title: sample_23

data_processing:
Adapter and polyA trimming with BBDuk (v38.9; options k=13, ktrim=r, forcetrimleft=11, useshortkmers=t, mink=5, qtrim=t, trimq=10, minlength=20)
Alignment and read counting with STAR aligner (v2.7.8a; default settings apart from --outFilterMismatchNoverLmax 0.1)
Library size normalization and vsd transformation in R v4.1.0 with DESeq2 v1.32.0
Weighted gene co-expression network analysis with WGCNA R package (v1.70) (soft threshold power = 10)
Pathway analysis for enrichment of GO biological pathways with clusterProfiler R package (v4.0.0)
Transcriptional deconvolution using the mMCPcounter package for R (v1.0.0)
Genome_build: GRCm38 and MT952602 (concatenated)
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

source_name: homogenized lung

tag: day post infection

tag: experimental reapeat

value: repeat 02


title: Dissociated lung from APOE knock-in mouse with COVID-19, sample 06

status: Public on Aug 07 2022

type: SRA

city: Berlin

first: Benjamin

middle: N

last: Ostendorf

address: Augustenburger Platz 1

country: Germany

institute: Charité University Medicine

library_source: transcriptomic

data_processing:
Adapter and polyA trimming with BBDuk (v38.9; options k=13, ktrim=r, forcetrimleft=11, useshortkmers=t, mink=5, qtrim=t, trimq=10, minlength=20)
Alignment and read counting with STAR aligner (v2.7.8a; default settings apart from --outFilterMismatchNoverLmax 0.1)
Library size normalization and vsd transformation in R v4.1.0 with DESeq2 v1.32.0
Weighted gene co-expression network analysis with WGCNA R package (v1.70) (soft threshold power = 10)
Assembly: GRCm38 and MT952602 (concatenated)
Supplementary files format and content: Tsv-formatted table with raw gene counts for every gene and every sample

molecule: total RNA

organism: Mus musculus

source_name: Lung

growth_protocol: APOE knock-in mice were intranasally infected with SARS-CoV-2 MA10

extract_protocol:
Lungs were resected on day 4 post infection, homogenized in Trizol using a Miltenyi homogenizer (program RNA_01), and RNA was extracted using Zymo's Direct-zol kit including DNAse digestion.
Lexogen 3' FWD kit

tag: Sex

value: female

tag: disease state

tag: agent

value: SARS-CoV-2 MA10

tag: tissue

value: Lung

tag: genotype

tag: timepoint

value: 4 dpi


title: sample_22


title: sample_21


title: sample_20


title: sample_24


title: CZ3, Wildtype, snRNA-seq

city: Baltimore

first: Jonathan

middle: Joseph

last: Augustin

state: MD

address: 733 N. Broadway Rm#332

country: USA

institute: Johns Hopkins Medical Institute

description: 10X Genomics

library_source: transcriptomic single cell

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were made using Kallisto-bustools (https://www.kallistobus.tools)
Assembly: mm10
Supplementary files format and content: comma-separated values files and matrix files

molecule: nuclear RNA

source_name: Dorsal Telencephalon

extract_protocol:
Day E15.5 embryonic brains were collected from both wildtype and Pantr2-/- mice. The dorsal telencephalon was dissected, flash frozen in liquid nitrogen, and stored at –80°C until ready for nuclei isolation. Nuclei were isolated using a modified version of the Salty-EZ Lysis Buffer method outlined on protocols.io (https://www.protocols.io/view/protocol-for-nuclei-isolation-from-fresh-and-froze-kqdg36ybzg25/v5). In brief, tissues were lysed in 300μl of ice cold Salty-EZ10 lysis buffer containing 10mM Tris HCL pH 7.5, 146mM NaCl, 1mM CaCl2, 21mM MgCl2, 0.03% Tween-20, 10% EZ-lysis Buffer (Sigma Aldrich), 1mM DTT and 0.2U/μl Protector RNase Inhibitor. The tissue was gently triturated using a P200 wide-bore pipette tip on ice until the tissue was completely in solution. Once homogenized, 700μl of ice cold Slaty-EZ10 buffer was added and each sample was gently mixed using a wide-bore pipette tip. Next, the solution was spun down at 4°C at 500G for 5 minutes to pellet the nuclei in suspension. The supernatant was aspirated off and 500μl of WRB2 buffer containing 10mM Tris-HCl pH 7.5, 10mM NaCl, 3mM MgCl2, 1% w/v bovine serum albumin (BSA), 1mM DTT and 0.2U/μl Protector RNase Inhibitor. Nuclei were washed three times in 500μl WRB2 buffer before being used as the input material for 10X Genomics Chromium Single Cell V2 kit following standard protocol included with the kit.
Library was performed according to the manufacter’s instructions (single cell 3’ v2 protocol, 10x Genomics). Briefly, nuclei were resuspended in the master mix and loaded together with partitioning oil and gel beads into the chip to generate the gel bead-in-emulsion (GEM). The poly-A RNA from the cell lysate contained in every single GEM was retrotranscripted to cDNA, which contains an Ilumina R1 primer sequence, Unique Molecular Identifier (UMI) and the 10x Barcode. The pooled barcoded cDNA was then cleaned up with Silane DynaBeads, amplified by PCR and the apropiated sized fragments were selected with SPRIselect reagent for subsequent library construction. During the library construction Ilumina R2 primer sequence, paired-end constructs with P5 and P7 sequences and a sample index were added.

value: Dorsal Telencephalon

tag: age

tag: strain

value: Wildtype


title: Dissociated lung from APOE knock-in mouse with COVID-19, sample 07


title: Dissociated lung from APOE knock-in mouse with COVID-19, sample 02


title: Dissociated lung from APOE knock-in mouse with COVID-19, sample 03


title: Dissociated lung from APOE knock-in mouse with COVID-19, sample 10


title: Dissociated lung from APOE knock-in mouse with COVID-19, sample 01


title: Dissociated lung from APOE knock-in mouse with COVID-19, sample 05


title: sample_14


title: sample_15


title: sample_07


title: sample_09


title: sample_08


title: sample_05


title: sample_19


title: sample_18


title: sample_04


title: sample_12


title: sample_17


title: sample_13


title: sample_06


title: sample_16


title: sample_03


title: mammary gland, age 20M, 8 months CYP19A1 transgene induction, letrozole 2 months exposure, biol rep 3

city: Cheonan

first: Keunsoo

last: Kang

address: 119 Dandae-ro

country: South Korea

institute: Dankook University

data_processing:
Sequencing using the Illumina NextSeq 550, SE 75 bp read length; minimum reads ?50M per sample
Sequencing quality was checked using FastQC (version 0.11.9; https://www.bioinformatics.babraham.ac.uk/projects/fastqc/).
Contaminated adaptor and/or row quality portions of sequenced reads were trimmed using Trim Galore (version 0.6.5; https://github.com/FelixKrueger/TrimGalore).
Trimmed reads were aligned to the reference mouse genome (mm10), using STAR (version 2.7.9a) (Dobin et al. Bioinformatics 2013).
Batch effect was normalized using RUVSeq (version 1.26.0) with the RUVg method (Risso et al. Nat Biotech. 2014).
Normalized expression levels were estimated by means of transcripts per million (TPM) using RSEM (version 1.3.1) (Li and Dewey, BMC Bioinformatics 2011)
Assembly: mm10
Supplementary files format and content: tab-delimited text file includes TPM values for each sample

source_name: mammary gland

extract_protocol: Indexed, single-index sequencing libraries prepared from total RNA (1 microgram/sample) using the Illumina TruSeq Stranded Total RNA Library Preparation Human/Mouse/Rat Kit (rRNA depletion and random-primed cDNA synthesis)

treatment_protocol: Treatment protocol: tamoxifen (25 mg/60 day release) or letrozole (2.5 mg/60 day release) implanted subcutaneously at age 18 months; mammmary tissue obtained at time of necropsy age 20 months

value: mammary gland

value: 20 months

tag: treatment

value: letrozole 2 months exposure


title: sample_11


title: sample_02


title: mammary gland, age 20M, 8 months CYP19A1 transgene induction, tamoxifen 2 months exposure, biol rep 3

value: tamoxifen 2 months exposure


title: sample_01


title: Endothelial cells, induced Sox17 deletion in adult, hypoxia 4

city: Seoul

first: Chan Soon

last: Park

state: Select

address: Department of Internal Medicine Seoul National University Hospital  101 Daehak-ro

institute: Seoul National University Hospital

data_processing:
Illumina Casava1.8 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence(using fastx_trimmer), then mapped to mm10 (UCSC) whole genome using Bowtie2
Read count extraction using bedtools.
Assembly: mm10
Supplementary files format and content: tab-delimited text files include read count values for each Sample

source_name: lung

growth_protocol: To activate Cre recombination in both groups, 2 mg of tamoxifen (Sigma-Aldrich) dissolved in corn oil was injected intraperitoneally to 8-week-old mice for 5 consecutive days.

extract_protocol:
Ribosome-associated transcripts from lung ECs were immunoprecipitated by capturing HA-tagged ribosomes. Then, RNA was harvested using Rneasy mini plus kit (Qiagen). Extracted mRNA was used for the construction of sequencing libraries.
The construction of library was performed using QuantSeq 3’ mRNA-Seq Library Prep Kit (Lexogen, Inc., Austria) according to the manufacturer’s instructions.

treatment_protocol: Control mice aged 10 weeks were placed in a room air and maintained for 3 weeks. Sox17i∆EC/hypoxia mice aged 10 weeks were exposed to 3-week hypoxia.

value: lung

tag: cell type

value: endothelial cells

value: hypoxia, induced Sox17 deletion in adult


title: Endothelial cells, induced Sox17 deletion in adult, hypoxia 3


title: repeat 1 sample_39

growth_protocol: APOE knock-in mice were either used in the absence of infection or intranasally infected with SARS-CoV-2 MA10

extract_protocol:
Lungs were resected in the absence of infection or on days 2 or 4 post infection, homogenized in Trizol using a Miltenyi homogenizer (program RNA_01), and RNA was extracted using Zymo's Direct-zol kit including DNAse digestion.
Lexogen 3' FWD kit

tag: experimental repeat

value: repeat 01


title: sample_10


title: repeat 1 sample_37


title: repeat 1 sample_38


title: Endothelial cells, induced Sox17 deletion in adult, hypoxia 2


title: repeat 1 sample_36


title: mammary gland, age 20M, 8 months CYP19A1 transgene induction, tamoxifen 2 months exposure, biol rep 2


title: repeat 1 sample_35


title: repeat 1 sample_33


title: Endothelial cells, control, normoxia 4

value: Cdh5-CreERT2; RPL22tm1.1Psam

value: normoxia (room air)


title: repeat 1 sample_34


title: repeat 1 sample_32


title: Endothelial cells, induced Sox17 deletion in adult, hypoxia 1


title: mammary gland, age 20M, 8 months CYP19A1 transgene induction, tamoxifen 2 months exposure, biol rep 1


title: mammary gland, age 20M, 8 months CYP19A1 transgene induction, biol rep 3


title: Endothelial cells, control, normoxia 3


title: mammary gland, age 20M, 8 months CYP19A1 transgene induction, biol rep 2


title: mammary gland, age 20M, 8 months CYP19A1 transgene induction, letrozole 2 months exposure, biol rep 2


title: Endothelial cells, control, normoxia 2


title: repeat 1 sample_28


title: repeat 1 sample_27


title: mammary gland, age 20M, 8 months CYP19A1 transgene induction, letrozole 2 months exposure, biol rep 1


title: repeat 1 sample_29


title: Endothelial cells, control, normoxia 1


title: mammary gland, age 20M, 8 months CYP19A1 transgene induction, biol rep 1


title: repeat 1 sample_26


title: repeat 1 sample_25


title: mammary gland, age 18M, 6 months CYP19A1 transgene induction, biol rep 3

value: 18 months


description: see README file

data_processing:
Sequences were aligned to GRCm38 and demultiplexed using the ParseBiosciences pipeline (v0.9.6p) at default settings for each individual sublibrary using the `split-pipe --mode all` and  `--sample ctrl_01_E2 A1-A2  --sample ctrl_02_E2 A3-A4  --sample ctrl_03_E2 A5-A6  --sample ctrl_04_E3 A7-A8  --sample ctrl_05_E3 A9-A10  --sample ctrl_06_E3 A11-A12  --sample ctrl_07_E4 B1-B2  --sample ctrl_08_E4 B3-B4  --sample ctrl_09_E4 B5-B6  --sample inf_01_E2 B7-B8  --sample inf_02_E2 B9-B10  --sample inf_03_E3 B11-B12  --sample inf_04_E3 C1-C2  --sample inf_05_E4 C3-C4  --sample inf_06_E4 C5-C6  --sample inf_07_E4 C7-C8  --sample inf_08_E2 C9-C10  --sample inf_09_E2 C11-C12  --sample inf_10_E2 D1-D2  --sample inf_11_E3 D3  --sample inf_12_E3 D4  --sample inf_13_E4 D5  --sample inf_15_E4 D6  --sample inf_16_E4 D7  --sample inf_17_E2 D8  --sample inf_18_E3 D9  --sample inf_19_E4 D10  --sample inf_20_E4 D11  --sample inf_23_E3 D12` to demultiplex the samples (sample/well-ID pairs are also noted in the file `sample_well_IDs.tsv`); the the output of the eight sublibraries was then combined using `split-pipe --mode combine`.
Downstream analysis was performed using the Seurat R package (v4.0.2) as described in the methods section of the corresponding manuscript.
Differentially expressed genes in clusters from infected mice with different APOE genotypes were determined using Seurat's `FindMarkers` function; enrichment of immune-related pathways of the hallmark gene set in these genes was assessed using the `GSEA` function of the clusterProfiler R package.
Assembly: GRCm38
Supplementary files format and content: all_genes.csv, cell_metadata.csv, and DGE.mtx represent the output of the Parse Biosciences pipeline after combining the output of all eight sublibraries and can be imported into Seurat
Supplementary files format and content: sample_well_Ids.tsv indicates the well-sample combinations for library generation (also see the command in the first data processing step)
Supplementary files format and content: cell_annotations.tsv contains metadata including cluster identities.

