title: Stereotaxic brain tumor biopsy, patient 43, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_43

value: brain

value: Diffuse glioma

value: male


title: Illumina-PHD1-KD-UT_R2

status: Public on Jul 23 2026

type: SRA

city: La Tronche

first: Christopher

last: Swale

address: Batiment Jean Roger

department: U1209 - Institute of Advanced Biosciences

country: France

institute: INSERM

description: Library name: Sample 11

library_source: transcriptomic

data_processing:
The quality of the raw sequencing reads was assessed using FastQC (www.bioinformatics. babraham.ac.uk/projects/fastqc/) and MultiQC. For the expression data quantification and normalization, the FASTQ reads were aligned  T. gondii ME49 genome using Subread version 2.0.1 with the following options ‘subread-align -d 50 -D 600 --sortReadsByCoordinates’. Read counts for each gene were calculated using featureCounts from the Subread package.
Assembly: TGME49 version 67
Supplementary files format and content: Count file or normalized TPM values in a .csv format

molecule: polyA RNA

organism: Toxoplasma gondii

source_name: RH

growth_protocol: HFF cells were grown to confluence and infected with RHKU80 GCN5b or PHD1 mAID HA strains. For every library, a T180 plate was infected with 2ml of freshly egressed tachyzoites. At the time of harvest, the cells were scrapped and washed in fresh DPBS.

extract_protocol:
Total RNA was prepared using TRIzol extraction followed by RNAeasy column cleaning (Qiagen).
Messenger RNAs (mRNA) were purified from total RNA using poly-T oligo-attached magnetic beads. After fragmentation, the first strand cDNA was synthesized using random hexamer primers. Then the second strand cDNA was synthesized using dUTP, instead of dTTP. The directional library was ready after end repair, A-tailing, adapter ligation, size selection, USER enzyme digestion, amplification, and purification. The library was checked with Qubit and real-time PCR for quantification and bioanalyzer for size distribution detection.The samples were sequenced on the Illumina NovaSeq platform (2 x 150 bp, strand-specific sequencing) and generated ~40 million paired-end reads for each sample.

treatment_protocol: Cells were left untreated or supplemented with indole-3-acetic acid (IAA).

tag: cell line

value: RH

tag: cell type

value: Intracellular tachyzoites

tag: genotype

tag: treatment

value: untreated sample


title: Skin, TCR+GFP+, Fc-IL-2 mutein 24, TCR

city: South San Francisco

first: Weiwen

last: Deng

address: 750 Gateway Blvd

country: USA

institute: Amgen

description:
Library name: kTr2_TCR
10x Genomics

data_processing:
Raw fastq data of GEX and HTO libraries was demultiplexed using a custom workflow of Cell Ranger (v7.1.0) multi
Demultiplexed bam files were convert to fastq using Cellranger bamtofastq
Converted fastq files of the GEX library as well as raw fastq files of TCR and ADT libraries were aligned on the mouse reference genome mm10 and GRCm38 V(D)J using Cellranger multi. Force-cell value set to the number detected by demultiplexing.
RDS files were generated with output of GEX and ADT libraries using Seurat R package (version 4.3.0)
Assembly: mm10
Assembly: GRCm38 V(D)J
Supplementary files format and content: GEX library: sample.rds files
Supplementary files format and content: ADT library: sample_ADT.rds files (GEX embedded)
Supplementary files format and content: TCR library: sample_HTO_filtered_contig_annotations.csv
Library strategy: CITE-seq

organism: Mus musculus

source_name: Skin

extract_protocol:
Skin cell suspensions were enriched with Percoll density gradient centrifugation using Ficoll-Paque™ PLUS density gradient media gradient. The interphase containing the enriched leukocytes was collected for downstream processing. Before staining, cells were preincubated for 20 min with anti-mouse CD16/32. For sample multiplexing, each individual sample from mice within the same treatment group were incubated with a cocktail of TotalSeq™-C Mouse Universal Cocktail, V1.0 plus one TotalSeq™-C Hashtag antibodies (Hashtags C0306, C0307, C0308, C0309, and C0310). Sample were washed with BD Pharmingen™ Stain Buffer (BSA) and were stained for CD45-BV786, TCRβ -PE and LIVE/DEAD™ Fixable Near-IR Dead Cell Stain Kit. Within each treatment group, samples were pooled and sorted on a FACS Aria into three single cell fractions: CD45+ TCRβ+ GFP+, CD45+ TCRβ + GFP-, CD45+ TCRβ-.
Libraries were prepared according to the manufacter's instructions (Chromium Next GEM Single Cell 5' Reagent Kits v2 (Dual Index) User Guide with Feature Barcode technology for Cell Surface Protein & Immune Receptor Mapping). Briefly, sorted cells were captured in droplet emulsions using Chromium X and libraries were constructed using the 10x Genomics Chromium Next GEM Single Cell 5' Kit v2 and Chromium Next GEM Single Cell 5' Gel Bead Kit v2. Amplified 5' Gene Expression, V(D)J and Cell Surface Protein libraries were profiled using the Agilent 4200 TapeStation System and pooled at 10:1:1 ratio. Pooled samples were sequenced on Illumina NovaSeq X using a 28-10-10-90 asymmetric read configuration. A median sequencing depth of 50,000 reads/cell was targeted for each Gene Expression Library.

tag: tissue

value: Skin

tag: strain

value: Treg

value: Fc-IL-2 mutein 24, s.c.

tag: library

value: VDJ-T


title: M21 cells, CTR, DMSO, 24h, rep1

city: Chengdu

first: Jing

last: Yu

state: Sichuan

address: No.17, Section 3, Renmin South Road

country: China

institute: Sichuan university

description: Library name: A21-1-1

data_processing:
Raw sequencing data underwent standardized quality control using trim_galore (v0.6.4) to remove low-quality bases and adapter sequences. Quality-verified clean reads were subsequently aligned to the reference genome (GRCh38) with HISAT2 (v2.1.0). Resultant SAM files were sorted and converted to BAM format via samtools (v1.15.1), followed by gene expression quantification with featureCounts (v2.0.0) based on Ensembl gene annotations.
Assembly: GRCh38
Supplementary files format and content: Excel fle includes TPM for each Sample

molecule: total RNA

organism: Homo sapiens

source_name: skin

growth_protocol: The human-derived melanoma cell lines M21 and A2058 were maintained in DMEM supplemented with 10% fetal bovine serum (FBS)and antibiotics in humidified atmosphere with 5% CO2 at 37℃

extract_protocol:
1） Samples were ground under liquid nitrogen in a mortar and pestle, and the resulting powder dissolved in TRIzol™ Reagent (Invitrogen, Carlsbad, CA, USA), then extracted total RNA. 2） The quality and purity of RNA were examined by a NanoDrop™ One/OneC spectrophotometer (Thermo Scientific, Waltham, MA, USA) and Life Invitrogen Qubit RNA BR (Broad-Range) Assay Kit. RNA integrity was analyzed using Agilent 4200 TapeStation system (Agilent, Santa Clara, CA, USA).
1）0.1-1 μg of total RNA per sample was prepared for library preparation. The ABclonal Poly(A)mRNA Capture Module and ABclonal Fast RNA-seq Lib Prep Kit V2 were used for mRNA isolation and library construction following the manufacturers’ protocols. 2）As for the quality control of library, Qubit dsDNA HS Assay Kit was used to measure the concentration of library, then Agilent 4200 was used to examine the distribution of segments in library. Finally, library molar concentration was determined using the KAPA Library Quant kit (illumina) universal qPCR Mix. 3）High-throughput transcriptome sequencing was performed on an Illumina NovaSeq xplus platform according to the manufacturer’ s instructions.

value: skin

value: melanoma epithelial cells

value: WT (Control)

value: DMSO


title: BR_H_0_R_R1

city: Bozeman

first: Jennifer

last: Lachowiec

state: MT

address: 119 Plant Biosciences Building

institute: Montana State University

description:
Library name: P12-R1-R
Column name in NormalizedCounts: BR_H_0_R_R1

data_processing:
Read quality was checked with FASTQC and MULTIQC software with an average phred score of 36 (Andrews, 2010). RNA sequence trimming, mapping, and read counting was performed with Trimmomatic v.0.36, STAR, and the Rsubread package v.1.5.2 with function featureCounts, respectively using default settings (Bolger et al., 2014; Dobin et al., 2013; Liao et al., 2014). Reads were aligned using the Triticum aestivum cv. Chinese Spring IWGSC RefSeq v2.1 genome assembly (Zhu et al., 2021). Genes with fewer than 10 total reads across all 123 remaining samples were removed were considered non-expressed. Differential gene expression analysis was performed via DESeq2 (Love et al., 2014). Two samples from different treatment groups were removed from analysis as they clustered far from the other biological replicates based on visual assessment of PCA. DESeq2 was additionally used to normalize read counts by length and read depth for data visualization with pheatmap and ggplot2 (Kolde, 2010; Wickham, 2016).
Assembly: Reads were aligned using the Triticum aestivum cv. Chinese Spring IWGSC RefSeq v2.1 genome assembly (Zhu et al., 2021) and STAR R package using default settings (Dobin et al., 2013)
Supplementary files format and content: NormalizedCounts.csv, inlcudes normalized count by length and read depth for each sample

organism: Triticum aestivum

source_name: Rachis

growth_protocol: Triticum aestivum  cultivar ‘Vida’ (PI 642366) seeds were germinated in petri plates in a total volume of 0.4 mL water per seed for 48 hours in in the dark. The seed were then planted approximately 2.5 cm deep in untreated Sunshine Mix #1 soil (Sun Gro Horticulture Distribution Inc.), with five seeds per pot. Pots were placed in growth chambers with a 16-hour photoperiod with fluorescent and LED bulbs at 27,000 Lux light intensity, kept at 22°C. Plants were watered as needed and fertilized once a week with a 0.3% solution of 20-20-20 Professional Peters fertilizer. Pot placement followed a randomized complete block design.

extract_protocol:
For RNAseq, 64 plants were grown, with four biological replicates per seed treatment, temperature, and time of sampling combinations. Prior to and at 8, 16, and 24 hours after heat exposure began, the unemerged heads were collected. The unemerged wheat heads were removed from the leaf sheath and dissected into rachis and floret tissues, flash frozen,  and stored at -70°C.  Tissue was ground with 3 mm sterilized glass beads in a bead beater, RNA extraction was performed with the RNeasy Plant Mini Kit (Qiagen, Cat #74903) using Beta-mercaptoethanol, and samples were treated with DNase I (Turbo DNA-free Kit, Invitrogen AM1907). Quality of the samples was confirmed via Bioanalyzer using the Agilent RNA 6000 Nano Kit (Agilent, 5067-1511). Five total samples were removed for poor or low RNA quality, leaving a minimum of three biological replications per treatment combination.
RNA-seq libraries were prepared using poly(A)-selected mRNA, and RNA-sequencing was performed by Genewiz.

treatment_protocol: Triticum aestivum cultivar ‘Vida’ (PI 642366) (Lanning et al., 2006) seeds were treated with 1 µM dimethyl sulfoxide (DMSO) control or 1 µM 2,4-epibrassinolide (EBR) during germination in petri plates. At the tetrad stage of growth on the Waddington 7.5 scale (booting), half of the pots from each treatment were moved to a 29° C continuous heat stress. The temperature chosen is the average high temperature in major wheat growing regions in Montana during the typical growing season and was previously determined to induce the canonical heat stress (Kothari et al., 2025)

value: Rachis

value: Vida

value: 2,4 EBR

tag: temperature

tag: time

tag: replication


title: BR_H_24_R_R4

description:
Library name: P11-R4-R
Column name in NormalizedCounts: BR_H_24_R_R4


title: Stereotaxic brain tumor biopsy, patient 41, Glioblastoma

type: genomic

city: Frankfurt

first: Katharina

last: Weber

address: Heinrich-Hoffmann-Straße 7

department: Institute of Neurology

country: Germany

institute: University Hospital Frankfurt

hyb_protocol: Bisulfite-converted DNA was hybridized to Illumina Infinium MethylationEPIC BeadChip (v1) following the manufacturer's standard protocol.

scan_protocol: EPIC array BeadChips were scanned using an iScan system (Illumina, San Diego, California, USA) and raw intensity data (IDAT files) were obtained.

