title: Human Tconv stim, Donor 2, CRISPRoff NS-, gRNA gJ-53

description: Library name: Donor2_human_Tconv_stim_CRISPRoff_NS-_gJ-53

data_processing:
Motif analysis was conducted as described in Freimer et al. 2022, with minor modifications. Fastq adapters were trimmed using cutadapt (v4.4), keeping reads with a minimum read length of 20 bps. Reads were aligned to the human genome GRCh38 (GENCODE, gencode.v43.primary_assembly.basic.annotation.gtf) using STAR (v2.7.10b) with the setting –outFilterMultimapNmax 1. Reads overlapping with genes were counted using featureCounts (Subread, v2.0.3) with the setting –s 2.
Assembly: hg38
Supplementary files format and content: Raw count file

extract_protocol:
To isolate RNA, cells were pelleted and resuspended in 300 µL RNA Lysis Buffer (Zymo Research, catalog no. R1060-1-50), vortexed vigorously, and frozen at -80°C until further processing. Lysed cells were thawed, and RNA was isolated using the QuickRNA Microprep Kit (Zymo Research, catalog no. R1050), according to the manufacturer’s protocol, but skipping the DNase I treatment. Eluted RNA in 15 µL was treated with 1.5 µL TURBO DNase in 1X TURBO DNase Buffer (Invitrogen, catalog no. AM2238) and incubated 20 minutes at 37°C. RNA samples were cleaned up using the RNA Clean & Concentrator-5 Kit (Zymo Research, catalog no. R1013), according to the manufacturer’s protocol. RNA integrity (RIN score > 8.0) and yield was assessed using Qubit RNA HS Assay Kit (Fisher Scientific, catalog no. Q32855) and Tapestation High Sensitivity RNA ScreenTape Analysis (Agilent, catalog nos. 5067-5579 and 5067-5580).
Libraries for RNA sequencing were prepared using the KAPA RNA HyperPrep Kit with RiboErase (HMR) (Roche, catalog no. 08098131702) and KAPA Unique Dual-Indexed Adapter Kit (Roche, catalog no. 08861919702), according to manufacturer’s protocol.

treatment_protocol: Human Tconvs from two donors were isolated, cultured, and modified with CRISPRoff using gRNAs targeting NS- and AAVS1 (2). At day 9, cells were restimulated with Immunocult Human CD3/CD28/CD2 T Cell Activator (STEMCELL Technologies, catalog no. 10970). At day 11 (48 hours-post restimulation, stimulated), 5E5-1E6 cells per condition were harvested for RNA isolation.

value: stim

value: Donor 2

tag: target

value: NS-

tag: grna


title: Mouse Tconv stim, Rep 2, Input ChIP, DSG/formaldehyde fixation

status: Public on Nov 13 2025

type: SRA

city: San Francisco

first: Alexander

last: Marson

state: CA

address: 1650 Owens St.

country: USA

institute: UCSF Gladstone Institutes

description: Library name: Rep2_mouse_Tconv_stim_Input_ChIP_dsg

library_source: genomic

data_processing:
Reads were mapped to the mm10 mouse 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 MACS3 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: GRCm38/mm10
Supplementary files format and content: Bigwig and MACS3 peak bed files.

molecule: genomic DNA

organism: Mus musculus

source_name: Spleen, Lymph nodes

growth_protocol: Spleens, mesenteric lymph nodes, and inguinal lymph nodes were harvested from Foxp3-Thy1.1 mice and mashed through a 70 µM filter in EasySep, and flow through cells were centrifuged at 300g for 5 min. Supernatant was aspirated, and cells were resuspended at 100E6 cells/mL. CD4+ T cells were isolated from washed splenocytes and lymph nodes cells using the EasySep Mouse CD4+ T Cell Isolation Kit (STEMCELL Technologies, catalog no. 19852), according to the manufacturer’s protocol. Murine Tregs were depleted from CD4+ T cells using the EasySep Mouse CD90.1 Positive Selection Kit (STEMCELL Technologies, catalog no. 18958). Tconvs were cultured in Kool Aid Complete Media (DMEM High Glucose, Custom “Kool Aid” media, catalog no. UCSFMP-RM-4591, obtained from UCSF Media Center) supplemented with 10% FBS (R&D Systems, catalog no. S11550), 100 µM 2-Mercaptoethanol (Sigma, catalog no. 63689-25ML-F), 10 mM HEPES (Sigma, catalog no. H0887-100ML), 1 mM Sodium Pyruvate (Fisher Scientific, catalog no. 11360070), 100 IU/mL Penicillin-Streptomycin (Sigma, catalog no. P4333-100ML), 2 mM L-Glutamine (Fisher Scientific, catalog no. 25030081) and 1X MEM (Fisher Scientific, catalog no. 11140050). Cells were cultured in NeutrAvidin (Fisher Scientific, catalog no.31,000) coated (5µg/mL in PBS) 24 well plates. Cells were stimulated with Biotin Hamster Anti-Mouse CD3e clone 145-2C1 (Becton Dickinson, catalog no. 553060) at a concentration of 0.25 µg/mL and Biotin CD28 clone 37.51 (Becton Dickinson, catalog no.553296) at a concentration of 1 µg/mL. Recombinant Human IL-2 (R&D Systems, catalog no. 202-GMP or BT-002-GMP) was supplemented at 50 IU/mL for Tconv cells. Additionally, recombinant Human IL-7 (Fisher Scientific, catalog no. 207-IL-050) at a concentration of 2.5 ng/mL and unconjugated Human IL-15 (Fisher Scientific, catalog no. 247-ILB-025) at a concentration of 50 ng/mL were added to enhance cell recovery, growth, and viability. Cells were cultured at 37°C and 5% CO2 and split to 1E6 cells/mL every ~2 days by topping off media and completely replacing the appropriate dose of IL-2, IL-7 and IL-15.

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. Mouse Tconv 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 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: 14 days after isolation and initial stimulation, a portion of cells were stimulated with soluble 0.25 µg/mL anti-CD3 (Bio-X-Cell, catalog no. BE0001-1) and 1 µg/mL anti-CD28 (Bio-X-Cell, catalog no. BE0015-5) antibodies. On day 15 post-initial stimulation, rested (Rest) and restimulated (Stim) cells were fixed for ChIP-seq.

tag: tissue

value: Spleen, Lymph nodes

tag: cell type

value: Tconv

tag: replicate

value: Rep 2

tag: condition

tag: genotype

tag: Sex

value: Male

tag: age

value: 10 weeks

tag: chip target

value: Input

tag: fixation method

value: DSG/formaldehyde

tag: chip antibody


title: COTC030 1022 tumor sample

city: Bethesda

first: Anjali

last: Garg

state: Maryland

address: Building 10, Room 1B53

department: Comparative Oncology Program,NCI

institute: National Institute of Health

description:
Library name: COTC030_1022_T
Column name in all_sample_raw_count.txt: COTC030_1022_T

library_source: transcriptomic

data_processing:
The HiSeq Real Time Analysis software (RTA v.3.4.4) was used for processing raw data files.
The Illumina bcl2fastq2.17 was used to demultiplex and convert binary base calls and qualities to fastq format.
The trimmed reads were mapped to the CanFam4 reference genome (GSD_1.0 from NCBI) using STAR aligner (version 2.7.0f) with two-pass alignment option.
RSEM (version 1.3.1) was used for gene and transcript quantification based on the CanFam4 GTF file.
Library complexity was measured regarding unique fragments in the mapped reads using Picard’s MarkDuplicate utility.
Assembly: Canis lupus familiaris
Supplementary files format and content: txt file include FPKM values for each sample

molecule: total RNA

organism: Canis lupus familiaris

source_name: Untreated bone primary osteosarcoma

extract_protocol:
RNA for mRNAseq was isolated from canine frozen tumor tissue in RNAlater using Qiagen Allprep DNA/RNA Mini Kit (Cat#80204).
RNA libraries were pooled and sequenced on NovaSeq S1 using a 2x150 cycle kit.