source_name: lungs from APOE knock-in mice with or without COVID-19 (see README file)

extract_protocol:
The pulmonary circulation of mice was flushed with PBS; lungs were resected and dissociated using the Miltenyi lung dissociation kit and a gentleMACS dissociator. Red blood cells were lysed using ACK buffer. Cells were fixed using Parse Bioscience's cell fixation kit (SB1001).
Parse Biosciences single cell whole transcriptome kit (SB2001).
scRNA-seq (combinatorial indexing based)

treatment_protocol: Mice were either left non-infected or they were infected 4 days prior to lung harvest with SARS-CoV-2 MA10 via intranasal injection.


title: E14_ESC_wt_2

first: Jae Hoon

last: Song

address: 201 Songpa-daero, Songpa-gu

institute: ENCell Co., Ltd.

data_processing:
Adapter trimming was performed with a skewer (v0.2.2) with -L 100 and -e options.
Trimmed reads were mapped to the mm10 UCSC Mus musculus genome using the STAR (v2.7.9a).
Mapped reads were quantified and normalized to FPKM values using the Cuffnorm of the Cufflinks (v2.2.1) toolset.
Assembly: mm10
Supplementary files format and content: genes.fpkm_table.txt: Tab-delimited text file of gene expression table in FPKM value generated with Cuffnorm (v2.2.1).

molecule: polyA RNA

source_name: embryonic stem cell

growth_protocol: Mouse E14 ES cell lines were maintained under DMEM supplemented with 15% fetal bovine serum, non-essential amino acids, penicillin-streptomycin-gentamycin, b-mercaptoethanol, and LIF. Cells were cultured on mitomycin C-inactivated C3H mouse embryonic fibroblast.

extract_protocol: Total RNA were extracted from the independent samples, and further purified mRNA containing poly A tail using poly T-attached beads. The library was prepared using purified mRNA using the Illumina TruSeq Stranded mRNA Sample Prep Kit (Illumina, Inc., San Diego, CA, USA, #RS-122-2101).

value: embryonic stem cell

tag: differentiation state

value: undifferentiated


title: repeat 1 sample_24


title: repeat 1 sample_31


title: repeat 1 sample_23


title: repeat 1 sample_22


title: repeat 1 sample_20


title: mammary gland, age 18M, 6 months CYP19A1 transgene induction, biol rep 2


title: repeat 1 sample_30


title: E14_ESC_wt_1


title: repeat 1 sample_21


title: repeat 1 sample_18


title: mammary gland, age 20M, 8 months Esr1 transgene induction, letrozole 2 months exposure, biol rep 2


title: Het2_S8, scRNAseq

city: Bethesda

first: Sergey

last: Leikin

state: Maryland

department: NICHD

institute: National Institutes of Health

description: 10x Genomics

data_processing:
Demultiplexing, barcode processing, and gene counting were performed with the Cell Ranger software (10X Genomics) v5.0.0 (S1-S4) and v6.0.1 (S5-S8)
Assembly: mm10-3.0.0
Supplementary files format and content: Tab-separated values files (barcodes and genes), matrix files, h5 files (filtered features matrix), and html files (10X Genomics web summaries)

source_name: parietal bone

extract_protocol:
Parietal bone cells were isolated as described in Gorrell et al, Adv Sci, 2022
Libraries were constructed with 10X Genomics kits (3' v3 and v3.1) following the manufacturer's protocol
10X Genomics, 3' v3 (S1-S4) and v3.1 (S5-S8)

value: Het G610C

value: parietal bone


title: mammary gland, age 20M, 8 months Esr1 transgene induction, letrozole 2 months exposure, biol rep 1


title: repeat 1 sample_15


title: repeat 1 sample_16


title: mammary gland, age 18M, 6 months CYP19A1 transgene induction, biol rep 1


title: WT1_S5, scRNAseq

description: 10x Genomics

value: WT


title: WT2_S6, scRNAseq

description: 10x Genomics


title: Het1_S7, scRNAseq

description: 10x Genomics


title: repeat 1 sample_13


title: repeat 1 sample_14


title: repeat 1 sample_12


title: mammary gland, age 20M, 8 months Esr1 transgene induction, tamoxifen 2 months exposure, biol rep 2


title: Het2_S4, scRNAseq

description: 10x Genomics


title: Het1_S3, scRNAseq

description: 10x Genomics


title: mammary gland, age 20M, 8 months Esr1 transgene induction, letrozole 2 months exposure, biol rep 3


title: repeat 1 sample_19


title: mammary gland, age 20M, 8 months Esr1 transgene induction, tamoxifen 2 months exposure, biol rep 1


title: repeat 1 sample_09


title: repeat 1 sample_08


title: repeat 1 sample_07


title: mammary gland, age 20M, 8 months Esr1 transgene induction, biol rep 3


title: WT1_S1, scRNAseq

description: 10x Genomics


title: repeat 1 sample_17


title: repeat 1 sample_05


title: repeat 1 sample_04


title: repeat 1 sample_02


title: repeat 1 sample_03


title: mammary gland, age 18M, 6 months Esr1 transgene induction, biol rep 3


title: repeat 1 sample_11


title: mammary gland, age 20M, 8 months Esr1 transgene induction, biol rep 2


title: repeat 1 sample_06


title: 7_WT_femur_S7, scRNAseq

description: 10x Genomics

data_processing:
Demultiplexing, barcode processing, and gene counting were performed with the Cell Ranger software (10X Genomics) v3.1 (Samples 1 and 2) and v4.0 (Samples 3-10)
Assembly: mm10-3.0.0
Supplementary files format and content: Tab-separated values files (barcodes and genes), matrix files, h5 files (filtered features matrix), and html files (10X Genomics web summaries)

source_name: femur & tibia

extract_protocol:
Femur and tibia cells were isolated from E18.5 embryos as described in Gorrell et al, Adv Sci, 2022
Libraries were constructed with 10X Genomics kits (3' v3 and v3.1) following the manufacturer's protocol
10X Genomics, 3' v3

value: femur & tibia


title: mammary gland, age 20M, 8 months Esr1 transgene induction, tamoxifen 2 months exposure, biol rep 3

value: mammary gland


title: mammary gland, age 20M, 8 months Esr1 transgene induction, biol rep 1

value: 20 months


title: repeat 1 sample_10


title: 400_1

city: Hangzhou

first: Caiyong

last: Chen

state: Zhejiang

address: 866 Yuhangtang Road

country: China

institute: Zhejiang University

data_processing:
Basecalling using Illumina Casava1.7 software.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to WBcel235 whole genome using bowtie v0.12.2 with parameters -q -p 4 -e 100 -y -a -m 10 --best --strata.
Reads per kilobase of exon per megabase of library size (RPKM) were calculated using a protocol from Chepelev et al., Nucleic Acids Research, 2009. Breifly, exons from all isoforms of a gene were merged to create one meta-transcript. The number of reads falling in the exons of this meta-transcript were counted and normalized by the size of the meta-transcript and by the size of the library.
Assembly: NCBI WBcel235 genome
Supplementary files format and content: tab-delimited text files include sequence counts and FPKM values for each sample

organism: Caenorhabditis elegans

source_name: worm intestines

growth_protocol: Worms were cultured in liquid C. elegans Habitation and Reproduction (mCeHR2) medium at 20°C.

extract_protocol:
Intestines from three adult worms were combined and lysed. The lysed samples were subjected to reverse transcription using with SMARTScribe Reverse Transcriptase.
The cDNA products were amplified by PCR. The sequencing library was constructed according to the manufacturer’s instructions (Illumina).

treatment_protocol: Synchronized L1 worms were treated with 2 μM, 20 μM, or 400 μM heme for one generation (4 days).

value: worm intestines

value: 400 μM heme


title: 400_2


title: 400_3


title: 5_Het_femur_S5, scRNAseq

description: 10x Genomics


title: WT2_S2, scRNAseq

description: 10x Genomics


title: 20_3

value: 20 μM heme


title: mammary gland, age 18M, 6 months Esr1 transgene induction, biol rep 1


title: 6_Het_femur_S6, scRNAseq

description: 10x Genomics


title: 20_1


title: 20_2


title: 4_WT_femur_S4 scRNAseq

description: 10x Genomics


title: MOLM-13 shLAMP5 #4

city: Cincinnati

first: Nathan

last: Salomonis

state: OH

address: 3333 Burnet Avenue

department: Biomedical Informatics

institute: Cincinnati Children's Hospital

description: MOLM-13 shLAMP5 #4

data_processing:
FASTQ sequence files were aligned to the human genome hg19 using Kallisto to compute isoform and gene level TPM values against the human EnsMart72 database. AltAnalyze version 2.1.1 was used to derive differential gene expression changes.
Genome_build: hg19
Supplementary_files_format_and_content: Gene-level Transcript Per Million (TPM) estimates.

organism: Homo sapiens

source_name: Leukemia cell line

growth_protocol: MOLM-13 cells were maintained in RPMI 1640 supplemented with 10% FBS

extract_protocol:
After treatment, total RNA was isolated using the QIAGEN RNEasy Mini kit according to manufacturer's protocols.
Library construction was performed using the TruSeq RNA Library Prep Kit (Illumina) according to the manufacturer's protocols. Purified and adapter-ligated cDNA was captured on a flowcell, and sequenced on an Illumina HiSeq 2500

treatment_protocol: Cells were transduced with lentiviral shRNA against LAMP5 or non-targeting control, and selected with puromycin prior to harvest at the indicated timepoints

value: Acute myeloid leukemia

tag: translocation

tag: shRNA

tag: cell line


title: 2_3

value: 2 μM heme


title: 8_Hom_femur_S8, scRNAseq

description: 10x Genomics

value: Hom G610C


title: repeat 1 sample_01

tag: day post infection

value: repeat 01


title: MOLM-13 shLAMP5 #3

description: MOLM-13 shLAMP5 #3


title: 2_2


title: 3_Het_femur_S3, scRNAseq

description: 10x Genomics


title: 2_1


title: mammary gland, age 18M, 6 months Esr1 transgene induction, biol rep 2


title: MOLM-13 shLAMP5 #2

description: MOLM-13 shLAMP5 #2


title: MOLM-13 shLAMP5 #1

description: MOLM-13 shLAMP5 #1


title: PD-1 agonist treated mouse_003

city: Los Angeles

first: Georges Doumet

last: HELOU

state: California

address: 1450 Biggy St, NRT5505, Keck School of Medicine

department: Molecular Microbiology and Immunology

institute: University of Southern California

description: 10x Genomics Chromium Single Cell

data_processing:
Single-cell sequencing data was processed and analyzed using Partek® Flow® software (v10.0.21.1116), where read pairs were first trimmed, according to the 10x Genomics Chromium Single Cell 3’ v3 prep kit used, to extract barcode and UMI information.
Assembly: mm10
Supplementary files format and content: Tab-separated values files and matrix files

extract_protocol:
Sorted live CD45+ cells (viability ≥95%) were stained using TotalSeq™-A antibodies and cell hashing reagents. Three samples from each group were mixed using the cell hashing technique (⁓50,000 cells/group).
The pooled library was loaded and sequenced on an Illumina HiSeq X Ten System (150bp x 8bp x 150bp).

