone treatment (2 hrs)


title: EPA cells 6h blue light, rep1

description:
Library name: Sample 22
column name in de novo analysis files: BLCF_6h_rep1


title: 956107_Saline_None_1

description:
Library name: 956107
3794.8755.956107.S33.W101102


title: GD2568 CUT&RUN seq RPMI 8226 CEBPB KO Dex 2h H3K27ac

description: Library name: Sample 8


title: EPA cells 3h blue light, rep3

description:
Library name: Sample 21
column name in de novo analysis files: BLCF_3h_rep3


title: 4_E1

description:
Library name: 180_E1
small RNA

value: leaf

value: Reglindis

value: production orchard

value: not infected


title: EPA cells 3h blue light, rep1

description:
Library name: Sample 19
column name in de novo analysis files: BLCF_3h_rep1


title: GD2566 CUT&RUN seq RPMI 8226 SCR DMSO 2h H3K27ac

description: Library name: Sample 6

value: Scramble control


title: GD2567 CUT&RUN seq RPMI 8226 CEBPB KO Dex 2h H3K27ac

description: Library name: Sample 7


title: 5_E2

description:
Library name: 181_E2
small RNA

value: Remo


title: GD2565 CUT&RUN seq RPMI 8226 SCR DMSO 2h H3K27ac

description: Library name: Sample 5


title: EPA cells 1h blue light, rep2

description:
Library name: Sample 17
column name in de novo analysis files: BLCF_1h_rep2


title: GD2563 CUT&RUN seq RPMI 8226 SCR Dex 2h H3K27ac

description: Library name: Sample 3


title: EPA cells 1h blue light, rep1

description:
Library name: Sample 16
column name in de novo analysis files: BLCF_1h_rep1


title: EPA cells 1h blue light, rep3

description:
Library name: Sample 18
column name in de novo analysis files: BLCF_1h_rep3


title: EPA cells 0h blue light, rep3

description:
Library name: Sample 15
column name in de novo analysis files: BLCF_0h_rep3

value: 0h (control)


title: EPA cells 0h blue light, rep2

description:
Library name: Sample 14
column name in de novo analysis files: BLCF_0h_rep2


title: GD2562 CUT&RUN seq RPMI 8226 SCR Dex 2h H3K27ac

description: Library name: Sample 2


title: EPA cells 0h blue light, rep1

description:
Library name: Sample 13
column name in de novo analysis files: BLCF_0h_rep1


title: GD2570 CUT&RUN seq RPMI 8226 CEBPB KO DMSO 2h H3K27ac

description: Library name: Sample 10


title: knockdown1

city: Wuhan

first: pengcheng

last: Li

state: HUbei

address: Tongji Hospital Research Building, Wuxian East Road, Caidian District, Wuhan City, Hubei Province

institute: Tongji medical colleage

description:
Library name: K1
line B

data_processing:
Illumina Casava 1.8 software used for basecalling
Raw data (raw reads) of fastp format were firstly processed through in-house perl scripts..In this step, clean data (clean reads) were obtained by removing reads containing adapter, reads containing ploy-N and low quality reads from raw data. At the same time, Q20, Q30 and GC content the clean data were calculated. All the downstream analyses were based on the clean data with high quality.
Reference genome and gene model annotation files were downloaded from genome website directly. Index of the reference genome was built using Hisat2 v2.0.5 and paired-end clean reads were aligned to the reference genome using Hisat2 v2.0.5.
featureCounts v1.5.0-p3 was used to count the reads numbers mapped to each gene.
Differential expression analysis of two conditions/groups (two biological replicates per condition) was performed using the DESeq2 R package (1.20.0).
Assembly: GRCh38/hg38
Supplementary files format and content: tab-delimited text files include FPKM values and gene count for each Sample

source_name: RKO

extract_protocol:
Total RNA was isolated using TRIzol (Invitrogen) according to the manufacturer’s instructions.
RNA libraries for RNA-seq were prepared using Fast RNA-seq Lib Prep Kit V2 （Cat.No.RK20306）following manufacturer's protocols.