data_processing: IDAT files were uploaded to molecularpathology.org (Heidelberg brain tumor classifier v11b4) to obtain calibrated scores for DNA methylation classes and subclasses, copy number variation profiles and MGMT promoter methylation status. Additional classification was performed using the Bethesda brain tumor classifier (v2) and EpiDiP. IDAT files were processed using R packages RnBeads for quality control and normalization (dasen method from wateRmelon package), and MethylCIBERSORT for reference-based tumor deconvolution. Genome build: hg19.

label: Cy3, Cy5

molecule: genomic DNA

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_41

label_protocol: DNA was further processed and hybridized to the Illumina Infinium MethylationEPIC BeadChip (v1) following protocols provided by the manufacturer.

growth_protocol: Not applicable (FFPE tissue from stereotaxic brain tumor biopsies)

extract_protocol: Selected FFPE tissue samples were transferred into Eppendorf tubes using biopsy punches.

treatment_protocol: Not applicable (tissue samples)

value: brain

tag: disease state

value: Glioblastoma

tag: age

tag: Sex

value: female

tag: sample type

value: FFPE stereotaxic biopsy

tag: dna concentration (ng/ul)


title: Illumina-GCN5b-KD-IAA48h_R3

description: Library name: Sample 09

value: RH

value: Intracellular tachyzoites

value: IAA treated for 48h


title: M21 cells, CTR, 100nM imatinib, 24h, rep3

description: Library name: A21-1+3

value: imatinib, 100nM, 24h


title: BR_H_24_R_R3

description:
Library name: P11-R3-R
Column name in NormalizedCounts: BR_H_24_R_R3


title: Stereotaxic brain tumor biopsy, patient 40, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_40

value: male


title: BR_H_24_R_R2

description:
Library name: P11-R2-R
Column name in NormalizedCounts: BR_H_24_R_R2


title: M21 cells, CTR, 100nM imatinib, 24h, rep2

description: Library name: A21-1+2


title: Illumina-GCN5b-KD-IAA48h_R2

description: Library name: Sample 08


title: Stereotaxic brain tumor biopsy, patient 39, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_39


title: M21 cells, CTR, 100nM imatinib, 24h, rep1

description: Library name: A21-1+1


title: Illumina-GCN5b-KD-IAA48h_R1

description: Library name: Sample 07


title: BR_H_24_R_R1

description:
Library name: P11-R1-R
Column name in NormalizedCounts: BR_H_24_R_R1


title: Illumina-GCN5b-KD-IAA24h_R3

description: Library name: Sample 06

value: IAA treated for 24h


title: BR_L_0_R_R4

description:
Library name: P10-R4-R
Column name in NormalizedCounts: BR_L_0_R_R4


title: Organoid_TNF_pegtarazimod_5

type: RNA

city: Freiburg

first: Geoffroy

last: Andrieux

address: Breisacherstr 153

institute: University clinics Freiburg

hyb_protocol: Labeled fragments were hybridized to Applied Biosystems Clariom S Human Arrays for 16h at 45°C with 60 rpm in an Affymetrix Hybridization Oven 645

scan_protocol: After washing and staining, the arrays were scanned with the Affymetrix GeneChip Scanner 3000 7G. CEL-files were produced from the raw data with Affymetrix GeneChip Command Console Software.

data_processing: CEL files were analysed using the oligo R package. RMA method, log2 transformed were used to normalized the probe intensities.

label: Biotin

source_name: small intestine organoids

label_protocol: We used the Applied Biosystems GeneChip WT PLUS Reagent Kit (catalog no.: 902280 and 902281) for cDNA synthesis, amplification, cRNA synthesis, second-cycle cDNA synthesis, as well as fragmentation and labeling with Biotin according to manufacturer's instructions.

growth_protocol: BALB/c small intestine organoids were cultured in StemCell Technologies IntestiCult organoid growth medium in 50 µl domes of a 1:1 mixture of Corning Matrigel GFR Basement Membrane at 37 °C and 5% CO2.

extract_protocol: RNA was isolated using the Rneasy Mini Kit (Quiagen) according to the manufacturer's instructions.

treatment_protocol: Organoids were challenged with 20 ng/ml recombinant murine TNF and treated with 5 mg/ml pegtarazimod or 2.625 mM PEG as vehicle for 48 h.

value: TNF + pegtarazimod


title: BR_L_0_R_R3

description:
Library name: P10-R3-R
Column name in NormalizedCounts: BR_L_0_R_R3


title: Organoid_TNF_pegtarazimod_4


title: Illumina-GCN5b-KD-IAA24h_R1

description: Library name: Sample 04


title: Skin, TCR+GFP+, Isotype Control, GEX

description:
Library name: kTr1_GEX
10x Genomics

value: Skin

value: Treg

value: Isotype Control, s.c.

value: GEX


title: Organoid_TNF_pegtarazimod_3


title: BR_L_0_R_R2

description:
Library name: P10-R2-R
Column name in NormalizedCounts: BR_L_0_R_R2

value: Rachis

value: Vida


title: Stereotaxic brain tumor biopsy, patient 35, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_35

value: brain

value: Diffuse glioma


title: Organoid_TNF_Vehicle_5

value: TNF + Vehicle


title: Illumina-GCN5b-KD-UT_R3

description: Library name: Sample 03

value: untreated sample


title: Illumina-PHD1-KD-IAA48h_R1

description: Library name: Sample 16

value: RH


title: Stereotaxic brain tumor biopsy, patient 34, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_34

value: Glioblastoma

value: FFPE stereotaxic biopsy


title: Illumina-GCN5b-KD-UT_R2

description: Library name: Sample 02

value: Intracellular tachyzoites


title: BR_L_0_R_R1

description:
Library name: P10-R1-R
Column name in NormalizedCounts: BR_L_0_R_R1

value: 2,4 EBR


title: Illumina-PHD1-KD-IAA24h_R3

description: Library name: Sample 15

value: IAA treated for 24h


title: BR_H_16_R_R3

description:
Library name: P9-R3-R
Column name in NormalizedCounts: BR_H_16_R_R3


title: Organoid_TNF_Vehicle_4


title: Stereotaxic brain tumor biopsy, patient 33, Hemangioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_33

value: Hemangioblastoma

value: female


title: Illumina-GCN5b-KD-UT_R1

description: Library name: Sample 01


title: Stereotaxic brain tumor biopsy, patient 44, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_44


title: BR_H_16_R_R2

description:
Library name: P9-R2-R
Column name in NormalizedCounts: BR_H_16_R_R2


title: Stereotaxic brain tumor biopsy, patient 32, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_32

value: male


title: Organoid_TNF_Vehicle_3


title: BR_H_16_R_R1

description:
Library name: P9-R1-R
Column name in NormalizedCounts: BR_H_16_R_R1


title: ATACseq-GCN5b-KD-IAA_replicate-3

description:
Library name: Sample 06
ATACseq_GCN5b-KD_IAA.bw

library_source: genomic

data_processing:
Following the demultiplexing of raw reads by bcl2fastq V3, trimming, quality control, alignment to the ME49 reference genome (using bwa2).
Aligned reads were used for peak calling of the ChIP enriched peaks using Generich with a cutoff p-value of 0.1. Reads were counted on annotated peaks by featureCounts and counts were processed by DeSeq2 to generate global statistical analysis of peak intensities in between conditions using biological triplicates.
Assembly: Toxoplasma gondii genome (ME49 strain) file "ToxoDB-13.0_TgondiiME49_Genome.fasta" dowloaded from the ToxoDB website : http://toxodb.org/common/downloads/release-13.0/TgondiiME49/fasta/data/
Supplementary files format and content: Data visualization:  Merged bam files were converted to bigWig file format using a bin size of 5 by bamCoverage (Deeptools).

growth_protocol: Toxoplasma gondii type I strains were maintained by serial passage Human foreskin fibroblast (HFF) monolayer under tachyzoite conditions in DMEM (Life) supplemented with 10% (vol/vol) FBS (Life) and 25 mM Hepes buffer, pH 7.2.

extract_protocol:
Diagenode ATAC-SEQ kit protocol (C01080002)
Illumina I5 I7 dual indexing

treatment_protocol: HFF cells were grown to confluence and infected with RHK80 GCN5b KD strain. Genome-wide Tn5 transposase accessiblity profiles in each strain left untreated (UT) and treated with IAA (indole-3-acetic acid) for 24 hours.


title: Stereotaxic brain tumor biopsy, patient 31, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_31


title: Stereotaxic brain tumor biopsy, patient 42, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_42


title: ATACseq-GCN5b-KD-IAA_replicate-2

description:
Library name: Sample 05
ATACseq_GCN5b-KD_IAA.bw


title: DM_L_24_R_R4

description:
Library name: P8-R4-R
Column name in NormalizedCounts: DM_L_24_R_R4

value: DMSO Control


title: Illumina-PHD1-KD-UT_R1

description: Library name: Sample 10


title: Stereotaxic brain tumor biopsy, patient 30, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_30


title: ATACseq-GCN5b-KD-IAA_replicate-1

description:
Library name: Sample 04
ATACseq_GCN5b-KD_IAA.bw


title: Organoid_TNF_pegtarazimod_2

value: TNF + pegtarazimod


title: DM_L_24_R_R3

description:
Library name: P8-R3-R
Column name in NormalizedCounts: DM_L_24_R_R3

value: Rachis


title: Stereotaxic brain tumor biopsy, patient 29, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_29


title: Organoid_TNF_Vehicle_2


title: ATACseq-GCN5b-KD-UT_replicate-3

description:
Library name: Sample 03
ATACseq_GCN5b-KD_UT.bw

source_name: RH


title: Organoid_TNF_Vehicle_1


title: Stereotaxic brain tumor biopsy, patient 28, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_28


title: ATACseq-GCN5b-KD-UT_replicate-2

description:
Library name: Sample 02
ATACseq_GCN5b-KD_UT.bw


title: DM_L_24_R_R2

description:
Library name: P8-R2-R
Column name in NormalizedCounts: DM_L_24_R_R2

value: Vida


title: DM_L_24_R_R1

description:
Library name: P8-R1-R
Column name in NormalizedCounts: DM_L_24_R_R1


title: Arsenite, 3

city: Vienna

first: Aleksandra

middle: S

last: Anisimova

address: Dr. Bohr Gasse 9

country: Austria

institute: Max Perutz Labs, Vienna BioCenter, University of Vienna, Medical University of Vienna

description:
Library name: S143_As2h_3
QuantSeq

data_processing:
Basecalling was performed using Real-Time Analysis (RTA) v1.18.54.4. BCL files were converted to demultiplexed fastq files with bcl2fastq v2.20.0.422.
The quality-, adapter-, and polyA-trimmed reads were aligned to human genome hg38 using GENCODE annotation (release 36) with STAR v2.7.5c. As the reads originate from cross-linked PAR-CLIP samples and have T to C transitions, mismatches were allowed.
Assembly: hg38
Supplementary files format and content: csv, gene counts

growth_protocol: HCT116 conditionally expressing Tet-OsTIR1 were obtained from Masato Kanemaki Lab and cultured in McCoy's 5A (Modified) Medium (Sigma) with 10% Fetal Bovine Serum (Gibco), 2 mM Glutamine (Sigma), 1% Pen/Step (Sigma).

extract_protocol:
Five 15 cm (diameter) dishes of 60 % confluent cells per condition were put on ice, washed with ice-cold PBS (Sigma) and crosslinked with the UV light at 365nm with 0.15 J/cm2 and pelleted. Cell pellets were lysed in 1250 μL of ice-cold lysis buffer (25 mM HEPES pH 7.3, 150 mM NaCl, 0.5% NP-40, 0.5 mM EDTA, 10% Glycerol, 0.1% SDS, 0.2% Sodium Deoxycholate, 1x protease inhibitors cocktail, 0.1mM DTT), by incubation on ice for 15 min with intermittent vortexing and passing three times through the 27G needle. The lysate was clarified by centrifugation for 20 min at 20,000g at +4ºC. An aliquot of the lysate was taken before the in-lysate RNase digest for total RNA isolation and sequencing. Total RNA was isolated using peqGOLD TriFast (Peqlab, VWR).
Transcriptome libraries were prepared with QuantSeq 3’ mRNA-Seq Library Prep Kit (Lexogen). Libraries were sequenced on a NextSeq2000 P2 at SR100 mode (Illumina) at the Vienna BioCenter NGS facility with 30 million reads per sample on average.

