title: ESR2_hypoxia_10_nM_17estradiol_rep3

type: RNA

city: Poznan

first: Marcin

last: Rucinski

address: Swiecicki 6

department: Histology and Embriology

country: Poland

institute: Poznan University of Medical Science

description: Gene expression data from HCT16, colon cancer cell line

hyb_protocol: Biotin-labeled fragments of cDNA (5.5 μg) were hybridized to Affymterix GeneChip Human Genome U219 ArrayStrip (16 h, 45°C).

scan_protocol: Microarrays were washed and stained according to the technical protocol, using AffymetrixGeneAtlasFuidics Station. The array strips were scanned employing Imaging Station of GeneAtlas System.

data_processing: Obtained CEL files were subjected for background correction and normalization using RobustMultiarrayAveraging (RMA) algorithm implemented in affy package of BioConductor.

label: biotin

source_name: ESR2+ hypoxia + 10 nM 17β-estradiol

label_protocol: The protocol involving transcription in vitro, biotin labelling and cDNA fragmentation for further hybridization was carried out using the Affymetrix GeneChip IVT Express Kit (Affymetrix, Santa Clara, CA, USA)

growth_protocol: HCT116 and DLD-1 colon cancer cells were obtained from the American Type Culture Collection (Rockville, MD). Cells were maintained at 37°C in a humidified chamber with 5% CO in McCoy’s 5A or RPMI-1640 medium (Gibco BRL) containing 10% heat-inactivated fetal bovine serum (FBS) and 2 mM glutamine, antibiotic antimycotic solution (A5955, Sigma Aldrich). They were also tested negative for Mycoplasma (LT07-318, Lonza).

extract_protocol: The total RNA was extracted using TRI Reagent (Sigma-Aldrich, St. Louis, MO, USA) then purified on columns (NucleoSpin Total RNA Isolation, Qiagen GmbH, Hilden, Germany). The amount of total RNA was determined by optical density at 260 nm and its purity was estimated by 260/280 nm absorption ratio (higher than 1.8) (NanoDrop spectrophotometer, Thermo Scientific, Waltham, MA, USA). The RNA integrity and quality were checked in a Bioanalyser 2100 (Agilent Technologies, Inc., Santa Clara, CA, USA). The resulting RNA integrity numbers (RINs) were between 8.5 and 10 with an average of 9.2. Each sample was diluted to the RNA concentration of 100 ng/μl, at the OD260/OD280 ratio of 1.8/2.0. From each RNA sample, 100 ng of RNA was taken for microarray experiments.

treatment_protocol: Before and during 10 nM 17β-estradiol (Sigma Aldrich) treatment, cells were cultured in phenol-red free Dulbecco′s Modified Eagle′s Medium (DMEM) supplemented with 10% charcoal-dextran stripped FBS. Hypoxic conditions were achieved using Forma Series II 3131 cell culture incubator, Thermo Fisher, modified to permit flushing the chamber with a humidified mixture of 5% CO2, 94% N2. Full-length ESR2 cDNA together with N-terminally located HaloTag was cloned into the pWPI-IRES-GFP-PGK-Puro lentiviral expression vector using the Gibson assembly mix (New England Biolabs). ESR2 lentiviruses were produced by transfection of the lentiviral vector into HEK293T packaging cells. Cells were selected using 1 μg/ml of puromycin. More than 80% of cells expressed GFP after three continuous infections were achieved and used for all experiments (HCT116ESR2+, DLD-1ESR2+). To investigate the binding of ERβ to estrogen element response (ERE) sequences on DNA cells were also transduced with ESR2 construct containing mutated DNA binding domain (DBD)- HCT116DBD, DLD-1DBD). Mutagenesis of crucial for binding amino acids E167A and G168A (11) was performed using a megaprimer method (12) and was confirmed by DNA sequencing and functionally by limited ERE luciferase activation compared with WT receptor.

tag: oxygen status

value: hypoxia

value: ESR2 overxpression


title: ESR2_hypoxia_10_nM_17estradiol_rep1

status: Public on Jun 30 2024

molecule: total RNA

organism: Homo sapiens

tag: cell line

tag: genotype

tag: treatment


title: ESR2_hypoxia_10_nM_17estradiol_rep2


title: siCtrl rep1

type: SRA

city: Jena

first: Bjoern

last: von Eyss

address: Beutenbergstr. 11

country: Germany

institute: Leibniz Institute on Aging

library_source: transcriptomic

data_processing:
Reads containing adapters/ with low quality were trimmed using cutadapt (v4.2), -m 30
Trimmed sequence reads were mapped against hg19 using bowtie2 (2.3.5.1)
Read count extraction was performed in R using countOverlaps (GenomicRanges)
Assembly: hg19

source_name: MCF10A cells

growth_protocol: MCF10A cells  were cultured in DMEM/F12 (Thermo Fisher Scientific) supplemented with 5 % horse serum (Thermo Fisher Scientific), 1 % penicillin-streptomycin (Sigma-Aldrich), 10 µg/ml human insulin (Sigma-Aldrich), 0.5 µg/ml hydrocortisone (Sigma-Aldrich), 0.1 µg/ml cholera toxin (Sigma-Aldrich) and 20 ng/ml human EGF (Biomol).

extract_protocol:
RNA was extracted using Rneasy columns (Qiagen). 0.5 µg of RNA were used for library prep.
3'Seq libraries were generated using the QuantSeq 3'-mRNA-Seq Library Prep Kit FWD for Illumina (Lexogen) according to the manufacturer's protocol.

treatment_protocol: MCF10A cells were transfected with siRNAs (20 nM, Dharmacon) using the Lipofectamine RNAiMAX reagent (Thermo) according to the manufacturer's recommendations.

value: MCF10A cells

tag: cell type

value: Mammary breast epithelial

value: Non-targeting control


title: ESR2_hypoxia_control_rep2

source_name: ESR2+ hypoxia control


title: ESR2_hypoxia_control_rep3


title: PACE_019

city: New York City

first: Matthew

last: Muller

state: NY

address: 550 First Avenue

department: Department of Medicine

country: USA

institute: New York University

data_processing:
Fastq qualtiy assessed via FastQC 0.11.7
Sample contamination assessed via fastqscreen v0.13.0
Fastq files trimmed via Trimmomatic v0.36
Fastq files aligned to the HG38 human genome via STAR v2.7.3a
Reads counted via featurecounts v1.6.3
Assembly: hg38
Supplementary files format and content: Raw count table as a .txt file.

molecule: polyA RNA

source_name: whole blood

extract_protocol:
18mLs of whole blood was drawn into 3.2% sodium citrate tubes (BD Vacutainer). Blood was centrifuged to obtain platelet rich plasma, and then depleted of leukocytes and RBCs using StemCell EasySep CD45 and RBC magnetic depletion kits. Purified platelets were pelleted and lysed with 500uL of Qiazol lysis buffer and stored at -80 until ready for RNA extraction. RNA extraction was done by utilizing Zymo Microprep kits (R2062), and stored again at -80 until ready for sequencing.
Sequencing libraries were barcoded and prepared using the Clontech SMART-Seq HT with Nxt HT kit (Takara Bio USA), and libraries sequenced single end on an Illumina NovaSeq 6000

tag: tissue

value: whole blood

value: platelet

tag: age

tag: Sex

value: Male


title: 85co rep3

city: Shenzhen

first: Yan

last: wei

address: shenzhenxueyuanroad

country: China

institute: Southern University of Science and Technology

data_processing:
Bowtie for alignment, v2, default parameter
Cufflink for expression profiling, V2.1.0, default parameter
Assembly: TAIR11
Supplementary files format and content: pyj_raw_data.xlsx represents FPKMs

organism: Arabidopsis thaliana

source_name: seedling

growth_protocol: Surface sterilized seeds were imbibed in water at 4 °C for 3 days and germinated on 1/2 MS agar at standard condition (16h ligth/8h dark, 22 °C) for 7 days

extract_protocol:
Total RNA extracted using Trizol (Invitrogen)
stander Illumina mRNA-seq protocol

treatment_protocol: HS was imposed by submersion of parafilm-sealed plates into a temperature_x0002_controlled water bath

tag: stress

value: 85 min with heat stress


title: SAM cells, sgJUN#1 rep2

source_name: MCF10A SAM

growth_protocol: SAM cells (MCF10A derivatives expressing  were cultured in DMEM/F12 (Thermo Fisher Scientific) supplemented with 5 % horse serum (Thermo Fisher Scientific), 1 % penicillin-streptomycin (Sigma-Aldrich), 10 µg/ml human insulin (Sigma-Aldrich), 0.5 µg/ml hydrocortisone (Sigma-Aldrich), 0.1 µg/ml cholera toxin (Sigma-Aldrich) and 20 ng/ml human EGF (Biomol).

treatment_protocol: MCF10A cells were infected with SAM components, dCas9-VP64 and MCP-p65-HSF1 (kind gift from Feng Zhang, Addgene cat.no. 61425 and 61426), and single clones (SAM cells) were picked. To overexpress JUN, cells were either infected with a non-targeting control (NTC) or a sgRNA targeting the JUN promoter leading to JUN oeverexpression.

value: MCF10A SAM

value: JUN overexpression


title: 85co rep2


title: PACE_020

value: Female


title: SAM cells, sgJUN#1 rep1


title: ESR2_hypoxia_control_rep1


title: PACE_013


title: PACE_014


title: ESR2_normoxia_10_nM_17estradiol_rep3

source_name: ESR2+ normoxia + 10 nM 17β-estradiol

value: normoxia


title: 85co rep1


title: SCC7-IP

city: Wuhan

first: An

last: Song

address: Luoyu Road 237

institute: Wuhan University

library_source: genomic

data_processing:
Raw sequencing data was first filtered by Trimmomatic (version 0.36), low-quality reads were discarded and the reads contaminated with adaptor sequences were trimmed.
The clean reads were used for protein binding site analysis. They were mapped to the reference genome of Mus using STAR software (version 2.5.3a) with default parameters.
The RSeQC（version 2.6）was used for reads distribution analysis.
The MACS2 software（Version 2.1.1）was used for peak calling. The bedtools（Version 2.25.0）was used for peaks annotation and peak distribution analysis.
bigWig files were generated using deepTools.
Assembly: GCF_000001635.26_GRCm38.p6
Supplementary files format and content: bigWig

molecule: genomic DNA

organism: Mus

growth_protocol: DMEM, 15% fetal bovine serum, 2mM glutamine, 1x penicillin/streptomycin. Cells were cultured at 37C at 5% CO2.

extract_protocol:
The tissue/cell was fixed in 1% formaldehyde for 10 min at room temperature by a vacuum pump, after which 0.125 M glycine was added and the mixture was sat for 5 min to terminate the crosslinking reaction. The tissue was then collected and frozen in liquid nitrogen, and grinded by tissue lyser. The grinded powder was treated with lysis buffer and nucleus was collected by centrifuging at 2000g for 5min. Then, nucleus was treated with nucleus lysis buffer and sonicated to fragment chromatin DNA.
The 10% lysis sonicated chromatin was stored and named “input”, and 80% was used in immunoprecipitation reactions with anti-STAT1 antibody（Cell Signaling Technology #14994）and named “IP”, and 10% was incubated with rabbit IgG (Cell Signaling Technology) as a negative control and named “IgG”，respectively. The DNA of input and IP was extracted by phenol-chloroform method. The high-throughput DNA sequencing libraries were prepared by using VAHTS Universal DNA Library Prep Kit for Illumina V3（Catalog NO. ND607, Vazyme）. The library products corresponding to 200-500 bps were enriched, quantified and finally sequenced on Novaseq 6000 sequencer (Illumina) with PE150 model.

value: cancer cell

tag: chip antibody


title: SAM cells, NTC rep3


title: PACE_007


title: PACE_022


title: SCC7-Input


title: SAM cells, NTC rep2


title: SAM cells, NTC rep1


title: SCC7 Input

extract_protocol:
The tissue/cell was fixed in 1% formaldehyde for 10 min at room temperature by a vacuum pump, after which 0.125 M glycine was added and the mixture was sat for 5 min to terminate the crosslinking reaction. The tissue was then collected and frozen in liquid nitrogen, and grinded by tissue lyser. The grinded powder was treated with lysis buffer and nucleus was collected by centrifuging at 2000g for 5min. Then, nucleus was treated with nucleus lysis buffer and sonicated to fragment chromatin DNA.
The 10% lysis sonicated chromatin was stored and named “input”, and 80% was used in immunoprecipitation reactions with anti-IRF-1 antibody（Cell Signaling Technology #8478）and named “IP”, and 10% was incubated with rabbit IgG (Cell Signaling Technology) as a negative control and named “IgG”，respectively. The DNA of input and IP was extracted by phenol-chloroform method. The high-throughput DNA sequencing libraries were prepared by using VAHTS Universal DNA Library Prep Kit for Illumina V3（Catalog NO. ND607, Vazyme）. The library products corresponding to 200-500 bps were enriched, quantified and finally sequenced on Novaseq 6000 sequencer (Illumina) with PE150 model.