value: CD45+ cells

value: Lung

value: HUMANIZED PD-1 MOUSE MODEL

value: 6-8 weeks old

value: PD-1 agonist


title: Isotype treated mouse_003

description: 10x Genomics Chromium Single Cell

value: isotype


title: PD-1 agonist treated mouse_001

description: 10x Genomics Chromium Single Cell


title: Isotype treated mouse_002

description: 10x Genomics Chromium Single Cell


title: PD-1 agonist treated mouse_002

description: 10x Genomics Chromium Single Cell


title: Wt_femur_S2, scRNAseq

description: 10x Genomics

value: WT


title: Isotype treated mouse_001

description: 10x Genomics Chromium Single Cell


title: MOLM-13 shNT non-targeting control #2

description: MOLM-13 shNT #2

value: shNT


title: Hom_femur_S1, scRNAseq

description: 10x Genomics

value: femur & tibia


title: MOLM-13 shNT non-targeting control #1

description: MOLM-13 shNT #1


title: sa001 iPSC-derived neurons

city: South San Francisco

first: Amy

last: Easton

state: CA

address: 1 DNA Way

department: Department of Neuroscience

institute: Genentech, Inc

data_processing:
NovaSeq Control Software 1.7.5
Sequence reads were trimmed for adaptor sequence/low-quality sequence using Trimmomatic (0.32)
Basecalling was performed using Real-Time Analysis (RTA) v3.4.4 and fastq generated with bcl2fastq2.20 v2.20.0.422
Mapping of the reads was performed with HISAT2
BAM files contains RNAseq reads  mapped using HISAT2 (v2.1.0) to the human genome assembly GRCh38 (hg38);  the mapped BAM file was used to generate a sashimi plot.
Assembly: GRCh38
Supplementary files format and content: The BAM (.bam) file contain a header section and an alignment section. The header contains information such as sample name, sample length, and alignment method. The alignments section contains read name, read sequence, read quality, alignment information, and custom tags.
Supplementary files format and content: The BAI (.bai) index file contains additional metadata for the corrisponding BAM file, providing a summary of the number of mapped and placed unmapped read-segments per reference sequence, and of any unplaced unmapped read-segments.

source_name: SA001 hESC

extract_protocol:
RNA isolation was performed using RNeasy Mini Kit  (Qiagen) following manufacture’s instruction.
Coding and non-coding RNA-Seq: RNA libraries for RNA-seq were prepared using TruSeq stranded total RNA library preparation protocol with RiboZero to remove rRNA (Illumina)

value: SA001 hESC

value: neuronal culture

value: wt


title: 2_WT_femur_S2, scRNAseq

description: 10x Genomics


title: 1_Hom_femur_S1, scRNAseq

description: 10x Genomics


title: BJAB-B1 Cells, DMS mock probed, DMS- replicate 1

status: Public on Aug 06 2022

city: Ames

first: Walter

middle: N.

last: Moss

state: IA

address: 2437 Pammel Dr

department: BBMB

institute: Iowa State University

description: 151 sequencing cycles

data_processing:
The StructureFold2 (SF2) package (10.1016/j.ymeth.2018.01.018) was used to process FASTQ files into react files, starting with trimming (utilizing Cutadapt (version 1.13) )
SF2 mapping utilizing Bowtie2 (version 2.3.4.1)
SF2 filtering utilizing SamTools (version 1.10)
SF2 conversion to RTSC files
SF2 conversion of RTSC files to REACT files
Assembly: hg38 Human Genome, EBV genome NC_009334.1
Supplementary files format and content: React files containing normalized DMS structure reactivity values, similar to fasta file format, but react values are tab deliminated and sequence header has no ">"

growth_protocol: BJAB-B1 cells were grown in suspension at 37°C and 5% CO2 using RPMI media supplemented with 2 mM l-glutamine, 1% penicillin–streptomycin, 10 mM HEPES, 1 mM sodium pyruvate (Life Technologies), and 10% FBS (Atlanta Biologicals).

extract_protocol:
TRIzol extraction, followed by standard ethanol precipitations
cDNA library preparation protocol outlined by Structure-seq2 (https://doi.org/10.1007/978-1-4939-9045-0_20), was followed

treatment_protocol: BJAB-B1 cells were treated with DMS (2% v/v) (or DPBS for DMS- Samples) for 2 minutes at room temperature prior to neutralization with DTT (powder at a 5 times molar ratio to DMS).

value: B-cell lymphoma

value: EBV infected

value: DPBS mock probed 2%(v/v)


title: BJAB-B1 Cells, DMS mock probed, DMS- replicate 3

description: 151 sequencing cycles


title: BJAB-B1 Cells, DMS probed, DMS+ replicate 3

description: 151 sequencing cycles

value: DMS probed 2%(v/v)


title: HeLa siTRESLIN 24 hours biol rep 2

city: Copenhagen N

first: Mads

last: Lerdrup

address: Blegdamsvej 3C

department: Department of Cellular and Molecular Medicine

country: Denmark

institute: University of Copenhagen

data_processing:
The quality of the sequenced reads was assessed using FastQC (v. 0.10.1; https://www.bioinformatics.babraham.ac.uk/projects/fastqc/), FastqScreen (v. 0.11.4; PMID: 30254741), and MultiQC (v. 1.7; PMID: 27312411).
Reads were trimmed by BGI prior to release and mapped to canonical chromosomes from hg38 using STAR (v. 2.5.1a; PMID: 23104886) using the setting “--outFilterMismatchNoverLmax 0.1 --outFilterMatchNmin 16 --outFilterMismatchNmax 5.
Transcriptome annotation was retrieved from Refseq (PMID: 26553804) and fed into HTseq-count to quantify reads within transcripts using the settings ‘htseq-count --mode=union --stranded=no’.
Assembly: hg38
Supplementary files format and content: Tab-delimited text files include raw count values wihtin genes for each sample

extract_protocol:
Total RNA was extracted from cells using manufacturer’s instructions (Cytiva Life Sciences). 1µg of total RNA was used for the construction of sequencing libraries.
Libraries were constructed by BGI according to the following procedure (All reagents were made by BGI unless stated): mRNA molecules were purified from total RNA using oligo(dT)-attached magnetic beads, and fragmented into small pieces. cDNA was generated using random hexamer-primed reverse transcription, followed by a second-strand cDNA synthesis. The synthesized cDNA was subjected to end-repair and was 3’ adenylated. Adapters were ligated to the ends of 3’ adenylated cDNA fragments. cDNA fragments were PCR amplified and purified with Ampure XP Beads (Agencourt), and dissolved in EB solution. Library quality was validated on an Agilent Technologies 2100 bioanalyzer, and double stranded PCR products were heat denatured and circularized by the splint oligo sequence. Single strand circle DNA was amplified with phi29 to make DNA nanoballs (DNBs) with more than 300 copies of one molecule. DNBs were loaded into the patterned nanoarray and single end 50 basepair reads were generated on a BGISEQ-500.

value: HeLa

value: Homo sapiens, female, epithelial cervical carcinoma

tag: sirna

value: siTRESLIN

value: RNAi transfection

tag: time

value: 24 hours


title: HeLa siTRESLIN 48 hours biol rep 2

value: 48 hours


title: HeLa siTRESLIN 48 hours biol rep 3


title: HeLa siCTR biol rep 2

value: Control


title: HeLa siTRESLIN 24 hours biol rep 3


title: HeLa siCTR biol rep 1


title: HeLa siCTR biol rep 3


title: HeLa siTRESLIN 24 hours biol rep 1


title: HeLa siTRESLIN 48 hours biol rep 1


title: BJAB-B1 Cells, DMS mock probed, DMS- replicate 2

description: 151 sequencing cycles


title: BJAB-B1 Cells, DMS probed, DMS+ replicate 2

description: 151 sequencing cycles


title: BJAB-B1 Cells, DMS probed, DMS+ replicate 1

description: 151 sequencing cycles


title: PIM1-FHH rep2

city: Wuhan

first: Shaofeng

last: Lin

address: Luoyu Road 1037

institute: Huazhong University of Science and Technology

data_processing:
Illumina Casava software was used for basecalling.
TopHat (version 2.2.1) was used to map raw reads to the reference genome.
Cufflinks (version 2.2.1) was used to assemble reads and calculate fragments per kilobase of exon per million fragments mapped (FPKM) values of mapped mRNAs for the estimation of their expression levels.
Genome_build: GRCh38
Supplementary_files_format_and_content: Tab-delimited text files include FPKM values for each Sample.

source_name: dermal fibroblast

growth_protocol: Human dermal fibroblasts (HDFs) were derived from human skin biopsy and cultured in human fibroblast medium (HFM).

extract_protocol:
Cell samples were collected with Ttizol Reagent and incubated at room temperature 5min. The homogenates was thoroughly mixed with chloroform and standby 3min at room temperature. After centrifugation of 12,000 g at 4 °C, aqueous phase was transferred and mixed with isopropanol. After centrifugation of the homogenates by 12,000 g at 4 °C, RNA sediment was washed two times with 75% ethanol and obtained the total RNA.
TruSeq® RNA LT Sample Prep Kit v2 (Illumina, USA) was used to treat 2 μg RNA for each sample. Then, mRNA was purified from total RNA with RNA Purification Beads, and fragmented with Elute, Prime, Fragment Mix (EPF). First and second strand cDNAs were synthesized based on random primers. End repair was performed with End Repair Mix 2. After Adenylating 3' ends, adaptors were ligated to RNA fragments. And then, the cDNA was amplified with PCR and quantified with Qubit (Invitrogen, USA).

treatment_protocol: HDFs were seeded on collagen-I-coated dishes and infected with lenti-viruses carrying the indicated genes. Cells were cultured in HFM on the first two days，and change human modified Block’s medium (HMM) every other day. Cells were collected for further analyses at different time points according to specific experiment.

value: Fibroblast

value: Skin

value: PIM1 overexpression at 5 days after infection of FOXA3, HNF1A and HNF4A


title: PIM1-FHH rep1


title: GFP-FHH rep3

value: GFP overexpression at 5 days after infection of FOXA3, HNF1A and HNF4A


title: FHH-5d

value: 5 days after infection of FOXA3, HNF1A and HNF4A


title: FHH-2.25d

value: 2.25 days after infection of FOXA3, HNF1A and HNF4A


title: GFP-FHH rep1


title: GFP

value: infection of GFP


title: 213691

type: genomic

city: NICE

first: Thibault

last: FABAS

address: 28 av Valombrose

country: France

institute: CHU Nice

hyb_protocol: DNA was restriction digested, PCR amplified, fragmented, labeled and hybridized to each array according to the manufacturer's instructions.

scan_protocol: The Arrays were then washed using Affymetrix fluidics stations, and scanned using the Gene Chip Scanner 3000.

data_processing:
The array image was acquired using Affymetrix GeneChip® Command Console Software (version 3.2.2). Copy number values for individual SNPs were extracted and converted from CEL files into signal intensities using CHAs (version 4.0). Genomic Smoothing analysis was performed by CHAs
The CNV value is the logarithm base 2 of the sample signal divided by a reference value from the OncoScan_CNV.FFPE.na33.r2.REF_MODEL library. The allelic status is determined accordind the B-Allelic Frequency (BAF = B/(A+B) ; AA : BAF = 0, BB : BAF = 1, AB : BAF = 0,5) or the Allele Difference (Diff = A - B where each A and B allele are 0,5 units. AA : Diff = 1, AB : Diff = 0, BB : Diff = -1)

label: Biotin

molecule: genomic DNA

source_name: Merkel cell carcinoma

label_protocol: As per manufacturer (Affymetrix)

extract_protocol: DNA was extracted from formol fixed-paraffin embedded fragments using Maxwell FFPE DNA LEV Kit (PROMEGA).

tag: condition

value: carcinoma


title: 213690


title: 213600


title: 213687


title: 213686


title: 213682


title: 213684


title: PIM1-FHH rep3


title: 213438


title: 213439


title: 213431


title: 213383


title: 213433


title: GFP-FHH rep2


title: 213432


title: 213440


title: 213595


title: 213437


title: 212510


title: 213384


title: 213442


title: 213378


title: 213381


title: 212478


title: 213430


title: 212512


title: 212436


title: 213795


title: 212474


title: 212476


title: 212475


title: 212472


title: 212477


title: 213382


title: 253 GE  #7

city: Bronx

first: XUSHENG

last: ZHANG

state: NY

address: 1300 Morris Park Ave

department: GENETICS

institute: ALBERT EINSTEIN COLLEGE OF MEDICINE

description: 10x Genomics

data_processing:
Cell Ranger Pipeline v6.0.1
Assembly: OryCun2.0
Supplementary files format and content: HDF5 Feature Barcode Matrix Format

organism: Oryctolagus cuniculus

source_name: dissected ventral ganglionic eminence from brain slices

extract_protocol: dissected brain tissue  dissociated to single cells with Neural Tissue dissociation Kit

treatment_protocol: rabbit kits with moderate IVH were treated with intracerebroventricular (ICV) Ad-GFP or Ad-SHH (CMV promoter 5ul volume ) 1mm ant. 4mm lateral and 3mm deep from Bregma under anesthesia

value: interneuron

value: Ad-GFP


title: 253 GE  #13

description: 10x Genomics

value: dissected ventral ganglionic eminence from brain slices

value: Ad-SHH


title: 254 GE  #2

description: 10x Genomics


title: K231#2 d3 no IVH

description: 10x Genomics

tag: group

value: no IVH


title: 212473


title: K232#9 d3 IVH

description: 10x Genomics

tag: tissue

value: IVH


title: 251 GE  #2

description: 10x Genomics


title: K231#3 d3 IVH

description: 10x Genomics


title: 254 GE  #3

description: 10x Genomics


title: sham 4 at 6H,spleen

type: RNA

city: Beijing

first: Liming

last: Liu

address: 16 Dongzhimen nei south street, Dongcheng District, Beijing, China

institute: Institute of Chinese Materia Medica，China Academy of Chinese Medical Sciences

hyb_protocol: Total RNA was reverse transcribed, amplified, labelled and hybridized to Rat Genome 1.0 arrays (Affymetrix).

