title: Multiome - ATAC - PH1302

description:
Library name: Multiome - ATAC - PH1302
PH1302_filtered_feature_bc_matrix.h5

source_name: Ovarian cancer

value: Ovarian cancer


title: Rv1, BPTFKD, rep1

status: Public on Nov 13 2025

type: SRA

city: Baltimore

first: Jianfei

last: Qi

state: MD

address: 655W baltimore street

department: Biochemistry and Molecular Biology

country: USA

institute: University of Maryland

description: Library name: Rv1-BPTFKD-rep1

library_source: genomic

data_processing:
Quality of ATAC-seq reads was checked using FastQC software
Trim Galore was used to remove adaptor sequences and reads were aligned to the human hg19 reference genome using Bowtie2 –very-sensitive option
SAM files were converted to BAM files, and mitochondrial reads and PCR duplicates were removed. BAM files were used either for peak calling with MACS2 callpeak or for generating RPGC-normalized bigwig files using Deeptools
Assembly: hg19
Supplementary files format and content: bigWig

molecule: genomic DNA

organism: Homo sapiens

growth_protocol: Cells were maintained in RPMI 1640 media supplemented with 10% fetal bovine serum, 1% penicillin/streptomycin. Cells were incubated at 37C and 5% CO2.

extract_protocol:
Two biological replicates (1x105 living cells per replicate) were used for sample preparation, following the ATAC-seq kit manual (Active Motif). In brief, cell nuclei were extracted using ATAC lysis buffer and incubated with 50 μl of Tagmentation Master Mix at 37° C for 30 minutes in a thermomixer. The transposed DNA was purified with spin column.
we prepared sequencing library using Nextera index kit. The library was purified with double-sided SPRI beads and library fragment size was evaluated using an Agilent Bioanalyzer.

tag: cell line

tag: cell type

value: Human prostate cancer cell line

tag: genotype

value: BPTF knock down

tag: treatment

value: shBPTF


title: SDS_D3-2+Topsym

city: Taipei

first: Kang-Yu

last: Tai

address: Yonglin Biomedical Engineering Hall, No.49, Fanglan Rd.

department: Genome and Systems Biology Degree Program

country: Taiwan

institute: National Taiwan University

description: Library name: SDS-3-2T

library_source: transcriptomic

data_processing:
Galaxy version 23.1.rc1
Sequence reads were trimmed for adaptors by Trim Galore ( ver0.6.7)
Trimmed sequence were mapped to mm10 using HISAT2 (ver 2.2.1)
Raw counts were extracted by featureCounts (ver 2.0.3)
Normalized counts were performed by DESeq2 (ver 2.11.40.8)
Assembly: mm10
Supplementary files format and content: tab-delimited file contains raw counts for each sample
Supplementary files format and content: tab-delimited matrix file contains normalized counts for all sample

molecule: total RNA

organism: Mus musculus

source_name: whole skin

growth_protocol: 7 week-age mice

extract_protocol:
RNeasy Micro Kit
Agilent SureSelect Strand-Specific RNA library Prep Cat#G9691B

treatment_protocol: Topical treatment as indicated for 4 days

tag: tissue

value: whole skin


title: Multiome - ATAC - PH1239

city: Rochester

first: Alexandre

last: Gaspar-Maia

state: MN

address: 200 First street SW

institute: Mayo Clinic

description:
Library name: Multiome - ATAC - PH1239
PH1239_filtered_feature_bc_matrix.h5

library_source: genomic single cell

data_processing:
Sequenced reads from the gene expression (GEX) and DNA accessibility (ATAC) droplet libraries were aligned and quantified using 10x Genomics Cell Ranger ARC v2.0.0. The reads were aligned to the pre-built human reference genome GRCh38 - v2020-A-2.0.0 (May 3, 2021) provided by 10X Genomics. After quality control, data from each sample was processed using Seurat v5.2.1 and Signac v1.14.0 functions. GEX and ATAC count matrices from each sample were merged independently using Seurat. GEX count matrix was log-normalized, scaled to mean 0 and variance 1, and dimensionality reduction was performed using PCA on the top 2000 highly variable genes. Uniform manifold approximation and projection (UMAP) for GEX was calculated using the top 50 principal components. Open chromatin peaks called per sample were merged using reduce function from GenomicRanges R package, then the ATAC fragment count matrix was recalculated using Signac. Merged peaks that were smaller than 20 base pairs or larger than 10000 base pairs were removed from analysis. The ATAC count matrix was normalized using Term Frequency - Inverse Document Frequency (TF-IDF) and dimensionality reduction performed using singular value decomposition (SVD) using only peaks with non-zero counts in at least 20 cells - together known as latent semantic indexing (LSI) that generates LSI components. The UMAP for ATAC was calculated using LSI components 2 to 50. Seurat’s weighted nearest neighbor (WNN) algorithm was used on principal components 1 to 50 (GEX) and LSI components 2 to 50 (ATAC) together to obtain a combined UMAP projection of both modalities. Cells with more than 20% of reads mapped to mitochondrial genes, those with less than 200 unique genes detected (GEX), those with less than 200 unique peaks detected (ATAC) and those with transcription start site (TSS) enrichment score (as calculated by Signac) less than 1 were removed for quality control.
Assembly: hg38
Supplementary files format and content: Cellranger output files: h5 files and ATAC fragments files (tsv)

extract_protocol:
Fresh tumor tissue obtained from ovarian cancer patients after debulking surgery was transferred in a sterile 10 cm culture dish and finely minced using scissors in KRB buffer. The sample was centrifuged at 500 g for 5 mins at 4°C, and the supernatant was gently removed without disrupting the pellet. Washes were repeated until the supernatant appeared clear. After the last centrifugation, tissue was aliquoted in cryovials and frozen in liquid nitrogen. On the day of the experiment, frozen tissue was cut into small pieces without thawing and used for the nuclei isolations.
The nuclei isolation was performed based on the 10x genomics suggested protocol for Complex Tissues for Single Cell snATAC + snRNA. After mincing, a small piece of tissue the size of rice grain was transferred to a 2 mL microcentrifuge tube containing 300 μL of NP-40 Lysis Buffer and homogenized on ice using a pellet pestle. After adding 1 mL of NP-40 Lysis Buffer, the samples were incubated on ice for either 5 or 30 minutes. After incubation, the samples were strained using a 70 μm cell strainer, and the flow through was then centrifuged 500 g for 5 mins at 4°C. After centrifugation, the nuclei were permeabilized by resuspending in 100 μL of 0.1X Lysis Buffer and incubating on ice for 2 minutes. After adding 1 mL of Wash Buffer, the nuclei were centrifuged at 500 g for 5 mins at 4°C, and the pellet was resuspended in Diluted nuclei buffer. The nuclei concentration was assessed by PI staining using a Cellometer K2 cell counter. Between 1000 and 8000 nuclei per sample were subjected to transposase assays (exposing buffered nuclei to Tn5 transposase) before proceeding to single-cell partitioning into gel beads in emulsion, barcoding, and pre-amplification. ATAC library construction and cDNA, followed by GEX library construction, were done following established 10x Genomics protocols. The libraries concentration was measured using Qubit High Sensitivity assays (Thermo Fisher Scientific), and library profiles were assessed in a fragment analyzer (Advance Analytical) before sequencing. The snATAC and snRNA libraries were sequenced for 50bp paired end sequencing, PE50 and 100PE, respectively on a HiSeq 4000, NextSeq 2000 or NovaSeq X 1.5B instrument (Illumina) before demultiplexing and alignment to the reference genome (GRCh38/hg38).

value: Untreated

tag: library type

value: ATAC


title: TSSB64 (MP65_siLuc)

city: Nice

first: Corine

last: Bertolotto

address: 151 route Saint-Antoine de Ginestière

country: France

institute: C3M - U1065 INSERM - CENTRE MEDITERRANEEN DE MEDECINE MOLECULAIRE

description: Library name: Sample3

data_processing:
Image analysis and base calling were performed using RTA 2.7.7 and bcl2fastq 2.20.0.422.
Assembly: hg38
Supplementary files format and content: excel file with number of reads

molecule: polyA RNA

extract_protocol:
Trizol RNA extraction
P2699 : Total RNA-Seq libraries were generated from 400 ng of total RNA using Illumina Stranded Total RNA Prep, Ligation with Ribo-Zero Plus kit and IDT for Illumina RNA UD Indexes, Ligation (Illumina, San Diego, USA), according to manufacturer's instructions. Briefly, abundant ribosomal RNAs were depleted by hybridization to specific DNA probes and enzymatic digestion. The depleted RNAs were purified and fragmented using divalent cations at 94oC for 2 minutes. After random hexamers annealing, fragmented RNAs were then reverse transcribed into first strand complementary DNA (cDNA). Second strand cDNA synthesis further generated blunt-ended double-stranded cDNA and incorporated dTTP in place of dUTP to achieve strand specificity by quenching the second strand during amplification. Following A-tailing of DNA fragments and ligation of pre-index anchors, PCR amplification was used to add indexes and primer sequences and to enrich DNA libraries (30 sec at 98oC; [10 sec at 98oC, 30 sec at 60oC, 30 sec at 72oC] x 13 cycles; 5 min at 72oC). Surplus PCR primers were further removed by purification using SPRIselect beads (Beckman-Coulter, Villepinte, France) and the final libraries were checked for quality and quantified using capillary electrophoresis.

value: uveal melanoma


title: SDS_D1-2+Topsym

description: Library name: SDS-1-2T


title: Rv1, control, rep2

description: Library name: Rv1-pLKO.1-rep2

value: WT

value: shControl


title: Multiome - ATAC - PH1238

description:
Library name: Multiome - ATAC - PH1238
PH1238_filtered_feature_bc_matrix.h5

value: Chemotherapy treated


title: Multiome - ATAC - PH1230

description:
Library name: Multiome - ATAC - PH1230
PH1230_filtered_feature_bc_matrix.h5


title: SDS_D1-1+Topsym

description: Library name: SDS-1-1T


title: Multiome - ATAC - PH1214

description:
Library name: Multiome - ATAC - PH1214
PH1214_filtered_feature_bc_matrix.h5


title: Rv1, control, rep1

description: Library name: Rv1-pLKO.1-rep1


title: Multiome - ATAC - PH27_C

description:
Library name: Multiome - ATAC - PH27_C
PH27_C_filtered_feature_bc_matrix.h5

source_name: Ovarian cancer in PDX

value: Ovarian cancer in PDX


title: Multiome - ATAC - PH1199

description:
Library name: Multiome - ATAC - PH1199
PH1199N_filtered_feature_bc_matrix.h5


title: SDS_D0-2+Topsym

description: Library name: SDS-0-2T


title: Multiome - GEX - PH626_T

description: Library name: Multiome - GEX - PH626_T

library_source: transcriptomic single cell

molecule: nuclear RNA

value: mRNA


title: SDS_D0-1+Topsym

description: Library name: SDS-0-1T


title: Multiome - GEX - PH27_T

description: Library name: Multiome - GEX - PH27_T


title: Multiome - ATAC - PH1303

description:
Library name: Multiome - ATAC - PH1303
1303_R_filtered_feature_bc_matrix.h5


title: P3 biol rep 3

city: Nijmegen

first: Sofia

last: Puvogel

address: Geert Grooteplein Zuid 10

country: Netherlands

institute: RadboudUMC

description: Library name: 105585-001-033

data_processing:
Quality Control: Fastp was used to eliminate PolyG artifacts and clip adapters, including a list of adapter sequences currently used by Illumina.
Read Alignment: Reads were mapped to the GRCh38 human reference genome using HISAT2. The library was reversely stranded, and the parameter --rna-strandness = RF was used. Resulting SAM files were sorted and indexed using SAMtools.
UMI Deduplication: Deduplication was performed with UMI-tools to account for unique molecular identifiers.
Assembly: GRCh38
Supplementary files format and content: Processed data are provided as a .txt count matrix generated through feature counting.

source_name: hiPSCs-derived astrocytes

growth_protocol: Astrocytes were differentiated from iPSCs using a previously described protocol (Schuurmans, I.M.E., Mordelt, A., Linda, K., et al., 2024).

extract_protocol:
Total RNA was extracted using the Quick-RNA Microprep Kit (Zymo Research, #ZY-R1051), following the manufacturer's protocol.
cDNA libraries were prepared using the NEBNext Ultra II Directional RNA Library Prep Kit. Sequencing was performed using paired-end reads on an Illumina NovaSeq 6000 platform at GenomeScan B.V., Leiden.

treatment_protocol: No treatment was applied. iPSC-derived astrocytes were grown for 35 days in vitro.