treatment_protocol: 4-thiouridine (4sU, Sigma) was added to the cell culture media 15 hours prior collection at 100 µM. ER stress was induced with 5 µg/mL tunicamycin (TM) for 4 hours prior to collection. DMSO in 1:1000 dilution was used a control. Oxidative stress was induced with 250 µM sodium arsenate (As) for 2 hours prior to collection. PBS in 1:1000 dilution was used a control (samples named "Control").

value: human colorectal carcinoma

value: 4-thiouridine (4sU) 100 μM 15h, sodium arsenate 250 μM 2h


title: Stereotaxic brain tumor biopsy, patient 27, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_27

value: brain


title: ATACseq-GCN5b-KD-UT_replicate-1

description:
Library name: Sample 01
ATACseq_GCN5b-KD_UT.bw

value: untreated sample


title: DM_H_0_R_R4

description:
Library name: P7-R4-R
Column name in NormalizedCounts: DM_H_0_R_R4

source_name: Rachis


title: Control for Arsenite samples, 3

description:
Library name: S143_Control_3
QuantSeq

value: 4-thiouridine (4sU) 100 μM 15h, PBS 1:1000 2h


title: Stereotaxic brain tumor biopsy, patient 38, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_38

value: Glioblastoma


title: Stereotaxic brain tumor biopsy, patient 26, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_26

value: FFPE stereotaxic biopsy


title: PHD1-KD-HA-GCN5b-Myc_Myc-antibody_IAA

description: Library name: Sample 20

data_processing:
Data Analysis: After receiving demultiplexed fastq files from the sequencing service provider (ArrayStar), reads were aligned to Toxoplasma gondii reference genome (TGME49) using BOWTIE2. Aligned reads were used for peak calling of the ChIP regions using MACS2. Statistically significant ChIP-enriched regions (peaks) were identified individually using a p-value threshold of 10-5. BigWig datasets were generated using  bedGraphToBigWig to convert the bedgraph file generated by MACS2
Data visualization: The mapped 100 bp reads represent enriched DNA fragments by ChIP experiment. Any region of interest in the raw ChIP-seq signal profile can be directly visualized in genome browser. We use 10-bp resolution intervals (10-bp bins) to partition the stacked reads region, and count the number of reads in each bin.  All the 10 bp resolution ChIP-seq profiles of each sample are saved as UCSC wig format files, which can be visualized in Toxoplasma gondii Genome Browser (http://protists.ensembl.org/Toxoplasma_gondii/Info/ Index).
Assembly: Toxoplasma gondii genome (ME49 strain) file "ToxoDB-13.0_TgondiiME49_Genome.fasta" to upload at ToxoDB website : http://toxodb.org/common/downloads/release-13.0/TgondiiME49/fasta/data/
Supplementary files format and content: Statistically significant ChIP-enriched regions (peaks) were identified individually, using a p-value threshold of 10-4. Te BigWig format file containing the ChIP-enriched regions and was generated for each sample. We use 10-bp resolution intervals (10-bp bins) to partition the stacked reads region, and count the number of reads in each bin.  All the 10 bp resolution ChIP-seq profiles of each sample are saved as UCSC wig format files, which can be visualized in Toxoplasma gondii Genome Browser (http://protists.ensembl.org/Toxoplasma_gondii/Info/ Index).

extract_protocol:
Crosslinked chromatin was lysed in ice-cold lysis buffer (50mM HEPES KOH pH7.5, 140mM NaCl, 1mM EDTA, 10% glycerol, 0.5%NP-40, 0.125% triton X-100, protease inhibitor cocktail) and sheared in shearing buffer (1mM EDTA pH8.0, 0.5mM EGTA pH8.0, 10mM Tris pH8.0, protease inhibitor cocktail) by sonication using a Diagenode Biorupter. Samples were sonicated, for 16 cycles (30 seconds ON and 30 seconds OFF), to 200-500 base-pair average size. Immunoprecipitation was carried out using sheared chromatin, 5% BSA, protease inhibitor cocktail, 10% triton X-100, 10% deoxycholate, DiaMag Protein A-coated magnetic beads (Diagenode) and antibodies (H4K5acK8acK12acK16ac, H4K31me1, HDAC3, and HA). A rabbit IgG antiserum was used as a control mock. After overnight incubation at 4°C on rotating wheel, chromatin-antibody complexes were washed and eluted from beads by using iDeal ChIP-seq kit for Histones (Diagenode) according to the manufacturer’s protocol. Samples were decrosslinked by heating for 4 hours at 65°C. DNA was purified by using IPure kit (Diagenode) and quantified by using Qubit Assays (Thermo Fisher Scientific) according to the manufacturer's protocol. For ChIP-seq, purified DNA was used to prepare libraries and then sequenced by Arraystar (USA, http://www.arraystar.com/).
ChIP-Sequencing library preparation was performed according to Illumina’s protocol Preparing Samples for ChIP Sequencing of DNA. Library Preparation: 10 ng DNA of each sample was converted to phosphorylated blunt-ended with T4 DNA polymerase, Klenow polymerase and T4 polymerase (NEB); An ‘A’ base was added to the 3' end of the blunt phosphorylated DNA fragments using the polymerase activity of Klenow (exo minus) polymerase (NEB); Illumina's genomic adapters were ligated to the A tailed DNA fragments; PCR amplification was performed to enrich ligated fragments using Phusion High Fidelity PCR Master Mix with HF Buffer (Finnzymes Oy). The enriched product of ~200-700 bp was cut out from gel and purified. Sequencing:  The library was denatured with 0.1M NaOH to generate single-stranded DNA molecules, and loaded onto channels of the flow cell at 8pM concentration, amplified in situ using TruSeq Rapid SR cluster kit (#GD-402-4001, Illumina). Sequencing was carried out by running 100 cycles on Illumina HiSeq 4000 according to the manufacturer’s instructions.

value: RH

value: Intracellular tachyzoites


title: Stereotaxic brain tumor biopsy, patient 37, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_37


title: Illumina-GCN5b-KD-IAA24h_R2

description: Library name: Sample 05

value: IAA treated for 24h


title: DM_H_0_R_R3

description:
Library name: P7-R3-R
Column name in NormalizedCounts: DM_H_0_R_R3


title: Arsenite, 2

description:
Library name: S143_As2h_2
QuantSeq


title: Stereotaxic brain tumor biopsy, patient 25, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_25


title: PHD1-KD-HA-GCN5b-Myc_Myc-antibody_UT

description: Library name: Sample 19

treatment_protocol: HFF cells were grown to confluence and infected with RHKU80 GCN5b mAID HA or RHKU80 PHD1 mAID HA strains. Genome-wide GCN5b , MORC, HDAC3, PHD1, H3K18ac, H3K4me3, H3K19ac, occupancy profiles in RHKU80 GCN5b mAID HA while GCN5b or PHD1 occupancies were obtained in a RHKU80 PHD1 mAID HA cell line. In all cases, untreated (UT) and treated with IAA (indole-3-acetic acid) for 24 hours conditions were prepared. Harvested intracellular parasites were crosslinked with formaldehyde (final concentration 1%) for 8 mins at room temperature and the crosslinking was stopped by addition of glycine (final concentration 0.125M) for 5 min at room temperature.


title: Control for Arsenite samples, 2

description:
Library name: S143_Control_2
QuantSeq


title: DM_H_0_R_R2

description:
Library name: P7-R2-R
Column name in NormalizedCounts: DM_H_0_R_R2


title: Stereotaxic brain tumor biopsy, patient 36, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_36


title: Stereotaxic brain tumor biopsy, patient 24, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_24


title: PHD1-KD-HA-GCN5b-Myc_HA-antibody_IAA

description: Library name: Sample 18


title: Arsenite, 1

description:
Library name: S143_As2h_1
QuantSeq


title: DM_H_0_R_R1

description:
Library name: P7-R1-R
Column name in NormalizedCounts: DM_H_0_R_R1


title: Stereotaxic brain tumor biopsy, patient 23, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_23

value: Diffuse glioma


title: PHD1-KD-HA-GCN5b-Myc_HA-antibody_UT

description: Library name: Sample 17


title: BR_H_8_R_R4

description:
Library name: P6-R4-R
Column name in NormalizedCounts: BR_H_8_R_R4

value: 2,4 EBR


title: Control for Arsenite samples, 1

description:
Library name: S143_Control_1
QuantSeq


title: GCN5b-KD-HA-PHD1-Myc_Myc-antibody_IAA

description: Library name: Sample 16


title: BR_H_8_R_R3

description:
Library name: P6-R3-R
Column name in NormalizedCounts: BR_H_8_R_R3


title: Stereotaxic brain tumor biopsy, patient 22, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_22

value: female


title: Stereotaxic brain tumor biopsy, patient 21, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_21


title: TM 4 hours, 2

description:
Library name: S142_TM4h_2
QuantSeq

value: 4-thiouridine (4sU) 100 μM 15h, 5 μg/mL TM 4h


title: TM 1 hour, 2

description:
Library name: S142_TM1h_2
QuantSeq

value: 4-thiouridine (4sU) 100 μM 15h, 5 μg/mL TM 1h


title: BR_H_8_R_R2

description:
Library name: P6-R2-R
Column name in NormalizedCounts: BR_H_8_R_R2

value: Rachis


title: KU80_GCN5b-KD-HA-PHD1-Myc_Myc-antibody_UT

description: Library name: Sample 15


title: Stereotaxic brain tumor biopsy, patient 20, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_20

value: male


title: GCN5b-KD-HA-PHD1-Myc_HA-antibody_IAA

description: Library name: Sample 14


title: Stereotaxic brain tumor biopsy, patient 19, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_19


title: DMSO control for TM samples, 2

description:
Library name: S142_DMSO_2
QuantSeq

value: human colorectal carcinoma

value: 4-thiouridine (4sU) 100 μM 15h, DMSO 1:1000 4h


title: BR_H_8_R_R1

description:
Library name: P6-R1-R
Column name in NormalizedCounts: BR_H_8_R_R1

value: Vida


title: GCN5b-KD-HA-PHD1-Myc_HA-antibody_UT

description: Library name: Sample 13

value: untreated sample


title: BR_L_24_R_R4

description:
Library name: P5-R4-R
Column name in NormalizedCounts: BR_L_24_R_R4


title: TM 4 hours, 1

description:
Library name: S142_TM4h_1
QuantSeq


title: BR_L_24_R_R3

description:
Library name: P5-R3-R
Column name in NormalizedCounts: BR_L_24_R_R3


title: GCN5b-KD-HA_H3K9K14ac-antibody_IAA24h

description: Library name: Sample 12


title: TM 1 hour, 1

description:
Library name: S142_TM1h_1
QuantSeq


title: Stereotaxic brain tumor biopsy, patient 17, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_17

value: brain


title: GCN5b-KD-HA_H3K9K14ac-antibody_UT

description: Library name: Sample 11


title: BR_L_24_R_R2

description:
Library name: P5-R2-R
Column name in NormalizedCounts: BR_L_24_R_R2


title: DMSO control for TM samples, 1

description:
Library name: S142_DMSO_1
QuantSeq


title: GCN5b-KD-HA_H3K4me3-antibody_IAA24h

description: Library name: Sample 10


title: Stereotaxic brain tumor biopsy, patient 16, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_16


title: BR_L_24_R_R1

description:
Library name: P5-R1-R
Column name in NormalizedCounts: BR_L_24_R_R1


title: TM 4 hours, 3

description:
Library name: S141_TM4h_3
QuantSeq


title: Stereotaxic brain tumor biopsy, patient 15, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_15


title: GCN5b-KD-HA_H3K4me3-antibody_UT

description: Library name: Sample 09

source_name: RH


title: TM 1 hour, 3

description:
Library name: S141_TM1h_3
QuantSeq


title: BR_L_8_R_R2

description:
Library name: P16-R2-R
Column name in NormalizedCounts: BR_L_8_R_R2


title: Stereotaxic brain tumor biopsy, patient 14, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_14


title: GCN5b-KD-HA_H3K18ac-antibody_IAA24h

description: Library name: Sample 08


title: DMSO control for TM samples, 3

description:
Library name: S141_DMSO_3
QuantSeq


title: DM_H_24_R_R3

description:
Library name: P4-R3-R
Column name in NormalizedCounts: DM_H_24_R_R3

source_name: Rachis

value: DMSO Control


title: Stereotaxic brain tumor biopsy, patient 13, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_13