value: Untreated bone primary osteosarcoma


title: ClRyeinput

city: Seeland OT Gatersleben

first: Anne

last: Fiebig

address: Correnstraße 3

department: BIT

country: Germany

institute: IPK Gatersleben

description:
Library name: ClRyeinput
The original chromatin DNA was a mixture of chromatin isolated from flowers of Chamaelirium luteum and leaves of Secale cereale inbred line Lo7 and sonicated to 150-400 bp in length

data_processing:
ChIP-Seq Mapper (Galaxy version 1.1.1.4) was used to map the ChIP- and input reads on RepeatExplorer-derived contig sequences of repeat clusters.
The paired-end reads of ChIP- and input-seq were quality-filtered by Trimmmomatic (Galaxy Version 0.39) and the resulting reads were mapped to the C. luteum genome assembly using Bowtie2 (Galaxy Version 2.5.3) with default parameters.
The deeptools bamCompare function (Galaxy Version 3.5.4) was used to generate the normalized ChIP-seq signal track of the average of read counts in ChIP over input in a genome-wide 1 kb windows.
Assembly: GCA_964656255
Supplementary files format and content: bigwig

organism: Chamaelirium luteum

source_name: leaf, flower

extract_protocol:
Genomic DNA of C. luteum was extracted from leaf tissue using the DNeasy Plant Mini kit (Qiagen, Germany)
ChIPseq libraries were prepared by NEBNEXT® UltraTM II DNA Library Prep Kit for Illumina (New England Biolabs) and sequenced using NovaSeq 6000 system (Illumina) by Novogene (UK) in the paired-end run (2×150 bp).

treatment_protocol: 0.65 g of C. luteum flower and 1.0 g of Secale cereale (inbred line Lo7) leaf tissue were ground with liquid nitrogen and homogenized separately in 10 ml nuclei isolation buffer (1 M sucrose, 5 mM KCl, 5 mM MgCl2, 60 mM HEPES pH 8.0, 5 mM EDTA, 0.6% Triton X-100, 0.4 mM PMSF, 1 µM pepstatin A, cOmplete protease inhibitor cocktail (Roche)). Nuclei fixation was performed in 1% PFA/ nuclei isolation buffer at RT, 12 rpm for 10 min and terminated by adding glycine to a final concentration of 130 mM. The nuclei suspension was filtrated through Miracloth (Millipore) twice and a 50-µm CellTrics filter (Sysmex) once and centrifuged at 4°C, 3,000 ×g for 10 min. The nuclei pellet was resuspended in 1 ml extraction buffer (0.25 M sucrose, 10 mM Tris-HCl pH 8.0, 10 mM MgCl2, 1% Triton X-100, 1 mM EDTA, 5 mM β-mercaptoethanol, 0.1 mM PMSF, 1 µM pepstatin A, cOmplete protease inhibitor cocktail), followed by centrifugation at 4°C, 12,000 ×g for 10 min. After removing the supernatant, nuclei were resuspended in 150 µl of nuclei lysis buffer (20 mM Tris-HCl pH 8.0, 10 mM EDTA, 1% SDS, 0.1 mM PMSF, 1 µM pepstatin A, cOmplete protease inhibitor cocktail). Chromatins were sonicated for 14 cycles of 30 s ON, 30 s OFF at high power, in a Bioruptor (Diagenode), followed by an addition of 100 µl ChIP dilution buffer (16.7 mM Tris-HCl pH 8.0, 167 mM NaCl, 1.1% Triton X-100, 1 mM EDTA, cOmplete protease inhibitor cocktail), and continued sonication to a total of 31 cycles under the same setting. The sonicated samples were diluted 10 times with ChIP dilution buffer, centrifuged at 4°C, 13,000 ×g for 5 min, and the supernatant of each samples were transferred to new tubes. To dilute the high proportion of the putative C. luteum centromeric repeat, sonicated chromatin of C. luteum and S. cereale was mixed in a 1:3 ratio. The mixed chromatins were incubated with the ClCENH3 antibody (10 mg/ml) to a final 1:500 dilution at 4°C by shaking at 14 rpm for 12 h. DynabeadsTM Protein A (Invitrogen) in ChIP dilution buffer, corresponding to 0.1× volume of the chromatin solution, was added to the antibody-prebound chromatins and incubated at 4°C by shaking at 14 rpm for 1.5 h. The collected beads were then washed twice in low salt buffer (150 mM NaCl, 0.1% SDS, 1% Triton X-100, 2 mM EDTA, 20mM Tris-HCl pH 8.0), followed by three washes in high salt buffer (500 mM NaCl, 0.1% SDS, 1% Triton X-100, 2 mM EDTA, 20 mM Tris-HCl pH 8.0), and another two washes in TE buffer at 4°C by shaking at 14 rpm for 5 min. The bead-bound chromatin was purified by using iPure kit v2 (Diagenode) following the manual and quantified using QubitTM dsDNA HS Assay kit (Invitrogen).

value: leaf, flower


title: Mouse Tconv rest, Rep 2, Input ChIP, DSG/formaldehyde fixation

description: Library name: Rep2_mouse_Tconv_rest_Input_ChIP_dsg

value: rest


title: Human Tconv rest, Donor 1, MBD2 ChIP, DSG/formaldehyde fixation

description: Library name: Donor1_human_Tconv_rest_MBD2_ChIP_dsg

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.

organism: Homo sapiens

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. 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 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:
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

tag: donor

value: Donor 1

value: MBD2 (Active Motif, 39547)


title: Sta phase_5primeTNET-seq_-TEX_rep2

city: Tianjin

first: Zhe

last: Sun

address: 32 West 7th Ave

country: China

institute: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences

library_source: other

data_processing:
Basecalling was performed by RTA 1.18.
Adaptor and barcode were removed by cutadapt 1.18.
For 5primeTNET-seq, duplicates were removed by bbmap 38.22.
5primeTNET-seq R2 and R1 reads were merged by FLASH 1.2.11.
Merged reads were aligned to E. coli MG1655 genome NC_000913.2 using bowtie 1.2.2 with parameters --best --strata -m 1.
For 5primeTNET-seq, the read coverage, the counts of all uniquely aligned reads whose 5’ end and 3prime end mapped were recorded and counted in each coordinate by bedtools 2.27.1.
Assembly: Escherichia coli str. K-12 substr. MG1655, NC_000913.2
Supplementary files format and content: wig files, the scores represent the counts of uniquely aligned reads whose 5prime end or 3prime end mapped at each coordinate in each strand.

molecule: other

organism: Escherichia coli str. K-12 substr. W3110

source_name: beta prime-WT

growth_protocol: The overnight cell cultures were diluted in 250 ml LB medium (OD600 = 0.02) and incubate at 37 ºC to mid-log phase (OD600 = 0.5) or stationary phase (OD600 = 2.0).

extract_protocol:
The cells collected were resuspended and lysed by 120 kU Ready-Lyse lysozyme (Lucigen) at room temperature for 10 min. The chromosomal DNA with RNAP binding was pelleted and treated with 6 U Turbo DNase (Invitrogen) and 100 U DNase I (Roche) by vortexing at room temperature for 10 min. After centrifugation, the supernatant containing ECs was transferred to mix with Ni-NTA beads for 1 hour at 4 ºC with continuous shaking (1000 rpm). The beads were washed 3 times and loaded onto 0.5 ml Ultrafree-MC centrifugal filters (Millipore). The immobilized ECs were washed one more time and eluted with wash buffer containing 0.5 M imidazole. The nucleic acids in the eluates were extracted once with phenol:chloroform:isoamylalcohol (PCI; 25:24:1) and once with chloroform. The top water phase was collected and the nucleic acids were precipitated by isopropanol at -80 ºC for 30 min. After centrifugation, the nucleic acids pellet was washed by 80% ethanol and air dried. The resultant pellet was dissolved in nuclease-free water. The DNA was removed by 2 U Turbo DNase and 10 U DNase I at 37 ºC for 15 min. The residual RNA was extracted by PCI and precipitated by isopropanol, and fractionated on 15% TBE-Urea gel. Tthe nascent RNA between 8–100-nt on the gel was extracted and purified.
Libraries were constructed based on the modifed protocol of NET-seq (Mayer, A. and Churchman, L. S. Nat Protoc. 2016). Briefly, a barcode DNA linker was ligated to 3prime end of nascent RNA. The resulting RNA-DNA chimera was treated with or without 2 U 5’-phosphate-dependent exonuclease (Epicentre) at 30ºC for 1h, and then reverse transcribed by using PrimeScript Reverse Transcriptase and a 5prime phosphorylated reverse transcription primer. The cDNA between 80-175 nt were excised after running 10% Urea-TBE gel, extracted and precipitated by isopropanol. The purified cDNA was circularized by ssDNA ligase (Takara) and applied to PCR the sequencing library using Illumina index primers and PrimeSTAR Max DNA polymerase (Takara). The DNA product excised from gel was extracted and purified for Illumina sequencing.