scan_protocol: GeneChips were scanned using the Hewlett-Packard GeneArray Scanner G3000 7G.

data_processing: The data were analyzed with RMA using Affymetrix default analysis settings and global scaling as normalization method. The trimmed mean target intensity of each array was arbitrarily set to 100.

label: biotin

organism: Rattus

source_name: sham 4 at 6H

label_protocol: M-MLV reverse transcription (Promega) was used to synthesize cDNA.

growth_protocol: All animals were housed individually at 22 ± 2 °C with a relative humidity of 50 ± 10% and a 12-h light/12-h dark cycle.The animals were kept in a pathogen-free environment with free access to food and water.

extract_protocol: Extraction of total RNA was performed according to the manufacturer's instructions of Qiagen RNA kit.

treatment_protocol: The rats were randomly divided into 3 groups according to a uniform design (n=3/group): 6h sham group, 6h vehicle control group, 6h YJ groups (25mg•kg−1). The sham operation group was subjected to only preoperative anesthesia and blood vessel separation; the other groups were subjected to the MCAO model. The animals were sacrificed to obtain brain, spleen and blood respectively. Furthermore, brains and spleens were made as homogenate and white blood cells were separated from whole blood for further tests.

value: Sprague Dawley

value: spleen

value: sham 4


title: sham 5 at 6H,blood

source_name: sham 5 at 6H

value: blood

value: sham 5


title: K266#1 d3 control

description: 10x Genomics

value: control


title: K226#7 d3 no IVH

description: 10x Genomics


title: sham 4 at 6H,brain

value: brain


title: 251 GE  #1

description: 10x Genomics


title: sham 5 at 6H,spleen


title: sham 3 at 6H,blood

source_name: sham 3 at 6H

value: sham 3


title: model 3 at 6H,blood

source_name: model 3 at 6H

value: model 3


title: sham 3 at 6H,brain


title: sham 5 at 6H,brain


title: sham 3 at 6H,spleen


title: sham 4 at 6H,blood


title: MH-3

data_processing:
Illumina Casava software was used for basecalling.
TopHat (version 2.2.1) was used to map raw reads to the reference genome.
Cufflinks (version 2.2.1) was used to assemble reads and calculate fragments per kilobase of exon per million fragments mapped (FPKM) values of mapped mRNAs for the estimation of their expression levels.
Genome_build: mm9
Supplementary_files_format_and_content: Tab-delimited text files include FPKM values for each Sample.

source_name: primary hepatocytes

value: hepatocyte


title: model 3 at 6H,spleen


title: model 2 at 6H,spleen

source_name: model 2 at 6H

value: model 2


title: model 3 at 6H,brain


title: model 2 at 6H,blood


title: MH-2


title: model 1 at 6H,spleen

source_name: model 1 at 6H

value: model 1


title: lpc-Hep-3

source_name: liver progenitor cells differentiate to ALBUMIN+ hepatocytes (lpc-Hep)


title: lpc-Hep-2


title: model 2 at 6H,brain


title: lpc-Hep-1


title: model 1 at 6H,brain


title: vehicle 5 at 6H,brain

source_name: vehicle 5 at 6H

value: vehicle 5


title: vehicle 5 at 6H,spleen


title: Dendritic Cell mRNA, UMOD Homozygous WT, 2

city: Little Rock

first: Annjanette

last: Stone

email: annjanettemc@yahoo.com, astone@uams.edu

state: AR

address: 4300 West 7th Street

department: Research

institute: CAVHS

description:
WT2
mRNA

data_processing:
Total number of bases, reads, GC (%), Q20 (%) Q30 (%) are calculated for each sample from raw data (FastQC v0.11.7).
Phred Quality Score is determined for each cycle of sequence and plotted with fastq.gz v0.11.7.
Trimmomatic 0.38 program is used to remove adapter sequence and bases with base quality lower than three from the ends. Trimmed reads are mapped to reference genome with HISAT2 version 2.1.0, splice-aware aligner. Bowtie2 v 2.3.4.1 is used for graphs of FM index (GFM).
Stringtie Version 2.1.3b was used for transcript assembly with aligned reads.
Expression profile was calculated for each sample and transcript/gene as read count and FPKM (Fragment per kilobase of transcript per Million mapped reads).
DEG (Differentially Expressed Genes) analysis was performed on comparison pairs (Test_vs_Control) (DESeq2).
Correlation analysis between samples was obtained through Pearson's coefficient of the normalized values.
Hierarchical clustering (Euclidean distance, complete linkage analysis) was performed to graphically represent expression patterns between samples and genes.
DEG list was further analyzed with gProfiler (https://biit.cs.ut.ee/gprofiler/orth) for gene set enrichment analysis per biological process (BP), cellular component (CC) and molecular function (MF).
Assembly: UCSC mm10
Supplementary files format and content: Express_Profile.mm10.transcript.xlxs: FPKM and raw counts as DESeq2 output (in xlsx format).

source_name: Dendritic Cells

growth_protocol: UMOD C106F Mutant mice and wt littermate controls were allowed to grow until 12-16 weeks at which point they were sacrificed.

extract_protocol:
Dendritic cell isolation: Mouse spleens were digested with 2 mg/ml collagenase IV (ThermoFisher) for 30 minutes at 37°C and passed through 70 um nylon filters. Dendritic cells were isolated by negative selection using a pan dendritic cell immunomagnetic kit (Miltenyi 130-100-875), according to the manufacturer’s directions.
The library preparation was done using a NEBNext Ultra II Directional RNA Library kit (New England Biolabs). 75 ng of total RNA was mRNA enriched using oligodT beads. The mRNA-stranded libraries were pre-amplified with PCR incorporating dual index primers and purified with magnetic beads.

treatment_protocol: To create a guanine to thymine point mutation in the mouse UMOD gene (TG mice only) resulting in cysteine to phenylalanine transition, a plasmid containing a CRISPR RNA guide was injected with Cas9 expression plasmid, along with a 153-bp single-stranded donor DNA fragment containing the mutation, into C57BL/6J mouse embryos at the single-cell stage. These embryos were then implanted into foster mothers. Mice carrying the C105F mutation were identified using a Taqman SNP genotyping assay that demonstrated clear allelic discrimination.

value: Dendritic Cells

value: C57BL/ 6J

tag: genotype

value: Wildtype (+/+)


title: Dendritic Cell mRNA, UMOD Homozygous WT, 3

description:
WT3
mRNA


title: Dendritic Cell mRNA, UMOD Homozygous WT, 1

description:
WT1
mRNA


title: vehicle 2 at 6H,brain

source_name: vehicle 2 at 6H

value: vehicle 2


title: vehicle 4 at 6H,spleen

source_name: vehicle 4 at 6H

value: Sprague Dawley

value: vehicle 4


title: vehicle 4 at 6H,blood


title: Dendritic Cell mRNA, UMOD C105F/+ mutant, 3

description:
M3
mRNA

value: Heterozygous Mutant (C105F/+)


title: Dendritic Cell mRNA, UMOD Homozygous WT, 6

description:
WT6
mRNA


title: Dendritic Cell mRNA, UMOD C105F/+ mutant, 6

description:
M6
mRNA


title: Dendritic Cell mRNA, UMOD C105F/+ mutant, 5

description:
M5
mRNA


title: Dendritic Cell mRNA, UMOD Homozygous WT, 4

description:
WT4
mRNA


title: Dendritic Cell mRNA, UMOD Homozygous WT, 5

description:
WT5
mRNA


title: vehicle 2 at 6H,spleen

value: spleen


title: vehicle 2 at 6H,blood

value: blood


title: model 1 at 6H,blood


title: vehicle 5 at 6H,blood


title: vehicle 4 at 6H,brain


title: Dendritic Cell mRNA, UMOD C105F/+ mutant, 2

description:
M2
mRNA


title: MH-1


title: PMs,ko vsv,0h

city: Tian Jin

first: Yang

last: Cao

address: No. 94 Weijin Road

institute: Nankai University

data_processing:
Illumina Casava1.7 software used for basecalling
Sequence reads were trimmed for adaptor sequence,and masked for low-complexity or low-quality sequence ，then mapped to mm10 whole genome using bowtie v0.12.2 with parameters
Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated using a protocol from Chepelev et al,Nucleic Acids Research,2009 .In short ,exons from all isforms.
Assembly: mm10
Supplementary files format and content: tab-delimited text files include RPKM values for each Sample

source_name: Peritoneal macrophages

extract_protocol:
Retinasn were removed,flash frozen on dry ice ,and RNA was harvested using Trizol reagent Illumina TruSeq RNA Sample Prep Kit(Cat#FC-122-1001) was used with 1ug of total RNA for the construction
RNA libraries for RNA-seq were prepared using standard Illumina protocols.

value: Peritoneal macrophages

value: lncRNA-BTX konckout

value: vsv( Vesicular stomatitis virus) infection

value: hour 0


title: PMs,wt vsv,8h

source_name: peritoneal macrophages

value: peritoneal macrophages

value: WT

value: hour 8


title: PMs,wt vsv,0h


title: PMs,ko vsv,8h


title: 12d OP50

first: An Na

institute: Seoul National University

description: Gene expression after 12 d exposal of E. coli OP50

hyb_protocol: Following the manufacturer's instructions, 1.5 ug of Cy3-labeled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 250 ml containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent. After the fragmentation process was complete, 250 ml of 2x Agilent hybridization buffer was added to the fragmentation mixture, which was then hybridized for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed for 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and for 1 minute at 37°C with GE Wash Buffer 2 (Agilent), followed by rapid centrifugation for drying.

scan_protocol: Immediately after washing, slides were scanned on the Agilent DNA Microarray Scanner with one color scan setting for 1x44k array slides.

data_processing: Background removed and spatially detrended Processed Signal intensities were obtained by analyzing the scanned pictures using Feature Extraction Software 9.1. Features identified as Feature Non-uniform outliers in Feature Extraction were eliminated.

label: cyanine 3

source_name: RNA extract, 12d OP50

label_protocol: cRNA tagged with cyanine-3 (Cy3) was synthesized using the One-Color Low RNA Input Linear Amplification PLUS kit and RNAeasy column purification from 0.5 ng RNA. Using a NanoDrop Spectrophotometer, the incorporation of dye and cRNA yield were analyzed.

growth_protocol: C. elegans fer-15;fem-1 was obtained from the CGC and maintained on NGM plates at 15℃

extract_protocol: Total RNA was isolated using TRIZOL reagent and purified using RNeasy Mini Kit with a mini bead beater according to the manufacturer’s instructions

treatment_protocol: L4 staged worms were randomly harvested at 6 and 12 days after exposure to OP50 to examine gene expressions differed during aging

tag: maturation stage


title: Dendritic Cell mRNA, UMOD C105F/+ mutant, 4

description:
M4
mRNA


title: 6d OP50

description: Gene expression after 6 d exposal of E. coli OP50

source_name: RNA extract, 6d OP50


title: Blood, restraint stress 6h, rep4

city: Jinan

first: Hao

last: He

state: Shandong

address: No.6699, Qingdao Road, Huaiyin District

institute: Shandong First Medical University

data_processing:
Raw data (raw reads) of fastq format were firstly processed through in-house perl scripts. Clean data (clean reads) were obtained by removing reads containing adapter, reads containing ploy-N and low quality reads from raw data.
Paired-end clean reads were aligned to the reference genome using Hisat2. Featurecount was used to count the reads numbers mapped to each gene.
Assembly: mm10
Supplementary files format and content: tab-delimited text files include count values for all Samples

source_name: peripheral blood

extract_protocol:
Blood were flash frozen on dry ice, and RNA was havested using Illumina TruSeq RNA Sample Prep Kit (Cat#FC-122-1001). 1 ug of total RNA was used for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols.

value: peripheral blood

tag: gender

value: Female

tag: age

value: 6-8 weeks age

value: blood cells

value: restraint stress, 6h


title: Blood, restraint stress 6h, rep3

tag: strain

tag: treatment


title: Blood, restraint stress 6h, rep2

tag: cell type


title: Blood, no stress, rep4

value: no restraint stress


title: Blood, restraint stress 6h, rep1


title: Blood, no stress, rep3


title: Blood, no stress, rep2


title: X2023462_Pdx1ZNKP_mesentery_met_het

city: Cambridge

first: Amir

last: Jassim

address: Robinson Way

country: United Kingdom

institute: Cancer Research UK Cambridge Institute

data_processing:
Single end 50bp RNA reads were aligned to GRCm38 with HISAT2 (with default parameters). Each sample was sequenced across several lanes; per-lane BAM files were merged into per-sample BAMs.
Quality control metrics were collected for each file, including duplication statistics and number of reads assigned to genes. Reads were counted on annotated features with sub-reads featureCounts, providing 'total', 'aligned to the genome', and 'assigned to a gene' (i.e. included in the analysis) counts.
Percentages of aligned bases were computed for several categories: coding, UTR, intronic, intergenic. Other quality control metrics were the percentage of reads on the correct strand, median coefficient of variation of coverage, median 5’ bias, median 3’ bias, and the ratio of 5’ to 3’ coverage.
Quality control also included an expression heatmap drawn using log2 transformed counts. Log2 transformed counts were generated from normalized counts using the log2 function in R and counts function from DEseq2.
Assembly: GRCm38
Supplementary files format and content: CSV file of raw counts

source_name: mesentery

extract_protocol:
Total RNA was extracted from tissues using Maxwell RSC miRNA Tissue Kit (AS1460, Promega). RNA quality was assessed using TapeStation System (5067-5579, Agilent).
The Illumina TruSeq stranded mRNA kit (20020595, Illumina) was used to prepare RNA libraries and RNA quality confirmed using Tapestation (Agilent) and quantified using Kapa qPCR library quantification kit for Illumina platforms (KK4873,Kapa Biosystems). Samples were normalised using the Agilent Bravo, pooled and sequenced on NovaSeq flow cell to generate single end 50bp reads at 20M reads per sample.