value: RKO

value: KMT5A knockdown


title: GD2561 CUT&RUN seq RPMI 8226 SCR Dex 2h H3K27ac

description: Library name: Sample 1

value: RPMI 8226


title: knockdown3

description:
Library name: K3
line D


title: knockdown2

description:
Library name: K2
line C


title: ChIP_SIX1-3 + DOX

city: Manchester

first: Georges

last: Lacaud

state: England

address: Wilmslow Raod

department: Stem Cell Biology

institute: Cancer Research UK Manchester Institute

description: Library name: 87 + SIX1 cell (3)

data_processing:
Raw fastqc sequencing files were run through nextflow on the CRUK-MI (Manchester institute) server to generate merged salmon gene counts and BigWig files
DESeq2 R package was run to infer differentially expressed genes accross samples ChIPpeakAnno bioconductor package was used in R for gene ontology as well as differential binding analysis to observe lost peaks following SIX1 knockdown.
Assembly: hg38
Supplementary files format and content: Salon merged gene counts were generared by nextflow. All counts concatenated on same tsv file BigWig files generated by nextflow - ChIP

source_name: Blood (Bone marrow)

growth_protocol: THP1 cells with a DOX inducible promotor for SIX1-sh RNA were grown in RPMI media (+/- DOX) in order to harvest 4x10^8 cells in T225 TC flasks for ChIP. KO and NT cells were cultured in RPMI media for 48hrs before sorting and RNA isolation.

extract_protocol:
Direct RNA isolation with RNeasy Mini Kit [Qiagen Ref. 74004] -RNA Following pulldown - DNA was extracted using Phenol:Chloroform:Isoamyl Alcohol (25:24:1, v/v), before being further purified with QIAquick PCR Purification Kit
RNA was sent to BMKGENE for library preperation and sequencing

value: Blood (Bone marrow)

value: monocytic


title: Control3

description:
Library name: C3
line G

value: wildtpye

value: ShNC


title: ChIP_SIX1-4 + DOX

description: Library name: 87 + SIX1 sigma (1)


title: ChIP_SIX1-2 + DOX

description: Library name: 87 + SIX1 cell (2)


title: Control1

description:
Library name: C1
line E


title: EPA cells 3h blue light, rep2

description:
Library name: Sample 20
column name in de novo analysis files: BLCF_3h_rep2

value: EPA

value: fibroblasts

value: blue light exposure


title: ChIP_SIX1-1 + DOX

description: Library name: 87 + SIX1 cell (1)


title: ChIP isotype + DOX

description: Library name: 87 + IgG


title: ChIP_SIX1-3

description: Library name: 87 - SIX1 sigma (1)


title: OV8_Tram, rep2

city: San Antonio

first: Yidong

state: Texas

address: 8403 Floyd Curl Drive, MSC 7784

department: Population Health Sciences

institute: UT Health Science Center at San Antonio

description: Library name: OV8_Tram_2

data_processing:
Illumina bcl2fastq software used for base-calling
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to hg38 whole genome using HISAT2 algorithm
gene count measurements were obtained using FeatureCounts using refseq annotation
Assembly: hg38
Supplementary files format and content: Tab-delimited text file with columns: 1) GeneID, 2) Chr; 3) transcript start position; 4) transcript end position; 5) Strand; 6) Length of transcript; 7) read counts

growth_protocol: OVCAR8 cells were cultured in RPMI medium supplented with 10% FBS.

extract_protocol:
Total RNA was extracted using RNeasy mini kit (Qiagen, CA) and quantified using nanodrop and Agilent bioanalyzer.
Total RNA samples  were used for sequencing library preparation according to Illumina stranded mRNA sample preparation guide. Each library was bar-coded and then pooled for sequencing run with 100bp paired-end sequencing protocol.

treatment_protocol: OVCAR8 cells treated with vehicle, EC359 (150 nM), trametinib (100 nM), or the combination of both agents for 7 hours

value: Human

value: Ovarian cancer

value: Trametinib


title: HEK Keratinocytes 64 3x100 mJ SSR

city: Hamburg

first: Leonardo

last: Elsbroek

address: Beiersdorfstraße 1-9

institute: Beiersdorf AG

description: Library name: 42_HEK64_3x100

data_processing:
For quality control Raw FASTQ reads were processed with fastp (adapter trimming, low-quality filtering, removal of poly-N reads).
Gene-level read counts were quantified using featureCounts v1.5.0-p3.
Assembly: GRCh38 / GENCODE v43.
Supplementary files format and content: Count matrices gene-level read counts csv files include raw RNA counts for each sample.

source_name: Epidermis

growth_protocol: Primary human keratinocytes from AK lesions and photo-exposed healthy facial epidermis (HEK) were cultured in vitro in Stem Cell Basal Keratinocyte expansion media with antibiotics in humidified atmosphere with 5% CO2 at 37C.