tag: strain

value: beta prime-WT

tag: growth

value: W3110 rpoC-6xHis

tag: treatment

value: Nascent RNA not treated with 5´-Phosphate-Dependent Exonuclease


title: COTC030 1019 tumor sample

description:
Library name: COTC030_1019_T
Column name in all_sample_raw_count.txt: COTC030_1019_T


title: Sta phase_5primeTNET-seq_+TEX_rep2

value: Nascent RNA treated with 5´-Phosphate-Dependent Exonuclease


title: Sta phase_5primeTNET-seq_-TEX_rep1


title: Human Tconv stim, Donor 2, CRISPRoff AAVS1, gRNA gJ-49

description: Library name: Donor2_human_Tconv_stim_CRISPRoff_AAVS1_gJ-49

value: stim

value: Donor 2


title: COTC030 1018 tumor sample

description:
Library name: COTC030_1018_T
Column name in all_sample_raw_count.txt: COTC030_1018_T


title: Mouse Tconv rest, Rep 1, Input ChIP, DSG/formaldehyde fixation

description: Library name: Rep1_mouse_Tconv_rest_Input_ChIP_dsg

value: Spleen, Lymph nodes

value: Tconv

value: Rep 1


title: Human Tconv stim, Donor 2, CRISPRoff AAVS1, gRNA gJ-48

description: Library name: Donor2_human_Tconv_stim_CRISPRoff_AAVS1_gJ-48


title: Human Tconv stim, Donor 1, CRISPRoff NS-, gRNA gJ-53

description: Library name: Donor1_human_Tconv_stim_CRISPRoff_NS-_gJ-53

value: NS-


title: Mouse Tconv stim, Rep 2, H3K27ac ChIP, DSG/formaldehyde fixation

description: Library name: Rep2_mouse_Tconv_stim_H3K27ac_ChIP_dsg

value: Rep 2

value: Male

value: 10 weeks

value: DSG/formaldehyde

value: H3K27ac (Active Motif, 39133)


title: Sta phase_5primeTNET-seq_+TEX_rep1


title: COTC030 1017 tumor sample

description:
Library name: COTC030_1017_T
Column name in all_sample_raw_count.txt: COTC030_1017_T

value: Untreated bone primary osteosarcoma


title: Human Tconv stim, Donor 1, CRISPRoff AAVS1, gRNA gJ-49

description: Library name: Donor1_human_Tconv_stim_CRISPRoff_AAVS1_gJ-49

value: Blood

value: Donor 1


title: Mouse Tconv rest, Rep 2, H3K27ac ChIP, DSG/formaldehyde fixation

description: Library name: Rep2_mouse_Tconv_rest_H3K27ac_ChIP_dsg

value: rest


title: Human Tconv rest, Donor 1, GATA3-abcam ChIP, DSG/formaldehyde fixation

description: Library name: Donor1_human_Tconv_rest_GATA3-abcam_ChIP_dsg

value: GATA3 (Abcam, ab199428)


title: Log phase_5primeTNET-seq_+TEX_rep2

value: beta prime-WT

value: W3110 rpoC-6xHis

value: Nascent RNA treated with 5´-Phosphate-Dependent Exonuclease


title: COTC030 1024 tumor sample

description:
Library name: COTC030_1024_T
Column name in all_sample_raw_count.txt: COTC030_1024_T


title: COTC030 1016 tumor sample

description:
Library name: COTC030_1016_T
Column name in all_sample_raw_count.txt: COTC030_1016_T


title: Log phase_5primeTNET-seq_-TEX_rep1

value: Nascent RNA not treated with 5´-Phosphate-Dependent Exonuclease


title: Human Tconv stim, Donor 1, CRISPRoff AAVS1, gRNA gJ-48

description: Library name: Donor1_human_Tconv_stim_CRISPRoff_AAVS1_gJ-48

value: Tconv

value: stim


title: COTC030 1015 tumor sample

description:
Library name: COTC030_1015_T
Column name in all_sample_raw_count.txt: COTC030_1015_T


title: Mouse Tconv stim, Rep 2, EGR2 ChIP, DSG/formaldehyde fixation

description: Library name: Rep2_mouse_Tconv_stim_EGR2_ChIP_dsg

value: Spleen, Lymph nodes

value: EGR2 (Abcam, ab43020)


title: COTC030 1014 tumor sample

description:
Library name: COTC030_1014_T
Column name in all_sample_raw_count.txt: COTC030_1014_T


title: C4-2B/MDVR cells, shAR, 2

city: Taoyuan

first: Ling-Yu

last: Wang

address: No.259, Wenhua 1st Rd., Guishan Dist.,

department: Biochemistry and Molecular Biology

country: Taiwan

institute: Chang Gung University

description: Library name: Sample 14

data_processing:
Raw sequencing data were processed using CASAVA, and quality checks were performed with FastQC and MultiQC. Trimmomatic was used to remove adapters and low-quality reads, yielding high-quality "clean reads." "Relative Log Expression" (RLE) normalization was performed using DESeq2.
Assembly: The reads were mapped to the GRCh38 reference genome using HISAT2, and gene-level counts were obtained with featureCounts.
Supplementary files format and content: csv files include normalized read counts for each sample

source_name: cancer cell line

extract_protocol:
Total RNA was extracted by TRIzol™ Reagent (Thermo Fisher Scientific).
1 μg total RNA per sample was used as input material for the library construction. RNA sequencing libraries were prepared using the KAPA mRNA HyperPrep Kit, following the manufacturer’s protocol.

value: cancer cell line

tag: cell line

value: Prostate cancer

value: shAR


title: C4-2B/MDVR cells, shAR, 1

description: Library name: Sample 13


title: Mouse Tconv rest, Rep 2, EGR2 ChIP, DSG/formaldehyde fixation

description: Library name: Rep2_mouse_Tconv_rest_EGR2_ChIP_dsg

value: Rep 2

value: Male

value: 10 weeks

value: DSG/formaldehyde


title: COTC030 1013 tumor sample

description:
Library name: COTC030_1013_T
Column name in all_sample_raw_count.txt: COTC030_1013_T

value: Untreated bone primary osteosarcoma


title: Log phase_5primeTNET-seq_+TEX_rep1


title: Tasor_1-IgG

city: Nanjing

first: Bowen

address: 12 Xue-fu Road, Pukou District

institute: Nanjing University

description: Library name: Sample 4

data_processing:
The high-quality reads were aligned to the Mus musculus (mm10) reference genome utilizing bowtie2. Post-alignment reads mapping to ENCODE mm10 blacklist regions were filtered out with samtools. The BAM file generated by the unique mapped reads was used as input for MACS2 to call peaks with cutoff q-value < 0.01.
Assembly: mm10
Supplementary files format and content: bigwig
Library strategy: CUT&Tag

extract_protocol:
Briefly, 1 × 10^5 cells were treated with 10 μL concanavalin A-coated magnetic beads, followed by incubation with primary antibodies. After removal of primary antibody using magnet stand, Goat Anti-Rabbit IgG H&L was diluted at 1:100 in dig wash buffer, and cells were incubated at RT for 1 hr. Then, pA-Tn5 adapter complex was added and incubated with samples at RT for 1 hr. Next, genomic DNA was purified using Tagment DNA Extract Beads.
The libraries were constructed by mixing 21μL DNA with 2μL of a universal i5 and a uniquely barcoded i7 primer.