title: STARR_10bp_mRNA_rep2

city: Lausanne

first: Bart

last: Deplancke

address: SV Station 19

country: Switzerland

institute: EPFL

data_processing:
Insert seqeunes were mapped to the 12bp barcodes located at the 3 prime end with respect to the TSS using a custom script. STARR activity for each insert sequence was computed by taking the log2 ratio of mRNA over input DNA counts.
Library strategy: STARR-seq

growth_protocol: MEC1 cells were cultured in IMDM media (Gibco) supplemented with 10% Fetal Bovine Serum and 1% Penicillin/Streptomycin.

extract_protocol: Cells were harvested 24h post transfection and lysed in Trizol. RNA was extracted by Chloroform extraction, followed by Isopropanol precipitation with a 1:1 ratio. RNA pellets were resuspended in RNAse-free water and digested with DNAseI for 20 min at room-temperature. DNAseI-treated RNA was purified with a RNA-clean&concentrator-25 kit (Zymo). A total of 10-12 μg RNA was reverse transcribed using the STARR-seq specific RT prime. For each μl of Maxima H minus RT 3 μg input was used. RT reactions were diluted 1:4 with RNAse & DNAse-free water before further amplification. The splice-junction PCR was run using the STARR-seq thio-splice-junction primer and a custom reverse primer spanning the SalI restriction site used for linearizing the STARR-seq vector and the read1 Nextera adapter sequence inserted with the library. About one third of the RT reaction was amplified for 19-22 cycles. Illumina barcodes were added by a final PCR with 8 cycles using the Illumina Nextera guidelines. After each amplification step, PCR products were cleaned up using AMPure XP magenetic beads (Beckman Coulter). Plasmid libraries were directly amplified with a 8-cycle PCR using NEXTERA-specific index primers.

treatment_protocol: Transfections of 5-bp and 10-bp libraries were done in duplicates using ~ 6 Mio cells and 30 μg plasmid per replicate. Transfections were carried out with the Neon Transfection System (Thermo Fisher) as follows: 3 pulses of 20 ms at 1200 V in R buffer using 100 ul tips.

value: CLL cell line

value: WT

value: not applicable


title: SCC7 IP


title: PACE_005


title: ESR2_normoxia_10_nM_17estradiol_rep2


title: PACE_003


title: STARR_10bp_mRNA_rep1


title: ESR2_normoxia_control_rep1

source_name: ESR2+ normoxia control


title: RS60punch

city: Montreal

first: Gilbert

last: Bernier

state: Quebec

address: 5690 rosemont boulevard

country: Canada

institute: HMR

description: 10x Genomics

library_source: transcriptomic single cell

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were made using the Cell Ranger software v7.0.1 (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger)
Assembly: hg38
Supplementary files format and content: loupe files produced by Cell Ranger 7.0.1
Supplementary files format and content: h5 files produced by Cell Ranger 7.0.1

source_name: differentiated iPSC cells

extract_protocol:
RS cells were freshly collected from the plates, resuspended in a dissociation buffer to obtain single cells, and then strained to ensure analysis of only single cells. Viability was determined by staining for live cells.
Library was performed according to the manufacter’s instructions (single cell 3’ v3.1 protocol, 10x Genomics). Briefly, RS cells were resuspended in the master mix and loaded together with partitioning oil and gel beads into the chip to generate the gel bead-in-emulsion (GEM). The poly-A RNA from the cell lysate contained in every single GEM was retrotranscripted to cDNA, which contains an Ilumina R1 primer sequence, Unique Molecular Identifier (UMI) and the 10x Barcode. The pooled barcoded cDNA was then cleaned up with Silane DynaBeads, amplified by PCR and the apropiated sized fragments were selected with SPRIselect reagent for subsequent library construction. During the library construction Ilumina R2 primer sequence, paired-end constructs with P5 and P7 sequences and a sample index were added.

value: differentiated iPSC cells

value: iPSC

value: retinal sheet

value: differentiated for 60 days, then selected area where analyzed


title: ESR2_normoxia_control_rep2

value: ESR2 overxpression


title: AcaN3

city: Changchun city

first: Tingdi

last: Zhang

state: Jilin Province

address: 2519 Jiefang Road

department: college of new energy and environment

institute: Jilin University

data_processing:
The clustering of the index-coded samples was performed on a cBot Cluster Generation System using TruSeq PE Cluster Kit v3-cBot-HS (Illumia) according to the manufacturer’s instructions. After cluster generation, the library preparations were sequenced on an Illumina Novaseq platform and 150 bp paired-end reads were generated
Rockhopper was used to identify novel genes, operon, transcription start site (TSS), transcription terminal site (TTS) and Cis-natural antisense transcripts
HTSeq v0.6.1 was used to count the reads numbers mapped to each gene. And then FPKM (Fragments Per Kilobase of transcript sequence per Millions base pairs) of each gene was calculated based on the length of the gene and reads count mapped to this gene.
Differential expression analysis of Acinetobacter calcoaceticus strain Aca13 induced with substrates was performed using the DESeq R package.
KOBAS software was used to test the statistical enrichment of differential expression genes in KEGG pathways.
Assembly: NZ_CP088954
Supplementary files format and content: bam file includes processed data file for each sample.
Supplementary files format and content: raw file includes raw data file for each sample.

organism: Acinetobacter calcoaceticus

source_name: strain Aca13

growth_protocol: LB

extract_protocol:
extraction of total RNA, detection of RNA integrity and total RNA content,  then remove Ribosome RNA (rRNA) from total RNA to obtain mRNA. Subsequently, a fragmentation buffer was added to randomly break the obtained mRNA into short fragments and construct the library in a chain specific manner.
The cDNA libraries for RNA sequencing were constructed according to the chain specific library by the NEBNext Ultra Directional RNA Library Prep Kit for (Illumina) as per manufacturer’s instructions (NEB #E7420S)

treatment_protocol: Acinetobacter calcoaceticus strain Aca13 was incubated in the fresh LB broth until logarithmic growth period, add hexadecane or naphthalene, respectively.  RNA protect-Bacteria- Reagent was used to protect RNA after 3 hours induction. Then bacteria were collected for total RNA extraction.

tag: strain

value: strain Aca13

value: naphthalene


title: ESR2_normoxia_control_rep3


title: RS60

description: 10x Genomics

value: differentiated for 60 days


title: CID44971_CHUNKS2_ADT

city: Darlinghurst

first: Alexander

last: Swarbrick

state: New South Wales

address: 384 Victoria Street

department: Cancer

country: Australia

institute: Garvan Institute of Medical Research

data_processing:
FASTQ's from RNA assay files were demultiplexed and mapped to reference genome GRCh38 and mm10 using 10X Genomics Cell Ranger v3.0.2/v3.1 to generate count matrices. FASTQ's  from ADT assay were demultiplexed and mapped to Biolegend TotalSeq-A Barcodes using CITE-seq-Count v1.4.4 (https://github.com/Hoohm/CITE-seq-Count) to generate count matrices.
Assembly: GRCh38, mm10
Supplementary files format and content: Processed data files in MM matrix format (*.mtx), genes and barcode IDs (*.tsv) and cellranger summary out (*.html)

source_name: Mammary Tumor

extract_protocol:
Single-cell sequencing was performed using the Chromium Single-Cell v2/v3 3’ Chemistry Library, Gel Bead, Multiplex and Chip Kits (10X Genomics) as per manufacturer's protocol. A total of 3,000 to 5,000 cells were targeted per patient.
Libraries were sequenced on the NextSeq 500 platform (Illumina) with pair-ended sequencing and dual indexing. For RNA libraries, a total of 28, 8 and 98 cycles were run for Read 1, i7 index and Read 2, respectively. For ADT libraries, a total of 28, 8, 24 cycles.

value: TNBC

tag: gender

value: female

value: Primary Breast Tumor


title: CID44971_CHUNKS2_RNA


title: AcaN2


title: RS30

description: 10x Genomics

value: differentiated for 30 days


title: ESR2_normoxia_10_nM_17estradiol_rep1

value: normoxia


title: CID4660_CID4664_CID4676_ADT


title: AcaN1


title: AcaH3

value: Hexadecane


title: RS5

description: 10x Genomics

value: differentiated for 5 days


title: hypoxia_10_nM_17estradiol_rep2

source_name: hypoxia + 10 nM 17β-estradiol

value: hypoxia

value: WT


title: CID4513_ADT


title: K562, CDAN1-shRNA-c2, without Doxycycline, Day4

city: Hong Kong

first: Margaret HL

last: Ng

address: Prince of Wales Hospital

department: Department of Anatomical and Cellular Pathology

country: Hong Kong

institute: The Chinese University of Hong Kong

data_processing:
Transcripts per million (TPM) were quantified by salmon (v1.5.2) with human transcriptome (ensembl annotation version GRCh37.75)
Data processing was performed using the Ubuntu 18.04.6 platform.
Assembly: hg19
Supplementary files format and content: tab-delimited text files include TPM values for each Sample

growth_protocol: K562 cells were maintained in RPMI 1640 supplemented with 10% fetal bovine serum (FBS), 1% GlutaMAX, and 100U/ml Penicillin-Streptomycin in humidified atmosphere with 5% CO2 at 37C

extract_protocol:
RNA was harvested using QIAamp RNA Blood Mini Kit (Qiagen). 1 ug of total RNA was used for the construction of sequencing libraries.
RNA libraries for RNA-seq were prepared using NEBNext Ultra RNA Library Prep Kit for Illumina following manufacturer's protocols.

treatment_protocol: Puromycin at 2ug/ml was used to maintain the clones. shRNA expression was induced with 1ug/ml Doxcycline treatment for 4 days.

value: erythroleukemic

value: without Doxycycline


title: AcaH1


title: hypoxia_control_rep2

source_name: hypoxia control


title: hypoxia_10_nM_17estradiol_rep3


title: CID4513_RNA


title: hypoxia_control_rep3


title: hypoxia_control_rep1


title: hypoxia_10_nM_17estradiol_rep1


title: K562, CDAN1-shRNA-c1, with Doxycycline, Day4

value: with Doxycycline


title: Aca3

value: control


title: normoxia_10_nM_17estradiol_rep3

source_name: normoxia + 10 nM 17β-estradiol


title: Aca2


title: K562, CDAN1-shRNA-c1, without Doxycycline, Day4


title: Aca1


title: K562, NT-shRNA, without Doxycycline, Day4

value: NT-shRNA


title: CID4660_CID4664_CID4676_RNA


title: AcaH2

value: strain Aca13


title: K562, NT-shRNA, with Doxycycline, Day4


title: CID3838_ADT


title: K562, CDAN1-shRNA-c2, with Doxycycline, Day4


title: MCF7_2011

type: genomic

city: Heidelberg

first: Thomas

middle: Georg Philipp

last: Grunewald

address: Im Neuenheimer Feld 280

department: Division of Translational Pediatric Sarcoma Research

institute: German Cancer Research Center (DKFZ)

hyb_protocol: not provided

scan_protocol: not provided

data_processing: The initial processing and quality control (QC) of the raw genotyping data was performed using PLINK version 1.9 (SNP call rate >95%, Hardy-Weinberg exact test <1e-6, and variants on Y chromosome were excluded). In total 526,610 variants out of 696,726 passed the QC filters.

label: Cy3, Cy5

source_name: human breast carcinoma cell, MCF-7, WT

label_protocol: not provided

growth_protocol: All cells were maintained in RPMI 1640 supplemented with 10% fetal bovine serum (FBS) and and 1% penicillin-streptomycin in humidified atmosphere with 5% CO2 at 37C

extract_protocol: DNA was extracted with the NucleoSpin Tissue kit (Macherey Nagel) following the manufacturer’s protocol

treatment_protocol: no treatment

value: breast carcinoma cell, MCF-7, WT

tag: timepoint

value: Initial


title: LS180_RiboSeq_Rep2

city: Debrecen

first: Beáta

last: Scholtz

address: Nagyerdei krt. 98

department: Department of Biochemistry and Molecular Biology

country: Hungary

institute: University of Debrecen, Faculty of Medicine

description: stranded library

data_processing:
All processing steps were done on the Galaxy platform (usegalaxy.eu). The sequencing quality of the raw data was first checked with FASTQC.
UMI-tools extract.  # UMI-tools version: 1.1.2 --extract-method=string --bc-pattern=NNNNNNNNNNNN
Cutadapt 1.  # Tool version 4.0.  3' (End) Adapters: Custom - AGATCGGAAGAGCACACGTCTGAA. Cut bases from reads before adapter trimming: 4
Cutadapt 2.  # Tool version 4.0. Cut bases from reads before adapter trimming: -10
Trimmomatic. # Tool version 0.32.1; Perform initial ILLUMINACLIP step? - Yes. Custom: GTGACTGGAGCACACGTCTGAACTCCAGTCACGAGTGGATCTCG; SLIDINGWINDOW:4:20; MINLEN:16
Quality control: FASTQC
Bowtie alignment. #Tool version:1.2.0. Reference genome from history: hg38 rRNA and tRNA sequences in fasta format. indexSettings="indexPreSet" --paired="single" --input1=file produced by Trimmomatic --params="full" --skip="0" --alignLimit="-1" --trimH="0" --trimL="0" --mismatchSeed="2" --mismatchQual="70" --seedLen="25" --rounding="round" --forwardAlign="forward" --reverseAlign="noReverse" --tryHard="noTryHard" --allValAligns="noAllValAligns" --valAlign="1" --suppressAlign="-1" --best="noBest" --maxBacktracks="125" --offrate="-1" --seed="-1"
HISAT2: Align unmapped reads from Bowtie to the built-in hg38 reference genome, default settings. #Tool version:2.2.1
Ribotaper part 1. #Tool version: 1.3.1a. Gtf file: Gencode v40 basic annotation file, hg38.  Fasta file: hg38, GRCh38.primary_assembly.genome.fa.gz.Use ccdsid? -True. Use appris? -False
Ribotaper part 2. #Tool version: 1.3.1a. Ribo-seq alignment file: from HISAT2. Start_stops_far: from Ribotaper part 1.
Custom Galaxy workflows to calculate P-site read counts and ribo-seq coverage for selected genes
Bam-to-wiggle conversion. # Tool version rseqc_bam2wig/2.6.4. Chromosome size table was copied from the HISAT2 bam file. Skip multihits; MAPQ:20; Do not normalize; Single; positive --> positive; negative --> negative
Wig/Bedgraph-to-bigwig converter. # Wig-to-bigwig, Tool version 1.1.1.  Default settings
Assembly: HISAT2:hg38. Bowtie:Human tRNA sequences were downloaded from http://tpsic.igcz.poznan.pl. Human rRNA sequences (LSU and SSU) were downloaded from https://www.arb-silva.de/. The three fasta files were concatenated in Galaxy, and used as the tRNA_rRNA reference genome file for the Bowtie alignment.
Supplementary files format and content: bigwig generated from HISAT2 bam file, forward reads
Supplementary files format and content: bigwig generated from HISAT2 bam file, reverse reads
Library strategy: ribosome profiling