value: FFPE stereotaxic biopsy


title: GCN5b-KD-HA_H3K18ac-antibody_UT

description: Library name: Sample 07

value: RH


title: DM_H_24_R_R2

description:
Library name: P4-R2-R
Column name in NormalizedCounts: DM_H_24_R_R2


title: Organoid_TNF_pegtarazimod_1

source_name: small intestine organoids

value: TNF + pegtarazimod


title: CD_3_CD14+_blood_monocytes_rna

city: London

first: Eve

last: Hornsby

address: 4 Newark St

department: Blizard Institue/CIB

country: United Kingdom

institute: QMUL

description: Single Cell Multiome

library_source: transcriptomic single cell

data_processing:
cellranger-arc version 2.0.0 was used for demultiplexing base call files (BCLs) from gene expression and atac sequenced libraries using cellranger-arc mkfastq, followed by cellranger-arc count for genome alignment, gene expression quantification and ATAC peak-calling.
Assembly: GRCh38
Supplementary files format and content: Tab-separated values files and .h5 files

molecule: nuclear RNA

source_name: Peripheral blood

extract_protocol:
Peripheral Blood Mononuclear Cells (PBMC) were obtained by centrifugation in Ficoll®Paque PLUS (GE Healthcare). CD14+ monocytes were isolated from PBMC using positive magnetic selection (CD14 MicroBeads, Miltenyi Biotec). Cells were then washed twice in Phosphate Buffered Saline(Ca2+ and Mg2+ -free) containing 0.04% Bovine Serum Albumin.
10x Genomics Chromium Next GEM Single Cell Multiome ATAC + Gene Expression Reagent Bundle, 4 rxns PN-1000285
Single Cell Multiome ATAC + Gene Expression

value: Peripheral blood

tag: diagnosis

value: Crohn's Disease

value: CD14+ Monocytes


title: Stereotaxic brain tumor biopsy, patient 12, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_12

value: Glioblastoma


title: GCN5b-KD-HA_MORC-antibody_IAA24h

description: Library name: Sample 06

value: Intracellular tachyzoites

value: IAA treated for 24h


title: CD_2_CD14+_blood_monocytes_rna

description: Single Cell Multiome


title: DM_H_24_R_R1

description:
Library name: P4-R1-R
Column name in NormalizedCounts: DM_H_24_R_R1

value: Rachis


title: Stereotaxic brain tumor biopsy, patient 11, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_11

value: Diffuse glioma

value: female


title: GCN5b-KD-HA_MORC-antibody_UT

description: Library name: Sample 05


title: Stereotaxic brain tumor biopsy, patient 10, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_10


title: CD_1_CD14+_blood_monocytes_rna

description: Single Cell Multiome


title: DM_H_8_R_R4

description:
Library name: P3-R4-R
Column name in NormalizedCounts: DM_H_8_R_R4


title: DM_H_8_R_R3

description:
Library name: P3-R3-R
Column name in NormalizedCounts: DM_H_8_R_R3


title: GCN5b-KD-HA_HDAC3-antibody_IAA24h

description: Library name: Sample 04


title: HEALTHY_3_CD14+_blood_monocytes_rna

description: Single Cell Multiome

value: Healthy Control


title: Stereotaxic brain tumor biopsy, patient 9, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_9

value: male


title: DM_H_8_R_R2

description:
Library name: P3-R2-R
Column name in NormalizedCounts: DM_H_8_R_R2

value: Vida


title: Stereotaxic brain tumor biopsy, patient 8, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_8

value: brain


title: GCN5b-KD-HA_HDAC3-antibody_UT

description: Library name: Sample 03

value: untreated sample


title: GCN5b-KD-HA_HA-antibody_IAA24h

description: Library name: Sample 02


title: DM_H_8_R_R1

description:
Library name: P3-R1-R
Column name in NormalizedCounts: DM_H_8_R_R1

value: DMSO Control


title: HEALTHY_1_CD14+_blood_monocytes_rna

description: Single Cell Multiome


title: Stereotaxic brain tumor biopsy, patient 7, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_7

value: FFPE stereotaxic biopsy


title: GCN5b-KD-HA_HA-antibody_UT

description: Library name: Sample 01

value: RH


title: BR_L_16_R_R3

description:
Library name: P2-R3-R
Column name in NormalizedCounts: BR_L_16_R_R3

value: 2,4 EBR


title: CD_3_CD14+_blood_monocytes_atac

description: Single Cell Multiome

value: Peripheral blood

value: Crohn's Disease


title: Stereotaxic brain tumor biopsy, patient 6, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_6


title: IV- SenNP+ CD8 T cells, Day 6 post exposure, batch 2

city: Atlanta

first: Mark

middle: Elliott

last: Williams

address: 201 Dowman Dr

institute: Emory University

description:
Library name: T-cell-pool-3
10x Genomics

data_processing:
Pre-processing of sequencing results to generate count matrices was performed using the 10x Genomics Cell Ranger v8.0.0 count pipeline and aligning to a custom GRCm39-2024-A reference genome.
Assembly: Custom GRCm39-2024-A reference genome with Sendai Virus NP and Luciferase genes added
Supplementary files format and content: 10x Genomics mRNA output files labeled for each batch: barcodes.tsv.gz, features.tsv.gz, matrix.mtx.gz

source_name: Nasal cavity

extract_protocol:
Nasal cavity samples were digested in 5 g/l of Collagenase D and 1e6 U/l of Dnase for 30 minutes at 37°C. T cells were FACS sorted on a FACSAria as Viability Dye negative, intravital label (anti-CD3e) negative, Sendai-NP (Kb 324-332) positive, and CD8 (clone 53-6.7) positive.
10x 5’ scRNA-seq libraries were generated using the Chromium Next GEM Single Cell 5’ Reagent Kit v2 (10x Genomics 1000263) according to the instructions provided.

value: Nasal cavity

value: IV- SenNP+ CD8+ T cells

tag: batch


title: Stereotaxic brain tumor biopsy, patient 5, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_5


title: CD_2_CD14+_blood_monocytes_atac

description: Single Cell Multiome

value: CD14+ Monocytes


title: BR_L_16_R_R2

description:
Library name: P2-R2-R
Column name in NormalizedCounts: BR_L_16_R_R2

value: Rachis


title: CD_1_CD14+_blood_monocytes_atac

description: Single Cell Multiome


title: IV- SenNP+ CD8 T cells, Day 3 post exposure, batch 2

description:
Library name: T-cell-pool-2
10x Genomics


title: BR_L_16_R_R1

description:
Library name: P2-R1-R
Column name in NormalizedCounts: BR_L_16_R_R1


title: Stereotaxic brain tumor biopsy, patient 4, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_4

value: Glioblastoma


title: IV- SenNP+ CD8 T cells, Day 0 post exposure, batch 2

description:
Library name: T-cell-pool-1
10x Genomics


title: DM_L_0_R_R4

description:
Library name: P1-R4-R
Column name in NormalizedCounts: DM_L_0_R_R4


title: HEALTHY_3_CD14+_blood_monocytes_atac

description: Single Cell Multiome


title: Stereotaxic brain tumor biopsy, patient 3, Diffuse glioma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_3

value: Diffuse glioma

value: female


title: IV- SenNP+ CD8 T cells, Day 6 post exposure, batch1

description:
Library name: D6-PR8-SenNPpos
10x Genomics


title: DM_L_0_R_R3

description:
Library name: P1-R3-R
Column name in NormalizedCounts: DM_L_0_R_R3


title: HEALTHY_2_CD14+_blood_monocytes_atac

description: Single Cell Multiome

value: Healthy Control


title: Stereotaxic brain tumor biopsy, patient 2, unclassified brain lesion

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_2

value: unclassified brain lesion


title: IV- SenNP+ CD8 T cells, Day 3 post exposure, batch 1

description:
Library name: D3-PR8-SenNPpos
10x Genomics


title: DM_L_0_R_R2

description:
Library name: P1-R2-R
Column name in NormalizedCounts: DM_L_0_R_R2

value: Vida


title: HEALTHY_1_CD14+_blood_monocytes_atac

description: Single Cell Multiome


title: Stereotaxic brain tumor biopsy, patient 1, Glioblastoma

source_name: FFPE stereotaxic brain tumor biopsy, patient STX_1

value: brain

value: male


title: IV- SenNP+ CD8 T cells, Day 0 post exposure, batch 1

description:
Library name: D0-PR8-SenNPpos
10x Genomics


title: DM_L_0_R_R1

description:
Library name: P1-R1-R
Column name in NormalizedCounts: DM_L_0_R_R1

source_name: Rachis

value: DMSO Control


title: Sample15_AIMT_Clone2

status: Public on Jul 22 2026

city: Prague

first: Juraj

last: Michalik

address: Vídeňská 1083

country: Czech Republic

institute: Institute of Molecular Genetics of the Czech Academy of Sciences

description: Library name: Sample15

data_processing:
Single-end sequencing was performed on an Illumina NextSeq 500 using the NextSeq 500/550 High Output Reagent Cartridge v2 (75 cycles) (#20024906), yielding reads of 76 bp.
Base calling and FASTQ generation were performed using the Illumina BaseSpace GenerateFASTQ workflow v1.37.0.
Reads were trimmed using TrimmomaticSE (Bolger et al., 2014) with the following parameters: ILLUMINACLIP:NexteraPE-PE.fa:2:30:10, LEADING:3, TRAILING:3, SLIDINGWINDOW:4:15, and MINLEN:36; all other parameters were left at their default values.
Reads were aligned to the murine reference genome from Ensembl release 102 (Howe et al., 2021) using STAR v2.7.9a (Dobin et al., 2013) with default parameters.
Reads mapping to the human NGFR sequence (reference generated from GRCh38, Ensembl release 102) were identified using Bowtie2 v2.4.5 (Langmead et al., 2012) and removed from BAM files.
The results were quantified in R 4.5.2. using packages tximport 1.36.0 and GenomicAlignments 1.44.0.
Downstream analyses were performed in R v4.5.2 using DESeq2 v1.48.1 (Love et al., 2014).
Assembly: GRCm38 v102 (Ensembl)
Supplementary files format and content: raw counts for whole bulk experiment without reads mapping to numan NFGR sequence in csv format

source_name: spleen

growth_protocol: Immortalized bone marrow hematopoietic stem cells from Vβ5 Rag2-/- mice were transduced with MSCV retroviral vectors encoding Clone 2 or Clone 12 TCRs and GFP or LNGFR selection markers as previously described {Ruedl et al., 2008; Drobek et al., 2018; Paprckova et al., 2022).

extract_protocol:
The progeny was FACS-isolated frrom the spleen.
The cDNA libraries were prepared using the SMARTer Stranded Total RNA-Seq Kit v2 – Pico Input Mammalian (Takara, #634418) according to the manufacturer's instructions.

treatment_protocol: FACS-sorted GFP+/LNGFR+ CD45.2+ cells were transplanted into irradiated (6 Gy) congenic CD45.1 recipient mice. At least 8 weeks after the transplantation, the GFP+ or LNGFR+ retrogenic T cells were FACS-sorted from the spleens and lymph nodes, FACS-sorted, and transferred into another congenic hosts followed by Lm-OVA infection day after.