value: mouse embryonic stem cells


title: Mouse Tconv rest, Rep 1, EGR2 ChIP, DSG/formaldehyde fixation

description: Library name: Rep1_mouse_Tconv_rest_EGR2_ChIP_dsg

value: Rep 1

value: rest


title: C4-2B/MDVR cells, pLKO1, 2

description: Library name: Sample 12


title: C4-2B/MDVR cells, pLKO1, 1

description: Library name: Sample 11


title: COTC030 1012 tumor sample

description:
Library name: COTC030_1012_T
Column name in all_sample_raw_count.txt: COTC030_1012_T


title: Mouse Tconv stim, Rep 2, CTCF ChIP, DSG/formaldehyde fixation

description: Library name: Rep2_mouse_Tconv_stim_CTCF_ChIP_dsg

value: CTCF

value: CTCF (Abcam, ab128873)


title: Tasor_1

description: Library name: Sample 3


title: COTC030 1009 tumor sample

description:
Library name: COTC030_1009_T
Column name in all_sample_raw_count.txt: COTC030_1009_T


title: Log phase_5primeTNET-seq_-TEX_rep2

value: beta prime-WT

value: W3110 rpoC-6xHis

value: Nascent RNA not treated with 5´-Phosphate-Dependent Exonuclease


title: C4-2B/MDVR cells, ITRI-148, 2

description: Library name: Sample 10


title: Mpp8_1-IgG

description: Library name: Sample 2


title: COTC030 1007 tumor sample

description:
Library name: COTC030_1007_T
Column name in all_sample_raw_count.txt: COTC030_1007_T


title: Mpp8-1

description: Library name: Sample 1


title: Mouse Tconv rest, Rep 2, CTCF ChIP, DSG/formaldehyde fixation

description: Library name: Rep2_mouse_Tconv_rest_CTCF_ChIP_dsg

value: Spleen, Lymph nodes

value: Tconv


title: Mouse Tconv rest, Rep 1, CTCF ChIP, DSG/formaldehyde fixation

description: Library name: Rep1_mouse_Tconv_rest_CTCF_ChIP_dsg


title: COTC030 1006 tumor sample

description:
Library name: COTC030_1006_T
Column name in all_sample_raw_count.txt: COTC030_1006_T


title: C4-2B/MDVR cells, ITRI-148, 1

description: Library name: Sample 9

value: cancer cell line

value: Prostate cancer


title: Mouse Tconv stim, Rep 2, YY1 ChIP, DSG/formaldehyde fixation

description: Library name: Rep2_mouse_Tconv_stim_YY1_ChIP_dsg

value: Rep 2

value: stim


title: Mouse Tconv rest, Rep 1, H3K27ac ChIP, DSG/formaldehyde fixation

description: Library name: Rep1_mouse_Tconv_rest_H3K27ac_ChIP_dsg

value: Male

value: 10 weeks

value: H3K27ac (Active Motif, 39133)


title: C4-2B/MDVR cells, ITRI-90, 2

description: Library name: Sample 8


title: Mouse Tconv rest, Rep 2, YY1 ChIP, DSG/formaldehyde fixation

description: Library name: Rep2_mouse_Tconv_rest_YY1_ChIP_dsg

value: DSG/formaldehyde


title: αSyn PM_PFF injection_PM nasal_rep3

city: Baltimore

first: Xiaodi

last: Zhang

state: MD

address: 733 N. Broadway, MRB 710

institute: Johns Hopkins School of Medicine

description: Library name: PMPFFPMA2738

data_processing:
Reads were initially quality-checked with FastQC  v0.11.8, followed by trimming with Trimmomatic v0.39 when necessary. The quality-checked reads were then aligned to the mouse genome (mm10)  using HISAT2 v2.2.1. Gene-level expression was quantified using featureCounts  based on GENCODE release M25.
Assembly: mm10
Supplementary files format and content: tab-delimited text file includes raw counts for each sample

source_name: anterior cingulate cortex

extract_protocol:
Sequencing libraries were prepared using the RNeasy Lipid Tissue Mini Kit, following the manufacturer's protocol.
NEBNext® Ultra II Directional RNA Library Prep Kit for Illumina®.

treatment_protocol: RNA was extracted from the anterior cingulate cortex of mice treated under each condition, with four replicates per condition. 2-month-old humanized WT αSyn mice (FVB; 129S6-Sncatm1Nbm Tg (SNCA)1Nbm/J, Strain: 010710), obtained from The Jackson Laboratory (Bar Harbor, ME, USA), were used for RNA sequencing experiments. The mice were divided into five groups: the PBS group (intranasal administration of PBS), the PM2.5 group (intranasal administration of PM2.5), the PFF group (PFF injection), the PM-PFF group (PM-PFF injection), and the PM-PFF with PM2.5 exposure group (PM-PFF injection followed by intranasal PM2.5 administration one week later). All treatments were administered for a duration of 2 months.

value: anterior cingulate cortex

value: humanized aSyn WT mice


title: αSyn PM_PFF injection_PM nasal_rep4

description: Library name: PMPFFPM2778


title: C4-2B/MDVR cells, ITRI-90, 1

description: Library name: Sample 7


title: COTC030 1003 tumor sample

description:
Library name: COTC030_1003_T
Column name in all_sample_raw_count.txt: COTC030_1003_T

value: Untreated bone primary osteosarcoma


title: αSyn PM_PFF injection_PM nasal_rep2

description: Library name: PMPFFPMA2729


title: C4-2B/MDVR cells, DMSO, 2

description: Library name: Sample 6

value: DMSO


title: Mouse Tconv rest, Rep 1, YY1 ChIP, DSG/formaldehyde fixation

description: Library name: Rep1_mouse_Tconv_rest_YY1_ChIP_dsg

value: Rep 1

value: rest


title: COTC030 1001 tumor sample

description:
Library name: COTC030_1001_T
Column name in all_sample_raw_count.txt: COTC030_1001_T


title: αSyn PM_PFF injection_PM nasal_rep1

description: Library name: PMPFFPMA2727


title: C4-2B/MDVR cells, DMSO, 1

description: Library name: Sample 5


title: HK2-OEHNRNPF-3

city: Wu Han

first: LAN

last: WANG

state: Hubei

address: 湖北省武汉市武昌区湖北中医药大学昙华林校区昙华林188号

institute: Hubei University of Chinese Medicin

data_processing:
For high-throughput sequencing, the libraries were prepared following the manufacturer's instructions and applied to Illumina Novaseq 6000 system for 150 nt paired-end sequencing.
First discarded were raw reads containing more than 2-N bases. Then adaptors and low-quality bases were trimmed from raw sequencing reads using FASTX-Toolkit (Version 0.0.13). The short reads less than 16nt were dropped as well.
Clean reads were aligned to the GRCh38 genome by HISAT2 allowing 4 mismatches.
Uniquely mapped reads were used for gene reads number counting and FPKM calculation (fragments per kilobase of transcript per million fragments mapped) .
The alternative splicing events (ASEs) and regulated alternative splicing events (RASEs) between the samples were defined and quantified using the ABLas pipeline.
Assembly: GRCh38
Supplementary files format and content: expressed_gene_reads.sort.txt
Supplementary files format and content: expressed_gene_FPKM.txt

source_name: HK-2 cells

growth_protocol: cell culture

extract_protocol:
Total RNA was extracted according to the Trizol method, and 1 μL of total RNA was used as the template, reverse transcribed to cDNA, and continued to be amplified by using cDNA as the template, sequentially 95 ℃ for 3 min, 1 cycle, 95 ℃ for 10 s, and 62 ℃ for 40 s, for a total of 40 cycles. The mRNA levels of each group were calculated using 2-ΔΔCt method with β-actin as internal reference.
Library preparation, sequencing information (eg. cDNA libraries were prepared using the SMARTer Stranded Total RNA-Seq Kit version 2 (634413; Takara/Clontech) according to the manufacturer's procedure. 150 nt double-end sequencing was performed on the Illumina NovaSeq system)
1 μg of total RNA was used for RNA-seq library preparation. mRNAs were captured by VAHTS mRNA capture Beads (Vazyme, N401).The purified RNA was treated with RQ1 DNase (Promega) to remove DNA before used for directional VAHTS Universal V8 RNA-seq Library Prep Kit for Illumina （NR605） Polyadenylated mRNAs were purified and fragmented. Fragmented mRNAs were converted into double strand cDNA. Following end repair and A tailing, the DNAs were ligated to Adaptor (N323). After purification of ligation product and size fractioning to 300-500bps, the ligated products were amplified and purified, then quantified and stored at -80℃ before sequencing. The strand marked with dUTP (the 2nd cDNA strand) is not amplified, allowing strand-specific sequencing

value: HK-2 cells

value: HNRNPF overexpression inHK-2 cells


title: Total: 5-AZA-CdR + ADAR1i-124

city: Philadelphia

first: Haoran

state: Pennsylvania

address: 415 Curie Boulevard

institute: University of Pennsylvania

description: Library name: MM45

data_processing:
snakePipes mRNA-seq (v2.8.0)
Assembly: mm10
Supplementary files format and content: tab-delimited text file includes raw counts for each Sample

extract_protocol:
RNA was harvested using TRIzol reagent (Invitrogen). At least 1.5 ug of total RNA was used for RNA-seq. For dsRNA immunoprecipitation, J2 anti-dsRNA antibody (Scicons, 10010500) and anti-Z-RNA antibody (Z22)(Absolute Antibody, Ab00783-3.3) were used to enrich the dsRNA species with IgG as control.
Input  RNA was converted into Illumina sequencing libraries using the Illumina Standard Total RNA Prep, Ribo-Zero Plus Ligation (Illumina, 20040525), following the manufacturer’s instructions with the following modifications. Input RNA, along with Illumina RNA UD Indexes Set A Ligation (Illumina, 20091655), were ligated to the resulting cDNA after second-strand synthesis and end-prepping.