growth_protocol: LS180 cells were cultured in DMEM/10% FBS with penicillin-streptomycin (100 U and 100 mg/mL, respectively). Cells were cultured in 75 cm2 tissue culture flasks at 37ºC in a humidified atmosphere containing 5% (v/v) CO2, and were refed every 3 days until reaching 60-80% confluency. The cultures were last refed 6 hours prior harvesting.  Culture flasks were removed from the incubator and placed on ice. The medium was removed by aspiration, and the cells were washed in the flask with freshly prepared 8 ml ice-cold PBS/100 ug/ml cycloheximide. The cells were scraped into 4 ml ice-cold PBS/cycloheximide, and centrifuged gently at 4 C to pellet the cells. The supernatant was aspirated, the cell pellet was snap-frozen in lN2, and stored at - 80C until sending to EIRNA Bio Ltd. for sequencing.

extract_protocol:
Two cell pellets (each from a T75 tissue culture flask) were lysed in ice-cold polysome isolation buffer supplemented with cycloheximide. Lysates were RNase treated before ribosomes were enriched by size exclusion chromatography. Following RNA purification and size selection of ribosome protected mRNA fragments on urea PAGE gels, contaminating rRNA was depleted from samples using EIRNA Bio custom biotinylated rRNA depletion oligos before the enriched fragments were converted into Illumina compatible cDNA libraries.
stranded

treatment_protocol: cycloheximide at the time of harvesting and during RNA isolation

value: colon adenocarcinoma

value: cycloheximide


title: MCF7_2004


title: CID4040_ADT

value: ER+

value: female

value: Primary Breast Tumor


title: CID4040_RNA


title: HELA_0_FTP

source_name: human cervix carcinoma cell, HeLa, WT

value: cervix carcinoma cell, HeLa, WT

value: 6 months


title: LS180_RiboSeq_Rep1

description: stranded library


title: Mev3 48

city: Miami

first: Marjana

last: Tomic-Canic

state: Fl

address: 1600 NW 10th Ave

institute: University of Miami

description: processed data file: mCanicRNAseq.xlsx

data_processing:
Raw sequence data was processed by the on-instrument Real Time Analysis software (v.2.4.11) to basecall files.
These were converted to de-multiplexed FASTQ files with the Illumina supplied scripts in the BCL2FASTQ software (v2.17).
Reads were aligned to the human reference genome (hg19) with the STAR aligner (v2.5.0a).
Gene count quantification for total RNA was performed using the GeneCounts function within STAR against the GENCODE v19 human transcript .gtf file.
Genome_build: hg19 (GRCh37)
Supplementary_files_format_and_content: mCanicRNAseq.xlsx: Excel file with raw read counts, normalized CPM, fold change, p-value and FDR.

source_name: Skin

growth_protocol: Primary human keratinocytes were grown to approximately 90% confluency and switched to basal media for 5 hours.

extract_protocol: RNA libraries were prepared for sequencing using standard Illumina protocols.

treatment_protocol: Cells were treated with 5 µM mevastatin 3x daily every 4 hours for 48 hours.

tag: diagnosis

value: Diabetic foot ulcers

value: Primary human keratinocytes

value: Mevastatin


title: normoxia_10_nM_17estradiol_rep2

value: normoxia

value: WT


title: normoxia_10_nM_17estradiol_rep1


title: LS180_RNASeq_Rep2

description: stranded library

data_processing:
All processing steps were done on the Galaxy platform (usegalaxy.eu). The sequencing quality of the raw data was first checked with FASTQC.
Trimmomatic. # Tool version 0.32.1; Perform initial ILLUMINACLIP step? - Yes. >read1 AGATCGGAAGAGCACACGTCTGAACTCCAGTCA >read2 AGATCGGAAGAGCGTCGTGTAGGGAAAGAGTGT; SLIDINGWINDOW:4:20; MINLEN:16
FASTQC
HISAT2: Align reads after Trimmomatic correction to the built-in hg38 reference genome, default settings. #Tool version:2.2.1
Bam-to-wiggle conversion. # Tool version rseqc_bam2wig/2.6.4. Chromosome size table was copied from the HISAT2 bam file. Skip multihits; MAPQ:20; Do not normalize; Pair-end RNA-seq; read1 (positive --> positive; negative --> negative), read2 (positive --> negative; negative --> positive)
Wig/Bedgraph-to-bigwig converter. # Wig-to-bigwig, Tool version 1.1.1.  Default settings
Assembly: hg38
Supplementary files format and content: bigwig generated from HISAT2 bam file, forward reads
Supplementary files format and content: bigwig generated from HISAT2 bam file, reverse reads

growth_protocol: LS180 cells were cultured in DMEM/10% FBS with penicillin-streptomycin (100 U and 100 mg/mL, respectively). Cells were cultured in 75 cm2 tissue culture flasks at 37ºC in a humidified atmosphere containing 5% (v/v) CO2, and were refed every 3 days until reaching 60-80% confluency. The cultures were last refed 6 hours prior harvesting.  Culture flasks were removed from the incubator and placed on ice. The medium was removed by aspiration, and the cells were washed in the flask with freshly prepared 8 ml ice-cold PBS/100 ug/ml cycloheximide. After washing, 4 ml Tri-Reagent was added to the flask, and the cells were lysed in situ. The lysate was transferred into a fresh 15 ml tube, was snap-frozen in lN2, and stored at - 80C until sending to EIRNA Bio Ltd. for sequencing.

extract_protocol:
For stranded mRNA-seq, total RNA was extracted from two Tri-Reagent lysed pellets, then mRNA was poly(A)-enriched, fractionated and converted into Illumina compatible cDNA libraries.
stranded

treatment_protocol: cycloheximide at the time of harvesting


title: normoxia_control_rep2

source_name: normoxia control


title: normoxia_control_rep3


title: MCF7_0_FTP


title: Veh3 48

description: processed data file: mCanicRNAseq.xlsx

value: Vehicle


title: Mev2 48

description: processed data file: mCanicRNAseq.xlsx


title: normoxia_control_rep1


title: HELA_0_ITP


title: CID3838_RNA


title: LS180_RNASeq_Rep1

description: stranded library


title: Veh2 48

description: processed data file: mCanicRNAseq.xlsx


title: Mev1 48

description: processed data file: mCanicRNAseq.xlsx


title: MCF7_0_ITP


title: igG

city: Trento

first: Erik

last: Dassi

state: TN

address: Via Sommarive, 9

department: CIBIO

country: Italy

institute: University of Trento

data_processing:
Illumina Casava 1.8 software was used for basecalling.
UMI tools was applied for UMIs removal (v0.5.5; parameters: --bc-pattern NNNNXXXXXNNN). Reads were demultiplexed and 5bp length barcodes were removed with Cutadapt (v1.18; parameters demultiplxing and barcode removal: --no-indels --times 1 -e 0.2 -m 18 ). Sequenced reads were trimmed for adaptor sequence (AGATCGGAAGAGCGGTTCAG) and low-quality nucleotides, and then quantified against the Gencode v29 genome using STAR. Peaks were called with ENCODE eCLIP standard pipeline ("Clipper" for peaks calling and ENCODE custom script "merge_peaks" for peaks normalization)
Genome_build: GRCh38.p12
Supplementary_files_format_and_content: Peaks called with Clipper. Bed8 file with columns: chromosome, genomic_start, genomic_stop, cluster_name (contains:gene name, the number of reads in the cluster and a counter to give the cluster a unique identifier), min_pval, strand, thick_start, thick_stop

source_name: CHP-212 cells

growth_protocol: CHP-212 cell line was cultured in 1:1 ratio of Minimum Essential Medium and Ham’s F12 supplemented with 10% FBS, 1% L-Glutamine, 1% Penicillin/Streptomycin,1% Non-essential Aminoacids.

extract_protocol:
Total RNA of cells was extracted with the RNeasy Plus Mini Kit (QIAgen). Poly(A)+ RNA was purified from 75ug of total  RNA using Dynabead Oligo(dT)25 kit (61002) and chemically fragmented with a fragmentation buffer (ThermoFisher Scientific). Poly(A)+ RNA was subjected to m6A-immunoprecipitation using a rabbit polyclonal anti-m6A antibody (Abcam, ab151230) and rabbit IgG as control, and UV crosslinkinked. To isolate proper fragments, RNA was NuPAGE separated and transferred to a nitrocellulose membrane.
The isolated RNA was ligated to a barcoded adaptor to enable multiplexing of samples. Reverse transcription was carried out using unique barcoded primers for each sample, and the respective cDNA was amplified and cleaned using AMPure XP beads (Beckman Coulter). QC analysis was carried out using Tapestation (Agilent) and QuantiFluor (Promega).

treatment_protocol: All recombinant plasmids were generated using GatewayTM Technology (ThermoFisher Scientific) according to the manufacturer’s instructions. METTL14 (NC_000004.12), and ALKBH5 (NC_000017.11) human genes were subcloned into a pENTR1A no ccDB entry vector (Addgene, w48-1) and recombined into a third-generation pLenti CMV Puro DEST (w118-1) (Addgene, #17452) to perform overexpression experiments.

value: igG

tag: antibody

value: rabbit IgG as control


title: h2O


title: Veh1 48

description: processed data file: mCanicRNAseq.xlsx


title: MHH-ES1_FTP

source_name: human EwS cell, MHH-ES-1, WT

value: EwS cell, MHH-ES-1, WT

value: 12 months


title: rcALKBH5  input RNA replicate 1

value: ALKBH5 rescue


title: PC24 organoids

city: Tokyo

first: Masayuki

last: Fujii

address: 35, Shinanomachi, Shinjuku-ku

country: Japan

institute: Keio University

hyb_protocol: Standard Affymetrix procedures were used for cDNA hybridization.

scan_protocol: GeneCips were scanned using GeneChip Scanner 3000 7G system.

data_processing: Raw CEL files were proccessed using the robust multi-array average expression measure using the rna function in the affy R Bioconductor package.

source_name: Patient-derived pancreatic cancer organoids

label_protocol: Biotinated RNA was prepared as per Affymetrix protocol.

growth_protocol: The organoids were plated as single cells and cutured with EGF, Noggin and R-spondin for seven days.

extract_protocol: Total RNA was collected using Rneasy Plus Mini Kt according to the manufacturer's instruction.

treatment_protocol: All the samples were equally maintained in the above condition from passage.

value: Pancreas


title: PC35 organoids


title: SKES_FTP

source_name: human EwS cell, SK-ES-1, WT

value: EwS cell, SK-ES-1, WT


title: PC24 with control shRNA infection

source_name: Genetically engineered pancreatic cancer organoids


title: oeMETTL14 input RNA replicate 1

value: METTL14 overexpression


title: PC35 with TP63 knockout


title: PC24 GATA6 knockdown TP63 knockout organoids with delta-Np63 overexpression


title: KCT#1 organoids

source_name: Genetically engineered normal-like duct organoids


title: TC71_FTP

source_name: human EwS cell, TC71, WT

value: EwS cell, TC71, WT


title: control input RNA replicate 1

value: wild type


title: A673_FTP

source_name: human EwS cell, A673, WT

value: EwS cell, A673, WT


title: SKNMC_FTP

source_name: human EwS cell, SK-N-MC, WT

value: EwS cell, SK-N-MC, WT


title: oeMETTL14  IP RNA replicate 3

value: rabbit polyclonal anti-m6A antibody (Abcam, ab151230)


title: STEC_OFF_51, rep2

city: Gold Coast

first: John

middle: Michael

last: Atack

state: QLD

address: Gold Coast Campus

department: Institute for Glycomics

institute: Griffith University

data_processing:
Illumina bcl2fastq 2.20.0.422 pipeline.
Bowtie aligner (2.3.2) was used to map reads to the Escherichia coli O104-H4 STEC strain 2011C-3493 complete genome; GenBank accession number CP003289.1.
edgeR version 3.18.1 was used to perform differential expression analysis.
Genome_build: Escherichia coli O104-H4 STEC strain 2011C-3493 (CP003289.1).
Supplementary_files_format_and_content: STECmod_gene_counts.txt: Tab-delimited text file includes abundance measurements.
Supplementary_files_format_and_content: STECmod_ON.VS.OFF-all_genes.txt: Tab-delimited text file includes abundance measurements and expression differences STECmod ON 52reps vs. STECmod OFF 51reps.