value: hiPSCs-derived astrocytes

value: iPSC

value: astrocytes

tag: time

value: Days in vitro 35


title: P3 biol rep 2

description: Library name: 105585-001-032


title: Multiome - GEX - PH27_C

description: Library name: Multiome - GEX - PH27_C


title: Multiome - GEX - PH626_C

description: Library name: Multiome - GEX - PH626_C


title: Rv1, BPTFKD, rep2

description: Library name: Rv1-BPTFKD-rep2


title: Multiome - ATAC - PH1252

description:
Library name: Multiome - ATAC - PH1252
PH1252_filtered_feature_bc_matrix.h5

value: Ovarian cancer


title: SDS_D3-1+Topsym

description: Library name: SDS-3-1T


title: Multiome - ATAC - PH1243

description:
Library name: Multiome - ATAC - PH1243
PH1243_filtered_feature_bc_matrix.h5


title: P3 biol rep 1

description: Library name: 105585-001-031


title: SDS_D5-2

description: Library name: SDS-5-2


title: Multiome - GEX - PH235_C

description: Library name: Multiome - GEX - PH235_C


title: SDS_D5-1

description: Library name: SDS-5-1


title: MS-2 rep 5

city: Cincinnati

first: Matthew

last: Weirauch

state: Ohio

address: 3333 Burnet Avenue

department: Center for Autoimmune Genomics and Etiology (CAGE)

institute: Cincinnati Children's Hospital Medical Center

description: Library name: MS-2 rep 5 CARR0059 E007524

data_processing:
library strategy: Massively Parallel Reporter Assay (MPRA)
Oligo-barcode association: Paired-end, 150bp reads were first quality filtered using Trimmomatic (v0.38) (flags: PE -phred33, LEADING:25, TRAILING:25, MINLEN:80). Read 1 was separated into the 20bp barcode region and the oligo-matching region. The trimmed oligo-matching regions of Read 1 and Read 2 were mapped back to the synthesized oligo sequences using Bowtie2 (v2.3.4.1) (flags: -X 250, -very-sensitive, -p 16). Barcodes were then associated with the oligo sequences using the read ID. Only uniquely mapped barcodes were used for downstream analysis.
Barcode counting: Single-end 150bp reads were first quality filtered using Trimmomatic (v0.38) (flags: PE -phred33, LEADING:3, TRAILING:3, MINLEN:70). Each read was then separated into the 20bp barcode region and the constant region. The trimmed constant regions of the reads were mapped back to the constant region within the eGFP 3′ UTR using Bowtie2 (v2.3.4.1) (flags: -very-sensitive, -p 16). Only reads with Levenshtein distance of 4 or less within the constant region and perfect matches to the two bases directly adjacent to the barcode were kept. Barcodes were then associated with the retained reads using the read ID. All barcodes were summarized at the oligo level.
Assembly: hg38
Supplementary files format and content: for BC-seq libraries: tab-delimited txt file containing oligo summaries for each sample
Supplementary files format and content: tab-delimited txt file containing barcode counts for each sample

growth_protocol: The GM12878, MS-1, and MS-2 cell lines were grown in RPMI medium supplemented with 10% FBS, 100ug/ml normocin, and 1X antibiotic-antimycotic

extract_protocol:
Total RNA was extracted by the RNeasy Midi Kit following the manufacturer’s instruction. Extracted RNA was subjected to DNase treatment. eGFP mRNA enrichment and elution were performed using biotinylated probes as in Lu et al (PMID: 33712590), and the mRNA was reverse transcribed to cDNA using SuperScript™ IV First-Strand Synthesis System with gene specific primer MPRA_v3_Amp2Sc_R, following the manufacturer’s instruction.
We assembled the MPRA libraries as in Lu et al  (PMID: 33712590) and Shook et al  (PMID: 38183988)

treatment_protocol: After transfection, cells were recovered in pre-warmed DMEM medium supplemented only with 10% FBS for 24hrs. Cells were then collected for preparation of the sequencing library for barcode counting.

value: EBV-transformed B cell


title: Multiome - GEX - PH1302

description: Library name: Multiome - GEX - PH1302


title: SDS_D3-2

description: Library name: SDS-3-2


title: MS-2 rep 4

description: Library name: MS-2 rep 4 CARR0059 E007523


title: P1 biol rep 2

description: Library name: 105585-001-029


title: SDS_D3-1

description: Library name: SDS-3-1


title: P1 biol rep 1

description: Library name: 105585-001-028


title: Multiome - GEX - PH1252

description: Library name: Multiome - GEX - PH1252


title: MS-2 rep 3

description: Library name: MS-2 rep 3 CARR0059 E007522


title: SDS_D1-2

description: Library name: SDS-1-2


title: Multiome - GEX - PH1243

description: Library name: Multiome - GEX - PH1243


title: SDS_D1-1

description: Library name: SDS-1-1


title: PDE KO biol rep 2

description: Library name: 105585-001-026


title: PDE KO biol rep 1

description: Library name: 105585-001-025


title: Multiome - GEX - PH235_T

description: Library name: Multiome - GEX - PH235_T


title: Multiome - GEX - PH1239

description: Library name: Multiome - GEX - PH1239


title: Multiome - GEX - PH1230

description: Library name: Multiome - GEX - PH1230


title: SDS_D0-1

description: Library name: SDS-0-1


title: P1 biol rep 3

description: Library name: 105585-001-030


title: Dot1l_KO_TT-seq_rep3

city: Montréal

first: Javier

middle: M

last: Di Noia

state: QC

address: 110 Pine Avenue West

country: Canada

institute: Institut de recherches cliniques de Montréal (IRCM)

description: Library name: KO3

data_processing:
*library strategy: TT(chem)-seq
Sequencing adapters were trimmed with Trimmomatic  (0.36) with (parameter- PE ILLUMINACLIP:adapters.fa:2:30:10: TRAILING:3 MINLEN:25).
Trimmed means were mapped to custom CH12F3 genome (modified mm10) and  S.cerevisiae R64 genomes with STAR (2.7.9a) following manual instructions for multi-sample 2-pass mapping.
Mapped reads to mouse genome were filtered for mapping quality (MAPQ>=1)  using samtools (1.16.1).
Scaling factors were calculated based on number of reads mapped to S. cerevisiae R64 genome (library size).
Bigwig files were generated using bamCoverage from deepTools (3.5.0) using the calculated scaling factors with a bin size of 10bp.
Assembly: custom reference genome reflecting the recombined V(D)J loci in CH12, produced by modifying mouse (mm10) reference genome based on the IgH and Igκ cDNA sequences from CH12F3 cells using Reform; and S.cerevisiae R64
Supplementary files format and content: bigwig (forward and reverse strands)

growth_protocol: CH12F3-G (a subclone with higher CSR activity derived by limiting dilution from CH12F3) were cultured in RPMI 1640 media (Wisent) supplemented with 10% fetal bovine serum (Wisent), 1% penicillin/streptomycin (Wisent) and 0.1 mM 2-mercaptoethanol (Bioshop).

extract_protocol:
4SU enriched RNA was extracted following TT(chem)-seq protocol (Gregersen et al 2020). Briefly, RNA was extracted by TRIzol (Thermo Fisher Scientific, 15596026) accordingly to the manufacturer’s instructions. As a control for equal sample preparation, we spiked-in S. cerevisiae (strain BY4741) 4-thiouracil (4TU)-labelled RNA. S. cerevisiae were grown in YPD medium overnight, diluted to an OD600 of 0.1, and grown to mid-log phase (OD600 of 0.8) and labelled with 5 mM 4TU (Sigma-Aldrich, 440736) for 5 min. Total RNA was extracted following a hot-phenol purification (Collin et al 2019.). For purification of 4SU labelled RNA, 100 μg mammalian 4SU labelled RNA was spiked-in 1/100 of 4TU-labelled S. cerevisiae RNA. The 101 μg RNA was fragmented by addition of 20 μL 1 M NaOH to fragment RNA and left on ice for 30 min. Fragmentation was stopped by addition of 80 μL 1 M Tris pH 6.8 and cleaned up twice with Micro Bio-Spin™ P-30 Gel Columns (BioRad, 7326223) according to the manufacturer’s instructions. Biotinylation of 4SU-residues were carried out in a total volume of 250 μl, containing 10 mM Tris-HCl [pH 7.4], 1 mM EDTA and 5mg MTSEA biotin-XX linker (Biotium, BT90066) for 30 min at room temperature in the dark. RNA was then purified by phenol:chloroform extraction, denatured by 10 min incubation at 65°C and added to 200 μL μMACS Streptavidin MicroBeads (Miltenyi, 130-074-101). RNA was incubated with beads for 15 min at room temperature and beads applied to a μColumn in the magnetic field of a μMACS magnetic separator. Beads were washed twice with pull-out wash buffer (100 mM Tris-HCl, pH 7.4, 10 mM EDTA, 1 M NaCl and 0.1% Tween20). 4SU-RNA was eluted twice by addition of 100 mM DTT and RNA cleaned up using the RNeasy MinElute kit (Qiagen, 74204).
Libraries were prepared using the strand-specific TruSeq total RNA kit (Illumina) using 5 min 65°C fragmentation incubation to anneal primers but to prevent further fragmentation of the samples.

value: unstimulated


title: Multiome - GEX - PH1214

description: Library name: Multiome - GEX - PH1214


title: Multiome - GEX - PH1238

description: Library name: Multiome - GEX - PH1238


title: MS-1 rep 5

description: Library name: MS-1 rep 5 CARR0082 E007663


title: MS-1 rep 2

description: Library name: MS-1 rep 2 CARR0082 E007660


title: MS-1 rep 3

description: Library name: MS-1 rep 3 CARR0082 E007661


title: Dot1l_KO_TT-seq_rep1

description: Library name: KO1


title: Multiome - GEX - PH1199

description: Library name: Multiome - GEX - PH1199


title: C4 biol rep 3

description: Library name: 105585-001-024


title: C4 biol rep 2

description: Library name: 105585-001-023


title: Multiome - GEX - PH1303

description: Library name: Multiome - GEX - PH1303


title: C1 biol rep 3

description: Library name: 105585-001-021


title: PDE KO biol rep 3

description: Library name: 105585-001-027


title: MS-1 rep 1

description: Library name: MS-1 rep 1 CARR0082 E007659


title: MS-2 rep 2

description: Library name: MS-2 rep 2 CARR0059 E007521


title: SDS_D0-2

description: Library name: SDS-0-2


title: WT_TT-seq_rep3

description: Library name: WT3


title: MS-2 rep 1

description: Library name: MS-2 rep 1 CARR0059 E007520


title: MS-1 rep 4

description: Library name: MS-1 rep 4 CARR0082 E007662


title: Treg FOXP3 high bin, Donor 2, CRISRPi screen

city: San Francisco

first: Alexander

last: Marson

state: CA

address: 1650 Owens St.

institute: UCSF Gladstone Institutes

description: Library name: D2_Treg_high_FOXP3_CRISPRi_screen

data_processing:
CRISPRi pooled screens were analyzed using MAGeCK (v0.5.9.5). A count file containing individual gRNA abundance across all donors was generated using the command mageck count with specification –norm-method none. Differential enrichment of gRNAs was determined using DESeq2 (v1.40.2), as described previously (Mowery et al., 2024). Guide RNAs with fewer than ten reads were excluded. Guide RNAs not mapping to the FOXP3 locus were excluded from visualization.
Assembly: GRCh38/hg38
Supplementary files format and content: Combined DESeq2 analysis output file for Tregs and Tconvs comparing gRNA high and low bin enrichment.
Library strategy: CRISPR Screen

source_name: Blood

growth_protocol: Primary human T cells subsets were isolated from peripheral blood mononuclear cells (PBMCs) sourced from consented, fresh Human Peripheral Blood Leukopaks (STEMCELL Technologies, catalog no. 70500). Leukopaks were washed twice with 1X volume of EasySep Buffer (DPBS, 2% fetal bovine serum [FBS], 1 mM pH 8.0 EDTA) using centrifugation and resuspended at 50e6-200e6 cells/mL. CD4+CD25highCD127low Tregs and CD4+CD25low Tconv were isolated from washed PBMCs using the EasySep Human CD4+CD127lowCD25+ Regulatory T Cell Isolation Kit (STEMCELL Technologies, catalog no. 18063) according to manufacturer’s protocol. For enhanced cell purity, Tregs were stained for CD4 (Biolegend, catalog no. 344634 or 344620), CD25 (Tonbo, catalog no. 20-0259-T100), and CD127 (Becton Dickinson, catalog no. 557938) and further sorted into a CD4+CD25highCD127low population via FACS on a BD FACSAria or BD FACSAria Fusion I. Tregs and Tconvs were cultured in X-VIVO15 media (Lonza, catalog no. 02-053Q) supplemented with 5% FBS, 55 µM 2-Mercaptoethanol, and 4 mM N-Acetyl-L-Cysteine. Recombinant Human IL-2 (R&D Systems, catalog no. 202-GMP or BT-002-GMP) was supplemented at 200 IU/mL, and cells were stimulated for 48 hours with CTS Dynabeads CD3/CD28 beads (Thermo Fisher Scientific, catalog no. 40203D) at a cell to bead ratio of 1:1. After 48 hours, beads were removed using magnetic separation. Cells were cultured at 37°C and 5% CO2 and split to 5E5-1.5E6 cells/mL every ~48 hours by topping off media and completely replacing the appropriate dose of IL-2.