value: mouse melanoma cells

value: WT

value: Total:5-AZA-CdR + ADAR1i-124


title: αSyn PM_PFF injection_rep4

description: Library name: PMPFFA2734


title: BAL CD4+ T cells CMO

city: Charlottesville

first: Arka

last: Sen Chaudhuri

state: VA

address: 345 Crispell Drive

department: Carter Immunology Center

institute: University of Virginia

description:
Library name: Sequencing 4
10x Genomics

library_source: transcriptomic single cell

data_processing:
The sequenced data were aligned using Cell Ranger. The curated data then underwent demultiplexing, quality assurance, filtering, differential gene expression analysis, and cluster identification, all conducted using the standard Seurat protocol with Seurat (v 5.1.0) in R (R-4.3.3).
Assembly: Mouse mm10
Supplementary files format and content: Hashtag Demultiplexed features and matrix

source_name: Bronchoalveolar lavage (BAL)

extract_protocol:
For library preparation, BAL cells or sorted BAL CD4+ T cells from immunized mice in each group were pooled together and each group was hashtagged using BioLegend TotalSeq™-A antibodies according to the provided protocol.
The library of hashtagged pooled groups was then constructed using the 10x Genomics Chromium Next GEM Single Cell 3’ Reagent Kit with the v3.1 (Dual Index) protocol. Sequencing was performed using the NovaSeq X Plus platform, a service provided by Novogene.

value: Bronchoalveolar lavage (BAL)


title: VCaP cells, ITRI-148, 2

description: Library name: Sample 4

value: cancer cell line

value: VCaP

value: Prostate cancer


title: HK2-OEHNRNPF-2

address: 湖北省武汉市武昌区湖北中医药大学昙华林校区昙华林188号


title: BAL CD4+ T cells

description:
Library name: Sequencing 3
10x Genomics


title: αSyn PM_PFF injection_rep3

description: Library name: PMPFFA2732


title: Total BAL cells CMO

description:
Library name: Sequencing 2
10x Genomics


title: VCaP cells, DMSO, 2

description: Library name: Sample 2


title: HK2-OEHNRNPF-1

address: 湖北省武汉市武昌区湖北中医药大学昙华林校区昙华林188号


title: VCaP cells, ITRI-148, 1

description: Library name: Sample 3


title: COTC030 1004 tumor sample

description:
Library name: COTC030_1004_T
Column name in all_sample_raw_count.txt: COTC030_1004_T


title: Total BAL cells

description:
Library name: Sequencing 1
10x Genomics


title: Ctrl_2

address: 湖北省武汉市武昌区湖北中医药大学昙华林校区昙华林188号

value: HK-2 cells (control plasmid)


title: αSyn PM_PFF injection_rep1

description: Library name: PMPFFA2730


title: Ctrl_3

address: 湖北省武汉市武昌区湖北中医药大学昙华林校区昙华林188号


title: VCaP cells, DMSO, 1

description: Library name: Sample 1


title: PM nasal_rep4

description: Library name: PMA2779


title: None_24h_1

city: Exeter

first: Rhys

last: Farrer

state: Devon

address: Stocker Road

department: MRC Center for Medical Mycology

country: United Kingdom

institute: University of Exeter

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).

organism: Aphanius dispar

source_name: Whole organism

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: Whole organism

value: Wild type

value: A. dispar without injection

tag: time point

value: 24 hours


title: PM nasal_rep3

description: Library name: PMA2778

value: anterior cingulate cortex

value: humanized aSyn WT mice


title: Ctrl_1

address: 湖北省武汉市武昌区湖北中医药大学昙华林校区昙华林188号


title: Mouse Chow 3

city: Los Angeles

first: Jenny

last: Cheng

address: 675 Charles E. Young Dr. S

institute: University of California, Los Angeles

description:
Library name: Chow-3
10X Genomics

data_processing:
Fastq files from each mouse aorta pool were individually aligned to the mus musculus genome assembly GRCm38 (mm10) using CellRanger version 6.0.1 (10X Genomics). CellBender was used to learn the background noise profile and differentiate cell-containing droplets from cell-free droplets. A digital gene expression matrix, in which each row represents the read count of a gene and each column corresponds to a unique single cell, was generated from each library’s filtered feature-barcode datasets and was subsequently analyzed using the R package Seurat version 4.3.0. The number of expressed genes, number of unique molecular identifiers (UMIs), percentage of mitochondrial genes, and percentage of hemoglobin genes were considered in identifying outliers. Single cells were selected based on a threshold of between 200 and 5000 genes expressed and between 500 and 20,000 UMIs counted to reduce the presence of doublets as well as a mitochondrial gene expression cutoff of <15% and a hemoglobin gene expression cutoff of <0.1% to remove poor quality or contaminated cells. Raw transcript counts of each gene were normalized relative to the total number of read counts in that cell, and the resulting values were multiplied by 10,000 and log-transformed.
Assembly: Mouse mm10
Supplementary files format and content: Tab-separated value files and matrix files

source_name: ascending aorta, aortic arch, thoracic aorta

extract_protocol:
Each treatment group consisted of 3 aorta pools with n=2 mice/pool, for a total of 9 independent aorta pools from 18 mice. Aorta samples from the above groups were collected from the lesion prone areas of aorta, including the ascending aorta, aortic arch, and thoracic aorta. Aorta cell dissociation protocol was based on Wirka et al. Briefly, the aorta samples were cut into <1 mm pieces and incubated in 1 ml of Hanks’ Balanced Salt solution containing 2 units of liberase and 24 units of elastase at 37°C for 1 hour. Fetal bovine serum (10% of total volume) was then added to inactivate the enzymes. The digested cells were then passed through a 70 µm cell strainer and rinsed with 4 ml PBS. After centrifugation at 300g, 4°C, for 10 min, the supernatant was discarded. The cells were then resuspended in 4 ml PBS/0.04% BSA, spun down again, and resuspended in 100 ul PBS/0.04% BSA.
Approximately 16,000 aortic cells pooled from two mice within the same treatment group were used for each single cell library construction. In total, 9 independent libraries (3 libraries/diet group) were made using the 10x Genomics Chromium Next GEM Single Cell 3' GEM, Library & Gel Bead Kit v3.1. The 9 libraries were then sequenced in one lane of NovaSeq S4 2x100bp at 2.4 billion reads.