value: mesentery

tag: tumour type

value: metastasis

tag: nalcn status

value: Nalcn-het

tag: Sex

value: male

tag: age (days)


title: X206237_VilCreerZNKP_small_intestine_mass_wt

source_name: small intestine

value: small intestine

value: primary tumour

value: Nalcn-wt

value: female


title: KI_293T_E1-59

city: Suita

first: Atsushi

last: Kumanogoh

state: Osaka

address: 2-2 Yamadaoka

country: Japan

institute: Osaka University

data_processing:
The adapter sequence was trimmed using trim_galore version 0.6.4. The remaining trimmed reads were aligned to the human genome hg19 by STAR version 2.7.1a. Alternative splicing events between wild-type (+/+) clones and mutant (+/G96V) clones was detected using rMATS version 4.0.2.
Assembly: hg19
Supplementary files format and content: The processed data files were generated by rMATS.

source_name: cultured cell

growth_protocol: The cells were cultured in a humidified incubator under hypoxic conditions (94% N2, 5% CO2, and 1% O2) at 37℃ in high glucose (4 g/L) Dulbecco's modified Eagle medium (nacalai tesque) supplemented with 10% fetal bovine serum (Gibco) and 1% Penicillin-Streptomycin Mixed Solution (nacalai tesque).

extract_protocol:
Toral RNA was extracted from the cells using RNeasy Mini Kit according to the manufacturer's protocol.
RNA-sequencing Library preparation was performed using a TruSeq stranded mRNA sample prep kit (Illumina, San Diego, CA) according to the manufacturer’s instructions.

tag: cell line

value: SNRPD3 +/G96V

tag: oxygen condition

value: hypoxia


title: KI_293T_E1-15


title: KI_293T_E1-155


title: X2023462_Pdx1ZNKP_LN_het

source_name: lymph node

value: lymph node


title: X40357_PcZNKP_gastric_mass_het

source_name: gastric

value: gastric


title: X206237_VilCreerZNKP_small_intestine_mass_wt_2


title: X2023462_Pdx1ZNKP_pancreas_mass_2_het

source_name: pancreas

value: pancreas


title: X2021514_Pdx1ZNKP_pancreas_mass_cko

value: Nalcn-null


title: KI_293T_E1-123


title: T84 cells_Combo_6

city: Worcester

first: Beth

last: McCormick

state: MA

address: 368 Plantation Street

institute: University of Massachusetts Chan Medical School

data_processing:
Raw Data Filtering performed by Novogene: Removal of reads containing adapters, removal of reads containing N > 10%, Removal of reads containing low quality (Qscore<=5) base
Filtering of ribosomal RNA reads using bowtie2 aligner (v2.3.5)
Alignment to human hg38_gencode_V34 reference genome using STAR (v2.6.1)
Count matrices produced using FeatureCounts
Assembly: hg38_gencode_V34
Supplementary files format and content: featureCounts output tsv file containing gene name, gene length, and count for each sample

source_name: T84 cells

growth_protocol: T84 cells grown in DMEM:F12 growth media with 15mM HEPES and 100units/mL penicillin/streptomycin, seeded onto collagen-coated 24-well tissue culture plates at 30,000 cells/well until >80% confluent.

extract_protocol:
Total RNA collected with Qiagen RNeasy mini kit with DNase gDNA removal step.  RNA integrity (RQN>9) measured by Agilent 5300 Fragment analyzer
Stranded mRNA library preparation with poly A enrichment performed by Novogene

treatment_protocol: Incubated with DMSO vehicle control "Control", 5mM butyrate "Butyrate", 50µM bile acids (lithocholic acid, deoxycholic acid, ursodeoxycholic acid) "BileAcid", or combination of 5mM butyrate and 50µM bile acids (lithocholic acid, deoxycholic acid, ursodeoxycholic acid) "Combo" for 4 hours prior to total RNA collection

value: wild-type

tag: treatment time

value: 4 hours

tag: replicate


title: KI_293T_E1-130


title: X2020797_Pdx1ZNKP_pancreas_mass_wt_2


title: X2021514_Pdx1ZNKP_pancreas_mass_cko_2


title: X2020797_Pdx1ZNKP_pancreas_mass_wt


title: Blood, no stress, rep1

value: peripheral blood

value: Female

value: 6-8 weeks age

value: blood cells

value: WT

value: no restraint stress


title: KI_293T_E1-302


title: T84 cells_BileAcids_6


title: T84 cells_Butyrate_6


title: T84 cells_Control_6


title: X2019013_Pdx1ZNKP_pancreas_mass_cko

value: primary tumour


title: X2019013_Pdx1ZNKP_pancreas_mass_cko_2


title: T84 cells_Combo_5


title: T84 cells_Butyrate_5


title: T84 cells_Control_5


title: X2019009_Pdx1ZNKP_pancreas_mass_cko

value: male


title: X2019009_Pdx1ZNKP_pancreas_mass_cko_2


title: T84 cells_Combo_4


title: X2018647_Pdx1ZNKP_pancreas_mass_het

value: Nalcn-het

value: female


title: X2018647_Pdx1ZNKP_mesentery_met_het

value: mesentery

value: metastasis


title: T84 cells_Butyrate_3


title: T84 cells_Control_4


title: X2018465_Pdx1ZNKP_pancreas_mass_wt

value: Nalcn-wt


title: T84 cells_Combo_3


title: T84 cells_BileAcids_4


title: T84 cells_Butyrate_4


title: X2018465_Pdx1ZNKP_pancreas_mass_wt_2

value: pancreas


title: T84 cells_BileAcids_3


title: X2018647_Pdx1ZNKP_pancreas_mass_het_2


title: X2018645_Pdx1ZNKP_pancreas_mass_wt


title: X2017226_Pdx1ZNKP_pancreas_mass_het_2


title: X2017226_Pdx1ZNKP_pancreas_mass_het


title: T84 cells_Combo_2


title: T84 cells_Control_3


title: X1951278_PcZNKP_gastric_mass_cko

value: gastric

value: Nalcn-null


title: T84 cells_BileAcids_5

value: wild-type

value: 4 hours


title: X1950700_VilCreerZNKP_small_intestine_mass_wt_2

value: small intestine


title: T84 cells_BileAcids_2


title: X2019009_Pdx1ZNKP_pancreas_mass_cko_2_2


title: T84 cells_Butyrate_2


title: T84 cells_Control_2


title: X1950700_VilCreerZNKP_small_intestine_mass_wt


title: X1950698_VilCreerZNKP_small_intestine_mass_wt_2


title: T84 cells_Combo_1


title: X1950698_VilCreerZNKP_small_intestine_mass_wt


title: T84 cells_BileAcids_1


title: X1950670_PcZNKP_lung_met_wt

source_name: lung

value: lung


title: T84 cells_Butyrate_1


title: X1950500_Pdx1ZNKP_pancreas_mass_het


title: T84 cells_Control_1


title: X1950500_Pdx1ZNKP_fat_pad_met_het

source_name: fat pad

value: fat pad


title: X1950500_Pdx1ZNKP_diaphragm_met_het

source_name: diaphragm

value: diaphragm


title: X1950500_Pdx1ZNKP_diaphragm_met_het_2


title: X1944273_pcZNKP_lung_mass


title: X1944272_PcZNKP_gastric_mass_cko


title: X1922502_PcZNKP_gastric_mass_het


title: X1936646_pcZNKP_lung_mass


title: X191078_VilCreerZNKP_small_intestine_mass_wt_2


title: X1926282_VilCreerZNKP_small_intestine_mass_het


title: X1936645_PcZNKP_gastric_mass_het


title: Patient 9 ROI 10 DSP9_10_CK

city: Lausanne

first: François

last: Kuonen

address: Av. de Beaumont 29

country: Switzerland

institute: CHUV

hyb_protocol: Immunoflourecent tissue profiling was performed using PanCK (488) cytokeratin, and Syto13 for DNA staining to enable selection of 48 regions of interest (ROIs), divided into 48 tumor (panCK-positive) and 47 stromal (panCK-negative) areas of interest (AOIs) for targeted spatial transcriptomics using the commercial Nanostring GeoMx.

scan_protocol: Nanostring GeoMx Digital Spatial Profiling (DSP) Platform

data_processing: Sequencing libraries were generated by PCR from the photo-released indexing oligos and AOI-specific Illumina adapter sequences. Unique i5 and i7 sample indexes were added. Each PCR reaction used 4 μl of indexing oligonucleotides, 4 μl of indexing PCR primers, 2 μl of Nanostring 5X PCR Master Mix. Thermocycling conditions were 37 °C for 30 min, 50 °C for 10 min, 95 °C for 3 min; 18 cycles of 95 °C for 15 s, 65 °C for 1 min, 68 °C for 30 s; and 68 °C for 5 min. PCR reactions were pooled and purified twice using AMPure XP beads (Beckman Coulter, A63881), according to the manufacturer’s protocol. Pooled libraries were paired-sequenced at 2 × 27 base pairs and with unique dual indexes workflow on an Illumina HiSeq 4000 instrument. HiSeq-derived FASTQ files for each sample were compiled for each compartment using the bcl2fastq program of Illumina, and then demultiplexed and converted to digital count conversion (DCC) files using the GeoMx DnD pipeline (v.1) of Nanostring according to manufacturer’s pipeline. DCC files were imported back into the GeoMx DSP instrument for QC and data analyses using GeoMx DSP analysis suite version 2.2.0.111 (Nanostring). A minimum of 10,000 reads were required for each sample. Probes were checked for outlier status by implementing a global Grubb’s outlier test with alpha set to 0.01. The counts for all remaining probes for a given target were then collapsed into a single metric by taking the geometric mean of probe counts. For each sample, an RNA-probe-pool-specific negative probe normalization factor was generated on the basis of the geometric mean of negative probes in each pool. To ensure good data quality, we calculated the 75th percentile of the gene counts (that is, geometric mean across all non-outlier probes for a given gene) for each AOI, and normalized to the geometric mean of the 75th percentile across all AOIs to give the upper quartile (Q3) normalization factors for each AOI. The distribution of these Q3 normalization factors were then checked for outliers.

label: NanoString

source_name: Basal cell carcinoma

label_protocol: The spatial transcriptomic panel included 1812 genes targeted cancer transcriptome atlas panel to encompass a comprehensive detection of tumor-host interactions.

growth_protocol: FFPE blocks

extract_protocol: For GeoMx DSP sample collections, entire slides were imaged at 20X magnification and morphologic markers were used to select ROI using organic shapes. Automatic segmentation of ROI based on panCKpos markers were used to define AOIs, allowing to separate tumor cells (panCKpos) from adjacent stromal cells (panCKneg). A total of 95 AOIs were exposed to 385 nm light (UV), releasing the indexing oligonucleotides which were collected with a microcapillary and deposited into a 96-well plate for subsequent processing. The indexing oligonucleotides were dried overnight and resuspended in 10 μl of DEPC-treated water.

treatment_protocol: For slide preparation, we followed the GeoMx DSP slide preparation user manual (MAN-10087-04). In brief, FFPE tissue section of 5-µm were mounted on positively charged histology slides by grouping 3 patients per slide. Sections were incubated at 65 °C for 1 hour. Slides were deparaffinized in 3 xylene baths of 5 min, then rehydrated in ethanol gradient: 2 baths of 5 min in 100% EtOH, followed by 5 min in 95% EtOH. Slides were then washed with PBS 1X. Antigen retrieval was performed in Tris-EDTA pH 9.0 at 100°C for 15 min at low pressure. Slides were first dived into hot water for 10 seconds, and then into Tris-EDTA buffer. Cooker vent stayed open during the procedure to ensure low pressure. Slides were then washed in PBS 1X, incubated in proteinase K in PBS (1ug/ml) for 15 min at 37°C and washed again with PBS 1X. Tissues were post-fixed in 10% neutral-buffered formalin (NBF) for 5 min, washed twice for 5 min in NBF stop buffer (0.1M Tris Base, 0.1M Glycine) and finally once in PBS 1X. The RNA probe mix (CTA (https://www.nanostring.com/products/geomx-digital-spatial-profiler/geomx-rna-assays/geomx-cancer-transcriptome-atlas/), a pool of in situ hybridization probes with UV photocleavable oligonucleotide barcodes) was placed on each section and covered with HybriSlip Hybridization Covers. Slides were then incubated for hybridization overnight at 37°C in a Hyb EZ II hybridization oven (Advanced Cell Diagnostics). The day after, HybriSlip covers were gently removed and 25-min stringent washes were performed twice in 50% formamide and 2X Saline Sodium Citrate (SSC) at 37 °C. Tissues were washed for 5 min in 2X SSC, then blocked in Buffer W (Nanostring Technologies) for 30 min at room temperature in a humidity chamber. Next, tissues were stained with panCK-532 (clone AE1+AE3) (Novus, NBP2-33200) at 1:20 and SYTO 13 at 1:10 (Thermo Scientific S7575) in Buffer W for 1 h at room temperature. Slides were washed twice in fresh 2X SSC then loaded on the GeoMx Digital Spatial Profiler (DSP).