organism: Escherichia coli

source_name: Shiga toxin-producing Escherichia coli strain FHI96

growth_protocol: Mid-log cultures in LB broth.

extract_protocol:
RNA was isolated using Trizol according to manufacturer's protocol.
RNA libraries were prepared for sequencing using standard Illumina TruSeq protocols.

tag: genotype/variation

value: STECmod OFF 51reps

tag: presence or absence of methyltransferase

value: Absent


title: STEC_OFF_51, rep3


title: SKNMC_MTP

value: 6 months


title: MHH-ES1_MTP


title: STEC_OFF_51, rep1


title: rcALKBH5  IP RNA replicate 3


title: SKES_MTP


title: control  IP RNA replicate 3


title: rcALKBH5  IP RNA replicate 2


title: KCT#1 GATA6 knockdown organoids


title: WT small intestine, E18, rep2

city: Houston

first: Frank

last: McKeon

state: TX

address: 3455 Cullen Blvd

department: Biology and Biochemistry

institute: University of Houston

description: Gene expression data from stem cells

hyb_protocol: GeneChip® Hybridization, Wash, and Stain Kit (affymetrix)

data_processing: Expression console, Partek Genome Suite 6.6

label: Biotin

organism: Mus musculus

source_name: WT small intestine, E18

label_protocol: 3' IVT express kit (affymetrix)

extract_protocol: Trizol (Life Technologies)

treatment_protocol: Tissues were collected from p63 KO and WT mice.

value: wildtype

value: small intestine

tag: developmental stage

value: Balb/c


title: STEC_ON_52, rep3

value: STECmod ON 52reps

value: Present


title: WT proximal stomach, E18, rep1

description: Gene expression data from stem cells

scan_protocol: GeneChip® Hybridization, Wash, and Stain Kit (affymetrix)

source_name: WT proximal stomach, E18

value: proximal stomach


title: KCT#1 GATA6 knockdown organoids with delta-Np63 overexpression replicate 1

value: Pancreas


title: oeMETTL14  IP RNA replicate 2


title: PC24 GATA6 knockdown organoids


title: STEC_ON_52, rep2


title: TC71_MTP


title: WT proximal stomach, E18, rep2

description: Gene expression data from stem cells


title: control  IP RNA replicate 2


title: WT large intestine, E18, rep2

description: Gene expression data from stem cells

source_name: WT large intestine, E18

value: large intestine


title: A673_MTP


title: A673/SHEF_ITP

source_name: human EwS cell, A673, shEWSR1-FLI1

value: EwS cell, A673, shEWSR1-FLI1

value: Initial


title: STEC_ON_52, rep1


title: oeMETTL14 IP RNA replicate 1

value: METTL14 overexpression


title: Spontaneous hypertension rat, treated with dapagliflozin(10mg/kg/d) for 8 weeks [SGLT2_3]

city: 重庆

first: Jianlin

last: Du

state: 重庆

address: 74 Lin Jiang Road

institute: Department of Cardiology, The Second Affiliated Hospital of Chongqing Medical University

data_processing:
SOAPnuke (v1.5.2)
Sequenced reads were trimmed for adaptor sequence by SOAPnuke (v1.5.2), afterwards clean reads were obtained and stored in FASTQ format.
Clean reads were mapped to Rattus genome GCF_000001895.5_Rnor_6.0 using HISAT2(v2.0.4)
Bowtie2 (v2.2.5) was applied to align the clean reads to Rattus genome GCF_000001895.5_Rnor_6.0
Sequenced Fragments Per Kilobase of exon per Million fragments mapped to Rattus genome (FPKM) were calculated using RESEM(v1.2.12).
Supplementary files format and content: tab-delimited text file include FPKM values for each sample.

organism: Rattus norvegicus

source_name: kidney

growth_protocol: Wistar-Kyoto rats (WKY) (n = 3) and spontaneously hypertensive rats (n = 6) aged three months and weighing 210-240g were housed at the Experimental Animal Centre of Chong Qing Medical University (CQMU, China) for more than one week before the following experimental procedures. Then, the spontaneously hypertensive rats were randomly divided into two groups: the SHR group (n=3) and the SGLT2 group (n=3).

extract_protocol:
Total RNA was extracted from the rat tissues using Trizol (Invitrogen, Carlsbad, CA, USA) according to the manufacturer’s instructions.1.3 ug of total RNA was used for the construction of sequencing libraries.
the final library was generated by heated denaturing and circularizing the above double stranded PCR products using the splint oligo sequence, then followed by formatting the single strand circle DNA (ssCir DNA). DNA nanoballs (DNBs) were generated by library amplification and loaded into the patterned nanoarray and single end 50 bases reads on BGIseq500 platform (BGI-Shenzhen, China)

treatment_protocol: We fed dapagliflozin 10 mg/kg/d to the SGLT-2i group and equal amounts of distilled water orally to the WKY and SHR groups for the next eight weeks.

value: Wistar-Kyoto

value: kidney

value: Spontaneous hypertension rat

value: Dapagliflozin

tag: time

value: 8 weeks


title: rcALKBH5  IP RNA replicate 1

tag: genotype

value: ALKBH5 rescue

value: rabbit polyclonal anti-m6A antibody (Abcam, ab151230)


title: WT large intestine, E18, rep1

description: Gene expression data from stem cells

value: wildtype


title: A673/SHCTRL_ITP

source_name: human EwS cell, A673, shControl

value: EwS cell, A673, shControl


title: WT distal stomach, E18, rep1

description: Gene expression data from stem cells

source_name: WT distal stomach, E18

value: distal stomach

value: Balb/c


title: WT small intestine, E18, rep1

description: Gene expression data from stem cells

value: small intestine


title: Spontaneous hypertension rat, treated with dapagliflozin(10mg/kg/d) for 8 weeks [SGLT2_2]

tag: treatment


title: SKNMC_ITP

value: EwS cell, SK-N-MC, WT


title: KCT#1 GATA6 knockdown organoids with delta-Np63 overexpression replicate 2

value: Pancreas


title: control IP RNA replicate 1

source_name: CHP-212 cells

value: wild type


title: Spontaneous hypertension rat, treated with dapagliflozin(10mg/kg/d) for 8 weeks [SGLT2_1]


title: WT bladder, E19, rep2

description: Gene expression data from stem cells

source_name: WT bladder, E19

value: bladder


title: MHH-ES1_ITP

source_name: human EwS cell, MHH-ES-1, WT

value: EwS cell, MHH-ES-1, WT

value: Initial


title: Gastric adenocarcinoma cells - BGC823 shNAT10-2

city: GUANGZHOU

first: Min

last: Deng

state: GUANGDONG

address: Affiliated Cancer Hospital & Institute of Guangzhou Medical University, NO.78, Hengzhigang Road.

data_processing:
Cutadapt (v2.5 ) was used to trim adapters and filter for sequences
150 bp paired-end reads were mapped to the reference genome by using hisat2 (v2.1.0 ).
The reads mapped the genome were calculated using HT-seq (v0.7.2).
Genome_build: GRCh38
Supplementary_files_format_and_content: The reads counts table

source_name: BGC823 shNAT10

extract_protocol:
Total RNA was isolated from AGS with or without NAT10 knockout and BGC823 cells expressing shNAT10 or the control shRNA using TRIzol reagent. Poly(A)+ mRNA was subsequently purified from total RNA via Dynabeads mRNA Purification Kit (Invitrogen),and then sequenced on the Illumina NovaSeq 6000 platform.
RNA libraries were prepared for sequencing using standard Illumina6000 protocols

tag: cell line

tag: cell type

value: Gastric adenocarcinoma cells


title: SKES_ITP

source_name: human EwS cell, SK-ES-1, WT

tag: tissue

value: EwS cell, SK-ES-1, WT


title: Gastric adenocarcinoma cells - BGC823 shNAT10-1

institute: Affiliated Cancer Hospital & Institute of Guangzhou Medical University


title: Spontaneous hypertension rat [SHR_2]

value: Wistar-Kyoto

value: kidney

value: Spontaneous hypertension rat

value: Saline


title: Gastric adenocarcinoma cells - BGC823 shCTR-2

source_name: BGC823 shCTR

value: shCTR


title: TC71_ITP

source_name: human EwS cell, TC71, WT

value: EwS cell, TC71, WT


title: Spontaneous hypertension rat [SHR_1]

value: 8 weeks


title: A673_ITP

source_name: human EwS cell, A673, WT

value: EwS cell, A673, WT


title: Gastric adenocarcinoma cells - BGC823 shCTR-1


title: Wistar-Kyoto rats [WKY_3]

value: normal control


title: Gastric adenocarcinoma cells - AGS KO NAT10-2

source_name: AGS KO NAT10

value: AGS

value: KO NAT10


title: WT distal stomach, E18, rep2

description: Gene expression data from stem cells

value: wildtype

value: distal stomach

value: Balb/c


title: Wistar-Kyoto rats [WKY_2]


title: A673 HK_ITP

value: Initial


title: Spontaneous hypertension rat [SHR_3]


title: A673 TG_ITP


title: Wistar-Kyoto rats [WKY_1]


title: Gastric adenocarcinoma cells - AGS CTR-2

source_name: AGS CTR

value: Gastric adenocarcinoma cells

value: CTR


title: Gastric adenocarcinoma cells - AGS KO NAT10-1


title: WT bladder, E18, rep2

description: Gene expression data from stem cells

source_name: WT bladder, E18

value: bladder


title: Gastric adenocarcinoma cells - AGS CTR-1


title: A673 EDA_ITP


title: WT bladder, E18, rep1

description: Gene expression data from stem cells


title: AJ21: RBP A C4 NC3 VSMC HIGH

city: Cambridge

first: Irina

last: Mohorianu

address: Puddicombe Way

department: Wellcome-MRC Cambridge Stem Cell Institute

country: United Kingdom

institute: University of Cambridge

data_processing:
Quality checks were performed using FastQC (v0.11.2), summarised with MultiQC (v1.8). Adapter trimming was performed using TrimGalore (v0.6.4_dev) and alignment to the human reference genome (build GRCh38.p13) was performed using STAR (STAR_2.5.2a) with default parameters. Density plots, violin plots, MA plots, Jaccard similarity index, dendrograms and PCA were performed as further quality checks
The data was normalised using quantile normalisation and noise was removed using a fixed threshold of 100 (only genes entirely under the threshold were discarded and abundances <100 were set to 100). An alternative noise removal method based on [Mohorianu et al, 2017; https://doi.org/10.1371/journal.pone.0182694] was also implemented. Differential expression analysiss was performed using edgeR (v3.28.1).
Enrichment analysis on the set of differentially expressed genes against a background of all genes expressed in the dataset using the g:profiler (https://biit.cs.ut.ee/gprofiler/gost) R package (v0.1.8).
Alternative splicing analysis was performed using rMATS [Shen et al, 2014; https://doi.org/10.1073/pnas.1419161111]
Genome_build: GRCh38.p13
Supplementary_files_format_and_content: Csv of raw and normalised abundances. Rows represent features and columns represent samples. Values are raw in the first instance, and normalised with noise removed in the second.

source_name: hES-VSMCs

growth_protocol: Human Embryonic stem cell derived Vascular Smooth Muscle Cells (neural crest lineage) were cultured in DMEM media supplemented with 10% FBS, Pen/Strep and L-GLutamine for 10-14 days post differentiation in 10cm dishes (BD Falcon) coated with 0.1% Gelatin in PBS. Protocols for differentiation from hES to Neural crest to VSMCs - refer Granata et al, 2017 (PMID 27893734).

extract_protocol:
Cells were harvested by trypsinization and resuspended in FACS buffer (1% FBS in PBS supplemented with 2mM EDTA. Cells were sorted (BD FACSAria III) for Low, medium and High intensity GFP and collected in FACS Buffer. Sorted cells were spun down and resuspended in RLT buffer (RNAeasy, Qiagen) and frozen. All samples were extracted simultaneously using the RNeasy kit and resuspended in RNAse free water.
NovaSeq, PolyA selection beforehand (NEBNext® Poly(A) mRNA Magnetic Isolation Module, E7490),  NEBNext® Ultra8482 II Directional RNA Library Prep Kit for Illumina (E7760)
bulk mRNAseq, paired-end, 150nt

treatment_protocol: VSMCs were grown either untreated (NO dox) or treated with 0.2ug/ml Doxycycline for 10-14 days post differentiation in media containing 10% FBS.