value: spleen

tag: cell line

value: CD8+ T cells

value: Progeny of Taim

value: Lm-OVA infection

value: Day 3


title: Sample13_AIMT_Clone12

description: Library name: Sample13

tag: cell type


title: Sample16_Memory_Clone2

description: Library name: Sample16

value: Progeny of Tm


title: Sample11_AIMT_Clone12

description: Library name: Sample12


title: Sample10_Naive_Clone12

description: Library name: Sample10

value: Progeny of Tn


title: Sample9_Memory_Clone12

description: Library name: Sample9


title: Sample8_AIMT_Clone12

description: Library name: Sample8


title: Sample6_Memory_Clone2

description: Library name: Sample6


title: Sample7_Naive_Clone12

description: Library name: Sample7


title: Sample4_Naive_Clone2

description: Library name: Sample4


title: Sample5_AIMT_Clone2

description: Library name: Sample5


title: Sample3_Memory_Clone2

description: Library name: Sample3


title: Sample2_AIMT_Clone2

description: Library name: Sample2


title: Sample1_Naive_Clone2

description: Library name: Sample1


title: KM_4A2cr_4_Totals

city: Glasgow

first: Joseph

middle: Alexander

last: Waldron

address: Switchback road

institute: Beatson Institute for Cancer Research

description:
Library name: Sample 16
Cytoplasmic RNA (Paired to RPFs)

data_processing:
Cutadapt (version 1.18) was used to remove adaptors, trim 3’ bases with Phred scores < 20, and discard reads fewer than 30 and more than 50 bases after trimming. UMI-tools (version 1.0.1) was used to extract unique molecular indexes (UMIs) (4 nt of random sequence at the start and end of every RPF read) from the reads and appended to the read name. Reads were aligned with BBmap (version 38.18), first to remove reads that align to either rRNA or tRNA sequences. Non-rRNA/tRNA reads were then aligned to a filtered protein-coding FASTA (see below), with only the most abundant transcript per gene (calculated from cytoplasmic RNA samples). Samtools (version 1.9) was used to sort and index the resulting BAM files and UMI tools used to deduplicated with the directional method. The number of reads with the 5’ end at each position of every transcript was then counted. Protein coding aligned read lengths peaked at 29-31 nt and lengths 28-33 were used in this analysis, which showed strong CDS enrichment and periodicity. The offset for each read length was determined to be 12 nt for read lengths 28-30 nt, and 13 nt for 31-33 nt. Total counts across the entire CDS, except the first 20 and last 10 codons, were summed together and used as input into DESeq2 to test for differential expression
The paired cytoplasmic RNA samples were processed as the RPFs but with the following exceptions: UMIs were 12 nt at the start of each read only. No maximum read length was set when trimming reads with Cutadapt. Reads were aligned to a filtered protein-coding transcriptome (see below) with Bowtie2 (version 2.3.5.1), using the parameters recommended for use with RSEM, which are --sensitive --dpad 0 --gbar 99999999 --mp 1,1 --np 1 --score-min L,0,-0.1. Gene and isoform level quantification was performed using RSEM (version 1.3.3). The isoform quantification was used to determine the most abundant transcript per gene (see below), but differential expression was measured at the gene level with DESeq2.
Assembly: The gencode.vM27.pc_transcripts.fa file was downloaded from https://www.gencodegenes.org/mouse/release_M27.html and filtered to include only transcripts that had been manually annotated by HAVANA and that have a 5’UTR, a 3’UTR, a CDS equally divisible by 3, an nUG start codon, and a stop codon. All PAR_Y transcripts were also removed. The cytoplasmic RNA-seq data was then aligned to this filtered FASTA and the most abundant transcript per gene was determined, based on the mean TPMs across all samples from the RSEM output. The RPF reads were then aligned to a FASTA containing only the most abundant transcript for each gene.
Supplementary files format and content: pc_final_counts_all_frames.csv files contain integer values that represent the number of RPF reads (not normalised) that have aligned within the coding region of each transcript, excluding the first 20 and final 10 codons.
Supplementary files format and content: isoforms.results files are one of the output files from RSEM quantification. These were used for input into DEseq2 for differential expression analysis. Although these files represent isoform level expression information, DESeq2 is able to collapse this down to the gene level. All DE analysis was performed at the gene level.

molecule: cytoplasmic RNA

source_name: cells derived from lung tumors

growth_protocol: KM cells were isolated from KrasLSL-G12D/WT; Rosa26LSL-MYC/LSL-MYC mouse tumors  and KMNTC/4A2cr lines were generated by CRISPR technology. Briefly Guide RNA (gRNA) sequences targeting Eif4a2 were cloned into a lentiCRISPR vector and lentivirus encoding Cas9 were produced in HEKFT cells together with the guides.Virus transduced cells were then selected using puromycin and lack of eIF4A2 expression was assessed by western blot

extract_protocol:
Cells were then gently lifted, pelleted in PBS and  lysed in 500 µl ice-cold lysis buffer (15 mM Tris-HCl pH 7.5, 15 mM MgCl2, 150 mM NaCl, 1% Triton X-100, 0.05% Tween 20, 2% n-Dodecyl β Maltoside (Thermo Fisher Scientific, 89903), 0.5 mM DTT, 100 µg/ml cycloheximide, 1× cOmplete EDTA-free Protease Inhibitor Cocktail (Merck, 11836170001), 200 U/ml RiboLock RNase Inhibitor (Thermo Fisher Scientific, EO0381)) on ice for 5 min with agitation every minute. Lysates were centrifuged at 4°C for 5 min at 12,000 g and 500 µl supernatant was pipetted into a fresh 1.5 ml tube. For cytoplasmic RNA samples, RNA was extracted from 25 µl of lysate with 975 µl TRIzol, as per the manufacturer’s instructions. A 20-µl aliquot of lysate was also added to 5 µl of 5× SDS gel-loading buffer for western blotting to check for depletion of p. The remaining lysate was digested with 5 µl Ambion RNase I (cloned, 100 U/µl; Thermo Fisher Scientific, AM2295) at 22°C for 15 min, with agitation at 600 rpm in a thermo-mixer. The digestion was stopped with 10 µl SUPERaseIn RNase Inhibitor (20 U/μl; Thermo Fisher Scientific, AM2696) and samples were loaded onto a 10–50% sucrose gradient, containing 15 mM Tris-HCl pH 7.5, 15 mM MgCl2, 150 mM NaCl, and 100 µg/ml cycloheximide, prepared using a BioComp gradient station and cooled to 4°C for at least 1 h prior to use. Samples were centrifuged in a Beckman XPN-90 Ultracentrifuge with an SW40Ti rotor at 38,000 rpm for 2 h at 4°C. Fractions of 0.5 ml were collected with a Biocomp gradient station and Gilson FC 203B fraction collector. Fractions containing the 80S ribosome peak were extracted with acid phenol:chloroform, followed by two chloroform washes. RNA was precipitated with 2 µl glycogen (Roche 10901393001), 300 mM NaOAc pH 5.2, and an equal volume of isopropanol overnight at -20 °C. RNA was pelleted by centrifugation at 12,000 g for 45 min at 4 °C. The supernatant was removed with a pipette, and the pellet was washed twice with 70% ethanol and dissolved in 10 µl RNase-free water.
RNA was diluted with 10 µl 2× urea buffer (Thermo Fisher Scientific, LC6876), heated at 80°C for 90 s, placed immediately on ice, and then loaded onto a pre-run 15% TBE-urea gel (Thermo Fisher Scientific, EC68852BOX) and ran at 200 V for 1 h alongside custom 28 nt (AGCGUGUACUCCGAAGAGGAUCCAACGU) and 34 nt (GCAUUAACGCGAACUCGGCCUACAAUAGUGACGU) RNA markers. The gel was stained with 1× SYBR gold (Thermo Fisher Scientific, S11494) and imaged on a Typhoon FLA 7000. An image was printed to size to allow bands, inclusive of 28 nt and exclusive of 34 nt markers, to be cut from the gel, placed into a 1.5 ml RNA low-binding microcentrifuge tube, and crushed with a scalpel. RNA was eluted from the crushed gel pieces in 500 µl extraction buffer (300 mM NaOAc pH 5.2, 1 mM EDTA, 0.25% SDS) overnight at 16°C at 600 rpm in a thermo-mixer. Gel pieces were filtered out with a Costar Spin-X centrifuge tube filter (0.45 µm; Scientific Laboratory Supplies Ltd, 8163) and RNA was precipitated with 2 µl glycogen and 500 µl isopropanol overnight at -20 °C. Precipitated RNA was pelleted, washed, and dissolved in 13.5 µl RNase-free water, as above. To deplete rRNA the following was added to the 13.5 µl RNA; 5 µl hybridisation buffer (10 mM Tris-HCl pH7.5, 1 mM EDTA, 2 M NaCl), 1 µl RNasin Plus Ribonuclease Inhibitor (Promega N2615) and 0.5 µl biotinylated DNA oligo pool (100 uM total DNA with rRNA_depl_1 and rRNA_depl_2 at a 3:1 molar ratio compared to all other oligos), that had been denatured at 95 °C for 3min and then placed immediately on ice. The mix was incubated for 10min at 68 °C at 1250 rpm in a thermo- mixer and then allowed to cool slowly to room temp by turning off the thermo-mixer. rRNA was depleted with 160 µl Dynabeads MyOne Streptavidin C1 (Thermo Scientific, 65001) as per manufactures’ instructions. The depleted non-RNA was precipitated with 2 µl glycogen and three times the volume of ethanol. Precipitated RNA was again pelleted, washed, and dissolved in 43 µl RNase-free water, as above. RNA was heated at 80°C for 90 s and immediately placed on ice before undergoing 5′ phosphorylation and 3′ dephosphorylation with 1 µl T4 PNK (NEB, M0201S), 5 µl 10× PNK buffer, and 1 µl SUPERaseIn RNase Inhibitor (20 U/μl) at 37°C for 35 min, with 5 µl 10 mM ATP added for the final 20 min. RNA was extracted with acid phenol:chloroform and isopropanol-precipitated as above. Purified RNA was input into the Bioscientific Nextflex small RNA v3 kit (NOVA-5132-06), using the alternative step F bead clean-up, 15 PCR cycles, and gel-extraction option. Cytoplasmic RNA samples were run on an Agilent TapeStation to check RNA integrity. RNA concentration and sample purity were measured on a NanoDrop spectrophotometer. rRNA was depleted from 1 µg cytoplasmic RNA with the RiboCop v2, and sequencing libraries were prepared from the rRNA-depleted RNA with the Corall Total RNA-Seq Library Prep Kit v1 (Lexogen, 095.96) using 13 PCR cycles. Final cytoplasmic RNA and RPF libraries were quantified on an Agilent TapeStation, with a High Sensitivity D1000 tape (Agilent), and sequenced single-end on an Illumina NextSeq500 instrument with a 75 cycles high-output kit.

treatment_protocol: Cell were untreated for this experiment

value: cells derived from lung tumors


title: KM_4A2cr_3_Totals

description:
Library name: Sample 15
Cytoplasmic RNA (Paired to RPFs)


title: KM_4A2cr_2_Totals

description:
Library name: Sample 14
Cytoplasmic RNA (Paired to RPFs)


title: KM_NTC_4_Totals

description:
Library name: Sample 12
Cytoplasmic RNA (Paired to RPFs)

value: KM


title: KM_NTC_3_Totals

description:
Library name: Sample 11
Cytoplasmic RNA (Paired to RPFs)


title: KM_NTC_2_Totals

description:
Library name: Sample 10
Cytoplasmic RNA (Paired to RPFs)


title: KM_NTC_1_Totals

description:
Library name: Sample 9
Cytoplasmic RNA (Paired to RPFs)


title: KM_4A2cr_4_RPFs

description:
Library name: Sample 8
Ribosome protected fragments

library_source: other

data_processing:
Cutadapt (version 1.18) was used to remove adaptors, trim 3’ bases with Phred scores < 20, and discard reads fewer than 30 and more than 50 bases after trimming. UMI-tools (version 1.0.1) was used to extract unique molecular indexes (UMIs) (4 nt of random sequence at the start and end of every RPF read) from the reads and appended to the read name. Reads were aligned with BBmap (version 38.18), first to remove reads that align to either rRNA or tRNA sequences. Non-rRNA/tRNA reads were then aligned to a filtered protein-coding FASTA (see below), with only the most abundant transcript per gene (calculated from cytoplasmic RNA samples). Samtools (version 1.9) was used to sort and index the resulting BAM files and UMI tools used to deduplicated with the directional method. The number of reads with the 5’ end at each position of every transcript was then counted. Protein coding aligned read lengths peaked at 29-31 nt and lengths 28-33 were used in this analysis, which showed strong CDS enrichment and periodicity. The offset for each read length was determined to be 12 nt for read lengths 28-30 nt, and 13 nt for 31-33 nt. Total counts across the entire CDS, except the first 20 and last 10 codons, were summed together and used as input into DESeq2 to test for differential expression
The paired cytoplasmic RNA samples were processed as the RPFs but with the following exceptions: UMIs were 12 nt at the start of each read only. No maximum read length was set when trimming reads with Cutadapt. Reads were aligned to a filtered protein-coding transcriptome (see below) with Bowtie2 (version 2.3.5.1), using the parameters recommended for use with RSEM, which are --sensitive --dpad 0 --gbar 99999999 --mp 1,1 --np 1 --score-min L,0,-0.1. Gene and isoform level quantification was performed using RSEM (version 1.3.3). The isoform quantification was used to determine the most abundant transcript per gene (see below), but differential expression was measured at the gene level with DESeq2.
Assembly: The gencode.vM27.pc_transcripts.fa file was downloaded from https://www.gencodegenes.org/mouse/release_M27.html and filtered to include only transcripts that had been manually annotated by HAVANA and that have a 5’UTR, a 3’UTR, a CDS equally divisible by 3, an nUG start codon, and a stop codon. All PAR_Y transcripts were also removed. The cytoplasmic RNA-seq data was then aligned to this filtered FASTA and the most abundant transcript per gene was determined, based on the mean TPMs across all samples from the RSEM output. The RPF reads were then aligned to a FASTA containing only the most abundant transcript for each gene.
Supplementary files format and content: pc_final_counts_all_frames.csv files contain integer values that represent the number of RPF reads (not normalised) that have aligned within the coding region of each transcript, excluding the first 20 and final 10 codons.
Supplementary files format and content: isoforms.results files are one of the output files from RSEM quantification. These were used for input into DEseq2 for differential expression analysis. Although these files represent isoform level expression information, DESeq2 is able to collapse this down to the gene level. All DE analysis was performed at the gene level.
Library strategy: Ribo-Seq