value: Chow

value: Ldlr-/- on the C57BL/6J background

value: female

value: 14 weeks


title: Water_24h_1

description: AK Yolksac

value: A. dispar with sham injection


title: PM nasal_rep2

description: Library name: PMA2776


title: Mouse High-cholesterol 3

description:
Library name: High-cholesterol-3
10X Genomics

value: ascending aorta, aortic arch, thoracic aorta

value: High-cholesterol

value: 22 weeks


title: αSyn PM_PFF injection_rep2

description: Library name: PMPFFA2731


title: PM nasal_rep1

description: Library name: PMA2775


title: Water_48h_3

description: AK Yolksac

value: 48 hours


title: Mouse High-cholesterol+TMAO 2

description:
Library name: High-cholesterol+TMAO-2
10X Genomics

value: High-cholesterol+TMAO


title: None_48h_3

description: AK Yolksac


title: adultMaleThalamus, control, replicate_4

city: Cambridge

first: Catherine

last: Dulac

address: 52 Oxford St

institute: Harvard University

description:
Library name: Control1
10x Genomics

data_processing:
Transcript sequences were demultiplexed, aligned to the mm10 mouse genome, counted, and assembled into gene-cell matrices using the 10x Genomics Cell Ranger pipeline with default parameters. The matrices were then analyzed in Seurat version 2.3.4  84. Gene expression for each nucleus was log-normalized and scaled (i.e., log-transformed, normalized by total transcript count, and multiplied by a factor of 10,000). To eliminate aberrant droplets, we filtered out nuclei based on mitochondrial content (> 10%), nuclear doublets (> 20,000 UMIs), red blood cells (>1% hemoglobin genes), and immediate early gene (IEG) responses (e.g., responses to the dissection/dissociation process; > 1% IEG content). Glial cells were filtered out based on the expression of established glial-specific genes.
Assembly: Mouse mm10
Supplementary files format and content: Coma-separated values files and matrix files

molecule: polyA RNA

source_name: Brain

extract_protocol:
Nuclear isolation, purification, and barcoding.  Nuclei were extracted following a custom protocol designed to minimize the amount of time from euthanasia to nuclear barcoding (approximately four hours). We prepared 10 biological samples for sequencing. Each biological sample was a pool of three mice (to obtain sufficient material for sequencing). Nine of the biological samples came from three hierarchy cohorts, each consisting of rank-1, rank-3, and control animals. The tenth biological sample was an additional control animal. Thus, the 10 biological samples were comprised of 30 animals. All animals were age-matched and processed in parallel. Mouse brains were quickly dissected on ice and 2mm thick coronal sections were cut on a brain matrix (Zivic Instruments, #5325). Sections were placed on a chilled surface under a dissecting microscope and tissue punches from the thalamus were collected using a 2mm diameter cylindrical corer (Fine Science Tools, #18035-02). Tissue punches were pooled in chilled Hibernate-A Medium (ThermoFisher, #A1247501), minced into approximately 0.52 mm pieces, and transferred to 3 mL lysis buffer (10 mM Tris-HCl, 10 mM NaCl, 3 mM MgCl2, 0.1% Nonidet P-40). Tissue was dounce homogenized in a glass tube 6-10 times with a grinder pestle (Potter-Elvehjem, #886000-0020) rotating at 300 rpm until opaque, transferred to a 15 mL tube, and pelleted in a swinging bucket centrifuge (Beckman Coulter GS-6R) at 50 RCF for 5 min at 4C. Supernatant was removed and tissue carefully resuspended in 1.5 mL PBS with BSA (1%) and RNase inhibitor (0.2 U/L, Promega, #N2615). To remove debris, the suspension was passed over a 70 M filter (ThermoFisher, #NC0120755) and then again over a 20 M filter (ThermoFisher, #130-101-812). With the suspension on ice, nuclei were labeled by adding DAPI (1.5 L, ThermoFisher, #D1306). DAPI-positive nuclei were sorted at 4C on a FACSAria flow sorter (BD Immunocytometry Systems) with a 70 mm nozzle on the four-way high speed purity setting until approximately 100,000 nuclei were collected.
Using the 10X Genomics droplet microfluidics Gemcode platform 83, nuclei from the suspension were loaded onto Chromium instrument. Libraries were generated using Chromium v2 reagents, and cDNA amplified according to the manufacturer’s instructions

value: Brain

tag: batch


title: Mouse High-cholesterol+TMAO 3

description:
Library name: High-cholesterol+TMAO-3
10X Genomics

tag: tissue


title: αSyn PFF injection_rep4

description: Library name: PFFA2749


title: None_48h_2

description: AK Yolksac

value: Whole organism


title: Water_48h_2

description: AK Yolksac

value: Wild type


title: Mouse High-cholesterol 2

description:
Library name: High-cholesterol-2
10X Genomics


title: adultMaleThalamus, dominant, replicate_3

description:
Library name: A3MD
10x Genomics


title: adultMaleThalamus, subordinate, replicate_2

description:
Library name: A2MS
10x Genomics


title: αSyn PFF injection_rep2

description: Library name: PFFA2746


title: adultMaleThalamus, control, replicate_3

description:
Library name: A3MC
10x Genomics


title: Mouse High-cholesterol 1

description:
Library name: High-cholesterol-1
10X Genomics


title: None_48h_1

description: AK Yolksac

value: A. dispar without injection


title: adultMaleThalamus, dominant, replicate_2

description:
Library name: A2MD
10x Genomics


title: αSyn PFF injection_rep1

description: Library name: PFFA2726


title: Control_PBS_rep4

description: Library name: Control8


title: Water_48h_1

description: AK Yolksac


title: Mouse Chow 1

description:
Library name: Chow-1
10X Genomics


title: adultMaleThalamus, control, replicate_2

description:
Library name: A2MC
10x Genomics


title: adultMaleThalamus, subordinate, replicate_3

description:
Library name: A3MS
10x Genomics


title: Mouse High-cholesterol+TMAO 1

description:
Library name: High-cholesterol+TMAO-1
10X Genomics


title: None_24h_2

description: AK Yolksac

value: 24 hours


title: adultMaleThalamus, subordinate, replicate_1

description:
Library name: A1MS
10x Genomics


title: Control_PBS_rep3

description: Library name: Control7

value: anterior cingulate cortex


title: αSyn PFF injection_rep3

description: Library name: PFFA2748

value: humanized aSyn WT mice


title: Water_24h_3

description: AK Yolksac


title: IP_IgG

description: Library name: MM24

tag: cell type

value: mouse melanoma cells

value: WT


title: Water_24h_2

description: AK Yolksac


title: IP_Z22: 5-AZA-CdR + ADAR1i-124

description: Library name: MM20

value: IP_Z22:5-AZA-CdR + ADAR1i-124


title: adultMaleThalamus, dominant, replicate_1

description:
Library name: A1MD
10x Genomics


title: Mouse Chow 2

description:
Library name: Chow-2
10X Genomics

value: Chow

value: Ldlr-/- on the C57BL/6J background

value: female

value: 14 weeks


title: Control_PBS_rep2

description: Library name: Control6


title: Control_PBS_rep1

description: Library name: Control5


title: adultMaleThalamus, control, replicate_1

description:
Library name: A1MC
10x Genomics


title: IP_Z22: 5-AZA-CdR

description: Library name: MM17


title: IP_Z22: ADAR1i-124

description: Library name: MM16


title: Whole heart, 14 dpf larvae, ISO 100 uM 2

city: Albacete

first: Aaron

last: Garcia-Blazquez

address: C/ Almansa 14

department: Facultad de Medicina de Albacete

country: Spain

institute: Universidad de Castilla-La Mancha

data_processing:
Raw reads in fastq format were quality processed with a phred score of 30 and adapter sequences were removed using fastp.
Clean reads were aligned to the latest zebrafish genome of reference Ensembl GRCz11 using Hisat2.
Mapped reads were assembled with StringTie.
FeatureCounts was used to obtain gene counts
Assembly: GRCz11
Supplementary files format and content: Data sheet file include raw counts for each sample

organism: Danio rerio

source_name: Whole heart

extract_protocol:
Total RNA was isolated from pools of approximately 40 dissected larval hearts at 14 dpf using the NZY Total RNA isolation kit (Nzytech) following the manufacturer’s instructions
Library preparation and quality control was performed by Novogene, followed by paired-end sequencing (2 x 150 bp) performed with Novaseq X Plus (Illumina, USA). Each pool was sequenced at an average depth of 40 million reads.

treatment_protocol: For chronic treatment experiments, larvae were exposed to 100 µM ISO from 3 up to 14 dpf.

value: Whole heart

value: mitfaw2/w2; mpv17a9/a9

value: Isoproterenol 100 uM


title: Whole heart, 14 dpf larvae, ISO 100 uM 1


title: midAna PHFDPCs in October, male (bio rep 2)

city: Shenyang

first: Huiling

last: Xue

address: 120 Dongling Road,110866 Shenyang,China

institute: Shenyang Agricultural University

description: Library name: Sample 8

data_processing:
Sequenced reads were trimmed for adaptor sequence, and mapped to goat (Capra hircus) transcriptomes (GCF_001704415.1_ARS1) using Kallisto (v.0.46.1) with parameters -b 100. The gene abundances were obtained by Kallisto-Sleuth (v.0.30.0) pipeline in TPM (transcripts per million) (Bray NL et al. 2016).
Assembly: GCF_001704415.1_ARS1
Supplementary files format and content: tab-delimited text files include TPM values for each Sample.