tag: tumor type

value: Basal cell carcinoma

tag: histology

value: Infiltrative

tag: compartment

value: Tumor


title: Patient 9 ROI 09 DSP9_09_Stroma

value: Stroma


title: X191078_VilCreerZNKP_small_intestine_mass_wt

tag: tissue

value: primary tumour

value: female


title: X191078_VilCreerZNKP_fat_pad_met_wt

value: metastasis


title: Patient 9 ROI 09 DSP9_09_CK


title: X185515_VilCreerZNKP_small_intestine_mass_cko

value: male


title: Patient 9 ROI 08 DSP9_08_CK


title: Patient 9 ROI 07 DSP9_07_Stroma


title: X191078_VilCreerZNKP_mesentery_met_wt

value: mesentery

value: Nalcn-wt


title: Patient 9 ROI 07 DSP9_07_CK


title: Patient 9 ROI 6 DSP9_06_CK


title: Patient 9 ROI 06 DSP9_06_Stroma


title: Patient 8 ROI 12 DSP8_12_Stroma


title: Patient 8 ROI 11 DSP8_11_CK


title: X1823481_VilCreerZNKP_small_intestine_mass_cko_2

value: Nalcn-null


title: X1820592_VilCreerZNKP_small_intestine_mass_cko


title: Patient 9 ROI 10 DSP9_10_Stroma


title: Patient 9 ROI 08 DSP9_08_Stroma


title: Patient 8 ROI 11 DSP8_11_Stroma


title: Patient 7 ROI 05 DSP7_05_Stroma


title: X181984_PcZNKP_liver_met_cko

source_name: liver

value: liver


title: X181984_PcZNKP_liver_met_cko_2


title: Patient 7 ROI 05 DSP7_05_CK


title: Patient 8 ROI 12 DSP8_12_CK


title: Patient 7 ROI 04 DSP7_04_Stroma


title: Patient 7 RO 04 DSP7_04_CK


title: X181984_PcZNKP_gastric_mass_cko_2

value: gastric


title: X181984_PcZNKP_mesentery_met_cko


title: X181984_PcZNKP_lung_met_cko


title: Patient 7 ROI 03 DSP7_03_Stroma


title: X181984_PcZNKP_gastric_mass_cko


title: Patient 7 ROI 03 DSP7_03_CK


title: Patient 7 RO 02 DSP7_02_Stroma


title: X1814829_VilCreerZNKP_small_intestine_mass_cko_2

value: small intestine


title: X1814826_VilCreerZNKP_small_intestine_mass_wt_2


title: X181984_PcZNKP_diaphragm_met_cko


title: X1814829_VilCreerZNKP_small_intestine_mass_cko


title: X181984_PcZNKP_fat_pad_met_cko


title: Patient 7 ROI 01 DSP7_01_CK


title: Patient 7 ROI 01 DSP7_01_Stroma


title: X1814824_VilCreerZNKP_small_intestine_mass_het

value: Nalcn-het


title: X1814824_VilCreerZNKP_mesentery_met_het


title: Patient 5 ROI 05 DSP5_05_CK

value: Nodular


title: Patient 5 ROI 06 DSP5_06_CK


title: Patient 5 ROI 06 DSP5_06_stroma


title: X1814824_VilCreerZNKP_fat_pad_met_het

value: fat pad


title: X1813484_VilCreerZNKP_small_intestine_mass_het_2


title: X181341_PcZNKP_lymph_node_met_het

value: lymph node


title: Patient 4 ROI 04 DSP4_04_stroma

value: Basal cell carcinoma


title: X181341_PcZNKP_diaphragm_met_het

value: diaphragm


title: X1810557_VilCreerZNKP_small_intestine_mass_cko_2


title: X181341_PcZNKP_gastric_mass_het


title: X1751756_VilCreerZNKP_small_intestine_mass_het_2


title: Patient 4 ROI 03 DSP4_03_stroma

value: Stroma


title: Patient 4 ROI 03 DSP4_03_CK

value: Tumor


title: X1749550_pcZNKP_prostatesv_mass

source_name: prostate

value: prostate

value: primary tumour


title: X1735539_PcZNKP_kidney_facia_met_cko

source_name: kidney facia

value: kidney facia

value: metastasis

value: male


title: X1735539_PcZNKP_mesentery_met_cko

value: mesentery

value: Nalcn-null


title: Patient 7 ROI 02 DSP7_02_CK

value: Infiltrative


title: X1735539_PcZNKP_gastric_mass_cko_2

value: gastric


title: Patient 5 ROI 05 DSP5_05_stroma


title: Patient 4 ROI 01 DSP4_01_stroma


title: Patient 4 ROI 02 DSP4_02_stroma


title: X1732658_pcznptp_lung_mass

value: lung


title: Patient 4 ROI 01 DSP4_01_CK

value: Nodular


title: Patient 6 RIO 12 DSP6_12_troma


title: Patient 4 ROI 02 DSP4_02_CK


title: Patient 4 ROI 04 DSP4_04_CK


title: Patient 6 ROI 12 DSP6_12_CK


title: Patient 6 ROI 11 DSP6_11_stroma


title: Patient 6 ROI 11 DSP6_11-CK


title: Patient 6 ROI 10 DSP6_10_stroma


title: X1731200_PcZNKP_lung_met_cko


title: X172710_PcZNKP_gastric_mass_wt

value: Nalcn-wt


title: Patient 6 ROI 09 DSP6_09_stroma


title: Patient 6 ROI 10 DSP6_10_CK


title: X1727092_VilCreerZNKP_Small_intestine_mass_wt

value: small intestine


title: X1735539_PcZNKP_fat_pad_met_cko

value: fat pad


title: X172565_pcznptp_liver_noidule

value: liver


title: X1731597_pcznptp_liver_nod_2


title: fibroblasts from normal scar_group2

city: Daejeon

first: Jennifer

middle: Hyunjong

last: Shin

state: Yuseong-gu

address: 291 Daehak-ro

institute: KAIST

data_processing:
Illumina Casava1.8 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence (using fastx_trimmer), then mapped to hg19 whole genome using Bowtie2.
Read count extraction and normalization were performed using edgeR.
Assembly: hg19
Supplementary files format and content: *.txt: Tab-delimited text files include read count values for each Sample.

source_name: Human Skin

growth_protocol: Human fibroblasts were taken from patients and cultured on T75 flask. Cells were expanded for 2 days in Dulbecco’s Modified Eagle’s medium (DMEM, Lonza) supplemented with 10% fetal bovine serum (FBS, Lonza) and 1% penicillin-streptomycin (PS, Invitrogen).

extract_protocol:
RNA was harvested using Trizol reagent (Invitrogen). 500 ng of total RNA was used for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using LEXOGEN Quant-Seq Library Prep Kit (Cat#001.24) standard protocols.

tag: sample type

value: normal scar

value: skin fibroblasts

tag: passages


title: fibroblasts from hypertrophic scar_group3

value: hypertrophic scar


title: fibroblasts from keloid scar_group2

value: keloid scar


title: X1717399_pcznptp_liver_nod

tag: strain

value: primary tumour

value: female


title: X1713337_pcZNKP_epididymis_mass

source_name: epididymis

value: epididymis

value: male


title: fibroblasts from keloid scar_group1


title: Patient 6 ROI 08 DSP6_08_CK

value: Basal cell carcinoma

value: Tumor


title: Patient 6 ROI 07 DSP6_07_stroma

value: Stroma


title: X1718092_PcZNKP_lung_met_wt

value: lung

value: metastasis


title: Patient 6 ROI 09 DSP6_09_CK

value: Nodular


title: Patient 6 ROI 07 DSP6_07_CK


title: Patient 3 ROI 12 DSP3_12_stroma


title: fibroblasts from keloid scar_group3


title: fibroblasts from normal scar_group3


title: Patient 6 ROI 08 DSP6_08_stroma


title: X1713666_PcZNKP_lung_met_wt

value: Nalcn-wt


title: LNCaP cells, siXRN2_2

city: Rome

first: Marco

last: Pieraccioli

address: Largo Francesco Vito 1

country: Italy

institute: Catholic University of the Sacred Heart rome

data_processing:
Adapter trimming from raw 3′READS performed using Cutadapt.
The trimmed reads were aligned to hg19 using bowtie2.
Reads with a mapping quality score (MAPQ) ≥10 were kept for further analysis.
Reads with ≥2 non-genomic 5’Ts after alignment were called polyA site supporting (PASS) reads.
pAs within 24 nt of each other were clustered.
pAs with less than 5 PASS reads in all the samples were first trimmed.
Assembly: hg19
Supplementary files format and content: xlsx file with raw isoform abundance, change of APA isoform abundance and significance score (APA), and RPM, gene expression log2FC and significance score (GE) for every sample.

source_name: LNCaP prostate cancer cell line

growth_protocol: LNCaP cells were maintained in RPMI 1640 (Euroclone) supplemented with 10% fetal bovine serum (GIBCO), 4500 mg/L glucose (Sigma-Aldrich), 10 mM Hepes (Eurocolone), 1mM sodium pyruvate (Euroclone), 100 U/mL penicillin (Euroclone) and 100 μg/mL streptomycin (Euroclone).

extract_protocol: TRIZOL. Poly(A)+ RNA in 0.115 g of total RNA was captured using 12 L of oligo(dT)25 magnetic beads (NEB) in 200 L 1_ binding buffer (10 mM Tris-Cl, pH 7.5, 150 mM NaCl, 1 mM EDTA, and 0.05% TWEEN 20) and fragmented on the beads using 1.5 U of RNase III (NEB) in 30 L RNase III buffer (10 mM Tris-Cl, pH 8.3, 60 mM NaCl, 10 mM MgCl2, and 1 mM DTT) at 37C for 15 min. After washing away unbound RNA fragments with binding buffer, poly(A)+ fragments were eluted from the beads with TE buffer (10 mM Tris-Cl, 1 mM EDTA, pH 7.5) and precipitated with ethanol, followed by ligation to 3 pmol of heat-denatured 5_ adapter (5_-CCUUGGCACCCGAGAAUUCCANNNN, Sigma) in the presence of 1 mM ATP, 0.1 L of SuperaseIn (Life Technologies), and 0.25 L of T4 RNA ligase 1 (NEB) in a 5 L reaction at 22C for 1 h. The ligation products were captured by 10 pmol of biotin-T15-(+TT)5 attached to 12 L of Dynabeads MyOne Streptavidin C1 (Life Technologies). After washing with washing buffer (10 mM Tris-Cl, pH 7.5, 1 mM NaCl, 1 mM EDTA, and 0.05% TWEEN 20), RNA fragments on the beads were incubated with 0.01 U/L of RNase H (Epicentre) at 37C for 30 min in 30 L of RNase H buffer (50 mM Tris-Cl, pH 7.5, 5 mM NaCl, 10 mM MgCl2, and 10 mM DTT). After washing with RNase H buffer, RNA fragments were eluted from the beads in elution buffer (1 mM NaCl, 1 mM EDTA, and 0.05% TWEEN 20) at 50C, precipitated with ethanol, and then ligated to 3 pmol of heat-denatured 5_ adenylated 3_ adapter (5_-rApp/NNNGATCGTCGGACTGTAGAACTCTGAAC/3ddC) with 0.25 L T4 RNA ligase 2 (truncated KQ version, NEB) at 22C for 1 h in a 5 L reaction containing 15% PEG 8000 (NEB) and 0.2 L of SuperaseIn (Life Technologies). The ligation products were then precipitated and reverse transcribed using M-MLV reverse transcriptase (Promega), followed by PCR amplification using Phusion high-fidelity DNA polymerase (NEB) and bar-coded PCR primers for 1218 cycles (12 cycles for 15 g input RNA, 13 cycles for 5 g input, 15 cycles for 1 g input, and 18 cycles for inputs <1 g). Both RT primer (5_-GTTCAGAGTTCTACAGTCCGACGATC) and PCR primers with indexes were described previously in Hoque et al. (2013). PCR products were size-selected twice with AMPure XP beads (Beckman Coulter), using 0.6 volumes of beads (relative to the PCR reaction volume) to remove large DNA molecules and an additional 0.4 volumes of beads to remove small DNA molecules. The eluted DNA was selected again with 1 volume of AMPure XP beads to further remove small DNA molecules. The size and quantity of the libraries eluted from the AMPure beads were examined using a high sensitivity DNA kit on an Agilent Bioanalyzer (Agilent). The library concentrations were further measured by qPCR using primers corresponding to 5_ and 3_ end regions of cDNAs.