molecule: other


title: KM_4A2cr_3_RPFs

description:
Library name: Sample 7
Ribosome protected fragments


title: KM_4A2cr_2_RPFs

description:
Library name: Sample 6
Ribosome protected fragments


title: KM_4A2cr_1_RPFs

description:
Library name: Sample 5
Ribosome protected fragments


title: KM_NTC_4_RPFs

description:
Library name: Sample 4
Ribosome protected fragments


title: KM_NTC_1_RPFs

description:
Library name: Sample 1
Ribosome protected fragments


title: KM_NTC_2_RPFs

description:
Library name: Sample 2
Ribosome protected fragments


title: Alb/TGF Mouse, Replicate 2

city: Bronx

first: Kith

last: Pradhan

state: ny

address: 1300 Morris Park Ave

institute: Einstein College of Medicine

description:
Library name: j1228_TGF_Library
10x Genomics single-cell 3′ gene expression; Cell Ranger filtered feature-barcode matrix (HDF5).

data_processing:
Standard Cellranger pipeline
Assembly: mm10
Supplementary files format and content: filtered matrix  of gene counts by cell.  Stored in .h5 format.

source_name: Bone marrow

extract_protocol:
Mice were humanely euthanized following approved institutional protocols. Bone marrow was harvested by carefully isolating and crushing the femur bones using sterile instruments. The resulting bone marrow suspension was filtered through a 40 µm cell strainer to remove debris and then treated with ammonium chloride potassium (ACK) lysis buffer to lyse and eliminate red blood cells. The remaining bone marrow suspension, free of red blood cells, was washed with phosphate-buffered saline (PBS) and resuspended in a single-cell solution.
For single-cell RNA sequencing (scRNA-seq), the prepared cell suspension was processed using the 10x Genomics Chromium Single Cell 3’ workflow, following the manufacturer’s protocol. Library preparation included reverse transcription, cDNA amplification, and barcoding of individual cells, enabling high-throughput sequencing and downstream analysis of gene expression at the single-cell level.

treatment_protocol:
Mice were not treated
Mice were not treated

value: Bone marrow

tag: genotype

value: C57BL/6-J/ Alb-TGF


title: Control Mouse, replicate 1

description:
Library name: J1197_Control
10x Genomics single-cell 3′ gene expression; Cell Ranger filtered feature-barcode matrix (HDF5).


title: Control Mouse, replicate 2

description:
Library name: j1228_Control_Library
10x Genomics single-cell 3′ gene expression; Cell Ranger filtered feature-barcode matrix (HDF5).


title: Ctrl_m_BAT

city: Montpellier Cedex 5

first: Jacques

last: Colinge

address: 208 avenue des Apothicaires

institute: IRCM

description: Library name: sample 2

data_processing:
alignment with STAR 2.5.2b against Mus musculus and Homo sapiens genomes
gene count extraction with HTSeq-count
Assembly: GRCm38.89 and GRCh38.83
Supplementary files format and content: text files with read counts in tabular format

source_name: murin BAT: Brown adipose tissue

growth_protocol: PDX models were established from fresh tumor fragments obtained from the Pathology Department at the Comprehensive Cancer Center of Montpellier (ICM) after informed consent of the patients. A fragment of tumor (~8 mm3) was implanted into the inter-scapular fat pads of 3-4-week-old female Swiss-nude mice. Tumors were passaged onto a further cohort of mice before graft volume reached 1,000 mm3.

extract_protocol:
Quick-RNA Miniprep Kit (Zymo Research, #R1055)
2X75pB_30 M Reads

value: murin BAT:Brown adipose tissue


title: Alb/TGF Mouse, Replicate 1

description:
Library name: J1197_TGF
10x Genomics single-cell 3′ gene expression; Cell Ranger filtered feature-barcode matrix (HDF5).


title: ISO+L-NAME_3

city: Kumamoto

first: Michio

last: Sato

address: 1-1-1 Honjyo Cyu-ou-ku

department: Molecular Genetics

country: Japan

institute: Kumamoto University

description: Library name: DK24070_06

data_processing:
FASTQ files were generated from raw sequencing reads using bcl2fastq (Illumina).
Assembly: GRCm39
Supplementary files format and content: Processed data files contain raw read counts and counts per million (CPM) values for each gene across all samples.
Supplementary files format and content: Excel files are provided in tab-delimited text format and contain gene-level read counts and normalized expression values as counts per million (CPM) for all samples.

source_name: Heart

extract_protocol:
Total RNA was isolated from heart tissues using Trizol Reagent (Thermo Fisher Scientific, Cat# 15596026) according to the manufacturer’s protocol, with glycogen (Roche, Cat# 10 901 393 001) added as a carrier.
Libraries were prepared using the QuantSeq 3′ mRNA-Seq V2 Library Prep Kit (FWD) with UDI (LEXOGEN, Cat# 193.384) according to the manufacturer’s instructions, for sequencing on the Illumina NextSeq platform.

value: Heart

value: WT

tag: treatment

value: ISO+L-NAME


title: control_3

description: Library name: DK24070_03

tag: tissue

value: untreated control


title: ISO+L-NAME_1

description: Library name: DK24070_04


title: HEALTHY_2_CD14+_blood_monocytes_rna

description: Single Cell Multiome

source_name: Peripheral blood

value: Peripheral blood

value: Healthy Control

value: CD14+ Monocytes


title: ISO+L-NAME_2

description: Library name: DK24070_05


title: control_2

description: Library name: DK24070_02


title: Sample_12_12h_mouse_C

city: Rehovot

first: Ariel

last: Ogran

address: 234 Herzl Street

department: Bimolecular Sciences

country: Israel

institute: Weizmann Institute of Science

description:
Library name: Sample_12_12h
NextSeq High Output - 75 cycles

data_processing:
Raw sequencing data in BCL format was demultiplexed and converted to FASTQ files using bcl2fastq (v2.20.0), allowing up to two mismatches in Index 1 and one mismatch in Index 2.
Preprocessing included separating 5′ and 3′ ends into sample-specific FASTQ files, trimming sequencing primers and Illumina adapters using Cutadapt (v1.16), removing poly(A) stretches caused by slippage during PCR amplification, and extracting 8 bp unique molecular identifiers (UMIs) from the 5′ ends using UMI-tools (v0.5.4).
Reads shorter than 20 bp were discarded.
Processed reads were aligned to the mouse genome (Mus_musculus.GRCm39, RepeatMasked primary assembly) using STAR (v2.5.3a) with parameters optimized for transcript isoform detection, including --alignIntronMax 200000 and --alignEndsType Extend5pOfRead1. Paired-end reads uniquely mapping to the same chromosome were identified, and PCR duplicates were removed using UMI-based de-duplication.
Transcript isoform (TIF) boundaries were defined as connections between 5′ transcription start site (TSS) clusters and 3′ polyadenylation site (PAS) clusters supported by at least four read pairs, excluding clusters with extreme mate-pair distances (<300 bp or >2 Mb).
Internal priming at the 3′ ends was mitigated by filtering tags with seven consecutive adenines in the downstream 10 nt. End clustering was performed using the CAGEr package, with normalization, spatial clustering, and annotation based on Gencode vM33. This comprehensive pipeline enabled the precise identification of TIF boundaries and regulatory features for downstream analyses.
TIF UTRs were extrapolated using the nearest reference transcripts as a guide (see Supplementary Figure S1C). For each TIF, the TSS and PAS coordinates were matched to their closest annotated transcripts from the Mus_musculus.GRCm39.108.chr.chr.M.gff3 genome annotation file.
Assembly: Mus_musculus.GRCm39.108.chr.chr.M.gff3
Supplementary files format and content: CSV files, Transcript isoforms (TIFs) genomic coordinates with totalCov (raw counts), peak_5'end, peak_3'end, TIF_length, TE,  annotation1, annotation2, seq_3'UTR and seq_5'UTR

source_name: mouse differentiated myotubes

growth_protocol: Primary myoblasts were extracted from the gastrocnemius muscle of female mice and plated onto 10 cm plates. Once the cells reached confluency, they were treated with 1 µM ERK inhibitors (ERKi; SCH772984, Cayman Chemicals)  to induce differentiation. Within 24 hours, the treated myoblasts fused into myotubes and were ready for conducting the mRNA and stability assays of this study. mRNA decay assay Following myotube formation, cells were treated with 7.5 µg/ml actinomycin D (Sigma) to inhibit transcription. Treatments were performed at four time points: 0, 3, 6, and 12 hours after actinomycin D addition. At the 12-hour time point, cells were lysed in 1 ml TRIzol reagent, incubated for 5 minutes, and scraped from the plates. Lysates were vortexed, incubated for an additional 10 minutes, and centrifuged to remove debris. The clarified supernatants were transferred to labeled tubes, snap-frozen, and stored at -80°C for subsequent RNA analysis.

extract_protocol:
For Polysome profling: Cells were treated with 100 µg/ml cycloheximide (CHX) for 5 minutes at room temperature to stabilize ribosome-mRNA complexes. Following treatment, cells were washed twice with ice-cold buffer containing 20 mM Tris (pH 8), 140 mM KCl, 5 mM MgCl₂, and 100 µg/ml CHX. The cells were then collected and lysed in 500 µl of the same buffer, supplemented with 0.5% Triton X-100, 0.5% sodium deoxycholate (DOC), 1.5 mM DTT, 150 units of RNase inhibitor (Eurx), and 5 µl of protease inhibitor cocktail (Sigma-Aldrich). The lysates were vortexed gently to ensure thorough mixing and then centrifuged at 12,000 × g for 5 minutes at 4°C to remove cell debris. The clarified lysates were layered onto a pre-formed 10–50% sucrose gradient and subjected to ultracentrifugation at 38,000 rpm for 105 minutes at 4°C using an SW41 rotor. Following centrifugation, gradients were fractionated using a piston gradient fractionator, and the optical density at 254 nm was continuously recorded with an ISCO absorbance detector (UA-6) to monitor ribosome-associated RNA distribution. For downstream analyses, fractions were categorized into polysome-free (Free), light polysome (2–5 ribosomes), and heavy polysome (6+ ribosomes). RNA was extracted from these fractions using TRIzol reagent and the Direct-zol RNA Miniprep Kit (Zymo Research) according to the manufacturer’s instructions. For mRNA decay assay: Following myotube formation, cells were treated with 7.5 µg/ml actinomycin D (Sigma) to inhibit transcription. Treatments were performed at four time points: 0, 3, 6, and 12 hours after actinomycin D addition. At the 12-hour time point, cells were lysed in 1 ml TRIzol reagent, incubated for 5 minutes, and scraped from the plates. Lysates were vortexed, incubated for an additional 10 minutes, and centrifuged to remove debris. The clarified supernatants were transferred to labeled tubes, snap-frozen, and stored at -80°C for subsequent RNA analysis.
To construct TIF-seq2 libraries, we followed a protocol optimized for generating full-length cDNA from capped and polyadenylated RNA. Specifically, 5.0 μg of total RNA was treated with Turbo DNase (Thermo Fisher Scientific) to eliminate genomic DNA contamination, followed by dephosphorylation with Calf Intestinal Alkaline Phosphatase (New England Biolabs) and decapping with Cap-Clip Acid Pyrophosphatase (Cell Script). A DNA/RNA chimeric oligonucleotide adaptor (From here on, all oligos, primers, and linkers are listed in Supplementary Table 1) containing a unique molecular identifier (UMI) was ligated to the 5’ end of the RNA, creating a universal anchor for subsequent amplification. Reverse transcription was performed using barcoded oligo-dT primers, introducing Illumina sequencing adapters and sample-specific barcodes. The resulting cDNA was purified and amplified by PCR (In which one primer was biotinylated). Circularization of PCR products was achieved through self-ligation under highly diluted conditions to favor intramolecular ligation. Unligated linear products were removed using a combination of plasmid-safe DNase (Lucigen) treatment and phenol-chloroform purification. Circularized cDNA was fragmented by sonication, and fragments containing both the 5’ and 3’ ends were captured using streptavidin-coated beads (Thermo Fisher Scientific). After end repair, dA tailing, and adapter ligation, the libraries were amplified by PCR, size-selected using a two-step beads-based protocol, and sequenced on an Illumina NextSeq 500 platform. Custom sequencing primers were used to facilitate paired-end sequencing, with read lengths of 76 bp for both read1 and read2, and 6 bp for each index read. This approach enabled the precise capture of transcript isoforms, including their exact 5’ and 3’ boundaries, providing high-resolution data for downstream analyses. All the oligos used in the TIF-seq2 library preparation protocol can be view in Additional info with relevant data.