organism: Capra hircus

source_name: Primary hair follicle dermal papilla cells from goat skin

extract_protocol:
Total RNA was extracted from the hair follicle dermal papilla cells using RNAiso Plus (TRIzol method, TaKaRa Bio, Dalian, China).
Paired-end sequencing of the cDNA library construction for transcriptome sequencing was performed according to the operating instructions for the NEB Next® Ultra™ RNA Library Prep Kit for Illumina® (NEB, USA).

value: male


title: IP_Z22: DMSO

description: Library name: MM15


title: midAna PHFDPCs in October, female (bio rep 1)

description: Library name: Sample 7

value: Primary hair follicle dermal papilla cells from goat skin


title: Whole heart, 14 dpf larvae, control 2

value: Control


title: IP_J2: 5-AZA-CdR + ADAR1i-124

description: Library name: MM11

value: IP_J2:5-AZA-CdR + ADAR1i-124


title: Whole heart, 14 dpf larvae, control 1


title: midAna SHFDPCs in October, male (bio rep 2)

description: Library name: Sample 6

source_name: Secondary hair follicle dermal papilla cells from goat skin


title: Esr1KOM

city: Seattle

first: Koichi

last: Hashikawa

state: WA

address: 1959 NE Pacific St. HSB J135

institute: University of Washington

description: 10x genomics V2 platform, MPOA tissue, P50 male, Esr1 deleted

data_processing:
the alignment of raw sequencing reads to the mouse genome was conducted using the 10x Genomics Cell Ranger pipeline (V3) to obtain cell by gene-count matrices for subsequent downstream analysis.
Genome_build: mm10
Supplementary_files_format_and_content: tsv files containing barcode, features and matrix.
Supplementary_files_format_and_content: rds files are Seurat objects (R object).

source_name: P50 male, Esr1KO

extract_protocol:
Male and female wild-type mice (n=3-4 per group) at P23 ± 1 (prepuberty group, hormonally intact), P35 ± 1 (mid-puberty group, hormonally intact), and P50 ± 2 (hormonally intact early adult or gonadectomy groups. Mice in gonadectomy groups were deprived of their gonads at P22-23) were singly housed 1-2 days prior to tissue dissection. We also prepared male and female Vgat-Flp :: Esr1lox/lox mice, which were bilaterally injected with 300 nl of AAV-frtFlex-splitCre at P14-18 in the MPOA, and their brain tissues were collected at P50 ± 2 (early adult) (n=3-4 per group). Subsequent steps largely followed previously published procedures (Hashikawa et al., 2020). Immediately after mice were removed from their home cage, they were deeply anesthetized with intraperitoneal injection of 0.2 mL of stock solution containing sodium pentobarbital (39 mg/mL) and phenytoin sodium (5mg/mL) and transcardially perfused with ice-old NMDG-aCSF containing inhibitor cocktails which was used throughout procedures unless noted. NMDG-aCSF: 96 mM NMDG, 2.5 mM KCl, 1.35 mM NaH2PO4, 30 mM NaHCO3, 20 mM HEPES, 25 mM glucose, 2 mM thiourea, 5 mM Na+ ascorbate, 3 mM Na+ pyruvate, 0.6 mM glutathione-ethyl-ester, 2 mM N-acetyl-cysteine, 0.5 mM CaCl2, 10 mM MgSO4; pH 7.35–7.40, 300-305 mOsm, oxygenated with 95% O2 and 5% CO2. Inhibitor cocktails: 500 nM TTX, 10 μM APV, 10 μM DNQX, 5 µM actinomycin, 37.7 µM anysomycin. The aCSF used for perfusion also contained transcription and translation inhibitors to minimize potential transcriptional evens introduced by pentobarbital injection, perfusion and brain extraction. Brains were extracted and coronal sections containing the MPOA were prepared at 300 µm using a vibratome (Leica, VT1200). Slices including the MPOA (3-4 slices/animal) were recovered in a chamber for 30 min on ice. MPOA tissues were then dissected using a micro scalpel (FEATHER) under the light microscope (LEICA, MZFL) from 3-4 animals/group (10-15 tissues/group) pooled and enzymatically digested with 1 mg/mL pronase (Sigma-Aldrich) for 30 min at room temperature, followed by mechanical trituration with fire-polished glass capillaries (tip diameter 200-300 µm) and filtered through strainers (pore size 40 µm) twice to remove cellular aggregates. Dead and dying cells in the cell suspension were removed using a dead cell removal kit (Miltenyi Biotec). After centrifugation, removal of supernatant and resuspension (20-30 ml), a fraction of cells (~5 ml) were mixed with trypan blue to manually determine the proportion of live cells and cell concentrations using a hemocytometer. Only samples of high viability (>80%) were used for subsequent cDNA library preparation. Final cell concentration was adjusted to 800-1,000 cells/µL. We typically collected 30,000-80,000 cells in total from 3-4 mice, which well exceeded the number of cells required for cDNA library generation, so that biological variability between subjects was reduced.
cDNA libraries were constructed following manufacture’s instruction (Chromium Single Cell 3’ Reagents Kits V2 User Guide, 10x Genomics). Briefly, to recover up to 10,000 single cells, ~17,000 dissociated cells were mixed with reverse transcription mix and loaded into the chip. The mRNAs of single cells were captured by barcoded beads in the droplets using a Chromium controller. Reverse transcribed cDNAs were then PCR amplified, fragmented, and ligated with adaptors followed by sample index PCR.  cDNA libraries were sequenced on an Illumina Nextseq 500 (v2.5) or Hiseq and the alignment of raw sequencing reads to the mouse genome was conducted using the 10x Genomics Cell Ranger pipeline (V3) to obtain cell by gene-count matrices for subsequent downstream analysis.

value: brain

value: all cells in the MPOA


title: OVX

description: 10x genomics V2 platform, MPOA tissue, overiectomized female

source_name: P50 female, overiectomized

value: overiectomized


title: P23F

description: 10x genomics V2 platform, MPOA tissue, P23 female

source_name: P23 female


title: Esr1KOF

description: 10x genomics V2 platform, MPOA tissue, P50 female , Esr1 deleted

source_name: P50 female, Esr1KO


title: IP_J2: 5-AZA-CdR

description: Library name: MM08

value: mouse melanoma cells


title: midAna SHFDPCs in October, female (bio rep 1)

description: Library name: Sample 5

value: Secondary hair follicle dermal papilla cells from goat skin


title: Sta phase_RNA-seq_rep3

data_processing:
Basecalling was performed by RTA 1.18.
Adaptor and barcode were removed by cutadapt 1.18.
RNA-seq reads were mapped to the genome by Bowtie2 2.2.8.
For RNA-seq, the read coverage was counted by bedtools 2.27.1 and the raw counts of aligned reads in specific regions were calculated by HTseq 0.11.2.
Assembly: Escherichia coli str. K-12 substr. MG1655, NC_000913.2
Supplementary files format and content: txt files, the scores represent the total reads count in each gene.

value: beta prime-WT

value: W3110 rpoC-6xHis


title: P35F

description: 10x genomics V2 platform, MPOA tissue, P35 female

source_name: P35 female

value: WT


title: IP_J2: ADAR1i-124

description: Library name: MM07


title: Sta phase_RNA-seq_rep2


title: P50F

description: 10x genomics V2 platform, MPOA tissue, P50 female

source_name: P50 female

value: female


title: earlyAna PHFDPCs in June, male (bio rep 2)

description: Library name: Sample 4

value: Primary hair follicle dermal papilla cells


title: IP_J2: DMSO

description: Library name: MM06


title: earlyAna PHFDPCs in June, female (bio rep 1)

description: Library name: Sample 3


title: Cast

description: 10x genomics V2 platform, MPOA tissue, Castrated male

source_name: P50 male, castrated

value: castrated


title: earlyAna SHFDPCs in June, male (bio rep 2)

description: Library name: Sample 2

value: Secondary hair follicle dermal papilla cells


title: 5-AZA-CdR

description: Library name: MM25


title: earlyAna SHFDPCs in June, female (bio rep 1)