treatment_protocol: For Sam68 and XRN2 RNAi experiments, LNCaP cells were silenced twice with 50 nM siRNAs using Lipofectamine RNAiMax (Thermo Fisher Scientific, Invitrogen) for 48 hours.

value: LNCaP

value: prostate cancer

tag: treatment

value: silenced with siXRN2 for 48 hours replicate 2


title: LNCaP cells, siCTRL_2

tag: cell type

value: silenced with siSCR for 48 hours replicate 2


title: LNCaP cells, siSAM68_1

value: silenced with siSAM68 for 48 hours replicate 1


title: LNCaP cells, siSAM68_2

value: silenced with siSAM68 for 48 hours replicate 2


title: fibroblasts from hypertrophic scar_group2

value: hypertrophic scar

value: skin fibroblasts


title: small intestine tumour, Nalcn-wt, rep3

description: 10x Genomics

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were made using the Cell Ranger software v2.1.1 (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger)
Raw RNA reads were processed with cellranger using mm10 from 10X as the reference genome to create filtered gene expression matrices. Cell barcodes detected by cellranger were used as input to CITESeq for hashtagged sequence data (solid organs) generating a counts matrix with cell barcodes and hashtag oligo sequences per cell.
The HTODemux function from Seurat was then used to identify clusters and classify cells according to their barcodes including negative and doublet cells.
Quality control metrics were generated using Scater followed by single cell object conversion to Seurat objects, merging of objects and then analyses run using the standard Seurat pipeline.
Assembly: mm10
Supplementary files format and content: Tab-separated values files and matrix files

source_name: small intestine tumour

extract_protocol:
Single-cell RNA-seq libraries were prepared using Chromium Single Cell 3′ Library & Gel Bead Kit v3, Chromium Chip B Kit and Chromium Single Cell 3' Reagent Kits v3 User Guide (Manual Part CG000183 Rev A; 10X Genomics). Cell suspensions were loaded on the Chromium instrument with the expectation of collecting gel-beads emulsions containing single cells.
RNA from the barcoded cells for each sample was subsequently reverse-transcribed in a C1000 Touch Thermal cycler (Bio-Rad) and all subsequent steps to generate single-cell libraries were performed according to the manufacturer’s protocol with no modifications (for most of the samples 12 cycles was used for cDNA amplification, 16 for samples with very low cell concentration).

value: sorted ZSG+ tumour cells

value: small intestine tumour

tag: age

value: 214 days

value: female


title: Patient 3 ROI 12 DSP3_12_CK

value: Basal cell carcinoma

value: Tumor


title: LNCaP cells, siCTRL_1

value: silenced with siSCR for 48 hours replicate1


title: Patient 3 ROI 11 DSP3_11_stroma

value: Nodular

value: Stroma


title: fibroblasts from hypertrophic scar_group1


title: muscle metastasis, Nalcn-het, rep2

description: 10x Genomics

source_name: skeletal muscle metastasis

value: skeletal muscle metastasis

value: 124 days

value: male


title: fibroblasts from normal scar_group1

value: normal scar


title: LNCaP cells, siXRN2_1

value: LNCaP

value: prostate cancer

value: silenced with siXRN2 for 48 hours replicate 1


title: IR-R MCF-7 cells_1

last: Huo

address: 27 Taiping Rd

institute: Beijing Institute of biotechnology

data_processing:
Quality control: Raw data (raw reads) of fastq 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.
Reads mapping to the reference genome: 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. We selected Hisat2 as the mapping tool for that Hisat2 can generate a database of splice junctions based on the gene model annotation file and thus a better mapping result than other non-splice mapping tools.
Novel transcripts prediction: The mapped reads of each sample were assembled by StringTie (v1.3.3b) (Mihaela Pertea.et al. 2015) in a reference-based approach. StringTie uses a novel network flow algorithm as well as an optional de novo assembly step to assemble and quantitate fulllength transcripts representing multiple splice variants for each gene locus.
Quantification of gene expression level feature: Counts v1.5.0-p3 was used to count the reads numbers mapped to each gene. And then FPKM of each gene was calculated based on the length of the gene and reads count mapped to this gene. FPKM, expected number of Fragments Per Kilobase of transcript sequence per Millions base pairs sequenced, considers the effect of sequencing depth and gene length for the reads count at the same time, and is currently the most commonly used method for estimating gene expression levels.
Differential expression analysis: (For DESeq2 with biological replicates) Differential expression analysis of two conditions/groups (two biological replicates per condition) was performed using the DESeq2R package (1.20.0). DESeq2 provide statistical routinesfor determining differential expression in digital gene expression data using a model based on the negative binomial distribution. The resulting P-values were adjusted using the Benjamini and Hochberg’s approach for controlling the false discovery rate . Genes with an adjusted P-value <0.05 found by DESeq2 were assigned as differentially expressed.
Assembly: mm10
Supplementary files format and content: Tab-separated values files and matrix files

extract_protocol:
RNA was harvested using Rneasy mini plus kit (Qiagen). 1.3 ug of total RNA was used for the construction of sequencing libraries.
RNA libraries for RNA-seq were prepared using SMARTER mRNA-Seq Library Prep Kit following manufacturer's protocols.

value: breast cancer cells

tag: genotype

value: IR-R

value: IR-treatment


title: IR-S MCF-7 cells_2

value: IR-S

value: non-treatment


title: IR-S MCF-7 cells_1


title: IR-R MCF-7 cells_2


title: Patient 3 ROI 10 DSP3_10_stroma

value: Basal cell carcinoma


title: small intestine tumour, Nalcn-het,rep2

description: 10x Genomics

value: sorted ZSG+ tumour cells

value: small intestine tumour

value: 214 days

tag: Sex


title: Patient 3 ROI 10 DSP3_10_CK

value: Nodular

value: Tumor


title: 46XX_1

city: Xiamen

last: Miao

state: Fujian

address: School of Medicine and School of Biomedical Sciences, Huaqiao University, 668 Jimei Road, Xiamen, Fujian

institute: Huaqiao University

library_source: genomic

data_processing:
we use FastQC to perform basic statistics on the quality of the clean data reads.Bismark software (version 0.16.3;) was used to perform alignments of bisulfite-treated reads to a reference genome (-X 700 --dovetail),Differentially methylated regions (DMRs) were identified using the DSS software
ATAC-seq data, sequencing adapters and poor quality bases were trimmed from the sequencing reads by using Skewer. Reads were mapped to the hg38 reference genome using Burrows-Wheeler Transform (BWA) (parameters -T 25 -k 18). Coverage tracks were computed as fragments per million per base pair (FPM) using deepTools bamCoverage (parameters -bs 10 –normalized Using CPM –extend Reads –ignore Duplicates).Peak-calling was performed by using Genrich (https://github.com/jsh58/Genrich) to analyze replicates
Genome_build: hg38
Supplementary_files_format_and_content: bam,bed and bigwig
Supplementary_files_format_and_content: KS_vs_NF.CG.DMR.bed: 47XXY vs 46XX group comparison
Supplementary_files_format_and_content: KS_vs_NM.CG.DMR.bed: 47XXY vs 46XY group comparison
Supplementary_files_format_and_content: NM_vs_NF.CG.DMR.bed: 46XY vs 46XX group comparison

source_name: 46XX group_amniotic fluid cells

extract_protocol:
DNA were extract from frozen blood, and A number of cells (50,000 cells) were centrifuged for 5 minutes (at 500 × g, 4°C) and suspended in cold PBS.
WGBS and ATAC libraries were prepared for sequencing using standard Illumina protocols

tag: tissue/cell type

value: amniotic fluid cells

tag: molecule subtype

value: TN5 input DNA


title: 46XX_2


title: 46XY_1

source_name: 46XY group_amniotic fluid cells


title: 46XY_2


title: mesentery metastatsis,Nalcn-het,rep1

description: 10x Genomics

source_name: mesentery metastasis of intestinal adenocarcinoma

value: male


title: 47XXY_2

source_name: 47XXY group_amniotic fluid cells


title: 47XXY_1


title: tCZC, Nalcn-het, rep2

description: 10x Genomics

source_name: sorted ZSG+ cells from peripheral blood

value: circulating tumour cells

value: sorted ZSG+ cells from peripheral blood

value: 124 days


title: PBMCx,Nalcn-wt, rep1

description: 10x Genomics

source_name: PBMCs

value: PBMCs

value: 120 days


title: Patient 2 ROI 06 DSP2_06_stroma

value: Stroma


title: Patient 2 ROI 06 DSP2_06_CK


title: NF_3

source_name: 46XX group_blood

value: blood


title: NF_2


title: Patient 3 ROI 11 DSP3_11_CK


title: NF_1


title: gastric tumour, Nalcn-het, rep 1

description: 10x Genomics

source_name: gastric tumour

value: gastric tumour


title: NM_3

source_name: 46XY group_blood


title: Patient 2 ROI 05 DSP2_05_stroma

source_name: Basal cell carcinoma

value: Basal cell carcinoma


title: Patient 2 ROI 05 DSP2_05_CK


title: KS_3

source_name: 47XXY group_blood


title: lymph node metastasis, Nalcn-wt, rep1

description: 10x Genomics

source_name: lymph node metastasis

value: sorted ZSG+ tumour cells

value: lymph node metastasis

value: 658 days

value: female


title: NM_2


title: tCZC, Nalcn-wt, rep1

description: 10x Genomics


title: mesentery metastatsis,Nalcn-het,rep2

description: 10x Genomics

source_name: mesentery metastasis of gastric adenocarcinoma


title: NM_1


title: KS_2


title: Patient 2 ROI 04 DSP2_04_stroma

value: Nodular


title: KS_1


title: JFH1_infected_5dpi_Huh7.5.1

city: Aurora

first: Daphne

middle: A.

last: Cooper

state: CO

department: Department of Microbiology

institute: University of Colorado

description: 2', 3'-cyclic phosphate cDNA library derived from JFH1-infected Huh7.5.1 cells at 5 dpi

data_processing:
Reads were trimmed for residual Illumina cDNA linker using Cutadapt.
Fastq datasets were trimmed for 8-base unique molecular identifier using umitools (github.com/brwnj/umitools).
Aligned to JFH1 and cellular RNA sequences using Bowtie2. UMIs counted using umitools. Alignment data converted to bedgraph format based on genomic (-) or anti-genomic (+) strand.
Aligned data normalized to the total number of aligned reads and depicted as a % of total aligned reads.
The 28S and 18S rRNA signal from uninfected Huh7.5.1 was subtracted from signal in 28S and 18S rRNA (1-5 dpi) (baseline.sample.txt). The 3'-dinucleotide from the read was obtained to characterize RNase specificity, and the % of total aligned reads was plotted for cleavage sites across the genome/sequence.
Genome_build: JFH1 HCV2A: AB047639.1
Genome_build: 45S pre-rRNA: NR_046235.1
Genome_build: 5S rRNA: NR_023371.1
Genome_build: U3 snoRNA: NR_006880.1
Genome_build: hg18 tRNA database (http://gtrnadb.ucsc.edu/Hsapi/hg18-tRNAs.fa)
Genome_build: Homo_sapiens_chr1.trna116-GluCTC
Genome_build: hg18 NCBI Build 36.1
Supplementary_files_format_and_content: Text files for reads [genome, cleavage site -1, cleavage site, # of UMI reads, normalized value (% total reads), 3'-dinucleotide] or bedgraph files. For negative strand HCV reads, data include reference sequence, cleavage site in positive orientation, cleavage site in negative orientation, # of UMI reads, normalized value (% total cDNA reads), and dinucleotide. Data where the normalized signal in 28S and 18S rRNA from uninfected Huh7.5.1 cells was removed from signal in 1-5 dpi samples are referred to as baseline.sample.txt, and include reference sequence, cleavage site -1, cleavage site, and corrected normalized value, and dinucleotide.

source_name: Human Hepatoma

extract_protocol:
Total RNA harvested from lysates using phenol chloroform extraction.
Illumina TruSeq RNA linker with 8-base Unique Molecular Identifier (UMI) was attached with Arabidopsis thaliana tRNA ligase (recognizes RNAs with terminal 2', 3'-cyclic phosphates). cDNA was generated using a DNA primer specific to the Illumina linker, and another Illumina TruSeq-specific DNA linker was attached to the 3'-end of the cDNA. cDNAs were PCR amplified, pooled, and sequenced.
2', 3'-cyclic phosphate cDNA synthesis and Illumina sequencing were used to identify and quantify metal-ion-independent endoribonuclease cleavage sites in host RNAs

treatment_protocol:
Huh7.5.1 cells were infected with HCV2a (JFH1) at an MOI of 0.1 FFU/mL.
Cell lysates were prepared for uninfected and 1-5 days post infection (dpi) samples.