value: Treatment

tag: gfp intensity

value: High


title: AJ20: RBP A C4 NC3 VSMC LOW

value: Low


title: p63 KO small intestine, E18, rep2

description: Gene expression data from stem cells

source_name: p63 KO small intestine, E18

value: p63 KO

value: small intestine


title: S2 cells RpS19a input 1

city: Canberra

first: Olga

last: Zaytseva

state: ACT

address: 131 Garran Road

institute: John Curtin School of Medical Research

data_processing:
Sequence quality was verified using FastQC
The first 10 bases were trimmed using Trimmomatic using sliding-window approach
Alignment was performed using HiSAT2
Gene-level counts were performed using featureCounts
Analysis of differential translation was performed using anota2seq package with normalize = TRUE and filterZeroGenes = TRUE options enabled. The anota2seq “batchVec” option was used to correct for batch effects between technical triplicates
Assembly: Drosophila melanogaster Ensembl genome BDGP6
Supplementary files format and content: Plain text (.txt) files containing raw gene-level counts for each sample
Supplementary files format and content: Microsoft Excel (.xlsx) file containing output of anota2seq (differential translation). Each column contains: ID, apvSlope, apvSlopeP, apvEff, apvRvmMSerror, apvRvmF, residRvmDf, apvRvmP, apvRvmPAdj, singleRegMode

organism: Drosophila melanogaster

source_name: S2 cells

growth_protocol: S2 cells grown in in Gibco Schneider’s S2 cell medium with 10% FBS, at 28°C

extract_protocol:
Extraction using a Promega Reliaprep mini extraction kit. RNA was eluted in 30 μl nuclease-free water, RNA quality was checked using an Agilent Bioanalyzer and RNA concentration was measured using Qubit RNA (broad range) reagents. 1 μg of RNA in 50 μl nuclease-free water was used to prepare sequencing libraries
RNA was prepared using the standard TruSeq Illumina protocol, with Oligo-dT beads used to enrich for mRNA

treatment_protocol: Transfection using effectene reagent with 7.5 μg of dsRNA for RpS19a for 72 hours before collection for polysome profiling.

value: S2 cells

value: Embryonic hemocytes

value: RpS19a knockdown

value: Effectene with RNAi

tag: fraction

value: total cell lysate


title: WT bladder, E15, rep1

description: Gene expression data from stem cells

source_name: WT bladder, E15

value: wildtype


title: A673 DS_ITP

value: EwS cell, A673, WT


title: A673 KS_ITP


title: S2 cells RpS19a polysomes 3

value: polysomes


title: AJ19: RBP A C4 NC3 VSMC NO DOX

value: No


title: p63 KO small intestine, E18, rep1

description: Gene expression data from stem cells

value: Balb/c


title: S2 cells RpS19a polysomes 2


title: AJ18: RBP A C4 NC1 VSMC HIGH


title: unassigned_2_WT_08

city: Suita

first: Yuta

last: Tachibana

state: Osaka

address: 3-1, Yamadaoka

department: Research Institute for Microbial Diseases (RIMD)

institute: Osaka University

description: Instrument model: DNBSEQ-G400RS

data_processing:
The gRNA sequencing reads were aligned to the gRNA library. The abundance of each gRNA was calculated and normalized to the total number of aligned reads.
Assembly: Unassigend_2 library for CRISPR screen
Supplementary files format and content: text file for each gRNA expresssion
Library strategy: DNA-seq

organism: Toxoplasma gondii RH

source_name: RH

extract_protocol:
Toxoplasma genomic DNA was extracted using the DNeasy Blood and Tissue kit (Qiagen).
Integrated gRNA sequences were amplified and barcoded by PCR with KOD FX Neo (TOYOBO). Genomic DNA (1μg) was used for the template. The resulting libraries were sequenced on a DNBSEQ-G400RS (MGI).

value: RH

value: tachyzoite


title: S2 cells RpS19a input 3


title: unassigned_2_WT_09

description: Instrument model: DNBSEQ-G400RS


title: A673 UTA_ITP

value: Initial


title: unassigned_2_WT_07

description: Instrument model: DNBSEQ-G400RS


title: p63 KO proximal stomach, E18, rep1

description: Gene expression data from stem cells

source_name: p63 KO proximal stomach, E18

value: proximal stomach


title: AJ17: RBP A C4 NC1 VSMC LOW

value: Treatment

value: Low


title: S2 cells control input 3

value: WT

value: Effectene alone


title: S2 cells RpS19a polysomes 1

value: S2 cells

value: Embryonic hemocytes


title: WT bladder, E19, rep1

description: Gene expression data from stem cells

value: bladder


title: p63 KO proximal stomach, E18, rep2

description: Gene expression data from stem cells

value: p63 KO


title: S2 cells control input 2

value: total cell lysate


title: AJ16: RBP A C4 NC1 VSMC NO DOX


title: WT bladder, E15, rep2

description: Gene expression data from stem cells

value: wildtype


title: p63 KO large intestine, E18, rep1

description: Gene expression data from stem cells

source_name: p63 KO large intestine, E18

value: large intestine


title: p63 KO large intestine, E18, rep2

description: Gene expression data from stem cells


title: unassigned_2_WT_05

description: Instrument model: DNBSEQ-G400RS


title: p63 KO distal stomach, E18, rep1

description: Gene expression data from stem cells

source_name: p63 KO distal stomach, E18

value: distal stomach


title: AJ15: RBP A C2 NC4 VSMC HIGH

value: High


title: p63 KO ballder, E19, rep2

description: Gene expression data from stem cells

source_name: p63 KO ballder, E19

value: Balb/c


title: unassigned_2_WT_06

description: Instrument model: DNBSEQ-G400RS

value: RH

value: tachyzoite


title: S2 cells control polysomes 3

value: polysomes


title: AJ14: RBP A C2 NC4 VSMC LOW

value: Treatment


title: unassigned_2_WT_03

description: Instrument model: DNBSEQ-G400RS


title: p63 KO distal stomach, E18, rep2

description: Gene expression data from stem cells


title: AJ13: RBP A C2 NC4 VSMC NO DOX

value: No


title: S2 cells control polysomes 1

value: WT


title: AJ11: RBP A C2 NC2 VSMC LOW

value: Low


title: AJ12: RBP A C2 NC2 VSMC HIGH


title: p63 KO ballder, E18, rep2

description: Gene expression data from stem cells

source_name: p63 KO ballder, E18


title: S2 cells control polysomes 2

value: S2 cells

value: Effectene alone


title: Human astrocyte, AB_12h#2

city: Seoul

first: Hoon

last: Ryu

state: Seoul

address: Seoul

department: Brain Science Institute

country: South Korea

institute: Korea Institute of Science and Technology

data_processing:
Raw reads were aligned to the human genome (GRCh37) according to the STAR as an accurate alignment tool for high-throughput RNA-seq data
Assembly: GRCh37
Supplementary files format and content: tab-delimited file includes CPM mean value, adjusted p-value, log2 fold change for each experiment group

source_name: Astrocyte

extract_protocol:
Total RNA was isolated from cells and brain tissues using a commercial extraction system (Qiagen, USA).
Sample libraries were prepared by Ultra RNA Library Prep Kit (#E7530, NEBNEXT, USA), Multiplex Oligos for Illumina (#E7335, NEBNEXT, USA) and poly(A) mRNA Magnetic Isolation Module (#E74900, NEBNEXT, USA) according to the manufacturer’s instructions.

value: Astrocyte

value: Treated Abeta_12hr


title: Human astrocyte, AB_12h#3


title: p63 KO ballder, E19, rep1

description: Gene expression data from stem cells

value: p63 KO

value: bladder


title: p63 KO ballder, E18, rep1

description: Gene expression data from stem cells


title: unassigned_2_WT_02

description: Instrument model: DNBSEQ-G400RS


title: p63 KO ballder, E15, rep2

description: Gene expression data from stem cells

source_name: p63 KO ballder, E15


title: Human astrocyte, AB_12h#1


title: unassigned_2_WT_01

description: Instrument model: DNBSEQ-G400RS


title: S2 cells control input 1

value: Embryonic hemocytes

value: total cell lysate


title: AJ27: VEC C6-1 NC P8->VSMC SD-4-1-19 +DOX 0.2 5-1-19 FACS GFP HI 18-1-19

value: Control

value: High


title: AJ10: RBP A C2 NC2 VSMC NO DOX


title: A549-BoM-3

city: Suzhou

first: zhao

last: WANG

address: No. 199 Ren'ai Road

department: Suzhou Medical College

institute: Soochow University

data_processing:
Libraries with different indices were multiplexed and loaded on an Illumina HiSeq instrument according to manufacturer’s instructions (Illumina, San Diego, CA, USA). Sequencing was carried out using a 2x150bp paired-end (PE) configuration; image analysis and base calling were conducted by the HiSeq Control Software (HCS) + OLB + GAPipeline-1.6 (Illumina) on the HiSeq instrument.
In order to remove technical sequences, including adapters, PCR primers, or fragments thereof, and quality of bases lower than 20, pass filter data of fastq format were processed by Cutadapt V1.9.1  to be high quality-clean data.
Firstly, reference genome sequences and gene model annotation files of relative species were downloaded from genome website. Secondly, Hisat2 v2.0.1 was used to index reference genome sequence. Finally, HTSeq V0.6.1 was used to estimate gene expression levels from the pair-end clean data.
In the beginning transcripts in fasta format are converted from known gff annotation file and indexed properly. Then, with the file as a reference gene file, HTSeq v0.6.1 estimated gene and isoform expression levels from the pair-end clean data.
Assembly: Homo_sapiens.GRCh37.91
Supplementary files format and content: tab-delimited text files include FPKM values for each Sample

growth_protocol: A549 cells were cultured in RPMI-1640 medium with 10% fetal bovine serum (FBS) at 37 °C in a 5% CO2 humidified incubator.

extract_protocol:
Total RNA of each sample was extracted using TRIzol Reagent (Invitrogen, USA). Total RNA of each sample was quantified and qualified by Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA, USA), NanoDrop (Thermo Fisher Scientific Inc.) and 1% agrose gel. 1 μg total RNA with RIN value above 6.5 was used for following library preparation.
Next generation sequencing library preparations were constructed according to the manufacturer’s protocol. The poly(A) mRNA isolation was performed using Poly(A) mRNA Magnetic Isolation Module or rRNA removal Kit. The mRNA fragmentation and priming were conducted using First-Strand Synthesis Reaction Buffer and Random primers. First-strand cDNA was synthesized using ProtoScript II Reverse Transcriptase and the second-strand cDNA was synthesized using Second-Strand Synthesis Enzyme Mix. The purified double-stranded cDNA by beads was then treated with the End Prep Enzyme Mix to repair both ends and add a dAtailing in one reaction, followed by a T-A ligation to add adaptors to both ends. Size selection of Adaptorligated DNA was then performed using beads, and fragments of ~420 bp (with the approximate insert size of 300 bp) were recovered. Each sample was then amplified by PCR for 13 cycles using P5 and P7 primers, with both primers carrying sequences which can anneal with flow cell to perform bridge PCR and P7 primer carrying a six-base index allowing for multiplexing. The PCR products were cleaned up using beads, validated using an Qsep100 (Bioptic，Taiwan， China), and quantified by Qubit3.0 Fluorometer (Invitrogen, Carlsbad, CA, USA).Then libraries with different indices were multiplexed and loaded on an Illumina HiSeq instrument according to manufacturer’s instructions (Illumina, San Diego, CA, USA).

value: lung adenocarcinoma cells

tag: group

value: bone metastasis


title: unassigned_2_lib

description: Instrument model: DNBSEQ-G400RS

value: RH

value: tachyzoite


title: unassigned_2_WT_04

description: Instrument model: DNBSEQ-G400RS


title: A549-BoM-2


title: unassigned_1_WT_10

description: Instrument model: DNBSEQ-G400

data_processing:
The gRNA sequencing reads were aligned to the gRNA library. The abundance of each gRNA was calculated and normalized to the total number of aligned reads.
Assembly: Unassigend_1 library for CRISPR screen
Supplementary files format and content: text file for each gRNA expresssion
Library strategy: DNA-seq


title: A549-BoM-1


title: AJ26: VEC C6-1 NC P8->VSMC SD-4-1-19 +DOX 0.2 5-1-19 FACS GFP LOW 18-1-19


title: Human astrocyte, AB_9h#1

value: Treated Abeta_9hr


title: Human astrocyte, AB_9h#2


title: A549-LuM-3

value: WT

value: lung metastasis


title: A549-LuM-2


title: AJ6: Vec C6-1 NC2 VSMC HIGH


title: unassigned_2_P4

description: Instrument model: DNBSEQ-G400RS


title: unassigned_1_WT_09

description: Instrument model: DNBSEQ-G400


title: Human astrocyte, AB_6h#3

value: Treated Abeta_6hr


title: A549-BoM-4


title: A549-LuM-1


title: Human astrocyte, AB_6h#2


title: unassigned_1_WT_07

description: Instrument model: DNBSEQ-G400


title: unassigned_1_WT_08

description: Instrument model: DNBSEQ-G400


title: Human astrocyte, AB_9h#3


title: AJ25: VEC C6-1 NC P8->VSMC SD-4-1-19 -DOX 18-1-19 unsort

value: No


title: unassigned_1_WT_06

description: Instrument model: DNBSEQ-G400


title: HCTRAD21D_mRNA_DMSO30m_r2

city: bethesda

first: Seolkyoung

last: Jung

state: MD

address: 10 Center Dr

department: NIAMS

institute: NIH

data_processing:
scRNAseq: The demultiplexing, barcoded processing, gene counting and aggregation were made using the Cell Ranger software v3.0.0 (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger)
base calling: illumina CASAVA 1.8.2
ChIP-Seq_density: alignment: bowtie-1.1.1/bowtie -S -m 1 -p 20 -a --best --strata -n 2 -l 50
ChIP-Seq_density: filtering (uniquely aligned reads): samtools-0.1.19-1.2/samtools view -S -b -F4
ChIP-Seq_density: normalized tag density: custom script, maximum 2 reads with the same start position allowed
mRNA-Seq_density:alignment: gsnap --use-sarray=0 --novelsplicing=0 --use-splicing refseq.splices.iit --format=sam -B 4
mRNA-Seq_density: filtering: samtools-1.2/samtools view -S -b -F132 -q 20
mRNA-Seq density: normalized tag density: bedtools-2.25/genomeCoverageBed -ibam -bg -split -scale normalization_factor, ucsc-314/bedGraphToBigWig
mRNA-seq half-life estimation: htseq count file generation from mRNA-seq aligned both for ERCC spike-in control and genomic sequence
mRNA-seq half-lfe estimation:estimate the decay rate constant (kdecay) by fitting the exponential curve on concentration changes upon timepoints using SSasymp R function. From the decay constant, we calculated the mRNA half-lives using the following equation: ln(2)/kdecay
Assembly: mm9, mm10 (only for scRNAseq), hg19
Supplementary files format and content: Tab-separated values files and matrix files
Supplementary files format and content: bigwig file