extract_protocol:
Forty-eight hours post-restimulation, Tregs and Tconvs were counted, washed, and stained with Ghost Dye Red 780 Live/Dead stain (Tonbo, catalog no. 13-0865-T500) and antibodies targeting CD4 (Biolegend, catalog no. 344634), FOXP3 (Biolegend, catalog no. 320112) and HELIOS (Biolegend, catalog no. 137216). Cells were sorted on lymphocytes, singlets, live cells, BFP+ (a marker of the gRNA library plasmid), and FOXP3 high and low bins that captured the top and bottom ~25% of FOXP3 expression in each cell type. Following sorting, cells were pelleted and lysed, and genomic DNA (gDNA) was extracted using phenol-chloroform gDNA extraction.
DNA regions containing the lentivirally integrated gRNA were amplified using PCR with custom primers. PCR reactions were conducted using 2 µg gDNA per 50 µL reaction with 0.5 µM forward and reverse primers, 0.075 U/µL TaKaRa Ex Taq DNA polymerase (Takara Bio, catalog no. RR001C), 0.2 mM dNTP, and 1X Ex Taq Buffer and amplified under the following cycling conditions: 1 min at 95°C, (30 s at 95°C, 30 s at 60°C, 30 s at 72°C) x 25 cycles, 10 min at 72°C, hold at 4°C. Following amplification, aliquots of individual samples were pooled, and primer dimers removed using a 1.4X SPRI bead cleanup. Proper amplification and concentration were assessed using Qubit quantification and DS1000 High Sensitivity Tapestation analysis, according to manufacturers’ protocols. Samples were pooled equimolarly and sequenced on an Illumina HiSeq4000 instrument using a custom sequencing primer.

treatment_protocol: Twenty-four hours post-isolation and stimulation, cells were infected with titered dCas9-ZIM3 lentivirus, targeting approximately 80% infection, by addition to culture flasks. Cells and lentivirus were gently pipetted once with a large serological pipette to mix. Forty-eight hours post-stimulation, cells were infected with gRNA library lentivirus, targeting approximately 60% infection, by addition to culture flasks and gently mixing. Twenty-four hours post-gRNA library lentivirus infection, viral media was removed, and cells were resuspended in media containing 1.5 ug/mL puromycin to select for cells with proper gRNA integration. Cells were cultured in puromycin-supplemented media for 48 hours. Nine days post-initial stimulation, cells were restimulated at a 1:1 cell to bead ratio with CTS Dynabeads CD3/CD28 (Thermo Fisher Scientific, catalog no. 40203D). Forty-eight hours post-restimulation, beads were separated from cells using magnetic isolation and prepared for phenotyping via fluorescence-activated cell sorting (FACS).

value: Blood

tag: donor

value: Donor 2

value: Treg

tag: crispr experiment

value: CRISPRi FOXP3 locus tiling

tag: group

value: FOXP3 high


title: C1 biol rep 2

description: Library name: 105585-001-020

value: WT


title: Dot1l_KO_TT-seq_rep2

description: Library name: KO2


title: C4 biol rep 1

description: Library name: 105585-001-022


title: C1 biol rep 1

description: Library name: 105585-001-019


title: GM12878 rep 5

description: Library name: GM12878 rep 5 E007519

tag: biomaterial provider


title: WT_TT-seq_rep2

description: Library name: WT2


title: Treg FOXP3 low bin, Donor 2, CRISRPi screen

description: Library name: D2_Treg_low_FOXP3_CRISPRi_screen

value: FOXP3 low


title: WT_TT-seq_rep1

description: Library name: WT1


title: Treg FOXP3 high bin, Donor 1, CRISRPi screen

description: Library name: D1_Treg_high_FOXP3_CRISPRi_screen

value: Donor 1


title: Treg FOXP3 low bin, Donor 1, CRISRPi screen

description: Library name: D1_Treg_low_FOXP3_CRISPRi_screen


title: GM12878 rep 3

description: Library name: GM12878 rep 3 E007517


title: GM12878 rep 1

description: Library name: GM12878 rep 1 E007515


title: GM12878 rep 2

description: Library name: GM12878 rep 2 E007516


title: GM12878 rep 4

description: Library name: GM12878 rep 4 E007518


title: S5P-RNAPII_WT_rep2

description: Library name: S5P_RNAPII_WT_2

data_processing:
Sequencing adapters and first 10bp were trimmed with fastp 0.23.1.
Trimmed reads were aligned with bowtie2 (2.4.4) to the spiked-in Drosophila melanogaster genome (dm6) and custom CH12F3 genome (modified mm10).
Duplicates reads were removed with MarkDuplicatesSpark from Picard tools through gatk (4.2.4.0) and were filtered for mapping quality (MAPQ>=1) using samtools (1.16.1).
Normalization was performed using DiffBind package (3.8.3), specifically by using DESeq2 (1.38.2) to calculate scaling factors based on “background bin” signals estimated by the csaw package (1.32.0).
These scaling factors were then used to generate bigwig files using bamCoverage and bamCompare from deepTools (3.5.0) with a bin size of 20bp.
Assembly: custom reference genome reflecting the recombined V(D)J loci in CH12, produced by modifying mouse (mm10) reference genome based on the IgH and Igκ cDNA sequences from CH12F3 cells using Reform; and dm6
Supplementary files format and content: bigWig

growth_protocol: CH12F3-G (a subclone with higher CSR activity derived by limiting dilution from CH12F3) were cultured in RPMI 1640 media (Wisent) supplemented with 10% fetal bovine serum (Wisent), 1% penicillin/streptomycin (Wisent) and 0.1umM 2-mercaptoethanol (Bioshop).

extract_protocol:
10 million CH12F3 cells cultured in 10mL of media were cross-linked with 1% formaldehyde for 8 minutes at room temperature, washed and resuspended in RIPA buffer and sonicated with Covaris E220 for 10 min with 10% Duty Factor, 140W PIP and 200 CBP and clarified by centrifugation. S2/Kc167 chromatin was prepared by crosslinking cells in 1X PBS containing 1% formaldehyde at RT for 20 min and quenched with 125 mM glycine for 5 min, washed and resuspended in 130 μL of RIPA buffer and fragmented using Covaris E220 for 150s with 140W PIP, 20% DF and 200 CPB and clarified by centrifugation. For spike-in, 1μg of S2/Kc167 chromatin was spiked to 24μg of CH12-F3 chromatin (1:24 ratio) before IP. Samples were pre-cleared with 20μL of Dynabeads Protein G for 1h (4°C). For the IP, 30μL Dynabeads Protein G were coupled with the target Ab (amounts as per manufacturer instructions) and 2μg of a spike-in Ab against Drosophila H2Av for 30 min and blocked 1h (4°C) with BT buffer. IP was performed by overnight incubation of the pre-cleared chromatin with blocked Ab-beads complexes. Beads were then washed and immunocomplexes were eluted and incubated overnight with proteinase K at 65°C. DNA was purified using QIAquick PCR purification kit (Qiagen), and DNA resuspended in 50μl Tris-HCl (pH 8).
Libraries were prepared using the KAPA HyperPrep library kit (Roche Diagnostics) as per manufacturer's instructions. Normalization of the samples quantities was done after quantification of the ligation products by qPCR and library size distribution was assessed on a 2100 bioanalyzer.


title: S5P-RNAPII_Dot1l_KO_rep1

description: Library name: S5P_RNAPII_KO_1


title: Tconv FOXP3 high bin, Donor 3, CRISRPi screen

description: Library name: D3_Tconv_high_FOXP3_CRISPRi_screen

value: Donor 3

value: Tconv


title: S5P-RNAPII_Dot1l_KO_rep2

description: Library name: S5P_RNAPII_KO_2


title: Tconv FOXP3 high bin, Donor 1, CRISRPi screen

description: Library name: D1_Tconv_high_FOXP3_CRISPRi_screen


title: HUVEC small RNA input, rep3

city: Boston

first: Ryan

last: Flynn

address: 1 Blackfan St

institute: Harvard University

data_processing:
Data was processed using the Basepair platform (https://app.basepairtech.com/) and the "Flynn Lab Human Alignment v1" automated pipeline therein.
Assembly: Hg38 and custom indexes for repetitive RNAs
Supplementary files format and content: The processed data is provided as an excel table of all mapped reads to each transcript unifed across all the indexes mapped to

source_name: endothelium

growth_protocol: Primary umbilical vein endothelial cells (HUVEC) (ATCC) were cultured in Vascular Cell Basal Medium (ATCC) supplemented with Endothelial Cell Growth Kit-VEGF (ATCC).

extract_protocol:
Cell pellets were directly lysed in 500 µL of RNAzol RT (Molecular Research Center). Samples were shaken at 50°C for 5 minutes. To phase separate the RNA, 0.4X volumes of water was added, vortexed, let to stand for 5 minutes at 25°C and lastly spun at 12,000x g at 4°C for 15 minutes. The aqueous phase was transferred to clean tubes and 1.1X volumes of isopropanol was added. The RNA was then purified over a Zymo column (Zymo Research). For all column cleanups, we followed the following protocol. First, 350 µL of pure water was added to each column and spun at 10,000x g for 30 seconds, and the flowthrough was discarded. Next, precipitated RNA from the RNAzol RT extraction (or binding buffer precipitated RNA, below) was added to the columns, spun at 10,000x g for 20 seconds, and the flowthrough was discarded. This step was repeated until all the precipitated RNA was passed over the column once. Next, the column was washed three times total: once using 400 µL of RNA Prep Buffer (3M GuHCl in 80% EtOH), twice with 400 µL of 80% ethanol. The first two spins were at 10,000x g for 20 seconds, the last for 30 seconds.
Libraries made following MSR-seq, PMID: 35513407

value: endothelium

value: HUVEC

value: endothelial


title: S2P-RNAPII_Dot1l_KO_rep2

description: Library name: S2P_RNAPII_KO_2


title: Tconv FOXP3 low bin, Donor 3, CRISRPi screen

description: Library name: D3_Tconv_low_FOXP3_CRISPRi_screen


title: HUVEC small RNA input, rep1


title: HUVEC small RNA input, rep2


title: Tconv FOXP3 low bin, Donor 1, CRISRPi screen

description: Library name: D1_Tconv_low_FOXP3_CRISPRi_screen


title: Treg FOXP3 high bin, Donor 2, CRISRP KO TF screen

description: Library name: D2_Treg_high_FOXP3_CRISPRKO_screen

data_processing:
CRISPR KO pooled screens were analyzed using MAGeCK (v0.5.9.5). A count file containing individual gRNA abundance across all donors was generated using the command mageck count with specification –norm-method none. Differentially enriched gRNAs and significantly enriched genes in high and low bins were identified using the command mageck test, specifying –sort-criteria pos.
Supplementary files format and content: Combined Treg and Tconv gene and gRNA level MAGeCK results.
Library strategy: CRISPR Screen

extract_protocol:
Forty-eight hours post-restimulation, cells were counted and stained with Ghost Dye Red 780 Live/Dead stain (Tonbo, catalog no. 13-0865-T500) and antibodies targeting FOXP3 (Biolegend, catalog no. 320116) and HELIOS (Biolegend, catalog no. 137216). Cells were sorted on lymphocytes, singlets (x2), live cells, GFP+ (a marker of the gRNA library plasmid), and FOXP3 high and low bins that captured the top and bottom 25% of FOXP3 expression. Following sorting, cells were pelleted and lysed, and gDNA was extracted using phenol-chloroform gDNA extraction.
DNA regions containing the lentivirally integrated gRNA were amplified using PCR with custom primers. PCR reactions were conducted using 1.75 µg gDNA per 50 µL reaction with 0.25 µM forward and reverse primers and 1X NEBNext Ultra II Q5 Master Mix (New England Biolabs, catalog no. M0544). Reactions were amplified under the following cycling conditions: 3 min at 98°C, (10 s at 98°C, 10 s at 63°C, 25 s at 72°C) x 23 cycles, 2 min at 72°C, hold at 4°C. Following amplification, aliquots of individual samples were pooled, and primer dimers removed using a 1.25X SPRI bead cleanup. Proper amplification and concentration were assessed using Qubit quantification and DS1000 High Sensitivity Tapestation analysis, according to manufacturers’ protocols. Samples were pooled equimolarly and sequenced on an Illumina HiSeq4000 instrument using a custom sequencing primer.

treatment_protocol: Twenty-four hours post-isolation and stimulation, cells were infected with titered gRNA library lentivirus, targeting approximately 85% infection, by addition to culture flasks. Cells and lentivirus were gently pipetted once with a large serological pipette to mix. Twenty-four hours post-gRNA library lentivirus infection, viral media was removed, and cells were washed with 1X volume pre-warmed media and plated at approximately 1E6 cells/mL. Cas9 RNPs were assembled using a Guide Swap crRNA method (Ting et al., 2018; Edit-R crRNA nontargeting Control 3, Dharmacon, catalog no. U-007503-01-05). Briefly, non-targeting crRNA (Dharmacon) and Edit-R CRISPR-Cas9 Synthetic tracrRNA (Dharmacon, catalog no. U-002005-20) were resuspended in Nuclease-Free Duplex Buffer (IDT, catalog no. 11-01-03-01) at 160 uM, complexed at a 1:1 molar ratio to create a 80 µM solution of RNA complexes, and incubated 30 min at 37°C. Single-stranded oligonucleotides (ssODN; 100 µM stock, sequence: TTAGCTCTGTTTACGTCCCAGCGGGCATGAGAGTAACAAGAGGGTGTGGTAATATTACGGTACCGAGCACTATCGATACAATATGTGTCATACGGACACG) were mixed at a 1:1 molar ratio with complexed RNA and incubated 5 min at 37°C. Cas9 protein (Berkeley Macrolab, 40 µM stock) was slowly added at a 1:1 molar ratio of Cas9 to RNA complexes, mixed thoroughly, and incubated 15 min at 37°C. RNPs were frozen at -80°C and thawed at room temperature prior to use. Seventy-two hours post-isolation and stimulation, cells were pelleted, and resuspended at 5E4 cells/µL (Treg) or 7.5E4 cells/µL (Tconv) in freshly supplemented P3 Primary Cell Nucleofector Solution (Lonza, catalog no. V4SP-3096). 20 µL of cells were mixed with 3.5 µL (50 pmol; Tregs) or 7 µL (100 pmol; Tconvs) RNP, and 22 µL (Tregs) or 25 µL (Tconvs) was transferred to a 96-well Nucleocuvette Plate (Lonza, catalog no. V4SP-3096). Cells were nucleofected using a Lonza 4D 96-well electroporation system with pulse code EO-115 (human Treg) or EH-115 (human Tconv). Immediately after electroporation, 80 µL pre-warmed media was added to each well, and the cells were incubated for 15 min at 37°C. Following incubation, cells were plated at approximately 1E6 cells/mL for culture. Nine days post-initial stimulation, cells were restimulated at a 1:1 cell to bead ratio with CTS Dynabeads CD3/CD28 (Thermo Fisher Scientific, catalog no. 40203D). Forty-eight hours post-restimulation, beads were separated from cells using magnetic isolation and prepared for phenotyping via fluorescence-activated cell sorting (FACS).