extract_protocol:
Total RNA was extracted using the RNeasy Mini Kit (Qiagen, cat. 74104) according to the manufacturer’s instructions. On-column DNase I digestion (Qiagen, cat. 79254) was performed to remove genomic DNA.  RNA integrity was assessed using the Bioanalyzer 2100 system (Agilent Technologies, CA, USA).
Messenger RNA was purified from total RNA using poly-T oligo-attached magnetic beads. After fragmentation, the first strand cDNA was synthesized using random hexamer primers, followed by the second strand cDNA synthesis using dTTP. Library quantification was performed via Qubit and real-time PCR; size distribution assessed with a Bioanalyzer

treatment_protocol: Cells received either a single exposure or repeated daily exposures for 3 consecutive days. The UVB-equivalent dose per exposure was 100 mJ/cm² (within the SSR spectrum). Cells were harvested 6 h after the final exposure for RNA extraction. Controls were processed in parallel. Each condition was profiled by RNA-seq in 7 biological replicates per group.

value: Epidermis

value: Keratinocyte

value: wt

value: 3x100 mJ SSR


title: OV8_Tram, rep1

description: Library name: OV8_Tram_1


title: OV8_EC359, rep3

description: Library name: OV8_EC359_3


title: GD2564 CUT&RUN seq RPMI 8226 SCR DMSO 2h H3K27ac

description: Library name: Sample 4

value: Scramble control

value: DMSO


title: HEK Keratinocytes 64 100 mJ SSR

description: Library name: 41_HEK64_100

value: 100 mJ SSR


title: HEK Keratinocytes Control 64

description: Library name: 40_HEK64_C

value: Control


title: OV8_Tram, rep3

description: Library name: OV8_Tram_3


title: HEK Keratinocytes 298 3x100 mJ SSR

description: Library name: 39_HEK298_3x100


title: OV8_EC359, rep2

description: Library name: OV8_EC359_2


title: HEK Keratinocytes Control 298

description: Library name: 37_HEK298_C


title: OV8_EC359, rep1

description: Library name: OV8_EC359_1


title: OV8_Control, rep3

description: Library name: OV8_Con_3


title: HEK Keratinocytes 298 100 mJ SSR

description: Library name: 38_HEK298_100


title: OV8_Control, rep2

description: Library name: OV8_Con_2


title: HEK Keratinocytes 295 3x100 mJ SSR

description: Library name: 36_HEK295_3x100


title: OV8_Control, rep1

description: Library name: OV8_Con_1


title: HEK Keratinocytes Control 295

description: Library name: 34_HEK295_C


title: Control2

description:
Library name: C2
line F

value: RKO


title: OV8_Combo, rep3

description: Library name: OV8_Combo_3

value: Trametinib + EC359


title: HEK Keratinocytes 295 100 mJ SSR

description: Library name: 35_HEK295_100


title: OV8_Combo, rep1

description: Library name: OV8_Combo_1


title: OV8_Combo, rep2

description: Library name: OV8_Combo_2


title: HEK Keratinocytes 275 3x100 mJ SSR

description: Library name: 30_HEK275_3x100


title: HEK Keratinocytes 291 100 mJ SSR

description: Library name: 32_HEK291_100


title: Patient 38 Baseline Transcriptomics, Male, Progressive Disease

city: Buffalo

first: Spencer

last: Rosario

address: Elm & Carlton Sts

institute: Roswell Park Comprehensive Cancer Center

description: Library name: Patient-38-BLBX

data_processing:
Raw reads that passed thequality filter were demultiplexed and pre-processed using FastQC for sequencing base quality control.
Genome alignment of raw reads to the human reference genome (GRCh38.p13) using STAR
Mapped reads were quantified at the gene level as a raw counts matrix using featureCounts from Subread
Assembly: GRCh38
Supplementary files format and content: Transcripomics_Processed .csv file includes raw counts for each sample

source_name: colorectal tumor

extract_protocol:
The FFPE samples were emulsified in Tissue Lysis Buffer in the presence of Proteinase K on the Covaris E220 sonicator. This was followed by an incubation at 56 °C for a short duration. The RNA-containing supernatant was separated from the DNA-containing tissue by a centrifugation step. RNA was then de-crosslinked, DNAse digested, and purified over an RNA-specific spin column. Sequentially, DNA was released from the DNA-containing tissue by the Covaris E220 sonicator. The resuspension was then put through an additional Proteinase K digestion, following a de-crosslinking step at 80 °C. DNA was then purified over a DNA-specific spin column.
Unique dual-indexed, molecular-barcoded libraries were purified with AMPureXP beads (Beckman Coulter) and validated for appropriate size (225-275bp) on a Tapestation 4200 DNA1000 screentape (Agilent Inc.). 200ng of purified library was then hybridized to the SureSelectXT Human All Exon V6+UTR Capture library (Agilent Inc.). The hybridized regions were then bound to streptavidin magnetic beads and washed to remove any non-specific bound products. Eluted library underwent a second 13 cycle PCR amplification to generate enough material for sequencing. Final libraries were purified, measured by Tapestation 4200 DNA1000 screentape (250-350bp), and quantitated using KAPA qPCR (KAPA Biosystems). Individual libraries were pooled in equimolar 2nM final concentration.