growth_protocol: HCT-116 degron cell lines: RAD21- mAID-mClover1 and HCT116-OsTIR1-MED26-AID2 were cultured in McCoy's 5A medium supplemented with L-Glutamine (Gibco), 10% FBS (Gemini), 100 U/ml penicillin and 100 µg/ml streptomycin (Gibco), at 37 °C; 5% CO2.  ES (E14) RAD213 and CTCF degron4 cell lines were cultured in knock-out DMEM (Gibco), ES cell qualified FBS (ATCC), 1:100 Glutamax (Gibco), 1:100 sodium pyruvate (Gibco), 1:100 NEAA (Gibco), 100 U/ml penicillin and 100 µg/ml streptomycin (Gibco), 1:1000 2-Mercaptoethanol (Gibco) and 1:10,000LIF (10,000x) on 0.1% gelatin coated dishes at 37 °C; 5% CO2.  Primary WT and Myc knockout B cells were derived from Myc flox/flox mice (WT) or RosaCreERTam/Myc flox/flox mice (KO) were treated with tamoxifen (Sigma) (three times of i.p. 1mg/mouse at 4, 3 and 1 day before sacrifice).  The deletion of floxed allele was checked as previously5 with PCR or intracellular staining of Myc protein. B cells were isolated from spleen of WT and KO mice, enriched with EasySep Mouse B Cell Isolation KIT (STEMCELL Technologies) and activated for 4 hours with 50 mg/ml LPS and 2.5 ng/ml IL4 in RPMI-1640 medium (Gibco) supplemented with 10% FBS (Gemini), 1:100 Glutamax (Gibco), 1:100 sodium pyruvate (Gibco), 1:100 NEAA (Gibco), 100 U/ml penicillin and 100 µg/ml streptomycin (Gibco), 1:1000 2-Mercaptoethanol (Gibco) and HEPES (Gibco), at 37 °C; 5% CO2 B220 positive primary bone marrow B cells were derived from C57BL/6J wild-type mice, enriched with CD45R (B220) MicroBeads Kit (Miltenyi Biotec) and Dead Cell Removal Kit (Miltenyi Biotec). Splenic B cells were derived from C57BL/6J wild-type mice immunized with a keyhole limpet haemocyanin (KLH; 25 µg/footpad, Sigma-Aldrich) in the presence of 10% CFA adjuvant (Sigma-Aldrich). After 5 days B cells were isolated from spleen, enriched with EasySep Mouse B Cell Isolation KIT (STEMCELL Technologies), and activated for 72 hours with 50 mg/ml LPS, 2.5 ng/ml IL4 and anti-CD180 in RPMI-1640 medium (Gibco) supplemented with 10% FBS (Gemini), 1:100 Glutamax (Gibco), 1:100 sodium pyruvate (Gibco), 1:100 NEAA (Gibco), 100 U/ml penicillin and 100 µg/ml streptomycin (Gibco), 1:1000 2-Mercaptoethanol (Gibco) and HEPES (Gibco),  at 37 °C; 5% CO2.  CH12 B lymphoma cells were cultured in RPMI-1640 media (Gibco) supplemented with 10% FBS (Gemini), 100 U/ml penicillin and 100 µg/ml streptomycin (Gibco), 1:1000 2-Mercaptoethanol (Gibco), at 37 °C; 5% CO2. LNK (NK cell line) cells were cultured in RPMI-1640 media (Gibco) supplemented with 10% FBS (Gemini), 1:100 Glutamax (Gibco), 100 U/ml penicillin and 100 µg/ml streptomycin (Gibco), 10 mM HEPES, 1:100 NEAA (Gibco), 1:100 sodium pyruvate (Gibco), 1:1000 2-Mercaptoethanol (Gibco) and 200 IU/ml IL-2 (Sigma-Aldrich). Skin cells for scRNA-Seq: whole epidermis from mouse neonates. Keratinocytes were isolated from Fvb neonate (P0-P2) skin (n = 2) as described in Lichti et al., 20086. Cells were subjected to MACS Dead Cell removal kit (Miltenyi Biotech) using the manufacturer’s protocol. Skin cells for mRNA half-lives estimation: primary keratinocyte. Skins from a pool of Fvb neonatal mice were collected at P0-P2 and incubated overnight in 2.0 U/mL Dispase II (Sigma) in KBM-Gold (Lonza) on an end-over-end rotator. The following day, skins were washed twice in 1x PBS (Sigma), and epidermises were separated from dermises. Epidermises were placed basal side down and floated in pre-warmed 0.25% Trypsin-EDTA (Thermo Scientific) for 20min on an orbital shaker at room temperature. An equal volume of pre-warmed 2.5mg/mL Soybean Trypsin Inhibitor (Thermo Scientific) was added to finish trypsinization, and epidermises were rubbed vigorously against bottom of plate using curved forceps to dislodge keratinocytes. Dislodged cells were pooled into a tube on ice in KBM-Gold. Pooled cells were pipetted ~40 times with a 10ml serological pipette and strained with a 100um filter before pelleting cells at 300xg for 10min at 4°C. Cells were resuspended in 4.5mL KBM-Gold per epidermis and cell count was obtained using Cellometer ViaStain AOPI Solution (Sigma). The cells were subjected to Percoll (GE LifeSciences; ratio of 10X PBS:Percoll:KBM-Gold media was 1:4:5, where the cells were at a concentration of 1 million per mL of final volume) gradient ultracentrifugation using a swinging bucket rotor (11,000 rpm for 42 minutes at 4°C) to separate the basal and suprabasal fractions. The cells at the top band were suprabasal cells and separated from the pellet at the bottom of the tube which were basal cells. The basal cell sample was carefully pipetted out, washed twice with 4°C 1x PBS, seeded onto collagen-coated (Collage Type I; Corning) 6-well plates at 1x106 cells per well, and incubated at 37°C overnight to allow cells to attach to the plate before proceeding with experiments.  Mast cells: Mouse bone marrow-derived mast cells (BMMC) were differentiated from the marrow of tibias and femurs of C57/BL6 mice and cultured for 8 weeks in RPMI 1640 containing 10% FBS, 1M HEPES, 100 U/mL penicillin, 100 μg/mL streptomycin, 4 mM l-glutamine, 1 mM sodium pyruvate, non-essential amino acids, 50 μM 2-mercaptoethanol, IL-3 (20 ng/mL) and stem cell factor (SCF) (20 ng/mL). The purity of mast cells was monitored by assessing the expression of the receptors for SCF (CD117;Kit) and for IgE (FcεRI) by flow cytometry. By the end of 6-8 weeks in culture more than 98% of the cultured population was FcεRI+/CD117+ double-positive mast cells.

extract_protocol:
RNA-Seq Cells (0.5M) were harvested and lysed in 100 µl of the lysis solution (RNAqueous™-Micro Total RNA Isolation Kit from Invitrogen). RNA was isolated and DNase treated according to manufacturer’s protocol. High RNA quality was confirmed using the Agilent TapeStation system before library preparation. mRNA polyA- purification, reverse transcription and library preparation were performed using NEBNext Poly(A) mRNA Magnetic Isolation Module. For spike-in control ERCC RNA Spike-In Mix (Invitrogen) was added before RNA isolation.   ChIP-Seq Cells were fixed with 1% formaldehyde (Sigma) for 10 min at 37°C, treated with 1/20 volume of 2.5 M Glycine, washed with PBS, snap-frozen on dry ice and stored at -80°C until further processing. After thawing cells were resuspended in RIPA buffer (10 mM Tris pH 7.6, 1 mM EDTA, 0.1% SDS, 0.1% sodium deoxycholate, 1% Triton X-100) supplemented with Complete Mini EDTA free proteinase inhibitor (Roche) and sonicated using ultrasonicator (Covaris) in 1 ml AFA tubes (Covaris): duration (s): 1200, peak power: 75, duty % factor: 15, cycles/burst:1000. Sonicated chromatin was pre-cleared with Dynabeads and incubated overnight at 4˚C with rotation with following antibodies: RAD21: ab992 (Abcam); MED26: 13641S (Cell Signaling Technology); TATA binding protein TBP: ab28175 (Abcam); BRD4: ab128874 (Abcam); RNA Pol II: ab26721 (Abcam). Next day samples were washed (10 min at 4˚C with rotation): 2x with RIPA buffer, 2x with RIPA buffer + 0.3 M NaCl, 2x with 1 ml of LiCl buffer (0.25M LiCl, 0.5% Igepal CA-630, 0.5% NaDOC, store at 4˚C), 1x with 1 ml of TE + 0.2 % Triton X-100, 1x with 1 ml of TE. After washes beads were resuspended with 100 ul of TE, followed by addition of 3 µl of 10% SDS and 5 µl of 20 mg/ml Proteinase K and incubated at 65˚C for 4h. After decrosslinking DNA was purified on columns using ChIP DNA Clean & Concentrator kit (Zymo Research). In the next step DNA was quantified using Qubit system (Invitrogen).  mRNA half-lives estimation  Cells were treated with 5 µg/mL actinomycin D (Sigma) and harvested (0.5M cells) at timepoint 0 hours and up to 24 hours: at 1, 2, 4, 8, 12 and 24 hours timepoints7. After each timepoint the viability of cells was checked with FACS, or Nexcelom cell counter after staining with Acridine Orange/Propidium Iodide (AO/PI) (Nexcelom Bioscience). Some cell types did not survive longer actinomycin D treatment than 8 hours, so the last collected timepoint was determined based on cell type viability (details in Extended Data Fig.1a). Pellets were lysed in 100 µl of the lysis solution (RNAqueous™-Micro Total RNA Isolation Kit from Invitrogen), ERCC RNA Spike-In Mix (Invitrogen) was added, RNA isolated (RNAqueous™-Micro Total RNA Isolation Kit from Invitrogen), and RNA-Seq was performed using NEBNext Poly(A) mRNA Magnetic Isolation Module.   scRNA-Seq Cells were washed twice with 1x PBS containing 0.04% weight/volume BSA, resuspended in the same buffer and encapsulated into droplets using 10X Genomics system.
RNA-Seq NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (New England Biolabs) was used for library construction. Single 50 cycles of sequencing data were acquired on NextSeq2000, NextSeq550, HiSeq3000 or NovaSeq6000 (Illumina).  ChIP-Seq Library was prepared using Ovation Ultralow System V2 (Tecan) according to manufacturer’s protocol. Before sequencing high quality was confirmed using the Agilent TapeStation system. Single 50 cycles of sequencing data were acquired on NextSeq550, NovaSeq6000 (Illumina).  scRNA-Seq Libraries were prepared using Chromium Single Cell 3′ Reagent Kits (10X Genomics) according to the manufacturer’s protocol. With double 10bp index cycles, 28 forward and 90 reverse cycles were run on NovaSeq6000 machine (Illumina).

treatment_protocol: HCT-116 degron cell lines: Starvation: cells were FBS depleted for 14 hours, followed by an additional incubation with FBS free medium for another 6 hours in the presence of DMSO (control), 500 µM auxin (indole-3-acetic acid, IAA) (Sigma-Aldrich) for degradation of the AID-tagged RAD21 and MED26 or 500nM JQ-1 (Sigma-Aldrich) for BRD4 displacement. Cells were analyzed after addition of FBS in the presence of DMSO or drugs at timepoints: 0h (RAD21 degron), 0.5h (RAD21 degron) and 2h (RAD21 and MED26 degron, JQ-1 treatment). ES (E14) For the AID-tagged proteins 500 µM auxin (indole-3-acetic acid, IAA) (Sigma-Aldrich) was added for 6 hours (RAD21) or 24 hours (CTCF). Splenic B cells To measure mRNA half-lives in activated B cells Actinomycin D was added after 2 days of stimulation with LPS, IL4 and anti-CD180 (more details: mRNA half-lives estimation). Skin cells To estimate mRNA half-lives in skin cells we used basal cells.