value: CRISPR KO TF screen


title: S2P-RNAPII_Dot1l_KO_rep1

description: Library name: S2P_RNAPII_KO_1


title: HUVEC small RNA, VEGFA165 IP, rep3


title: S2P-RNAPII_WT_rep2

description: Library name: S2P_RNAPII_WT_2


title: Treg FOXP3 low bin, Donor 2, CRISRP KO TF screen

description: Library name: D2_Treg_low_FOXP3_CRISPRKO_screen


title: HUVEC small RNA, VEGFA165 IP, rep2


title: S2P-RNAPII_WT_rep1

description: Library name: S2P_RNAPII_WT_1


title: HUVEC small RNA, VEGFA165 IP, rep1


title: Treg FOXP3 high bin, Donor 1, CRISRP KO TF screen

description: Library name: D1_Treg_high_FOXP3_CRISPRKO_screen


title: Treg FOXP3 low bin, Donor 1, CRISRP KO TF screen

description: Library name: D1_Treg_low_FOXP3_CRISPRKO_screen


title: HUVEC small RNA, noVEGFA165 IP, rep3


title: Tconv FOXP3 high bin, Donor 3, CRISRP KO TF screen

description: Library name: D3_Tconv_high_FOXP3_CRISPRKO_screen


title: HUVEC small RNA, noVEGFA165 IP, rep2


title: RNAPII_Dot1l_KO_rep1

description: Library name: RNAPII_KO_1


title: RNAPII_WT_rep1

description: Library name: RNAPII_WT_1


title: HUVEC small RNA, noVEGFA165 IP, rep1


title: Tconv FOXP3 high bin, Donor 2, CRISRP KO TF screen

description: Library name: D2_Tconv_high_FOXP3_CRISPRKO_screen


title: Nipbl_Dot1l_KO_rep2

description: Library name: Nipbl_KO_2


title: V_ypd_3

city: Exeter

first: Rhys

last: Farrer

state: Devon

address: Stocker Road

department: MRC Center for Medical Mycology

country: United Kingdom

institute: University of Exeter

data_processing:
Reads were trimmed with fastp v.0.23.1 to remove reads < 75 bases and trim based from the 3’ end with q-score < 22. Alignment and calculation of normalised gene expression were performed with the Trinity pipeline v2.15.1.
Reads were aligned to the GJEY01 AK de novo adult gill transcriptome or to the B8441 v2 C. auris reference genome.
Assembly: Aphanius dispar GJEY00000000.1, Candida auris Cand_auris_B8441_V2
Supplementary files format and content: Tab-separated text files include Fragments per Kilobase per Million (FPKM).

organism: Candidozyma auris

source_name: Yeast cells only

growth_protocol: Arabian Killifish were hatched overnight and reared at 28 °C for 72 h with day and night cycles before microinjection, after which point they were kept at 37 °C in 35 ppt artificial seawater.

extract_protocol:
Total RNA was extracted using the Monarch® Total RNA Miniprep Kit (NEB, USA). Embryos were flash frozen in liquid nitrogen, re-suspended in DNA/RNA Protection Reagent and disrupted with an autoclaved plastic micropestle. Bead beating was performed in a FastPrep 24 homogenizer (MP Biomedicals) with 0.5 mm zirconia/silica beads for 8 cycles of 35 s plus 45 s on ice. Lysate was recovered and incubated for 5 min. at 55 °C with proteinase K before addition of lysis buffer and RNA purification following manufacturer’s instructions.
Samples were normalised and prepared using the NEBNext Ultra II Directional RNA Library Prep - Poly(A) mRNA Magnetic Isolation Module following the standard protocol.

treatment_protocol: Yolk sac microinjection with 10 nl solution of ~500 yeasts in phenol red and deionised water, or sham injection, or no injection.

value: Yeast cells only

value: V

value: C. auris grown in YPD

tag: time point

value: 24 hours


title: Tconv FOXP3 low bin, Donor 2, CRISRP KO TF screen

description: Library name: D2_Tconv_low_FOXP3_CRISPRKO_screen

value: Blood

value: Donor 2


title: I_ypd_3

value: I


title: Tconv FOXP3 high bin, Donor 1, CRISRP KO TF screen

description: Library name: D1_Tconv_high_FOXP3_CRISPRKO_screen

value: FOXP3 high


title: Nipbl_WT_rep2

description: Library name: Nipbl_WT_2

value: WT

value: unstimulated


title: Tconv FOXP3 low bin, Donor 1, CRISRP KO TF screen

description: Library name: D1_Tconv_low_FOXP3_CRISPRKO_screen

value: Donor 1

value: FOXP3 low


title: Tconv FOXP3 low bin, Donor 3, CRISRP KO TF screen

description: Library name: D3_Tconv_low_FOXP3_CRISPRKO_screen

value: Donor 3

value: Tconv


title: Nipbl_WT_rep1

description: Library name: Nipbl_WT_1


title: S5P-RNAPII_WT_rep1

description: Library name: S5P_RNAPII_WT_1


title: RHP5836_TE

city: Singapore

first: Nicholas

last: Ang

address: 8a Biomedical Grove, Singapore 138648

country: Singapore

institute: Singapore Immunology Network (SIgN)

data_processing:
Paired-end 2 x 150 bp reads from Illumina sequencing were quality-checked using fastqc. Abundances of human transcripts obtained from gencode version 27 were estimated from the sequencing reads using Salmon version 0.8.2. Transcript abundances were summarized into gene counts using the tximport R/Bioconductor package. The gene abundances were normalized to transcript per million (TPM) and then log-transformed as log 2 (TPM + 1.0) with a pseudo-count of 1.0 added to prevent negative values. The normalized log transformed abundances were displayed in heatmaps where they were scaled to a Z-score in each row for coloring. Gene set enrichment analysis was performed using Broad Institute’s GSEA tool version 3.0 with immunologic signature gene sets (C7) collection of the MSIG database version 6.2 as the reference. The connectivity map (CMAP) algorithm was implemented using custom R scripts. Gene signatures used in CMAP analysis were defined by performing differential gene expression analysis using DESeq2 (for RNA-seq data) or limma (for microarray data) R/Bioconductor packages and then selecting differentially expressed genes based on a false discovery rate of 0.05. TCR repertoire analysis was performed with MiXCR where the same number of initial reads (8 million) was taken for analysis from each sample to normalize the results.
Assembly: The paired-end reads were mapped to Human GRCh38 reference genome using the STAR alignment tool.
Supplementary files format and content: Paired-end 2 x 150 bp reads from Illumina sequencing were quality-checked using fastqc. Abundances of human transcripts obtained from gencode version 27 were estimated from the sequencing reads using Salmon version 0.8.2. Transcript abundances were summarized into gene counts using the tximport R/Bioconductor package. The gene abundances were normalized to transcript per million (TPM) and then log-transformed as log 2 (TPM + 1.0) with a pseudo-count of 1.0 added to prevent negative values. The normalized log transformed abundances were displayed in heatmaps where they were scaled to a Z-score in each row for coloring. Gene set enrichment analysis was performed using Broad Institute’s GSEA tool version 3.0 with immunologic signature gene sets (C7) collection of the MSIG database version 6.2 as the reference. The connectivity map (CMAP) algorithm was implemented using custom R scripts. Gene signatures used in CMAP analysis were defined by performing differential gene expression analysis using DESeq2 (for RNA-seq data) or limma (for microarray data) R/Bioconductor packages and then selecting differentially expressed genes based on a false discovery rate of 0.05. TCR repertoire analysis was performed with MiXCR where the same number of initial reads (8 million) was taken for analysis from each sample to normalize the results.

source_name: peripheral blood mononuclear cells

extract_protocol:
Total RNA was extracted following the double extraction protocol: RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction (TRIzol, Thermo Fisher Scientific, Waltham, MA, USA) followed by a Qiagen RNeasy Micro clean-up procedure (Qiagen, Hilden, Germany). All human RNAs were analyzed on Agilent Bioanalyzer for quality assessment with RNA Integrity Number (RIN) ranging from 6.2 to 9.6 and median RIN 8.9 (Agilent, Santa Clara, CA, USA).
cDNA libraries were prepared using 1ng of total RNA and 0.5ul of a 1:50,000 dilution of ERCC RNA Spike in Controls (Ambion Thermo Fisher Scientific, Waltham, MA, USA) using SMARTSeq v2 protocol except for the following modifications: 1, Use of 20µM TSO; 2, Use of 250pg of cDNA with 1/5 reaction of Illumina Nextera XT kit (Illumina, San Diego, CA, USA). The length distribution of the cDNA libraries was monitored using a DNA High Sensitivity Reagent Kit on the Perkin Elmer Labchip (Perkin Elmer, Waltham, MA, USA). All samples were subjected to an indexed PE sequencing run of 2x51 cycles on an Illumina HiSeq 2000 (16 samples/lane).

value: peripheral blood mononuclear cells


title: IV_ypd_3

value: IV


title: II_ypd_3

value: II


title: Rad21_Dot1l_KO_rep2

description: Library name: Rad21_KO_2


title: RHP5835_Naive


title: RNAPII_Dot1l_KO_rep2

description: Library name: RNAPII_KO_2


title: Rad21_Dot1l_KO_rep1

description: Library name: Rad21_KO_1


title: I_ypd_2

value: Yeast cells only


title: RHP5834_CMV NLPV CD45RA+


title: II_ypd_2


title: RNAPII_WT_rep2

description: Library name: RNAPII_WT_2


title: RHP5833_CMV NLPV CD27+


title: III_ypd_2

value: III


title: Rad21_WT_rep2

description: Library name: Rad21_WT_2


title: Rad21_WT_rep1

description: Library name: Rad21_WT_1


title: III_ypd_3

value: C. auris grown in YPD


title: RHP5832_CMV NLPV


title: IV_ypd_2

value: 24 hours


title: H3K79me3_rep2

description: Library name: H3K79me3_2


title: V_ypd_2

value: V


title: V_ypd_1


title: RHP5829_CMV NLPV_4


title: RHP5830_Naive_1


title: H3K79me2_rep2

description: Library name: H3K79me2_2


title: RHP5831_TE_2


title: IV_ypd_1


title: RHP5828_EBV EBNA3B


title: H3K79me2_rep1

description: Library name: H3K79me2_1


title: III_ypd_1


title: Primary Tumor 2, No substrate, Normoxia, Replicate 3

city: Marseille

first: Ghislain

last: BIDAUT

address: 27 Bd Leï Roure

department: Bioinformatics

institute: Centre de Recherche en Cancérologie de Marseille

description:
Library name: P2_plastic_N_3_S39
N_P2plastic_S39

data_processing:
Trimming and adapter removal with Fasp 0.32 (cutoff quality 21)
Mapping to reference genome mm10 with Subread 1.6.4
Gene count with featureCounts from Subread 1.6.4 package
Assembly: mm10
Supplementary files format and content: Tab delimited file with raw gene counts profiles annotated with gene symbols

source_name: Pancreas

growth_protocol: PK4A cells were maintained in human plasma-like medium (HPLM) supplemented with 10% fetal calf serum (FCS), 100units/ml of penicillin, 100μg/ml of streptomycin, and 0.25μg/ml of amphotericin B in a humidified incubator with 5% CO2 at 37C.

extract_protocol:
RNA from three independent experiments was isolated using a RNeasy Plus Kit (Qiagen, Netherlands) and the RNA Integrity Number was determined with an RNA analysis kit (Agilent, Santa Clara, CA).
mRNA library preparation was performed with 50ng of RNA per sample using an Illumina Stranded mRNA Prep (Illumina, San Diego, CA) following the manufacturer’s instructions.

treatment_protocol: Cells were seeded on previously decellularized extracellular matrix from PDAC cancer-associated fibroblasts or on matrix-free cutlure vessels in human plasma-like medium supplemented with 10% fetal calf serum (FCS), 100units/ml of penicillin, 100μg/ml of streptomycin, and 0.25μg/ml of amphotericin B in a humidified incubator with 5% CO2 at 37C for 48h.