treatment_protocol: Whole transcriptome analysis of FFPE samples was performed using SureSelect XT HS2 RNA Target Enrichment System (Agilent Inc). 200ng of RNA was fragmented and converted to cDNA followed by end repair, duplex UMI adaptor ligation, and 12 cycles of PCR to complete the dual-index adapters.

value: colorectal tumor

tag: Sex

value: Male

tag: disease status

value: Progressive Disease

value: RAS wt

value: Baseline


title: HEK Keratinocytes 291 3x100 mJ SSR

description: Library name: 33_HEK291_3x100

tag: cell line

tag: cell type


title: Patient 21 Baseline Transcriptomics, Male, Stable Disease

description: Library name: Patient-21-BLBX

tag: tissue

value: Stable Disease


title: Patient 16 Baseline Transcriptomics, Female, Stable Disease

description: Library name: Patient-16-BLBX

value: Female

tag: genotype


title: Patient LH-06 Baseline Transcriptomics, Male, Stable Disease

description: Library name: Patient-LH06-BLBX

tag: time point


title: HEK Keratinocytes 275 100 mJ SSR

description: Library name: 29_HEK275_100

value: Epidermis

tag: treatment


title: HEK Keratinocytes Control 275

description: Library name: 28_HEK275_C

value: Keratinocyte


title: Patient 32 Baseline Transcriptomics, Male, Partial Regression

description: Library name: Patient-32-BLBX

value: Partial Regression


title: HEK Keratinocytes 27 100 mJ SSR

description: Library name: 26_HEK27_100

value: wt


title: Patient LH-04 Baseline Transcriptomics, Male, Stable Disease

description: Library name: Patient-LH04-BLBX


title: HEK Keratinocytes Control 291

description: Library name: 31_HEK291_C


title: HEK Keratinocytes Control 27

description: Library name: 25_HEK27_C


title: Patient 39 Post-Treatment Transcriptomics, Male, Stable Disease

description: Library name: Patient-39-POST-TX

value: Post-Treatment


title: Patient LH-02 Baseline Transcriptomics, Male, Stable Disease

description: Library name: Patient-LH02-BLBX


title: HEK Keratinocytes BCS54 3x100 mJ SSR

description: Library name: 24_BCS54_3x100

value: 3x100 mJ SSR


title: Patient 39 Baseline Transcriptomics, Male, Stable Disease

description: Library name: Patient-39-BLBX


title: HEK Keratinocytes BCS54 100 mJ SSR

description: Library name: 23_BCS54_100

value: 100 mJ SSR


title: Patient LH-05 Baseline Transcriptomics, Male, Stable Disease

description: Library name: Patient-LH05-BLBX


title: Patient 37 Post-Treatment Transcriptomics, Female, Progressive Disease

description: Library name: Patient-37-POST-TX


title: HEK Keratinocytes Control BCS54

description: Library name: 22_BCS54_C

value: Control


title: Patient 37 Baseline Transcriptomics, Female, Progressive Disease

description: Library name: Patient-37-BLBX


title: Patient 35 Post-Treatment Transcriptomics, Male, Stable Disease

description: Library name: Patient-35-POST-TX


title: AK Keratinocytes 59 100 mJ SSR

description: Library name: 20_AK59_100

value: AK Keratinocyte


title: AK Keratinocytes Control 59

description: Library name: 19_AK59_C


title: AK Keratinocytes 59 3x100 mJ SSR

description: Library name: 21_AK59_3x100


title: AK Keratinocytes 55 3x100 mJ SSR

description: Library name: 18_AK55_3x100


title: Patient 22 Baseline Transcriptomics, Male, Progressive Disease

description: Library name: Patient-22-BLBX


title: HEK Keratinocytes 27 3x100 mJ SSR

description: Library name: 27_HEK27_3x100


title: Patient 19 Post-Treatment Transcriptomics, Male, Stable Disease

description: Library name: Patient-19-POST-TX


title: AK Keratinocytes Control 55

description: Library name: 16_AK55_C


title: AK Keratinocytes 54 3x100 mJ SSR

description: Library name: 15_AK54_3x100


title: Patient 19 Baseline Transcriptomics, Male, Stable Disease

description: Library name: Patient-19-BLBX


title: AK Keratinocytes 54 100 mJ SSR

description: Library name: 14_AK54_100


title: AK Keratinocytes 49 3x100 mJ SSR

description: Library name: 12_AK49_3x100