value: colorectal carcinoma cell

value: 10% FBS 14h, noFBS+DMSO 6h, FBS + DMSO 30m


title: unassigned_1_WT_05

description: Instrument model: DNBSEQ-G400


title: AJ3: Vec C-1 NC2 VSMC HIGH


title: Human astrocyte, AB_3h#2

value: Astrocyte

value: Treated Abeta_3hr


title: Human astrocyte, AB_1h#3

value: Treated Abeta_1hr


title: AJ5: Vec C6-1 NC2 VSMC LOW

value: Low


title: HCTRAD21D_mRNA_DMSO30m_r1


title: AJ2: Vec C-1 NC2 VSMC LOW

value: Control


title: Human astrocyte, AB_1h#2


title: AJ1: Vec C-1 NC2 VSMC NO DOX


title: HCTRAD21D_mRNA_DMSO2h_r2

value: 10% FBS 14h, noFBS+DMSO 6h, FBS + DMSO 2h


title: p63 KO ballder, E15, rep1

description: Gene expression data from stem cells

value: p63 KO

value: bladder

tag: strain

value: Balb/c


title: unassigned_1_WT_04

description: Instrument model: DNBSEQ-G400

value: RH

value: tachyzoite


title: unassigned_1_WT_03

description: Instrument model: DNBSEQ-G400


title: HCTRAD21D_mRNA_DMSO2h_r1


title: Human astrocyte, AB_1h#1


title: Human astrocyte, Control#3

value: No treatment


title: HCTRAD21D_mRNA_DMSO0h_r2

value: 10% FBS 14h, noFBS+DMSO 6h


title: unassigned_1_WT_02

description: Instrument model: DNBSEQ-G400


title: AJ4: Vec C6-1 NC2 VSMC NO DOX


title: HCTRAD21D_mRNA_DMSO0h_r1


title: Human astrocyte, Control#2


title: unassigned_1_WT_01

description: Instrument model: DNBSEQ-G400


title: unassigned_1_P4

description: Instrument model: DNBSEQ-G400


title: Human astrocyte, Control#1


title: D_sc_3

city: Norwich

first: Wenbo

last: Ma

address: Norwich Research Park

institute: The Sainsbury Laboratory

description: Biological replicate 3

data_processing:
The quality of the raw sequencing data was checked with FastQC v0.11.9
reads were aligned with the indexed CLas-Psy62 isolate genome assembly (NC_012985.3) and parsed to it using HISAT2 v2.2.1
The number of reads mapped to each gene was counted using the feature counts function of the Rsubread package
Read counts were normalized for each sample for differential expression analysis using DESeq2 v1.30.1
Assembly: NC_012985.3
Supplementary files format and content: Matrix table with raw gene counts for every gene and every sample
Supplementary files format and content: Matrix table with normalized read counts for every genes and every sample

organism: Citrus x paradisi

source_name: Citrus paradisi Macfadyenand (Ri Red grapefruit)

extract_protocol:
100 mg of frozen seed coat vasculature samples were ground under freezing conditions using a GenoGrinder and tungsten beads at 1500 rpm for 30 seconds twice. The RNA was extracted from the homogenized tissue for untreated samples using the Qiagen RNeasy Plant Mini Kit according to the handbook instructions which included an on column Dnase I treatment. Samples were eluted with 50 uL of RNase-free water. The RNA was extracted from the bacterial pellet obtained after the above bacterial isolation resusepndedn in 1 mL of TLE extraction buffer. Then, 60 µL of 500 mM CaCl2 was added, mixed thoroughly, and placed on ice for 10 min. Centrifugation at 12,000 x g for 10 min at 4 °C was performed, followed by two rounds of separation with 500 µl of phenol:Chloroform:isoamyl alcohol (25:25:1) and centrifugation at 15,000 x g for 10 min at 4 °C. The supernatant was mixed with 2 M LiCl and incubated overnight at -20 °C. RNA was precipitated by centrifugation at 15,000 x g for 30 min, washed twice with ice-cold 75% ethanol, centrifuged at 15,000 x g for 15 min and re-suspended in 20 µl of RNAse-free water.
RNA was converted into sequencing-ready libraries for sequencing using TruSeq Stranded Total RNA Library Prep Kit (Illumina) for the bacterial isolation sample or Meta-transcriptome Library (Novogene) for untreated samples.  cDNA libraries were subjected to RNASeq, using an Illumina HiSeq-4000 system  with 100-bp strand-specific single-end reads for the bacterial isolation sample or Illumina NovaSeq-6000 system with 150-bp strand-specific paired-end reads for untreated samples.

treatment_protocol: pre-enrichment (bacterial isolation) performed according to De Francesco et al. 2022. All samples underwent ribodepletion using Illumina Ribo-Zero Plus kit

value: seed coat


title: kidney organoid (patient)

city: Parkville

first: Belinda

last: Phipson

state: VIC

address: 1G Royal Parade

department: Bioinformatics

institute: WEHI

data_processing:
The 10x Genomics Cell Ranger pipeline (v3.0.2) was used to align, filter and de-duplicate reads before counting those overlapping genes to produce count matrices for each sample.
The resulting datasets consist of matrix files (corrected_matrix.mtx.gz, null_matrix.mtx.gz and patient_matrix.mtx.gz) containing the UMI count values, barcode files (corrected_barcodes.tsv.gz, null_barcodes.tsv.gz, patient_barcodes.tsv.gz) containing the detected cell barcodes and genes files (corrected_features.tsv.gz, null_features.tsv.gz, patient_features.tsv.gz) containing the gene symbols of the detected genes. The dense matrix of the full dataset is provided in CSV format (all-counts.csv), with the associated cell information in CSV format (cell_info.csv).
Genome_build: hg38
Supplementary_files_format_and_content: Cell Ranger output files

source_name: kidney organoid

growth_protocol: iPSC were maintained in an undifferentiated state using Essential 8 culture media (Thermo Fisher Scientific) on 6-well plates coated with Matrigel (BD Biosciences). Formation of kidney organoids was performed as described previously (Howden 2019, EMBO Reports PMID: 30858339), and identical protocols were used for each of the iPSC clones.

extract_protocol:
Per clone, 4 organoids were dissociated together using a 1:1 mix of TrypLE and Accutase (STEMCELL Technologies) preheated at 37˚C. Organoids were incubated at 37˚C in dissociation mix for 20-25 minutes pipetting up and down every 3 minutes. Dissociation was inactivated using DMEM/F12 (Life Technologies, California, U.S.A.) with 10% fetal bovine serum and cells were placed on ice. To remove debris, cells were filtered through a 40μm cell strainer and subsequently through a 35μm FACStube strainer (Corning), counted using a hemocytometer and resuspended at a density of 1200cells per μl, filtered through a second FACStube strainer and a final cell count was performed before library preparation.
Chromium Single Cell Library Kit V3

value: kidney organoid

tag: day of differentiation

tag: media

value: E6; retinoic acid d12-d14


title: kidney organoid (corrected)


title: kidney organoid (null)


title: Human astrocyte, AB_3h#3


title: HCTRAD21D_mRNA_AUXIN30m_r2

value: colorectal carcinoma cell

value: 10% FBS 14h, noFBS+IAA 6h, FBS + IAA 30m


title: HCTRAD21D_mRNA_AUXIN30m_r1


title: unassigned_1_lib

description: Instrument model: DNBSEQ-G400


title: iSC_DEB135-10_ITGA6_POS_R2

city: Stanford

first: Anthony

last: Oro

state: CA

address: 269 Campus Drvie

department: Dermatology

institute: Stanford University

data_processing:
Tophat used for alignment.
Homer for raw counts and RPKM values
DESEQ2 used for differential expression
Assembly: hg38
Supplementary files format and content: text file, raw counts

growth_protocol: Following the 7 day differentiation, defined Keratinocyte SFM Media is given up to Day 45 at which time the cells are enriched with ITGA6 antibody for the keratinocytes. iSC population is the positive fraction while the reduced population is the negative fraction. Both Pos and Neg Fractions were expanded in Defined Keratinocyte SFM.

extract_protocol:
RNA was isolated using Trizol per manufacturer protocol. The aqueous phase was transferred  and wahsed using an RNAeasy spin colunm. DNA was removed with a Qiagen Dnase kit on the Rneasy column.
Libraries were constructed with the KAPA kit for PolyA enriched mRNA per manufacterer instructions.

treatment_protocol: iPSCs were expanded in StemFit Basic 02 media before differentiated in Essential 6 media treated with BMP-4 and Retinoic Acid for 7 days to differentiate the cells toward keratinocytes.

value: iSC population; positive fraction

value: iPSC derived keratinocytes sorted based on CD49f/ITGA6


title: D_sc_2

description: Biological replicate 2


title: iSC_DEB135-10_ITGA6_POS_R1


title: AAV replicate 2

city: Nantes

first: Jean-Baptiste

last: Dupont

address: 22 boulevard Benoni Goullin

department: INSERM

country: France

institute: Nantes Université

description: Test

data_processing:
Quantification cycle (Cq) values calculated with the Applied Biosystems SDS software version 2.3 by using automatic baseline with a threshold set at 0.1
In the manuscript where this data was included, only the raw Cq data are interpreted and no normalization step was performed.

label: FAM

organism: unclassified Adeno-associated dependoparvovirus A

source_name: AAV8 GFP vector

label_protocol: Reverse Transcription with the Megaplex Primer Pools A and B

extract_protocol: miRNeasy Serum/Plasma kit

treatment_protocol: Purification with CsCl gradient

value: AAV8 GFP vector


title: HCTRAD21D_mRNA_AUXIN2h_r2

value: 10% FBS 14h, noFBS+IAA 6h, FBS + IAA 2h


title: AAV replicate1

description: Test


title: HCTRAD21D_mRNA_AUXIN2h_r1


title: HCTRAD21D_mRNA_AUXIN0h_r2

value: 10% FBS 14h, noFBS+IAA 6h


title: iSC_DEB135-10_ITGA6_NEG_R2

value: reduced population; negative fraction


title: D_sc_1

description: Biological replicate 1


title: HEK293

description: Control

source_name: Producer cell line

growth_protocol: DMEM 10% Fetal Bovine Serum + 1% Pen Strep

extract_protocol: miRNeasy mini kit


title: iSC_DEB135-10_ITGA6_NEG_R1


title: HCTRAD21D_mRNA_AUXIN0h_r1


title: HUVECs treated with 5 mM of L-glucose for 6 hours under hypxia/serum starvation conditions 2

status: Public on Jun 29 2024

city: Augusta

first: Brian

last: Annex

state: Georgia

address: 1120 15th Street

department: VBC

institute: Augusta university

data_processing:
The BCL files were demultiplexed and converted to FASTQ format with bcl2fastq2 (v2.20.0).FastQC(v0.12.1) was used to evaluate reads quality. Reads were aligned to the reference genome (GENCODE:GRCh38.p13) usingSTAR(v2.7.10b) aligner. HTSeq-counts was used to count reads for each gene. Differential expression analysis was done usingDESeq2package. Also, theapeglmpackage was used for effect size estimation and lfcShrink log2 fold-change values.
Assembly: GENCODE:GRCh38.p13
Supplementary files format and content: processed data files format and content tab-delimited text file includes raw counts for each Sample The complete results of DE analysis is in DE_Analysis_Results.csv file.

source_name: Human umbilical vein endothelial cells

growth_protocol: HUVECs were endothelial cell growth media (cell application 211-500) in 37 degrees celisius in CO2 incubator

extract_protocol:
RNA was harvested using Direct-zol RNA MiniPrep, Cat.No: R2052, 800 ng of total RNA was used for the construction of sequencing libraries.
ORNA libraries for RNA-sea were prepared using SMARTER mRNA-Seq Library Prep Kit flowing manufacture's protocols.

treatment_protocol: HUVECs were treated with L-Glucose (5 mM) vs. glucosamine (5 mM) for 6 hours under 2% hypoxia and serum starvation

value: Human umbilical vein endothelial cells

value: wild type

value: L-Glucose


title: HUVECs treated with 5 mM of L-glucose for 6 hours under hypxia/serum starvation conditions 1


title: HUVECs treated with 5 mM of glucosamine for 6 hours under hypxia/serum starvation conditions 2

value: Glucosamine


title: HUVECs treated with 5 mM of glucosamine for 6 hours under hypxia/serum starvation conditions 3


title: HUVECs treated with 5 mM of L-glucose for 6 hours under hypxia/serum starvation conditions 3


title: S2 cells RpS19a input 2

value: S2 cells

value: Embryonic hemocytes

value: RpS19a knockdown

value: Effectene with RNAi

value: total cell lysate


title: HUVECs treated with 5 mM of glucosamine for 6 hours under hypxia/serum starvation conditions 1


title: ALDH_Low_replicate2

city: Edinburgh

last: Patton

address: Western General Hospital, Crewe Road South

department: Institute of Genetics and Cancer

institute: MRC Human Genetics Unit

data_processing:
Sequencing was performed using the NextSeq 500/550 High-Output v2 (150 cycle) Kit (# FC-404-2002) on the NextSeq 550 platform (Illumina Inc, #SY-415-1002). Libraries were combined in an equimolar pool based on the library quantification results and run across a single High-Output Flow Cell.
Raw FASTQ sequence reads were quality checked using FastQC (v. 0.11.3) and aligned to the human genome (GrCh38) assembly using STAR (v. STAR_2.5.1b) software with default parameters.
Raw counts of reads covering the transcriptome (Ensembl annotation version GRCh38.91) were obtained using htseq-count (0.6.1) with the “-s reverse” option.
Normalisation and Differential analysis between ALDHhHgh and ALDHLow cells was performed by DESeq2
Assembly: GRCh38
Supplementary files format and content: Comma-separated values text file including raw counts of each sample
Supplementary files format and content: Comma-separated values text file including TPM of each sample

source_name: melanoma

growth_protocol: DMEM high glucose, 10% FCS, 2 mM L-glutamine

extract_protocol:
total RNA were extracted using total RNA was extracted and purified from live cells using TRIzol (Invitrogen) and Direct-Zol RNA Miniprep Kits (ZYMO Research)
Libraries were prepared from 500 ng of each total-RNA sample using the TruSeq Stranded mRNA Library Kit (Illumina) according to the provided protocol and purified using AMPure XP beads (Beckman Coulter).