value: Pancreas

value: Murine PDAC cells


title: RHP5826_Naive_2


title: Primary Tumor 2, No substrate, Normoxia, Replicate 2

description:
Library name: P2_plastic_N_2_S23
N_P2plastic_S23


title: RHP5825_CMV NLPV_3


title: H3K79me3_rep1

description: Library name: H3K79me3_1


title: Negative Control

organism: blank sample

value: Negative control


title: RHP5824_CMV NLPV_2


title: Primary Tumor 2, No substrate, Normoxia, Replicate 1

description:
Library name: P2_plastic_N_1_S7
N_P2plastic_S7


title: Primary Tumor 2, ECM substrate, Normoxia, Replicate 3

description:
Library name: P2_ECM_N_3_S35
N_P2ECM_S35

value: Culture on extracellular matrix


title: Nipbl_Dot1l_KO_rep1

description: Library name: Nipbl_KO_1

value: unstimulated


title: I_ypd_1

value: I


title: Primary Tumor 2, ECM substrate, Normoxia, Replicate 1

description:
Library name: P2_ECM_N_1_S3
N_P2ECM_S3


title: HET_Male_input_R4

city: Philadelphia

first: Marissa

last: Maroni

state: PA

address: 3400 Civic Center Blvd

institute: University of Pennsylvania

description: Library name: Sample_16

data_processing:
Paired-end sequencing reads from the Illumina Nextseq 1000 were pre-processed using Illumina BCL2Fastq.
Fastq files were aligned by Bowtie2
bam files were subset for 18 millions reads for each biological rep using SAMTools for downstream analysis. Biological replicates were merged for either entire condition (i.e. R1-R4, 2 male, 2 females) or for each sex (i.e. R1-R2, 2 females and R3-R4, 2 mles) depending on the analysis.
NGSplot was used to generate metaplots
deepTools was used to generate bigwig files and raw counts
Assembly: mm10
Supplementary files format and content: bigwig files generated by deepTools from bam files aligned to bowtie2

source_name: Cortex

extract_protocol:
Cortical tissue was homogenized with a douncer in homogenization buffer supplemented with 1 mM DTT, 0.15 mM spermine, 0.5 mM spermidine, RNasin ® Plus Ribonuclease Inhibitor (Promega N2611), and EDTA-free protease inhibitor (Roche). Nuclei were isolated using an iodixanol density gradient and ultradentrifugation.
TruSeq ChIP Library Preparation Kit

value: Cortex

tag: Sex

value: Male

value: DOT1L floxed/+; CMV-Cre +/-


title: Primary Tumor 2, ECM substrate, Normoxia, Replicate 2

description:
Library name: P2_ECM_N_2_S19
N_P2ECM_S19


title: Primary Tumor 1, No substrate, Normoxia, Replicate 3

description:
Library name: P1_plastic_N_3_S37
N_P1plastic_S37


title: RHP5823_CMV #73

value: peripheral blood mononuclear cells


title: RHP5819_EBV EBNA3B_1


title: RHP5820_CMV NLPV_1


title: HET_Male_input_R3

description: Library name: Sample_14


title: IgG_control_R1

description: Library name: Sample_17

value: pooled male and female

value: Both CMV-Cre +/- and DOT1L floxed/+; CMV-Cre +/-


title: RHP5822_CMV NLPV CD27-_2


title: RHP5818_CMV NLPV CD27-_1


title: Control_Male_input_R4

description: Library name: Sample_15

value: CMV-Cre +/-


title: RHP5827_TE_1


title: II_ypd_1

value: Yeast cells only

value: II


title: Control_Male_input_R3

description: Library name: Sample_13


title: IgG_control_R2

description: Library name: Sample_18


title: Primary Tumor 1, ECM substrate, Normoxia, Replicate 3

description:
Library name: P1_ECM_N_3_S33
N_P1ECM_S33


title: Primary Tumor 1, ECM substrate, Normoxia, Replicate 2

description:
Library name: P1_ECM_N_2_S17
N_P1ECM_S17


title: HET_Female_input_R2

description: Library name: Sample_12

value: Female


title: RHP5821_CMV NLPV CD27+_2


title: Primary Tumor 1, No substrate, Normoxia, Replicate 1

description:
Library name: P1_plastic_N_1_S5
N_P1plastic_S5


title: Primary Tumor 1, No substrate, Normoxia, Replicate 2

description:
Library name: P1_plastic_N_2_S21
N_P1plastic_S21


title: Primary Tumor 1, ECM substrate, Normoxia, Replicate 1

description:
Library name: P1_ECM_N_1_S1
N_P1ECM_S1


title: Control_Female_input_R2

description: Library name: Sample_11


title: HET_Female_input_R1

description: Library name: Sample_10


title: RHP5816_CMV NLPV Mix


title: RHP5815_EBV EBNA3B


title: HFD_LFPKO_Fast_rep3

city: NEW YORK

first: Domenico

last: Accili

state: NY

address: 1150 ST NICHOLAS AVENUE, RM238

department: Medicine

institute: Columbia University

description:
Library name: DA024
DA024

data_processing:
Sequenced reads from the RNA-seq experiment were aligned to mouse genome mm10 using kallisto
Assembly: Mouse genome assembly GRCm38 (Genome Reference Consortium Mouse Build 38). Alignment and annotation were performed using Ensembl release 102.
Supplementary files format and content: tab-deliminated text files including TPM values for each sample

source_name: liver

extract_protocol:
Isolated liver tissues were snap frozened in liquid nitrogen, and RNA was harvested using Trizol reagent.
Libraries were prepared using the TruSeq Stranded mRNA Sample Prep Kit (Illumina)

treatment_protocol: Regular diet or high-fat diet was given for 3 weeks before tissue harvested

value: liver

value: FoxO1-/-; PPAR{alpha}-/-

value: High-fat diet


title: Control_Female_input_R1

description: Library name: Sample_9


title: HFD_LFPKO_Fast_rep1

description:
Library name: DA022
DA022


title: RHP5813_CMV NLPV CD27+


title: HFD_LFPKO_Fast_rep2

description:
Library name: DA023
DA023


title: HET_Male_H3K79me2_R4

description: Library name: Sample_8


title: RHP5814_CMV NLPV CD27-


title: HET_Male_H3K79me2_R3

description: Library name: Sample_6


title: HFD_LPKO_Fast_rep3

description:
Library name: DA021
DA021

value: PPAR{alpha}-/-


title: RHP5812_CMV NLPV Mix


title: HFD_LPKO_Fast_rep2

description:
Library name: DA020
DA020


title: Control_Male)H3K79me2_R4

description: Library name: Sample_7


title: Control_Male_H3K79me2_R3

description: Library name: Sample_5


title: RHP5811_CMV NLPV CD27-


title: HFD_LPKO_Fast_rep1

description:
Library name: DA019
DA019


title: RHP5809_CMV NLPV Mix


title: Control_Female_H3K79me2_R2

description: Library name: Sample_3


title: RHP5810_CMV NLPV CD27+


title: HET_Female_H3K79me2_R2

description: Library name: Sample_4


title: HFD_LFKO_Fast_rep2

description:
Library name: DA017
DA017


title: RHP5808_EBV EBNA3B


title: RHP5817_CMV NLPV CD27+_1


title: RHP5806_EBV EBNA4 CD27-


title: HFD_WT_Fast_rep2

description:
Library name: DA014
DA014

value: FoxO1lox/lox; PPAR{alpha}lox/lox


title: RHP5805_EBV EBNA4 CD27+


title: HFD_WT_Fast_rep3

description:
Library name: DA015
DA015


title: HFD_LFKO_Fast_rep1

description:
Library name: DA016
DA016


title: Dot1l HET

data_processing:
Paired-end sequencing reads from the Illumina Nextseq 1000 were pre-processed using Illumina BCL2Fastq and 10X Genomics Cellranger v5.0.1
Each cDNA read with nuclear barcode and unique molecular identifier was  aligned to the mm10 genome using using STAR v2.7.1 with MAPQ adjustment
SoupX was used to regress out predicted free droplet RNAdimensionality reduction and clustering was performed prior to processing using SoupX
Raw expression matrices were filtered for RNA count, detected genes, mitochondrial read percent, and ribosomal read percent before dimensionality reduction and clustering using Seurat v5.0
DE analysis was done using the Seurat R package
Assembly: mm10 optimized for single cell sequencing through hybrid intronic read recovery approach
Supplementary files format and content: _barcodes.tsv.gz tsv files containing nuclei barcodes for Cellranger either raw or filtered with Cellranger default parameters (downstream analyses utilized raw)
Supplementary files format and content: _features.tsv.gz tsv files containing gene features for Cellranger either raw or filtered with Cellranger default parameters (downstream analyses utilized raw)
Supplementary files format and content: _matrix.mtx.gz matrix file containing gene count information for Cellranger either raw or filtered with Cellranger default parameters (downstream analyses utilized raw)
Supplementary files format and content: Dot1l_scNuc.rds .rds file containing seurat object used for the analysis

extract_protocol:
Cortical tissue was homogenized with a douncer in homogenization buffer supplemented with 1 mM DTT, 0.15 mM spermine, 0.5 mM spermidine, RNasin ® Plus Ribonuclease Inhibitor (Promega N2611), and EDTA-free protease inhibitor (Roche). Nuclei were isolated using an iodixanol density gradient and ultradentrifugation.
Chromium Next GEM Single Cell Multiome ATAC + Gene Expression


title: HET_Female_H3K79me2_R1

description: Library name: Sample_2


title: RHP5807_EBV EBNA4 CD27+ high+


title: HFD_LFKO_Fast_rep3

description:
Library name: DA018
DA018


title: HFD_WT_Fast_rep1

description:
Library name: DA013
DA013


title: RHP5804_EBV EBNA4 Mix


title: Control


title: RD_LPKO_Fast_rep3

description:
Library name: DA009
DA009

value: Regular diet


title: RD_LFPKO_Fast_rep2

description:
Library name: DA011
DA011


title: Control_Female_H3K79me2_R1

description: Library name: Sample_1


title: RHP5802_EBV EBNA3B CD27+ high+


title: RD_LFPKO_Fast_rep3

description:
Library name: DA012
DA012


title: RHP5803_EBV EBNA4


title: RHP5801_EBV EBNA3B CD27-


title: RHP5800_EBV EBNA3B CD27+


title: RD_LPKO_Fast_rep2

description:
Library name: DA008
DA008


title: RHP5799_EBV EBNA3B


title: wild type 3

city: Shanghai

first: Aili

last: Cao

state: Shanghai

address: 600 Yishan Road

country: China

institute: Shanghai Sixth People's Hospital Affiliated to Shanghai Jiao Tong University School of Medicine

data_processing:
With the help of HISAT2 (version 2.2.1), the sequenced reads were mapped to the human genome. We used HTSeq-count (version 0.9.1) to create and quantify the raw counts. The DESeq2 (version 1.20.0) program was used to conduct the differential gene expression study. We used a combination of specific thresholds to identify the genes that showed significant differences in expression levels. By imposing these limits, we could be sure that we were only looking at differentially expressed genes (DEGs) when the fold change was 1.5 or more and the q-value was less than 0.05.
Assembly: hg38
Supplementary files format and content: tab-delimited text file includes raw counts for each sample

source_name: primary urinary epithelial cells

growth_protocol: rine-derived epithelial cells from the patient and four healthy controls were cultured. Briefly, urine samples were centrifuged at 400 g for 10 minutes at room temperature. The cell pellets were washed with PBS containing penicillin, streptomycin, and amphotericin B, then spun again at 200 g for 10 minutes. The pellets were resuspended in 2 ml of DMEM-F12 (Sigma-Aldrich), supplemented with 10% fetal bovine serum, antibiotics, and renal epithelial cell growth supplements (Lonza). This suspension was plated on a gelatin-coated 12-well plate and incubated at 37°C with 5% CO2. Colonies appeared in 4-5 days and were switched to a proliferation medium (a 1:1 mix of renal epithelial and mesenchymal cell media). Once cells reached 70-80% confluence, they were passaged using TrypsinLE (Gibco).

extract_protocol:
Total RNA was extracted using TRIzol reagent (Invitrogen, CA, USA) following the manufacturer’s protocol. RNA quality and quantity were measured with a NanoDrop 2000 spectrophotometer (Thermo Scientific, USA),
Using the Agilent 2100 Bioanalyzer, a cDNA library was constructed and its quality was evaluated.
Libraries were prepared with the VAHTS Universal V6 RNA-seq Library Prep Kit and sequenced on an Illumina Novaseq 6000 platform (150 bp paired-end reads) by OE Biotech Co., Ltd. (Shanghai, China).

value: primary urinary epithelial cells

value: wild type


title: Wild type 2


title: RD_LPKO_Fast_rep1

description:
Library name: DA007
DA007


title: RD_LFKO_Fast_rep3

description:
Library name: DA006
DA006


title: RHP5798_EBV #41


title: Wild type 1


title: RHP5797_CMV #73


title: RD_LFKO_Fast_rep2

description:
Library name: DA005
DA005


title: RHP5796_EBV EBNA4


title: RHP5795_Flu


title: RD_LFKO_Fast_rep1

description:
Library name: DA004
DA004


title: RD_WT_Fast_rep3

description:
Library name: DA003
DA003


title: Mutant 2

value: mutant


title: RD_LFPKO_Fast_rep1

description:
Library name: DA010
DA010


title: RHP5794_CMV NLP


title: Mutant 1


title: RHP5793_CMV NLP


title: Human Treg rest, Donor 2, SATB1 ChIP, Formaldehyde fixation

description: Library name: D2_Treg_rest_SATB1_ChIP_form

data_processing:
Reads were mapped to the hg38 human genome using Bowtie2 with default parameters. Duplicate reads were removed using SAMtools. Matching input control was used to call peaks. Peak calling was performed using MACS2 (Tregs) or MACS3 (all other samples) with significance set between e-2 to e-8. Bigwig tracks were generated using deepTools bamCoverage with RPKM normalization. Blacklisted regions (downloaded from ENCODE) were excluded from called peaks using bedtools.
Assembly: GRCh38/hg38
Supplementary files format and content: Bigwig and MACS2/MACS3 peak bed files.