tag: cell line

tag: experimental condition

value: Infected - 5 days post infection


title: Patient 2 ROI 04 DSP2_04_CK

value: Tumor


title: Patient 2 ROI 03 DSP2_03_stroma


title: mesentery metastatsis,Nalcn-wt,rep2

description: 10x Genomics

value: mesentery metastasis of intestinal adenocarcinoma


title: lymph node metastasis, Nalcn-het, rep1

description: 10x Genomics

value: 259 days


title: JFH1_Infected_4dpi_Huh7.5.1

description: 2', 3'-cyclic phosphate cDNA library derived from JFH1-infected Huh7.5.1 cells at 4 dpi

value: Infected - 4 days post infection


title: NASH liver

city: Palo Alto

first: Laetitia

last: Coassolo

address: 300 Pasteur Drive

department: Pathology

institute: Stanford University

data_processing:
Read1 (28bp), Index i7 (10bp), Index i5 (10bp) and Read2 (90bp)
Sequencing results were processed using 10x Genomics' Cell Ranger pipeline (Version 3.1.0).
Seurat was used to perform all downstreatm analysis.
Assembly: mm10
Supplementary files format and content: Matrix table with raw gene counts for every gene and every cells in Chow and NASH group
Supplementary files format and content: Tab separated value files with barcodes of each cells in Chow and NASH group
Supplementary files format and content: Tab separated value files with barcodes and matching genes of each cells in Chow and NASH group

source_name: Mouse primary liver cells

extract_protocol:
Liver cells were isolated from mouse liver tissue by collagenase digestion followed by modified percoll gradient centrifugation method (90% percoll with 100g and 1000g and 25% percoll with 100g and 1000g), which collects all non-parenchymal cells and various density hepatocytes.
The libraries were prepared using the 10X Genomics 3' version 3 single cell gene expression kit.

treatment_protocol: Mice were put on either with NASH (high fructose, high fat and high cholesterol) or chow diet for 6.5 weeks.

value: Liver

value: All liver cell type

value: NASH diet for 6.5 weeks


title: tCZC, Nalcn-het, rep1

description: 10x Genomics

value: circulating tumour cells

value: sorted ZSG+ cells from peripheral blood


title: Chow liver

value: Chow diet for 6.5 weeks


title: Patient 2 ROI 03 DSP2_03_CK

value: Basal cell carcinoma


title: Patient 2 ROI 02 DSP2_02_stroma

value: Stroma


title: Patient 2 ROI 02 DSP2_02_CK


title: small intestine tumour, Nalcn-wt, rep 3

description: 10x Genomics

source_name: small intestine tumour

value: sorted ZSG+ tumour cells

value: small intestine tumour


title: peritoneum metastasis, Nalcn-het, rep1

description: 10x Genomics

source_name: peritoneum metastasis

value: peritoneum metastasis

value: female


title: JR

city: Dallas

first: Lei

last: Guo

state: TX

address: 5323 Harry Hines Blvd

department: Department of Population and Data Sciences

institute: University of Texas Southwestern Medical Center

data_processing:
Fastq files were aligned to the human hg19 genome using STAR v2.7.3a
Gene expression was quantified using featureCounts in Subread v 1.6.3
Assembly: hg19
Supplementary files format and content: raw counts

source_name: JR

growth_protocol: RMS cells were maintained in DMEM supplemented with 10% fetal bovine serum (FBS), 1% Glutamine, and 1% penicillin/streptomycin at 5% CO2 at 37C. HSMMs were maintained in SkGM-2 medium (Lonza) at 5% CO2 at 37C.

extract_protocol:
Done using commercial reagents e.g. Qiagen, Invitrogen
Samples are run on the Agilent 2100 Bioanalyzer to determine level of degradation thus ensuring only high quality RNA is used (RIN Score 8 or higher). The Qubit fluorometer is used to determine the concentration prior to staring library prep. 4 µg of total DNAse treated RNA are then prepared with the TruSeq Stranded Total RNA LT Sample Prep Kit from Illumina. Poly-A RNA is purified and fragmented before strand specific cDNA synthesis. cDNA are then a-tailed and indexed adapters are ligated. After adapter ligation, samples are PCR amplified and purified with Ampure XP beads, then validated again on the Agilent 2100 Bioanalyzer. Samples are quantified by Qubit before being normalized and pooled, then run on the Illumina HiSeq 2500 using SBS v3 reagents.

treatment_protocol: For differentiation, HSMMs were maintained in differentiation medium containing DMEM with 2% horse serum and 10ug/ml insulin.

value: JR

value: RMS cell line

value: WT

tag: time


title: JFH1_infected_3dpi_Huh7.5.1

description: 2', 3'-cyclic phosphate cDNA library derived from JFH1-infected Huh7.5.1 cells at 3 dpi

value: Infected - 3 days post infection


title: JFH1_infected_2dpi_Huh7.5.1

description: 2', 3'-cyclic phosphate cDNA library derived from JFH1-infected Huh7.5.1 cells at 2 dpi

value: Infected - 2 days post infection


title: small intestine tumour, Nalcn-het, rep1

description: 10x Genomics

value: 259 days


title: RD-L

source_name: RD-L

value: RD-L


title: pancreas metastasis, Nalcn-het, rep1

description: 10x Genomics

source_name: pancreas metastasis

value: pancreas metastasis


title: Human Skeletal Muscle Myoblasts (HSMM)

description: D0_ATCACG_L001 and D0_ATCACG_L002

source_name: HSMM

value: HSMM

value: Skeletal muscle myoblasts

value: Growth medium

value: Day 0


title: RH18


title: Patient 2 ROI 01 DSP2_01_stroma

tag: tumor type

value: Basal cell carcinoma

value: Nodular

value: Stroma


title: RH28


title: Uninfected_Huh7.5.1

description: 2', 3'-cyclic phosphate cDNA library derived from uninfected Huh7.5.1 cells

value: Uninfected


title: RH30


title: Patient 2 ROI 01 DSP2_01_CK

tag: histology

value: Tumor


title: mesentery metastatsis,Nalcn-wt,rep1

description: 10x Genomics

source_name: mesentery metastasis of intestinal adenocarcinoma

value: sorted ZSG+ tumour cells

value: 132 days

value: female


title: Patient 1 ROI 09 DSP1_09_stroma

tag: compartment


title: small intestine tumour, Nalcn-wt, rep2

description: 10x Genomics

tag: tissue

value: small intestine tumour


title: Patient 1 ROI 08 DSP1_08_stroma


title: hashtagged tissue samples, Nalcn-wt, rep 5

description: 10x Genomics

source_name: uterus, kidney, liver, lung, stomach

value: mixed cell types

value: uterus, kidney, liver, lung, stomach

value: 654 days


title: JFH1_Infected_1dpi_Huh7.5.1

description: 2', 3'-cyclic phosphate cDNA library derived from JFH1-infected Huh7.5.1 cells at 1 dpi

value: Infected - 1 day post infection


title: WAC_PolII

city: Shenzhen

first: Peitao

last: Xu

address: No.3025, Shennan Middle Street

institute: The Eighth Affiliated Hospital of Sun Yat-sen University

data_processing:
Raw reads were filtered and trimmed to remove sequencing adaptors and low-quality reads
Clean reads were mapped to the human genome version hg38 using Bowtie2 (v2.5.1)
Peaks were called using MACS2 (v.2.1.1) and normalization (reads per kilobase per million or RPKM) of the CUT&Tag-seq data and heatmap plotting were performed using deepTools (v2.3.6.0)
The normalized CUT&Tag-seq data were visualized on Integrative Genomics Viewer (IGV)
Assembly: Grch38
Supplementary files format and content: bw files were generated by deepTools (v2.3.6.0)
Library strategy: CUT&Tag

source_name: Mesenchymal stem cells

extract_protocol:
The DNA fragments were extracted using phenol chloroform.
The CUT&Tag assay was performed using NovoNGS® CUT&Tag 2.0 High-Sensitivity Kit (for Illumina®) (Novoprotein Scientific Inc., Cat# N259-YH01-01A) according to the manufacturer’s instructions. Briefly, cells were harvested with trypsin and enriched by ConA-magnetic beads. A total of 50,000 cells were re-suspended and washed twice with 100 μL Dig-wash Buffer (20 mM HEPES pH 7.5; 150 mM NaCl; 0.5 mM spermidine; 1× protease inhibitor cocktail; 0.05% digitonin). The samples were incubated with primary BRD4 antibody (1:100, 4°C, 18 h) and secondary antibody (1:200, 25°C, 1 h). After incubation, the beads were washed three times in Dig-Hisalt Buffer (0.05% digitonin, 20 mM HEPES, pH 7.5, 300 mM NaCl, 0.5 mM spermidine, 1× protease inhibitor cocktail). Cells were incubated with the protein A-Tn5 transposome at 25°C for 1 h and washed three times in Dig-Hisalt buffer to remove unbound protein A-Tn5. The cells were re-suspended in 50 μL Tagmentation buffer (10 mM MgCl2 in Dig-Hisalt Buffer) and incubated at 37°C for 1 h. The tagmentation was terminated with the addition of 1 μL of 10% SDS at 55°C for 10 min.
Cleavage under targets and tagmentation (CUT&Tag)

value: Mesenchymal stem cells

value: Sh-WAC

tag: target

value: PolII


title: ntCZC,Nalcn-het, rep3

description: 10x Genomics

source_name: ZSG+ sorted peripheral blood

value: non-tumour circulating zsgreen+ cells

value: ZSG+ sorted peripheral blood

value: 649 days


title: WAC_H2Bub1


title: hashtagged tissue samples, Nalcn-wt,rep 4

description: 10x Genomics

value: 611 days


title: RNF40_PolII


title: Patient 1 ROI 07 DSP1_07_stroma

value: Basal cell carcinoma

value: Nodular

value: Stroma


title: Patient 1 ROI 08 DSP1_08_CK

value: Tumor


title: ntCZC,Nalcn-wt rep 3

description: 10x Genomics

value: 608 days

value: male


title: Patient 1 ROI 07 DSP1_07_CK


title: RNF40_H2Bub1


title: muscle metastasis, Nalcn-het, rep1

description: 10x Genomics

source_name: skeletal muscle metastasis

value: sorted ZSG+ tumour cells

value: skeletal muscle metastasis

value: 259 days

value: female


title: NC_PolII

value: Mesenchymal stem cells

value: Sh-NC


title: Patient 1 ROI 09 DSP1_09_CK

source_name: Basal cell carcinoma


title: Patient 11 ROI 07 DSP11_07_Stroma

value: Infiltrative


title: hashtagged tissue samples, Nalcn-wt,rep 3

description: 10x Genomics

source_name: epididymis, kidney, liver, lung, stomach

value: mixed cell types


title: ntCZC,Nalcn-null, rep4

description: 10x Genomics

value: non-tumour circulating zsgreen+ cells

value: ZSG+ sorted peripheral blood

value: 613 days


title: Patient 11 ROI 06 DSP11_06_Stroma

value: Basal cell carcinoma


title: RD

source_name: RD

value: RD

value: RMS cell line

value: WT


title: Patient 11 ROI 07 DSP11_07_CK

value: Tumor


title: NC_H2Bub1


title: ntCZC,Nalcn-null, rep3

description: 10x Genomics

value: 612 days


title: hashtagged tissue samples, Nalcn-null, rep 1

description: 10x Genomics

value: uterus, kidney, liver, lung, stomach

value: 647 days


title: Patient 11 ROI 06 DSP11_06_CK


title: Patient 11 ROI 05 DSP11_05_Stroma

value: Stroma


title: hashtagged tissue samples, rep 3

description: 10x Genomics

value: 611 days

value: female


title: Patient 11 ROI 05 DSP11_05_CK


title: Patient 11 RO 04 DSP11_04_CK


title: hashtagged tissue samples, Nalcn-null, rep 2

description: 10x Genomics

value: epididymis, kidney, liver, lung, stomach

value: 552 days

value: male


title: ntCZC,Nalcn-null, rep2

description: 10x Genomics

value: non-tumour circulating zsgreen+ cells


title: ntCZC,Nalcn-null, rep1

description: 10x Genomics

value: ZSG+ sorted peripheral blood

value: 551 days


title: hashtagged tissue samples, Nalcn-het, rep 3

description: 10x Genomics

value: mixed cell types

value: 610 days


title: MADM wildtype; small clones; mouse #6 [19313_SmallClones]

city: Charlottesville

first: Kevin

last: Janes

state: VA

address: Box 800759 Health System

department: Department of Biomedical Engineering

institute: University of Virginia

description: Nextera XT DNA Library Preparation

data_processing:
Adapters were trimmed using fastq-mcf with the following options:  -q 10 -t 0.01 -k 0 (quality threshold 10, 0.01% occurrence frequency, no nucleotide skew causing cycle removal).
Datasets were aligned to the mouse (GRCm38.95) transcriptome using RSEM. RSEM options  --bowtie2 --single-cell-prior (Bowtie2 transcriptome aligner, single-cell prior to account for dropouts)
Assembly: GRCm38.95
Supplementary files format and content: .csv file contains the RSEM estimated abundances for each gene per sample

source_name: Fallopian tube

extract_protocol:
Laser-guided microdissection followed by proteinase K digestion and poly(A) amplification
Nextera XT DNA Library Preparation Kit

value: Fallopian tube

value: Distal region

value: MADM RG;GR alleles, mixed background


title: hashtagged tissue samples, Nalcn-het, rep 2

description: 10x Genomics


title: Patient 11 ROI 03 DSP11_03_CK

value: Basal cell carcinoma

value: Infiltrative

value: Tumor


title: MADM wildtype; large clones; mouse #6 [19313_LargeClones]

description: Nextera XT DNA Library Preparation