treatment_protocol: A375 melanoma cells were stained with Aldefluor for ALDH activity following manufacturer's instructions and sorted for cells with top and bottome 5% Aldefluor signal by FACS Aria II (BD Biosciences)

value: melanoma

value: Aldefluor sorted ALDH bottom 5%


title: H3K27ac_KD_R1

data_processing:
Sequencing was performed using the NextSeq 500/550 High-Output v2.5 (150 cycle) Kit (#20024907) on the NextSeq 550 platform (Illumina Inc, #SY-415-1002). PhiX Control v3 (Illumina, #FC-110-3001) was spiked into the library pool at a concentration of ~1% to enable troubleshooting in the event of any issues with the run.
Basecall data produced by the NextSeq 550 is uploaded to BaseSpace and converted into FASTQ and then mapped to the human genome (GRCh38) and mouse genome (mm9) using bowtie2
The mapped read counts splitting between target species (human) and spike-in control (mm9) were then used for calculating the scaling factors.
Target species (human) mapped reads were filtered using SAMtools for the flag of PCR duplication
MACS2 calling broadpeak algorithm to identify histone H3K23 and H3K27 acetylation regions respectively, with each BAM file paired with the IgG control from the matching experiment group, (cut off -- mfold 2 50, band width --bw 300, FDR --qvalue 0.05).
Assembly: GRCh38
Supplementary files format and content: bigwig files after scaling normalisation for each CUTandTAG sample

source_name: MAPKi resistant human melanoma

growth_protocol: RPMI 1640, 10% FCS, 2 mM L-glutamine

extract_protocol:
CUTandTAG was performed on 0.1% PFA (1 min on ice) briefly treated cells
CUTandTAG DNA were released from the concanavalin A coated magnetic beads and library construction was perfomed by in-tube PCR using uniquely barcoded i5 primer and i7 primers, followed by Ampure XP beads post-PCR cleanup

value: MAPKi resistant human melanoma

tag: cutandtag antibody

value: Acetyl-Histone H3 (Lys27) (D5E4) RRID:AB_2798743


title: H3K27ac_Ctrl_R2

value: scramble


title: H3K23ac_KD_R1

value: Acetyl-Histone H3K23Ac antibody RRID:AB_2855229


title: IgG_KD_R1

value: Rabbit (DA1E) mAb IgG XP Isotype Control (CUT&RUN) RRID:AB_2924329


title: IgG_Ctrl_R1


title: IgG_Ctrl_R2


title: ALDH_High_replicate2

value: Aldefluor sorted ALDH top 5%


title: IgG_KD_R2


title: H3K27ac_KD_R2


title: H3K27ac_Ctrl_R1


title: ALDH_High_replicate1


title: A375_ALDH1A3KO_input_rep2

data_processing:
Sequencing was performed using the NextSeq 500/550 High-Output v2.5 (150 cycle) Kit (#20024907) on the NextSeq 550 platform (Illumina Inc, #SY-415-1002). PhiX Control v3 (Illumina, #FC-110-3001) was spiked into the library pool at a concentration of ~1% to enable troubleshooting in the event of any issues with the run.
Basecall data produced by the NextSeq 550 is automatically uploaded to BaseSpace, a cloud-based data management and analysis service provided by Illumina. Here it is converted into FASTQ filesand mapped to the human genome (GRCh38) using bowtie2 (bwa-0.7.9) applicationse.
Mapped regions extended beyond the end of the chromosomes were removed using SAMtools with chromosome lengths for GRCh38. MACS2 calling broadpeak algorithm was used to identify histone H3 acetylation regions, with each ChIP BAM file paired with the input control, cut off -- mfold 5 50, band width --bw 500, FDR --qvalue 0.05.
To visualise the histone H3 acetylation sites on the genome browser UCSC122 mapped reads were converted to bigWig files (.bw) using the bamCoverage (deepTools2) algorithm, with replicants for each condition merged.
Assembly: GRCh38
Supplementary files format and content: bed files of gapped peaks in each ChIP sample from MACS2 calling peaks

extract_protocol:
Cells were fixed in 1% formaldehyde in PBS for 10 minutes. After quenching and cell lysis, chromatin samples were sonicated using on ice for 8x 30 sec on-30 sec off burst cycles with a probe sonicator (SoniPrep150) in a chilled ice-water bath (12 Amplitude). 10% of total samples before IP were used as input control.
ChIP-seq libraries were prepared using NEBNext Ultra II DNA Library Prep Kit for Illumina and NEBNext Multiplex Oligos for Illumina (Index Primers Set 1) following manufacturer’s instructions. Libraries were combined in a single equimolar pool based on fragment size information from the Bioanalyser traces and the Qubit assay quantification results.

value: ALDH1A3 ko

tag: chip antibody


title: H3K23ac_Ctrl_R2


title: A375_ALDH1A3KO_input_rep1


title: ALDH_Low_replicate3


title: H3K23ac_Ctrl_R1


title: ALDH_Low_replicate1


title: ALDH_High_replicate3


title: A375_ALDH1A3KO_H3Ac_ChIP_rep1

value: anti-acetyl-Histone H3 (Millipore, RRID:AB_2115283)


title: A375_ALDH1A3KO_H3Ac_ChIP_rep2


title: A375_Ctrl_input_rep2

value: Cas9 control


title: H3K23ac_KD_R2


title: A375_Ctrl_H3Ac_ChIP_rep1


title: A375_Ctrl_H3Ac_ChIP_rep2


title: KhES1_D4ng_sample4

city: Abu Dhabi

first: Ken-ichiro

last: Kamei

address: Saadiyat Campus, #C1-032

country: United Arab Emirates

institute: New York University Abu Dhabi

description:
mRNA-seq (polyA+) data from blastoids derived from KhES1 cultured in DMEM without hydrogel, sample 4
KhES1_D4ng_2

data_processing:
Chromium barcodes were used for demultiplexing by using CellRanger program (10x Genomics; v6.1.1, https://support.10xgenomics.com/single-cell-gene-expression/software/overview/welcome).
Alignment and UMI counting were performed using CellRanger to map 373,641,788 sequencing reads to a custom version of the Ensembl GRCh38 reference genome.
Assembly: GRCh38
Supplementary files format and content: Gene expression values were calculated with CellRanger program.

growth_protocol: hESCs were used following the guidelines provided by the ethical committee of Kyoto University (Approved #ES3-9 and ES3-29). Prior to culturing, 1.3% (v/v) hESC-certified Matrigel (Corning, NY, USA) in DMEM/F12 medium was coated on a culture dish. Matrigel was incubated in a dish for 24 h at 4˚C. Then, excess Matrigel was removed, and the coated dish was washed with a fresh DMEM/F12 medium. A mTeSR-1 medium was used for the daily culturing of hPSCs. For passaging, cells were dissociated with TrypLE Express (Life Technologies, Carlsbad, CA, USA) for 3 min at 37 ˚C, and then harvested. A cell strainer was used to remove undesired cell aggregates from the cell suspension. The cells were centrifuged at 200 × g for 3 min and resuspended in the mTeSR-1 medium. The cells were counted using a NucleoCounter NC-200 (Chemetec, Baton Rouge, LA, USA). A mTeSR-1 medium containing 10 µM of Y27632 was used to prevent the apoptosis of the dissociated hPSCs, on day 1. mTeSR-1 medium without Y27632 was used on the subsequent days, with daily medium change. The cells were maintained in an incubator at 37°C with 5% (v/v) CO2.

extract_protocol:
Dissociated cells were applied to Chromium controller (10x Genomics, Pleasanton, CA, USA) to encapsulate cells, following the manufacture’s instruction (Chromium Next GEM Single Cell 3’ Reagent Kits v3.1 (Dual Index)).
StepOnePlus (Applied Biosystems)

treatment_protocol: hESCs cultured on Matrigel-coated dishes with a mTeSR-1 medium were harvested with TrypLE Express for 3 min at 37°C and transferred in a 15-mL tube. The dissociated hPSCs were resuspended in mTeSR-1 medium supplemented with 10 µM of Y27632, and then transferred to Aggrewell 400 (Stem Cell Technologies) at 1.5 × 10^5 cells per well. After culturing in an incubator for 24 h at 37°C with 5% (v/v) CO2, the hPSC aggregates were resuspended in the HG solution {DMEM with 10%[w/v] PNIPAAm-β-PEG hydrogel at 4°C. 260 µL of an hPSC aggregate suspension with HG was transferred into the well of a 24-well plate, and then DMEM supplemented with 10%(v/v) FBS was added at 37°C. After 3-day culture, iBlastoid medium was changed on a daily basis, and the cells were maintained at 37°C with 5% (v/v) CO2 in an incubator.

tag: condition

value: DMEM without hydrogel

tag: derived tissue

value: Embryonic stem cells


title: KhES1_D6ng_sample7

description:
mRNA-seq (polyA+) data from blastoids derived from KhES1 cultured in iBlastoid medium without hydrogel, sample 3
KhES1_D6ng_1

value: iBlastoid medium without hydrogel


title: SW1990-Gomisin G rep2

city: wenzhou

first: Changrui

last: Qian

state: zhejiang

address: wenzhou medical university

institute: wenzhou medical university

data_processing:
The RNA-seq data of 4 samples was filtered with fastp.
The clean reads were mapped to the reference genome using HISAT2.
Expression level of gene was calculated by featureCounts.
Assembly: hg38
Supplementary files format and content: tab-delimited text file includes raw counts for each Sample

growth_protocol: SW1990 cells were cultured in DMEM medium at 37℃ incubator.

extract_protocol:
SW1990 cells were pelleted and flash frozen on dry ice. Pellets were shipped to Genewiz for RNA extraction, Illumina Ribo-Zero rRNA removal, Illumina library construction, and sequencing.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: Methods: The SW1990 cells were treated with vehicle or 10 µM Gomisin G for 24 h.

value: pancreatic cancer cells

value: Gomisin G


title: SW1990-Vehicle rep2

value: Vehicle


title: KhES1_D4_sample2

description:
mRNA-seq (polyA+) data from blastoids derived from KhES1 cultured in DMEM with hydrogel, sample 2
KhES1_D4_2

value: DMEM with hydrogel


title: SW1990-Gomisin G rep1


title: Sperm, Control1

city: College Station

first: MICHAEL

middle: C.

last: GOLDING

state: Texas

address: 4466 TAMU

department: Physiology

institute: Texas A&M UNiversity

data_processing:
TESmall
Assembly: mirbase
Supplementary files format and content: Count Matrix

source_name: Sperm

extract_protocol:
Trizol
Qiaseq
small RNA seq

value: Sperm

value: control


title: Sperm, Control2


title: Sperm, Control3


title: Sperm, Ethanol2

value: ethanol


title: Sperm, Control4


title: Sperm, Ethanol4


title: SW1990-Vehicle rep1


title: Sperm, Ethanol3


title: Sperm, Ethanol1


title: KhES1_D6_sample6

description:
mRNA-seq (polyA+) data from blastoids derived from KhES1 cultured in iBlastoid medium with hydrogel, sample 2
KhES1_D6_2

value: iBlastoid medium with hydrogel

value: Early blastoid


title: KhES1_D6_sample5

description:
mRNA-seq (polyA+) data from blastoids derived from KhES1 cultured in iBlastoid medium with hydrogel, sample 1
KhES1_D6_1


title: KhES1_D0_sample4

city: Kyoto

state: Kyoto

address: Yoshida-Ushinomiya-cho, Sakyo-ku

department: iCeMS

institute: Kyoto University

description: mRNA-seq (polyA+) data from blastoids derived from KhES1 cultured in iBlastoid medium with hydrogel, sample 4

value: Blastoid


title: A375_Ctrl_input_rep1

value: melanoma


title: KhES1_D0_sample2

description: mRNA-seq (polyA+) data from KhES1 cultured in mTeSR-1 medium, sample 2


title: KhES1_D4_sample1

description:
mRNA-seq (polyA+) data from blastoids derived from KhES1 cultured in DMEM with hydrogel, sample 1
KhES1_D4_1


title: KhES1_D0_sample3

description: mRNA-seq (polyA+) data from blastoids derived from KhES1 cultured in iBlastoid medium with hydrogel, sample 3


title: KhES1_D0_sample1

description: mRNA-seq (polyA+) data from KhES1 cultured in mTeSR-1 medium, sample 1