growth_protocol: Primary human T cells subsets were isolated from peripheral blood mononuclear cells (PBMCs) sourced from consented, fresh Human Peripheral Blood Leukopaks (STEMCELL Technologies, catalog no. 70500). Leukopaks were washed twice with 1X volume of EasySep Buffer (DPBS, 2% fetal bovine serum [FBS], 1 mM pH 8.0 EDTA) using centrifugation and resuspended at 50e6-200e6 cells/mL. CD4+CD25highCD127low Tregs and CD4+CD25low Tconv were isolated from washed PBMCs using the EasySep Human CD4+CD127lowCD25+ Regulatory T Cell Isolation Kit (STEMCELL Technologies, catalog no. 18063) according to manufacturer’s protocol. For enhanced cell purity, Tregs were stained for CD4 (Biolegend, catalog no. 344634 or 344620), CD25 (Tonbo, catalog no. 20-0259-T100), and CD127 (Becton Dickinson, catalog no. 557938) and further sorted into a CD4+CD25highCD127low population via FACS on a BD FACSAria or BD FACSAria Fusion I. Tregs and Tconvs were cultured in X-VIVO15 media (Lonza, catalog no. 02-053Q) supplemented with 5% FBS, 55 µM 2-Mercaptoethanol, and 4 mM N-Acetyl-L-Cysteine. For CRISPRi screening, CRISPRi validation, and CRISPRn screening, recombinant Human IL-2 (R&D Systems, catalog no. 202-GMP or BT-002-GMP) was supplemented at 200 IU/mL, and cells were stimulated for 48 hours with CTS Dynabeads CD3/CD28 beads (Thermo Fisher Scientific, catalog no. 40203D) at a cell to bead ratio of 1:1. After 48 hours, beads were removed using magnetic separation. For all other experiments, cells were cultured using recombinant Human IL-2 (R&D Systems, catalog no. 202-GMP or BT-002-GMP) supplemented at 300 IU/mL and stimulated with 12.5 uL/mL Immunocult Human CD3/CD28/CD2 T Cell Activator (STEMCELL Technologies, catalog no. 10970), to enhance cell recovery, growth, and viability. Cells were cultured at 37C and 5% CO2 and split to 5E5-1.5E6 cells/mL every ~48 hours by topping off media and completely replacing the appropriate dose of IL-2.

extract_protocol:
Up to 2.5E7 cells were cross-linked in PBS with 1% methanol-free formaldehyde (Thermo, catalog no. 28908) for 10 min at room temperature followed by quenching in glycine at 125 mM final concentration. Tconv and selected Treg samples were cross-linked in 2 mM disuccinimidyl glutarate (Thermo, catalog no. 20593) in PBS for 30 min at room temperature prior to formaldehyde treatment. Cross-linked cell pellets were snap-frozen in liquid nitrogen and stored at –80ºC. Nuclei were isolated from thawed, cross-linked cells via sequential lysis in LB1 (50 mM HEPES-KOH pH 7.5, 140 mM NaCl, 1 mM EDTA, 10% glycerol, 0.5% IGEPAL CA-360, and 0.25% Triton X-100), LB2 (10 mM Tris-HCl pH 8, 200 mM NaCl, 1 mM EDTA, and 0.5 mM EGTA), and LB3 (10 mM Tris-HCl pH 8, 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 1% sodium deoxycholate [NaDOC], and 0.5% N-laurylsarcosine) supplemented with 0.5 mM phenylmethylsulfonyl fluoride (PMSF, Sigma, catalog no. P7626) and 0.5X protease inhibitor cocktail (PIC, Sigma, catalog no. P8340). Chromatin was sheared on a Covaris E220 focused ultrasonicator using 1 mL milliTubes (Covaris, catalog no. 520128) with 140W peak incident power, 5% duty factor, 200 cycles per burst, 6ºC temperature setpoint (minimum 3ºC, maximum 9ºC), fill level 10, and time 12-14 min to obtain target size 200-700 bp. Triton X-100 was added to final concentration of 1% prior to immunoprecipitation for 16 h at 4ºC with 2-8 µg of indicated antibodies bound to a 1:1 mixture of protein A and protein G magnetic beads (Thermo, catalog nos. 10001D and 10003D). Bead-bound antibody-chromatin complexes were sequentially washed three times with Wash Buffer 1 (20 mM Tris pH 8, 150 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 1% Triton X-100, 0.1% sodium dodecyl sulfate [SDS], and 0.1% NaDOC), twice with Wash Buffer 2 (20 mM Tris-HCl pH 8, 500 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 1% Triton X-100, 0.1% SDS, and 0.1% NaDOC), twice with Wash Buffer 3 (20 mM Tris-HCl pH 8, 250 mM LiCl, 1 mM EDTA, 0.5% IGEPAL CA-360, and 0.5% NaDOC), twice with TET (10 mM Tris-HCl pH 8, 1 mM EDTA, 0.2% Tween-20), and once with TE0.1 (10 mM Tris-HCl pH 8, 0.1 mM EDTA, 0.5 mM PMSF, and 0.5X PIC) supplemented with 0.5 mM PMSF and 0.5X PIC. Beads were resuspended in TT (10 mM Tris-HCl pH 8, 0.05% Tween-20) prior to on-bead library preparation using the NEBNext Ultra II DNA Library Prep Kit (NEB, catalog no. E7370L) as described previously (Texari et al., 2021). ChIP-seq libraries were multiplexed for paired-end sequencing on an Illumina NextSeq 2000 or NovaSeq X instrument.
On-bead library preparation was conducted using the NEBNext Ultra II DNA Library Prep Kit (NEB, catalog no. E7370L) as described previously (Texari et al., 2021). ChIP-seq libraries were multiplexed for paired-end sequencing.

treatment_protocol: Nine days after isolation and initial stimulation, a portion of cells were stimulated with 12.5 µL/mL Immunocult Human CD3/CD28/CD2 T Cell Activator (STEMCELL Technologies, catalog no. 10970). Eleven days post-initial stimulation, rested (Rest) and restimulated (Stim) cells were fixed for ChIP-seq.

value: Blood

value: Treg

tag: condition

value: rest

value: Donor 2

tag: chip target

tag: fixation method

value: Formaldehyde

tag: chip antibody

value: SATB1 (Abcam, Ab109122)


title: RD_WT_Fast_rep1

description:
Library name: DA001
DA001


title: Human Treg rest, Donor 2, Input ChIP, Formaldehyde fixation

description: Library name: D2_Treg_rest_Input_ChIP_form

value: Input


title: TTseq_dTAG_rep3

city: Copenhagen

first: Zhong

last: Han

address: Blegdamsvej 3B

department: ICMM, Institut for Cellulær og Molekylær Medicin

country: Denmark

institute: University of Copenhagen

data_processing:
pair-end reads were aligned with STAR, Version 2.7.11a
bigwig files were produced with bamcoverage from deeptools, Version 3.5.4
mean bigwig files were produced with bigwigaverage from deeptools, Version 3.5.4
Assembly: GRCh38
Supplementary files format and content: bigwig
Supplementary files format and content: annotation .bed files used in this study
Library strategy: TTseq

source_name: Flp-In T-REx HEK293

growth_protocol: Flp-In T-REx HEK293 cells (R78007, Thermo Fisher Scientific, human embryonic kidney epithelial, female origin) were cultured in DMEM, hihg glucose, GlutaMAX (10566016, Thermo Fisher Scientific) supplemented with 10% v/v FBS, 100 U/mL penicillin, 100 μg/mL streptomycin, at 37°C with 5% CO2 and routinely passaged 2-3 times a week.

extract_protocol:
The TRIzol-cell mixture was collected, and total RNA was extracted via TRIzol/chloroform extraction followed by isopropanol precipitation. RNA concentration was measured using the Qubit RNA BR Assay Kit (Thermo Fisher, Q10210). For each sample, 1 µg of S. cerevisiae 4TU-labeled RNA was mixed with 100 µg of purified 4SU-labeled RNA. The total volume was adjusted to 100 µL with RNase-free water, and 20 µL of 1 M NaOH was added per sample. Samples were incubated on ice for 20 minutes to fragment the RNA, and RNA fragmentation was stopped by adding 80 µL of 1 M Tris-HCl, pH 6.0. The samples were purified using Micro Bio-Spin P-30 gel columns (Bio-Rad, 7326250) to neutralize NaOH and prevent further fragmentation. The column-purified RNA was biotinylated in a 250 µL reaction containing 10 mM Tris-HCl, pH 7.4, 1 mM EDTA, and 5 mg of MTSEA biotin-XX linker (Biotium, BT90066) for 30 minutes at room temperature in the dark. Biotinylated RNA was purified by phenol-chloroform extraction and isopropanol precipitation. The RNA was then denatured at 65°C for 10 minutes and purified using the μMACS Streptavidin Kit (Miltenyi, 130-074-101) according to the manufacturer’s instructions. The purified nascent RNA was further cleaned using the RNeasy MinElute kit (QIAGEN, 74204) and used to prepare sequencing libraries.
TTchem-seq libraries were generated using the KAPA RNA HyperPrep Kit (Roche, 08098093702) with the fragmentation step omitted. A total of 75 ng of nascent RNA was denatured at 65°C for 5 minutes. The rest of the library preparation followed the manufacturer’s instructions, with two SPRI bead purifications (Beckman Coulter, A63881) performed using bead-to-sample ratios of 0.95x and 1x.

treatment_protocol: Briefly, 3 million cZH9.1: Hek293 Flp-In T-REx SETX-dTAG cells were seeded in 10 cm dishes. The following day, treat the cells with DMSO or 1 µM dTAG for 4 hours. After treatment, initiate 4SU labeling by adding 4SU directly to the culture medium to a final concentration of 1 mM. Cells were incubated with 4SU for 15 minutes, after which the medium was removed, and 1 mL of TRIzol was added per dish to terminate labeling.

tag: cell line

value: Flp-In T-REx HEK293

value: SETX-dTAG


title: RD_WT_Fast_rep2

description:
Library name: DA002
DA002

value: liver


title: TTseq_dTAG_rep2


title: RHP5792_CMV NLP

value: peripheral blood mononuclear cells


title: TTseq_dTAG_rep1


title: RHP5791_CMV NLP


title: Mutant 3

tag: cell type


title: TTseq_DMSO_rep3

tag: genotype


title: Human Treg stim, Donor 3, SRF ChIP, DSG/formaldehyde fixation

description: Library name: D3_Treg_Stim_SRF_ChIP_dsg

value: stim

value: Donor 3

value: SRF

value: DSG/formaldehyde

value: SRF (ActiveMotif, 61385)


title: Human Treg stim, Donor 3, SATB1 ChIP, DSG/formaldehyde fixation

description: Library name: D3_Treg_Stim_SATB1_ChIP_dsg


title: PC35

city: London

first: LMS

last: Bioinformatics Core

address: Hammersmith Hospital Campus, Du Cane Road

department: Bioinformatics Core

institute: MRC London Institute of Medical Sciences

data_processing:
Reads were processed using cutadapt v4.7 (Martin 2011) to remove Illumina adapters, trim, and filter on quality with the settings: -a AGATCGGAAGAGCACACGTCTGAACTCCAGTCA, -A AGATCGGAAGAGCGTCGTGTAGGGAAAGAGTGT, -n 1, -m 31, --nextseq-trim 20, --max-n 0
Processed reads were aligned with HISAT2 v2.2.1 (Kim et al. 2019) to GRCm39 with settings adjusted to report up to 100 multiple alignments for each read pair: --rna-strandness RF --no-discordant --no-mixed --no-unal -k 100 --score-min L,0,-0.66 --pen-noncansplice 20 --dta
Gene and TE locus-level counts were obtained with TElocal (TEtranscripts (Jin et al. 2015)) using settings to allow distribution of multimapping reads using maximum likelihood estimation: --stranded reverse --mode multi
GRCm39.110 (Ensembl) annotations were used for genes. TE annotations were produced using RepeatMasker v4.1.5 (Smit, Hubley, and Green 2015) configured with HMMER v3.3.2 (Wheeler and Eddy 2013) to scan the GRCm39 genome using Dfam3.7 (Storer et al. 2021) HMM TE models in sensitive mode: -s -no_is -norna -spec "mus musculus"
Within R v4.3.3, gene count data was used assess inter-sample relationships using hierarchal clustering and PCA. Additionally, the impact of technical experimental factors was assessed using variance partitioning (variancePartition v1.30.2 (Hoffman and Schadt 2016)). Following these assessments, the RUVs method (RUVseq, (Risso et al. 2014)) was used to remove the replicate batch effect by subsequently integrating the returned correction factors into the statistical model.
Within DESeq2 v1.40.2 (Love, Huber, and Anders 2014), independent hypothesis weighting was conducted to optimise the power of p value filtering using IHW v1.28.0 (Ignatiadis et al. 2016) and log2 fold-change shrinkage was performed using ashr v2.2-63 (Stephens 2017). Multiple correction testing adjustments were optimised by passing FDR and log2FC thresholds at the point of results generation.
Assembly: GRCm39
Supplementary files format and content: Gene- and TE-level expression estimates

growth_protocol: Mouse E14 ESCs were grown in serum/LIF conditions on 0.2% gelatin.