value: mouse differentiated myotubes

value: 12h_actinomycin D  (ActD)

tag: assay

value: mRNA decay


title: control_1

description: Library name: DK24070_01


title: Sample_11_12h_mouse_B

description:
Library name: Sample_11_12h
NextSeq High Output - 75 cycles


title: Sample_10_12h_mouse_A

description:
Library name: Sample_10_12h
NextSeq High Output - 75 cycles


title: Sample_9_6h_mouse_C

description:
Library name: Sample_9_6h
NextSeq High Output - 75 cycles

value: 6h_actinomycin D  (ActD)


title: Sample_8_6h_mouse_B

description:
Library name: Sample_8_6h
NextSeq High Output - 75 cycles


title: Sample_7_6h_mouse_A

description:
Library name: Sample_7_6h
NextSeq High Output - 75 cycles


title: Sample_6_3h_mouse_C

description:
Library name: Sample_6_3h
NextSeq High Output - 75 cycles

value: 3h_actinomycin D  (ActD)


title: Sample_4_3h_mouse_A

description:
Library name: Sample_4_3h
NextSeq High Output - 75 cycles


title: Sample_2_0h_mouse_B

description:
Library name: Sample_2_0h
NextSeq High Output - 75 cycles

value: 0h_actinomycin D  (ActD)


title: Sample_3_0h_mouse_C

description:
Library name: Sample_3_0h
NextSeq High Output - 75 cycles


title: Sample_5_3h_mouse_B

description:
Library name: Sample_5_3h
NextSeq High Output - 75 cycles


title: Sample_1_0h_mouse_A

description:
Library name: Sample_1_0h
NextSeq High Output - 75 cycles


title: Mouse_2_HEAVY_fraction_(polysome_profiling)

description:
Library name: Sample_6
NextSeq High Output - 75 cycles

value: cycloheximide (CHX)

value: Polysome profiling


title: Mouse_2_FREE_fraction_(polysome_profiling)

description:
Library name: Sample_4
NextSeq High Output - 75 cycles


title: Mouse_2_LIGHT_fraction_(polysome_profiling)

description:
Library name: Sample_5
NextSeq High Output - 75 cycles


title: Mouse_1_LIGHT_fraction_(polysome_profiling)

description:
Library name: Sample_2
NextSeq High Output - 75 cycles


title: Mouse_1_HEAVY_fraction_(polysome_profiling)

description:
Library name: Sample_3
NextSeq High Output - 75 cycles


title: Mouse_1_FREE_fraction_(polysome_profiling)

description:
Library name: Sample_1
NextSeq High Output - 75 cycles


title: PLT_pos5_S5

city: Oxford

first: Claus

last: Nerlov

address: John Radcliffe Hospital, University of Oxford

department: MRC Molecular Haematology Unit

institute: Weatheral Institute of Molecular Medicine

data_processing:
RNAseq: Raw reads were trimmed using trimmomatic software and inspected using FastQC
RNAseq: Trimmed reads were aligned to the mouse genome (mm10) using STAR software.
RNAseq: The featureCounts software was used to count reads on the basis of the RefSeq gene model
Assembly: mm10
Supplementary files format and content: Raw count matrix containing all samples was stored in Bulk_RNAseq_platelet.txt

extract_protocol:
Platelets from peripheral blood were sorted by FACS and cDNA was generated according to Smart-seq+ protocol
Illumina sequencing libraries were constructed using the Nextera XT DNA Sample Preparation kit according to manufacturer's protocol.

value: Tomato+ platelets


title: PLT_pos4_S4


title: PLT_pos3_S3


title: PLT_pos1_S1


title: PLT_neg4_S9

value: Tomato- platelets


title: PLT_neg5_S10


title: PLT_pos2_S2


title: PLT_neg3_S8


title: PLT_neg1_S6


title: Sample14_Naive_Clone2

description: Library name: Sample14

value: spleen

value: CD8+ T cells

value: Progeny of Tn

value: Lm-OVA infection

value: Day 3


title: PLT_neg2_S7

value: Peripheral blood


title: siNPM1-2

city: Changsha

first: caiping

last: ren

address: 110 Xiangya Road,

institute: Central South University

description: Library name: sample 5

data_processing:
After thawing the samples on ice, mix them thoroughly, centrifuge the mixture, and take an appropriate amount of the supernatant for detection. Extract RNA using the DNF-471 kit and perform quality control; once the quality control criteria are met, construct sequencing libraries, conduct sequencing on platforms such as Illumina, and carry out subsequent gene expression quantification and differential expression analysis.
Assembly: GRCh38
Supplementary files format and content: Process.gene.TPM.matrix.annot.xlsx

source_name: Cell line

extract_protocol:
After thawing the cell samples on ice, mix thoroughly and centrifuge; then take an appropriate amount of the sample for detection using the DNF-471 kit.
RNA Quality Assessment; RNA Enrichment/Fragmentation: Reverse Transcription： End Repair/A-Tailing； Adapter Ligation； PCR Amplification； Library Quality Control

value: Cell line

value: hPSCs


title: siNPM1-3

description: Library name: sample 6


title: siControl-3

description: Library name: sample 3

value: Control


title: siNPM1-1

description: Library name: sample 4


title: siControl-1

description: Library name: sample 1


title: siControl-2

description: Library name: sample 2


title: 09062023_G4-1B, Biol rep B 2

city: Mexico city

first: Aarón

last: Vázquez-Jiménez

state: Mexico city

address: Periferico Sur 4809

country: Mexico

institute: INMEGEN

description:
Library name: 09062023_G4-1B
FFPE

data_processing:
Raw sequencing data were processed using 10x Genomics space ranger 2.0 pipeline to generate fastq files
Raw sequencing data were processed using 10x Genomics space ranger 2.0 pipeline to generate fastq files
FASTQ files were processed to generate the count matrix using Space Ranger's default settings
Assembly: HGR38
Supplementary files format and content: barcodes.tsv.gz: list of spatial barcodes
Supplementary files format and content: features.tsv.gz: list of gene IDs
Supplementary files format and content: matrix.mtx.gz: gene expression count data in Matrix Exchange Format
Supplementary files format and content: aligned_fiducials.jpg: aligned fiducials of the tissue image
Supplementary files format and content: detected_tissue_image.jpg: image of tissue and spots
Supplementary files format and content: scalefactors_json.json: scale factors un json format
Supplementary files format and content: tissue_hires_image.png: hi-res image of tissue
Supplementary files format and content: tissue_lowres_image.png: low-res image of tissue
Supplementary files format and content: tissue_positions_list.csv: list of spatial barcodes and coordinates specifying spota
Supplementary files format and content: spatial_enrichment: Moran's I values for each feature.
Library strategy: Spatial Transcriptomics

source_name: pediatric medulloblastoma, Group 4

extract_protocol:
RNA Extraction  were prepared according to manufacturer's manuals and recommendations.
Spatial transcriptome sequencing libraries were prepared with 10xGenomics  Visium Spatial Gene expression Slide V1 & Reagent kit according to manufacturer's construction

value: pediatric medulloblastoma, Group 4


title: SHH-3B, Biol rep A 2

description:
Library name: SHH-3B
FFPE

source_name: pediatric medulloblastoma, SHH

value: pediatric medulloblastoma, SHH


title: SHH-3A, Biol rep A 1

description:
Library name: SHH-3A
FFPE


title: 09062023_SHH-2

description:
Library name: 09062023_SHH-2
FFPE


title: G4-1A, Biol rep B 1

description:
Library name: G4-1A
FFPE


title: 09062023_SHH-1

description:
Library name: 09062023_SHH-1
FFPE


title: 09062023_G3-1

description:
Library name: 09062023_G3-1
FFPE

source_name: pediatric medulloblastoma, Group 3

value: pediatric medulloblastoma, Group 3


title: PCOS

city: GuangZhou

first: Tengfei

last: Wang

state: GuangDong

address: No. 9 Jinsui Road, Tianhe District

institute: Guangzhou Women and Children's Medical Center

description: Library name: Sample2

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-cellgene-expression/software/pipelines/latest/what-is-cell-ranger)
Assembly: GRCh38
Supplementary files format and content: integrated_1027.rds

source_name: endometrium

extract_protocol:
The isolated endometrial cells were resuspended in digestion buffer  and incubated at 37 °C for 30 min. Cells were washed, re-suspended in PBS, and stained with a viability dye. After washing, cells were resuspended in FACS buffer . Live endometrial cells were sorted using a FACS Jazz (Becton Dickinson) based on the presence or absence of mCherry signal, using a C57BL/6 mouse as a negative control.
Library was performed according to the manufacter's instructions (single cell 3' v2 protocol, 10x Genomics). Briefly, GCs 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 llumina 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 llumina R2 primer sequence, paired-end constructs with P5 and P7 sequences and a sample index were added.

value: endometrium

value: primary cells

tag: subject status

value: PCOS


title: Normal

description: Library name: Sample1

value: Normal


title: HC5 PBMC, Blood, Healthy control

city: Beijing

first: Jin-Wen

last: Song

state: Beijing

address: No. 100, West Fourth Ring Road, Fengtai District

institute: Beijing 302 Hospital

description:
Library name: HC5
10×Genomics

data_processing:
Made by Cell Ranger (v7.1.0) (https://www.10xgenomics.com/support/software/cell-ranger/latest/analysis/running-pipelines/cr-flex-multi-frp)
Assembly: GRCh38
Supplementary files format and content: Tab-separated values files and matrix files

source_name: blood

extract_protocol:
PBMCs were isolated from ethylene diamine tetraacetic acid (EDTA) anticoagulated venous blood by Ficoll-Hypaque (Merck KGaA, Cat# GE17-1440-02) density gradient centrifugation. Briefly, blood samples were centrifuged at 450 xg for 10 minutes. The supernatant (plasma) was collected and preserved at -80°C. PBMCs were then isolated by Ficoll through density gradient centrifugation at 700 xg, with the acceleration set to 4 and the deceleration set to 1. Following centrifugation, cells from the plasma-Ficoll interface were collected and subsequently transferred to a new tube, then washed twice with phosphate-buffered saline (PBS) at 450 xg for 10 minutes. The resulting precipitates were resuspended in the cryopreservation liquid containing 90% fetal bovine serum (FBS) (Thermo Fisher Scientific, Cat# 100010023) and 10% dimethyl sulfoxide (DMSO) (Merck KGaA, Cat# 200-664-3).
Gene expression libraries were generated using the Chromium Fixed RNA Profiling workflow with the Human Transcriptome Probe Set, a probe-hybridization/ligation–based assay that targets around 18,532 genes. The method for cell counting and viability detection involves sample staining with a fluorescent dye (AO/PI Staining Solution kit, CS2-0106-5mL, Nexcelom Biosciences) and cell quantification with an automated fluorescent cell counter (Cellaca counter, MX-112-0127, Nexcelom Biosciences). Subsequently, probe hybridization was conducted, followed by partitioning in Gel Bead-in-Emulsion (GEM), ligation, and extension within the GEMs, culminating in library construction. Libraries were then loaded onto a high-throughput sequencing platform NextSeq500 (Illumina) to generate large amounts of RNA sequencing data.