extract_protocol:
Samples sorted FACS by 2C-GFP fluorescence and 2C-GFP positive and negative samples collected separately.
RNA was extracted, purified and DNAse-I treated using the RNEasy-mini kit, according to the manufacturers protocol (Qiagen).
Purified RNA was used for RNA-seq library preparation by polyA enrichment using a strand-specific method by Novogene.

treatment_protocol: Cells were transfected with control sgRNAs (sgGAL4), MERVL sgRNAs or Dux sgRNAs.

value: Rosa^dcas9-VPR/+, 2C-GFP/CD4

tag: treatment

tag: gfp

value: neg

tag: replicate


title: RHP5789_EBV EBNA4

tag: tissue


title: PC34

value: pos


title: PC36


title: PC33


title: Human Treg stim, Donor 3, Input ChIP, DSG/formaldehyde fixation

description: Library name: D3_Treg_Stim_Input_ChIP_dsg

value: Blood

value: Treg

tag: donor


title: TTseq_DMSO_rep1


title: Human Treg rest, Donor 3, SATB1 ChIP, DSG/formaldehyde fixation

description: Library name: D3_Treg_rest_SATB1_ChIP_dsg

value: rest


title: U87MG cells, DoubleKO-3

first: Chunlong

last: Zhong

address: 150 Jimo Road

department: Department of Neurosurgery

institute: Shanghai East Hospital, School of Medicine, Tongji University

data_processing:
fastp 0.20.1
RseQC 4.0.0
fastqc v.0.11.9
hisat2 2.1.0
Assembly: mm10
Supplementary files format and content: tab-delimited text files include raw counts for each sample

growth_protocol: U87MG glioblastoma cells were maintained in DMEM supplemented with 10% FBS and antibotics in humidified atmosphere with 5% CO2 at 37 ℃

extract_protocol:
Total RNA was extracted using the TRIzol reagent (Invitrogen, CA, USA). 1.3μg of total RNA was used fir the construction of sequencing libraries.
The libraries were constructed using VAHTS Universal V6 RNA-seq Library Prep Kit according to the manufacturer’s instructions.

value: glioblastoma cells

value: ATRX and TOP4B knockdown


title: Human Treg rest, Donor 3, Input ChIP, DSG/formaldehyde fixation

description: Library name: D3_Treg_rest_Input_ChIP_dsg

value: Input


title: RHP5790_EBV #41


title: RHP5786_CMV #13


title: RHP5788_EBV EBNA3B


title: Human Treg rest, Donor 2, FOXP3 ChIP, Formaldehyde fixation

description: Library name: D2_Treg_rest_FOXP3_form

value: Donor 2

value: Formaldehyde

value: FOXP3 (Novus, NB600-242)


title: U87MG cells, DoubleKO-2

value: ATRX and TOP3B knockdown


title: TTseq_DMSO_rep2

value: Flp-In T-REx HEK293

value: SETX-dTAG


title: PC30


title: RHP5787_EBV EBNA3B

value: peripheral blood mononuclear cells


title: Human Treg stim, Donor 3, FOXP3 ChIP, DSG/formaldehyde fixation

description: Library name: D3_Treg_Stim_FOXP3_ChIP_dsg

value: stim

value: Donor 3

value: DSG/formaldehyde


title: RHP5785_CMV NLP


title: Human Treg rest, Donor 3, FOXP3 ChIP, DSG/formaldehyde fixation

description: Library name: D3_Treg_rest_FOXP3_ChIP_dsg


title: U87MG cells, DoubleKO-1

value: ATRX and TOP2B knockdown


title: Hap1_DRBR30m_dTAG_rep3

description: Library name: sample18

data_processing:
single-end reads were aligned with STAR, Version 2.7.9
bigwig files were produced with bamcoverage from deeptools, Version 3.5.5
mean bigwig files were produced with bigwigaverage from deeptools, Version 3.5.5
Assembly: GRCh38
Supplementary files format and content: bw (mean of replciates)

growth_protocol: Hap1 cells (Horizon discovery ) were cultured in IMDM, GlutaMAX (31980030, Thermo Fisher Scientific) supplemented with 10% v/v FBS, 100 U/mL penicillin, 100 μg/mL streptomycin, at 37°C with 5% CO2 and routinely passaged 2-3 times a week.

extract_protocol:
Cells were incubated with 4SU for 15 minutes, after which the medium was removed, and 1 mL of TRIzol was added per dish to terminate labeling. The TRIzol-cell mixture was collected, and total RNA was extracted via TRIzol/chloroform extraction followed by isopropanol precipitation. RNA concentration was measured using the Qubit RNA BR Assay Kit (Thermo Fisher, Q10210). For each sample, 1 µg of S. cerevisiae 4TU-labeled RNA was mixed with 100 µg of purified 4SU-labeled RNA. The total volume was adjusted to 100 µL with RNase-free water, and 20 µL of 1 M NaOH was added per sample. Samples were incubated on ice for 20 minutes to fragment the RNA, and RNA fragmentation was stopped by adding 80 µL of 1 M Tris-HCl, pH 6.0. The samples were purified using Micro Bio-Spin P-30 gel columns (Bio-Rad, 7326250) to neutralize NaOH and prevent further fragmentation. The column-purified RNA was biotinylated in a 250 µL reaction containing 10 mM Tris-HCl, pH 7.4, 1 mM EDTA, and 5 mg of MTSEA biotin-XX linker (Biotium, BT90066) for 30 minutes at room temperature in the dark. Biotinylated RNA was purified by phenol-chloroform extraction and isopropanol precipitation. The RNA was then denatured at 65°C for 10 minutes and purified using the μMACS Streptavidin Kit (Miltenyi, 130-074-101) according to the manufacturer’s instructions. The purified nascent RNA was further cleaned using the RNeasy MinElute kit (QIAGEN, 74204) and used to prepare sequencing libraries.
Hap1_TTchem-seq libraries were produced by NEBNext Ultra II Directional RNA Library Prep Kit (NEB, E7760L) with NEBNext Multiplex Oligos (Unique Dual Index UMI Adaptors RNA Set 1, NEB E7416S) and sequenced with single-end 50 nt reads on the Illumina NextSeq 2000, generating approximately 20 million reads per sample at the Genomic Platforms, University of Copenhagen.

treatment_protocol: Cells were treated with DMSO or 1 µM dTAGv1 for one hour, followed by a 3-hour pre-treatment with 100 µM DRB (5,6-dichloro-1-β-D-ribofuranosylbenzimidazole). After this, cells were released from DRB inhibition for 15, 20, or 30 minutes, which included a 15-minute pulse with 1 mM 4-thiouridine, while still under DMSO or dTAG.


title: C5HetVav.3

first: Probir

last: Chakravarty

address: 1 Midland Road

institute: Francis Crick Institute

description: Library name: C5HetVav.3

data_processing:
Raw FASTQ reads were initially processed by the Cell Ranger version 9.0.0 pipeline, aligning to the mm10 (GRCm38, version 93) transcriptome
All subsequent analyses were performed in R (version 4.4.1)  using the Seurat (version 5.2.1) package
Assembly: mm10 [GRCm38]
Supplementary files format and content: Tab-separated barcodes ID's, features files and count files
Supplementary files format and content: Tab-separated meta.data from Seurat object with annotation, sample and UMAP embeddings per cell.

source_name: small intestine lamina propria

extract_protocol:
Small intestines  were cut into small pieces. Digested and isolated following the instructions provided in Miltenyi Biotec Lamina propria dissociation kit protocol, 130-097-410. Cells were resuspended in FACS buffer after centrifugation. Single cell suspensions were stained with CD45 antibody FACS sorted on an Aria Fusion (BD) with a 100um nozzle.
scRNA-seq

value: small intestine lamina propria

tag: gender

value: male

value: WT


title: Hap1_DRBR30m_dTAG_rep2

description: Library name: sample17


title: PC28

value: Rosa^dcas9-VPR/+, 2C-GFP/CD4

value: pos


title: C5HetVav.4

description: Library name: C5HetVav.4


title: U87MG cells, siTOP2B-3

value: glioblastoma cells

value: TOP4B Knockdown


title: PC27

value: neg


title: PC31


title: C5HetVav.2

description: Library name: C5HetVav.2

value: female


title: U87MG cells, siTOP2B-2

value: TOP3B Knockdown


title: Hap1_DRBR30m_dTAG_rep1

description: Library name: sample16


title: PC26


title: U87MG cells, siATRX-3

value: ATRX knockdown


title: Hap1_DRBR30m_DMSO_rep2

description: Library name: sample14

value: SETX-dTAG


title: C5deltaVav.4

description: Library name: C5deltaVav.4

value: KO


title: Hap1_DRBR30m_DMSO_rep3

description: Library name: sample15


title: PC25


title: PC24


title: U87MG cells, siATRX-2


title: U87MG cells, siATRX-1


title: Hap1_DRBR30m_DMSO_rep1

description: Library name: sample13


title: C5deltaVav.3

description: Library name: C5deltaVav.3


title: PC29


title: PC23


title: U87MG cells, Ctrl-3


title: PC22


title: U87MG cells, siTOP2B-1

value: TOP2B Knockdown


title: U87MG cells, Ctrl-2


title: PC32


title: Hap1_DRBR20m_dTAG_rep2

description: Library name: sample11


title: C5HetVav.1

description: Library name: C5HetVav.1


title: Hap1_DRBR20m_dTAG_rep3

description: Library name: sample12


title: C5deltaVav.1

description: Library name: C5deltaVav.1


title: PC21


title: Hap1_DRBR20m_dTAG_rep1

description: Library name: sample10


title: PC20


title: III_24h_2

description: AK Yolksac

organism:
Candidozyma auris
Aphanius dispar

source_name: Whole organism + yeast cells

value: Whole organism + yeast cells

value: Wild type

value: III

value: A. dispar infected with C. auris

value: 24 hours


title: II_24h_2

description: AK Yolksac

value: II


title: U87MG cells, Ctrl-1

value: WT


title: scStereo_age42

first: Yuxuan

last: Zheng

address: 825 Zhangheng Rd.

institute: Fudan University

data_processing:
The raw single-cell BCR sequencing data were processed with subcommand vdj in 10× Genomics software CellRanger (version: 6.1.2), in which process the sequencing data were aligned into the mouse reference VDJ database (GRCm38).
Assembly: Homo sapiens (GRCh38)
Supplementary files format and content: "barcodes.tsv" files include snRNA-seq barcode information, "features.tsv" files include snRNA-seq gene features and "matrix.mtx" files include snRNA-seq gene expression. "tissue.gef" files include scStereo-seq data.

molecule: cytoplasmic RNA

source_name: ovary

extract_protocol:
BD Vacutainer K2 EDTA tube (10ml) were used for blood collection. We obtained peripheral venous blood samples by venipunction from six participants. And then used Ficoll density gradient centrifugation (TBD, China) to separate PBMCs. We pre-added 4 mL Ficoll to each 15 mL centrifuge tube. For each participant, equal volumes of blood were carefully layered onto Ficoll and centrifuged at 800 g and RT for 25 min (3 acceleration and 1 brake). After centrifugation, we collected PBMCs with pipette into a 15-ml tube, and diluted up to 15 ml with pre-warmed PBS (TBD, China). After centrifuging for 10 min, at 250 g and RT, PBMC were resuspended in 2 mL Red Blood Cell Lysis Buffer (Solarbio, China). After incubating the cells on ice for 10 min, 8 mL PBS was added, gently mixed, and centrifuged at 4℃ 300 g for 10 min. We resuspended the centrifuged cells in 10ml of pre-prepared washing buffer (1× PBS with 0.04% BSA). The cleaning step was performed at 250 g and 4℃ for 10 min. The second time the cells were resuspended by the cleaning buffer and counted with the Dual Fluorescence Cell Counter (LunaTM fl). After centrifugation, the cells were resuspended in the cell cryopreservation system recommended by 10× Genomics. With about 2×106 cells per milliliter, one-milliliter aliquots were gradually frozen in a freezing box over the course of 24 h at -80℃ before being kept in a vapor-phase liquid nitrogen tank.
The cells were first thawed according to the 10× Genomics protocol (Demonstrated Protocol, CG00039). The thawed cells were then stained with trypan blue and placed under a light microscope to observe the cell state and background. The cells were counted using the Dual Fluorescence Cell Counter (LunaTM fl), and subsequent procedures were performed after confirming that the survival rate was not less than 90%. The ChromiumTM Single cell 5’ Reagent Kit, Chromium Single Cell Human BCR Amplification Kit, and Chromium Single Cell Human TCR Amplification Kit of the Chromium platform (10× Genomics, California, USA) were used to create the scRNA-seq and paired single-cell BCR and TCR libraries in accordance with the Chromium Single Cell 5’ Reagent Kits User Guide (v2 Chemistry Dual Index). On the Illumina HiSeq X Ten platform, generated scRNA-seq libraries were sequenced. The preprocessing of the PE150 Illumina sequencing reads was done using the 10× Genomics software CellRanger (version: 6.1.2).

value: ovary

tag: library

value: scStereo-seq

tag: age


title: Hap1_DRBR20m_DMSO_rep3

description: Library name: sample9


title: I_24h_2

description: AK Yolksac

value: I

tag: time point


title: scStereo_age34


title: PC19

value: Rosa^dcas9-VPR/+, 2C-GFP/CD4

value: neg


title: C5deltaVav.2

description: Library name: C5deltaVav.2

value: small intestine lamina propria

value: female


title: Hap1_DRBR20m_DMSO_rep2

description: Library name: sample8

value: SETX-dTAG


title: scStereo_age32


title: I_24h_1

description: AK Yolksac


title: Hap1_DRBR20m_DMSO_rep1

description: Library name: sample7


title: scStereo_age12


title: PC17


title: V_24h_1

description: AK Yolksac

value: V


title: Hap1_DRBR15m_dTAG_rep3

description: Library name: sample6


title: PC16

value: pos


title: snRNA_age42

value: snRNA-seq


title: Hap1_DRBR15m_dTAG_rep2

description: Library name: sample5


title: V_48h_3

description: AK Yolksac

value: Whole organism + yeast cells

value: Wild type

value: 48 hours


title: IV_24h_1

description: AK Yolksac

value: IV

value: A. dispar infected with C. auris

value: 24 hours


title: snRNA_age34

value: ovary


title: PC15


title: PC14


title: snRNA_age32


title: IV_48h_3

description: AK Yolksac


title: PC13


title: III_48h_3

description: AK Yolksac

value: III


title: 3q_Gain_cell_subclone_3

city: West Haven

first: Wenxue

last: Li

state: CT     Connecticut

address: 300 Heffernernan Dr Bldg A42,, Bldg 840, Room ABC 367

institute: Yale University

data_processing:
Reads are trimmed to remove low quality basse-calls upto 45 reads
HISAT2 was used for alignment of trimmed reads.
StringTie was used to generate gene counts
Assembly: hg38
Supplementary files format and content: Tab-delimited text files include FPKM values for each gene and each sample

source_name: lung epithelial cells

extract_protocol:
Cells were washed with PBS three times, snap-frozen, and then lysed with the QIAShredder columns (#79654, QIAGEN) according to the manufacturer’s instructions. Total RNA was isolated with the Qiagen RNeasy Mini Kit (#74104, QIAGEN), including the DNA digestion step using the RNAase-Free DNAase set kit (#79254, QIAGEN).
Library preparation was performed using the KAPA Biosystems mRNA HyperPrep Kit

value: lung epithelial cells


title: PC12

value: Rosa^dcas9-VPR/+, 2C-GFP/CD4


title: II_48h_3

description: AK Yolksac

value: II


title: PC18


title: I_48h_3

description: AK Yolksac

value: I


title: 3q_Gain_cell_subclone_1_


title: PC11

value: neg


title: TTseq_dTAG_rep3

description: Library name: sample6

data_processing:
single-end reads were aligned with STAR, Version 2.7.9
bigwig files were produced with bamcoverage from deeptools, Version 3.5.5
mean bigwig files were produced with bigwigaverage from deeptools, Version 3.5.5
Assembly: GRCh38
Supplementary files format and content: bw (mean of replicates)

extract_protocol:
The TRIzol-cell mixture was collected, and total RNA was extracted via TRIzol/chloroform extraction followed by isopropanol precipitation. RNA concentration was measured using the Qubit RNA BR Assay Kit (Thermo Fisher, Q10210). For each sample, 1 µg of S. cerevisiae 4TU-labeled RNA was mixed with 100 µg of purified 4SU-labeled RNA. The total volume was adjusted to 100 µL with RNase-free water, and 20 µL of 1 M NaOH was added per sample. Samples were incubated on ice for 20 minutes to fragment the RNA, and RNA fragmentation was stopped by adding 80 µL of 1 M Tris-HCl, pH 6.0. The samples were purified using Micro Bio-Spin P-30 gel columns (Bio-Rad, 7326250) to neutralize NaOH and prevent further fragmentation. The column-purified RNA was biotinylated in a 250 µL reaction containing 10 mM Tris-HCl, pH 7.4, 1 mM EDTA, and 5 mg of MTSEA biotin-XX linker (Biotium, BT90066) for 30 minutes at room temperature in the dark. Biotinylated RNA was purified by phenol-chloroform extraction and isopropanol precipitation. The RNA was then denatured at 65°C for 10 minutes and purified using the μMACS Streptavidin Kit (Miltenyi, 130-074-101) according to the manufacturer’s instructions. The purified nascent RNA was further cleaned using the RNeasy MinElute kit (QIAGEN, 74204) and used to prepare sequencing libraries.
Hap1_TTchem-seq libraries were produced by NEBNext Ultra II Directional RNA Library Prep Kit (NEB, E7760L) with NEBNext Multiplex Oligos (Unique Dual Index UMI Adaptors RNA Set 1, NEB E7416S) and sequenced with single-end 50 nt reads on the Illumina NextSeq 2000, generating approximately 20 million reads per sample at the Genomic Platforms, University of Copenhagen.

treatment_protocol: Briefly, 2.5 million cZH36.5: Hap1 SETX-dTAG cells were seeded in 10 cm dishes. The following day, treat the cells with DMSO or 1 µM dTAG for 4 hours. After treatment, initiate 4SU labeling by adding 4SU directly to the culture medium to a final concentration of 1 mM. Cells were incubated with 4SU for 15 minutes, after which the medium was removed, and 1 mL of TRIzol was added per dish to terminate labeling.

value: SETX-dTAG


title: PC10


title: V_48h_2

description: AK Yolksac

value: V


title: 3p Loss_cell_subclone_3


title: PC9


title: Hap1_DRBR15m_dTAG_rep1

description: Library name: sample4


title: 3p Loss_cell_subclone_2


title: Hap1_DRBR15m_DMSO_rep3

description: Library name: sample3


title: TTseq_dTAG_rep1

description: Library name: sample4


title: PC8

value: pos


title: TTseq_DMSO_rep3

description: Library name: sample3


title: 3p Loss_cell_subclone_1


title: III_48h_2

description: AK Yolksac


title: PC7


title: Hap1_DRBR15m_DMSO_rep2

description: Library name: sample2


title: II_48h_2

description: AK Yolksac


title: Hap1_DRBR15m_DMSO_rep1

description: Library name: sample1


title: 3q_Gain_cell_subclone_2


title: TTseq_DMSO_rep2

description: Library name: sample2


title: TTseq_DMSO_rep1

description: Library name: sample1


title: PC5


title: V_48h_1

description: AK Yolksac


title: PC4


title: TTseq_dTAG_rep2

description: Library name: sample5


title: RChIP_DRB2h_rep2

data_processing:
single-end reads  were aligned with STAR, Version 2.7.11a
bigwig files were produced with bamcoverage from deeptools, Version 3.5.4
mean bigwig files were produced with bigwigaverage from deeptools, Version 3.5.4
Assembly: GRCh38
Supplementary files format and content: bigwig (mean value of replicates, and log2 fold change of mean bigwig files)

growth_protocol: Flp-In T-REx HEK293 cells (R78007, Thermo Fisher Scientific, human embryonic kidney epithelial, female origin) were cultured in DMEM, high glucose, GlutaMAX (10566016, Thermo Fisher Scientific) supplemented with 10% v/v FBS, 100 U/mL penicillin, 100 μg/mL streptomycin, at 37°C with 5% CO2 and routinely passaged 2-3 times a week.

extract_protocol:
Nuclei were isolated and sonicated using a Bioruptor Pico (Diagenode). RNaseH1Mut-V5 was immunoprecipitated, and the cross-linked DNA was reversed by overnight incubation at 65°C.
After treatment with RNase A and proteinase K, DNA was recovered by ethanol precipitation. Phi29 DNA polymerase was used to synthesize second-strand DNA, followed by dA tailing and adaptor ligation. The library was amplified and sequenced with single-end 50 bp reads on the Illumina NextSeq 2000, generating approximately 50 million reads per sample.

treatment_protocol: HEK293 Flp-In T-Rex SETX-dTAG: RNaseH1Mut(D210N)-V5 cells (cZH21) were seeded at a density of 4 million cells per 15 cm plate. The next day, 100 ng/mL doxycycline was added to induce the expression of RNaseH1Mut(D210N)-V5. On the third day, cells were treated with DMSO or 1 µM dTAG for 2, 4, and 8 hours. Cells were crosslinked on the plate by adding formaldehyde to a final concentration of 1% and incubating at room temperature for 10 minutes. Crosslinking was quenched by adding glycine to a final concentration of 0.125 M.

value: Flp-In T-REx HEK293

value: SETX-dTAG, flpin RNaseH1Mut(D210N)-V5


title: I_48h_2

description: AK Yolksac

data_processing:
Reads were trimmed with fastp v.0.23.1 to remove reads < 75 bases and trim based from the 3’ end with q-score < 22. Alignment and calculation of normalised gene expression were performed with the Trinity pipeline v2.15.1.
Reads were aligned to the GJEY01 AK de novo adult gill transcriptome or to the B8441 v2 C. auris reference genome.
Assembly: Aphanius dispar GJEY00000000.1, Candida auris Cand_auris_B8441_V2
Supplementary files format and content: Tab-separated text files include Fragments per Kilobase per Million (FPKM).

extract_protocol:
Total RNA was extracted using the Monarch® Total RNA Miniprep Kit (NEB, USA). Embryos were flash frozen in liquid nitrogen, re-suspended in DNA/RNA Protection Reagent and disrupted with an autoclaved plastic micropestle. Bead beating was performed in a FastPrep 24 homogenizer (MP Biomedicals) with 0.5 mm zirconia/silica beads for 8 cycles of 35 s plus 45 s on ice. Lysate was recovered and incubated for 5 min. at 55 °C with proteinase K before addition of lysis buffer and RNA purification following manufacturer’s instructions.
Samples were normalised and prepared using the NEBNext Ultra II Directional RNA Library Prep - Poly(A) mRNA Magnetic Isolation Module following the standard protocol.

value: Whole organism + yeast cells

value: Wild type


title: IV_48h_2

description: AK Yolksac

value: IV

value: 48 hours


title: PC3


title: III_24h_1

description: AK Yolksac

value: A. dispar infected with C. auris

value: 24 hours


title: RChIP_DRB2h_rep1


title: RChIP_dTAG8h_rep2


title: IV_48h_1

description: AK Yolksac


title: HK2 cell，HG+OE-FGF9-2

city: Guangzhou

first: wen

middle: qing

last: chen

address: Huangpu Avenue, Tianhe District, Guangzhou City

institute: Jinan University

data_processing:
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence,and masked for low-complexity or low-quality sequence,then mapped to mm8 whole genome using bowtie v0.12.2 with parameters -q -p 4 -e 100 -y -a -m 10 --best --strata.
TPM were calculated using a protocol. In short, exons from all isoforms of a gene were merged to create one meta-transcript. The number of reads falling in the exons of this meta-transcript were counted and normalized by the size of the meta-transcript and by the size of the library.
Assembly: mm8
Supplementary files format and content: tab-dalimited text files include TPM values for each Sample.

source_name: kidney

extract_protocol:
Total RNA was extracted from the samples, and the concentration and purity of the extracted RNA were detected by Nanodrop2000, RNA integrity was detected by agarose gel electrophoresis, and RIN value was determined by Agilent2100. The total amount of RNA required for a single library construction is 1ug, the concentration is ≥50ng/μL, and the OD260/280 is between 1.8 and 2.2.
RNA library were prepared for sequencing using standard illumina protocols

treatment_protocol: HK2 cells (Human renal tubular epithelial cells ) were procured from Chinese Academy of Sciences Cell Bank (Shanghai，China). These HK2 cells were cultured in DMEM/F-12(Dulbecco's Modified Eagle Medium/Nutrient Mixture F-12)  from Gibco (Gaithersburg, MD, USA), supplemented with 5.5mM glucose, 10% fetal bovine serum obtained from ExCell Bio, 100 U/ml of penicillin, and 100 μg/ml of streptomycin. The cells were maintained in a humidified incubator at 37°C with 5% CO2 and cultured in six-well plates at a concentration of 1×106 cells/ml. They were subjected to 50mM glucose (designated as the High Glucose group, HG) treatment 26. For gene transfection, the HK2 cells were transfected using control vector (NG) or FGF9-overexpressing lentivirus vector (HG+OE-FGF9), and si-FGF9 (5＇-CUAUGUUGCAUUAAAUAAATT-3) with the assistance of lipofectamine 3000 (Invitrogen, Carlsbad, CA, USA)  26, 29. The transfection procedure was conducted following the manufacturer's instructions.

value: kidney

value: HK2 cell

value: Human renal proximal tubular epithelial cells


title: WT_cell_subclone_3

value: lung epithelial cells


title: III_48h_1

description: AK Yolksac

value: III


title: PC2

value: Rosa^dcas9-VPR/+, 2C-GFP/CD4


title: RChIP_dTAG8h_rep1


title: II_48h_1

description: AK Yolksac

value: II


title: PC1

value: neg


title: HK2 cell，HG+OE-FGF9-1


title: PC6

value: pos


title: I_48h_1

description: AK Yolksac

value: I


title: WT_cell_subclone_2


title: RChIP_DMSO8h_rep2

value: Flp-In T-REx HEK293

value: SETX-dTAG, flpin RNaseH1Mut(D210N)-V5


title: WT_cell_subclone_1