value: blood

value: PBMC


title: KM_4A2cr_1_Totals

description:
Library name: Sample 13
Cytoplasmic RNA (Paired to RPFs)

value: cells derived from lung tumors


title: HC3 PBMC, Blood, Healthy control

description:
Library name: HC3
10×Genomics


title: HC4 PBMC, Blood, Healthy control

description:
Library name: HC4
10×Genomics


title: HC2 PBMC, Blood, Healthy control

description:
Library name: HC2
10×Genomics


title: HC1 PBMC, Blood, Healthy control

description:
Library name: HC1
10×Genomics


title: PLWH17 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-1274
10×Genomics

value: cART


title: PLWH16 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-1234
10×Genomics


title: KM_NTC_3_RPFs

description:
Library name: Sample 3
Ribosome protected fragments

value: KM


title: PLWH14 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-1226
10×Genomics


title: DM_H_24_R_R4

city: Bozeman

first: Jennifer

last: Lachowiec

address: 119 Plant Biosciences Building

institute: Montana State University

description:
Library name: P4-R4-R
Column name in NormalizedCounts: DM_H_24_R_R4

data_processing:
Read quality was checked with FASTQC and MULTIQC software with an average phred score of 36 (Andrews, 2010). RNA sequence trimming, mapping, and read counting was performed with Trimmomatic v.0.36, STAR, and the Rsubread package v.1.5.2 with function featureCounts, respectively using default settings (Bolger et al., 2014; Dobin et al., 2013; Liao et al., 2014). Reads were aligned using the Triticum aestivum cv. Chinese Spring IWGSC RefSeq v2.1 genome assembly (Zhu et al., 2021). Genes with fewer than 10 total reads across all 123 remaining samples were removed were considered non-expressed. Differential gene expression analysis was performed via DESeq2 (Love et al., 2014). Two samples from different treatment groups were removed from analysis as they clustered far from the other biological replicates based on visual assessment of PCA. DESeq2 was additionally used to normalize read counts by length and read depth for data visualization with pheatmap and ggplot2 (Kolde, 2010; Wickham, 2016).
Assembly: Reads were aligned using the Triticum aestivum cv. Chinese Spring IWGSC RefSeq v2.1 genome assembly (Zhu et al., 2021) and STAR R package using default settings (Dobin et al., 2013)
Supplementary files format and content: NormalizedCounts.csv, inlcudes normalized count by length and read depth for each sample

extract_protocol:
For RNAseq, 64 plants were grown, with four biological replicates per seed treatment, temperature, and time of sampling combinations. Prior to and at 8, 16, and 24 hours after heat exposure began, the unemerged heads were collected. The unemerged wheat heads were removed from the leaf sheath and dissected into rachis and floret tissues, flash frozen,  and stored at -70°C.  Tissue was ground with 3 mm sterilized glass beads in a bead beater, RNA extraction was performed with the RNeasy Plant Mini Kit (Qiagen, Cat #74903) using Beta-mercaptoethanol, and samples were treated with DNase I (Turbo DNA-free Kit, Invitrogen AM1907). Quality of the samples was confirmed via Bioanalyzer using the Agilent RNA 6000 Nano Kit (Agilent, 5067-1511). Five total samples were removed for poor or low RNA quality, leaving a minimum of three biological replications per treatment combination.
RNA-seq libraries were prepared using poly(A)-selected mRNA, and RNA-sequencing was performed by Genewiz.

value: Rachis

value: Vida

value: DMSO Control

tag: temperature


title: PLWH15 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-1230
10×Genomics


title: PLWH13 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-1200
10×Genomics


title: PLWH11 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-1156
10×Genomics


title: PLWH12 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-1180
10×Genomics


title: PLWH10 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-1111
10×Genomics


title: PLWH9 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-1019
10×Genomics


title: PLWH8 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-1013
10×Genomics


title: PLWH7 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-939
10×Genomics


title: PLWH1 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-551
10×Genomics


title: PLWH6 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-889
10×Genomics


title: PLWH5 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-846
10×Genomics


title: PLWH17 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-504
10×Genomics

value: untreated


title: PLWH16 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-493
10×Genomics


title: PLWH15 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-420
10×Genomics


title: PLWH4 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-675
10×Genomics


title: PLWH2 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-623
10×Genomics


title: PLWH3 PBMC, Blood, HIV-infected (cART)

description:
Library name: PLWH-669
10×Genomics


title: PLWH14 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-316
10×Genomics


title: PLWH13 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-235
10×Genomics


title: PLWH11 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-162
10×Genomics


title: PLWH10 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-152
10×Genomics


title: PLWH12 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-225
10×Genomics


title: PLWH8 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-108
10×Genomics


title: PLWH9 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-139
10×Genomics


title: PLWH7 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-106
10×Genomics


title: PLWH2 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-9
10×Genomics


title: PLWH5 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-31
10×Genomics


title: PLWH1 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-5
10×Genomics


title: 0175_TAZ

city: Milan

first: Raffaella

last: Di Micco

address: Via Olgettina, 60

country: Italy

institute: San Raffaele Telethon Institute for Gene Therapy

description: Library name: 0175-TAZ

data_processing:
Reads quality control checked with FASTQC
Read alignment to reference genome with STAR
Reads per gene quantification with featurecounts
Normalization and differential analysis with DESeq2
Assembly: hg38
Supplementary files format and content: ReadsPerGene_TAZ_CTRL.txt

source_name: bone marrow

extract_protocol:
miRNeasy Mini Kit (QIAGEN)
NEBnext ultra II RNA directional library preparation kit, with respective rRNA depletion module

value: bone marrow

value: Patient-derived acute myeloid leukemia blasts

value: acute myeloid leukemia blasts

value: treated with tazemetostat


title: PLWH3 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-13
10×Genomics


title: 0826_TAZ

description: Library name: 0826-TAZ


title: PLWH4 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-29
10×Genomics


title: 0175_CTRL

description: Library name: 0175-CTRL


title: PLWH6 PBMC, Blood, HIV-infected (untreated)

description:
Library name: PLWH-36
10×Genomics


title: 0826_CTRL

description: Library name: 0826-CTRL


title: UPN05_TAZ

description: Library name: UPN05-TAZ


title: FFPE, LateProgressive,nr110

city: Bergen

first: Tony

last: Chen

address: Haukelandsveien 22

country: Norway

institute: Haukeland University Hospital

description: Human pancancer immune profiling panel primers and IDT-designed spike-in oligonucleotides targeting selected genes were added in the analysis

scan_protocol: Fluorescent detection system using nCounter MAX/FLEX Analysis System.

data_processing: Corresponding RCC files were generated for downstream analysis.

label: Fluorescently barcoded reporter probes

source_name: Kidney

label_protocol: Four ng/ul was used for complementary cDNA synteshization using reverse transcription and amplification, and was thereafter subjected to hybridization with capture and reporter probes.

extract_protocol: RNAwas extracted from five to six microtome sliced, 3 um thick sections. miRNeasy extraction kit was used for RNA isolation.

tag: gender

value: Male


title: UPN05_CTRL.

description: Library name: UPN05-CTRL


title: FFPE, LateProgressive,nr100


title: FFPE, LateProgressive,nr101


title: PDAC_ex_vivo_slice, OT146, Selenite_15uM_(24-48h), 48h

city: Huddinge

first: Tímea

last: Szekerczés

address: Alfred Nobels allé 8

department: LabMed

country: Sweden

institute: Karolinska Institutet

description:
Library name: OT146_48h_Se_15uM
Ex vivo culture 48 h; sodium selenite 15 uM applied for 24 h during the second day (24-48 h)

data_processing:
FASTQ processing was performed in the Chipster environment (CSC, Finland). Read quality was assessed with FastQC and summarized with MultiQC.
Adapter and quality trimming was performed with Trimmomatic v0.39 using TruSeq3-PE-2 adapters and a 4-bp sliding window (average Phred >= 15); reads shorter than 30 bp after trimming were discarded.
Trimmed reads were aligned to GRCh38 using STAR v2.7.10a.
Gene-level read counts were obtained with HTSeq-count v0.13.5 against Homo_sapiens.GRCh38.109 using paired-end mode and strandedness = reverse.
Differential expression was performed in edgeR (Chipster) using TMM normalization and a paired design (patient + group). Significance was defined as FDR < 0.05 (Benjamini-Hochberg).
Assembly: GRCh38; gene annotation: Homo_sapiens.GRCh38.109
Supplementary files format and content: Processed data files are tab-delimited TSV count tables generated by HTSeq-count (gene_id and raw read counts per sample)
Supplementary files format and content: Raw data are paired-end FASTQ files (R1/R2). Lane-level FASTQ files were merged per sample and read (R1 and R2) prior to submission.

source_name: pancreatic ductal adenocarcinoma tumor tissue

growth_protocol: Fresh PDAC tumor tissue was received in the pathology laboratory within ~20 min after surgery. Tumor tissue was sampled by a specialist pancreatic pathologist and cut into 350 um thick slices using a vibrating-blade microtome (Leica VT1200S) with a sapphire blade. Tissue slices were handled on ice in transport medium during preparation. The first slice was immediately formalin-fixed and FFPE-embedded as the 0 h baseline. Remaining slices were placed on 0.4 um pore, 30 mm Millicell membrane inserts in 35 mm culture dishes containing glutamine-free CMRL 1066 medium (Gibco 11530037) supplemented with 25 mM HEPES, 1 mM sodium pyruvate, ZnSO4, insulin-transferrin-selenite solution, 2.5% human AB serum, 1X Pen/Strep, and 100 nM diphenyl diselenide, and incubated at 37 C.

extract_protocol:
Total RNA was isolated from 10 um DNase-treated FFPE sections using the Maxwell RSC FFPE RNA Kit (Promega AS1440) according to the manufacturer. RNA quantity/quality was assessed using Qubit 4.0 with the RNA HS Assay Kit (Thermo Fisher Q32852).
Up to 50 ng total RNA per sample was used for library preparation with the SMARTer Total RNA-Seq Kit v2.5 Pico Input Mammalian (Takara 634411). RNA was thermally fragmented (94 C, 3 min) and reverse-transcribed with random N6 hexamers. Illumina adapters and sample barcodes were introduced during amplification; ribosomal cDNA was depleted by RNase H with mammalian R-probes; remaining fragments were re-amplified with universal adapter primers. Libraries were quantified (Qubit DNA HS, Q33230), normalized, pooled and sequenced on an Illumina NextSeq 500/550 Mid Output v2.5 (150 cycles) to generate 2 x 75 bp paired-end reads.

treatment_protocol: Slices were cultured for 48 h. Untreated slices were maintained to 48 h (48 h treatment control, no selenite). For treatment conditions, sodium selenite was added only during the second 24 h (24-48 h) at 5 uM or 15 uM.

value: sodium selenite 15 uM (24-48 h)


title: FFPE, LateProgressive,nr99


title: PDAC_ex_vivo_slice, OT146, Selenite_5uM_(24-48h), 48h

description:
Library name: OT146_48h_Se_5uM
Ex vivo culture 48 h; sodium selenite 5 uM applied for 24 h during the second day (24-48 h)

value: pancreatic ductal adenocarcinoma tumor tissue

value: sodium selenite 5 uM (24-48 h)


title: PDAC_ex_vivo_slice, OT146, Culture_control, 48h, no_treatment

description:
Library name: OT146_48h_TC
Ex vivo culture 48 h; no selenite (treatment control)

value: no selenite (culture control)


title: PDAC_ex_vivo_slice, OT146, Baseline, 0h

description:
Library name: OT146_0h
0 h baseline; collected immediately after surgery; no ex vivo culture

value: none (baseline)


title: PDAC_ex_vivo_slice, OT142, Selenite_15uM_(24-48h), 48h

description:
Library name: OT142_48h_Se_15uM
Ex vivo culture 48 h; sodium selenite 15 uM applied for 24 h during the second day (24-48 h)


title: FFPE, LateProgressive,nr96


title: FFPE, LateProgressive,nr97

value: Female


title: PDAC_ex_vivo_slice, OT142, Selenite_5uM_(24-48h), 48h

description:
Library name: OT142_48h_Se_5uM
Ex vivo culture 48 h; sodium selenite 5 uM applied for 24 h during the second day (24-48 h)