description: Library name: Sample 1


title: ADAR1i-124

description: Library name: MM27


title: DMSO

description: Library name: MM01

value: DMSO


title: Log phase_RNA-seq_rep3

source_name: beta prime-WT


title: Log phase_RNA-seq_rep2


title: P35M

description: 10x genomics V2 platform, MPOA tissue, P35 male

source_name: P35 male

value: male


title: P23M

description: 10x genomics V2 platform, MPOA tissue, P23 male

source_name: P23 male


title: NAIO3

city: Guangzhou

first: Tian

last: Zhou

state: Guangdong

address: NO.7 Jinsui Road

department: State Key Laboratory of Ophthalmology

institute: Zhongshan Ophthalmic Center

description: single cell suspension

data_processing:
10X Genomics Cellranger 3.0.2
Assembly: mm10
Supplementary files format and content: These output files of each sample are in standard output file formats, created by the 10X Genomics Cellranger pipeline, compatible with any downstream single-cell RNA analysis pipeline

source_name: RPE-choroidal complex

extract_protocol:
Eyes of 6 mice per group from PBS or NaIO3 treated C57BL/6J mice were collected and digested in 2% dispase II for 30 minutes in 37℃. Then the anterior structures with neural retina were removed. The RPE choroidal cells were shed off by gently patting on the back of the sclera. All the hexagonal brownish cells were collected in PBS under the microscopy and were resuspended in PBS. Single cells were processed on 10X genomics platform, and Illumina HiSeq X Ten was used for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: the mice were injected intravenously with 20 mg/kg of NaIO3 (Sigma-Aldrich Corp., St. Louis, MO, USA), which was dissolved in phosphate-buffered saline (PBS). The mice receiving the same volume of PBS worked as controls.

value: RPE-choroidal complex

value: RPE-choroidal cells

value: NAIO3 treatment for 7 days


title: PBS

description: single cell suspension

value: PBS treatment for 7 days


title: CML shOxr1+alum-5

status: Public on Nov 12 2025

city: Chicago

first: Priyam

last: Patel

state: IL

address: 750 North Lake Shore Drive, Rubloff 11-146

department: Quantitative Data Science Core

institute: Northwestern University

data_processing:
The quality of reads, in FASTQ format, was evaluated using FastQC (Andrews S, 2010) version 0.11.7.
Reads were trimmed to remove Illumina adapters from the 3’ ends using cutadapt version 1.14 (Martin, 2011) with a minimum Q score cutoff of 25.
Trimmed reads were aligned to the amplicon sequence using the Burrows Wheeler Aligner (BWA version 0.7.12) (Li H et al., 2009).
Resulting alignment files in BAM format were cleaned, sorted, and marked for duplicates using the Picard Tools version 1.85 CleanSam, SortSam, and Mark Duplicates, respectively.
Variants were detected from the processed BAM files using BCFtools (Li H et al., 2011) mpileup with minimum base quality and maximum depth parameters set to 20 and 8000, respectively.
Assembly: Transcript: ENST00000318560.6 ABL1-201 (cgagttggttcatcatcatt caacggtggc cgacgggctc atcaccacgc tccattatcc agccccaaagcgcaacaagc ccactgtcta tggtgtgtcc cccaactacg acaagtggga gatggaacgcacggacatca ccatgaagca caagctgggc gggggccagt acggggaggt gtacgagggcgtgtggaaga aatacagcct gacggtggcc gtgaagacct tgaaggagga caccatggaggtggaagagt tcttgaaaga agctgcagtc atgaaagaga tcaaacaccc taacctggtgcagctccttg gggtctgcac ccgggagccc ccgttctata tcatcactga gttcatgacctacgggaacc tcctggacta cctgagggag tgcaaccggc aggaggtgaa cgccgtggtgctgctgtaca tggccactca gatctcgtca gccatggagt acctggagaa gaaaaacttcatccacagag atcttgctgc ccgaaactgc ctggtagggg agaaccactt ggtgaaggtagctgattttg gcctgagcag gttgatgaca ggggacacct acacagccca tgctggagccaagttcccca tcaaatggac tgcacccgag agcctggcct acaacaagtt ctccatcaagtccgacgtct gggcatttgg agtattgctt tgggaaattg ctacctatgg catgtccccttacccgggaa ttgacctgtc ccaggtgtat gagctgctag agaaggacta ccgcatggagcgcccagaag gctgcccaga gaaggtctat gaactcatgc gagcatgttg gcagtggaatccctctgacc ggccctcctt tgctgaaatc caccaagcct ttgaaacaat gttccaggaatccagtatct cagacgaagt gga)
Supplementary files format and content: VCF Fiiles

source_name: Bone marrow

growth_protocol: mBMCs from secondary transplanted CML mice, which had been transduced with either shOxr1 (Oxr1 knockdown) or shControl

extract_protocol:
RNA was isolated from the transplanted mBMCs using the RNeasy plus micro kit (Qiagen 74034). Nested PCR was performed to enrich the bcr-abl kinase domain (KD) with bcr-abl specific primer set 1 (bcr-F: ACAGCATTCCGC TGACCATCAATAAG; abl-R1: ATGGTCCAGAGGATCGCTCTCT) and abl kinase domain primer set 2 (abl-F:  TGGTTCATCATCATTCAACGGTGG; abl-R2: TCCACTTCGTCTGAGATACTGGATT).
The library was constructed with Nextera XT DNA Library Prep kit( Illumina, FC-131-1024  ) and purified with SPRIselect beads (Beckman Coulter, A66514).
The multiplexed libraries were loaded for next-generation sequencing (NGS) on Illumina HiSeq 4000 using single-ended 50bp mode.

treatment_protocol: Mice were treated with imatinib, and subjected to 2-3 cycles of alum or control saline injections

value: Bone marrow

value: BMCs

value: IM+alum injection


title: CML shOxr1+saline-5


title: CML shOxr1+saline-4


title: CML shOxr1+alum-6


title: CML shOxr1+alum-2


title: Log phase_RNA-seq_rep1


title: CML shOxr1+alum-1


title: CML shOxr1+alum-3


title: CML shOxr1+alum-4


title: P50M

description: 10x genomics V2 platform, MPOA tissue, P50 male

source_name: P50 male

value: brain

value: all cells in the MPOA


title: CML shOxr1+saline-2

value: IM+saline injection


title: CML shOxr1+saline-1


title: CML shControl+alum-5


title: CML shControl+alum-4


title: CML shControl+alum-3


title: CML shControl+alum-2


title: CML shControl+alum-1


title: Sta phase_RNA-seq_rep1

value: beta prime-WT

value: W3110 rpoC-6xHis


title: CML shControl+saline-4


title: CML shControl+saline-1


title: CML shControl+saline-2


title: FF2_133

city: Santa Monica

first: Lucy

middle: K

last: Bicks

address: 1033 3rd street, apartment 106, apartment 106

department: Neurology

institute: UCLA School of Medicine

description: RNA-seq, 100-base-pair paired end reads on an Illumina HiSeq 4000

data_processing:
The reads were mapped to the human genome (hg38) with Gencode v25 annotations using STAR (v.2.5.2b) and gene expression was quantified using rsem (v1.3.0)
Genes that were expressed at very low levels (less that 10 reads in 30% of samples from a given day) were removed from the analysis
Samples with standardized sample network connectivity Z scores below –2 in each mutation were defined as outliers and removed
To control for technical variation due to the sequencing and library prep we calculated the principal components of the Picard sequencing metrics (http://broadinstitute.github.io/picard/) using the CollectAlignmentSummaryMetrics, CollectRnaSeqMetrics, and MarkDuplicates modules and included them in our model
To infer genetic ancestry, we called SNPs from the aligned reads using the GATK (v3.3) Haplotype caller. Sites with more than 5% missing samples, with rare minor allele frequency (less than 0.05) and Hardy–Weinberg disequilibrium (less than 1 × 10–6) were removed using plink (v1.09). We then used the remaining high-quality SNPs to run MDS together with HapMap3.3 (hg38). The first two MDS values, referred to as genetic ancestry PC1/2, were then included in our model.
Assembly: (hg38) with Gencode v25
Supplementary files format and content: counts

source_name: cortical organoid

extract_protocol:
ribosomal RNA depleted (RiboZero Gold, Illumina)
Truseq stranded RNA RiboZero Gold (Illumina)

value: cortical organoid

tag: source tissue

value: fibroblast

tag: differentiation day

tag: differentiation


title: FF2_132

tag: genotype


title: FF2_131


title: IBD

city: Seoul

first: Mi-Kyung

last: Oh

address: 1 Gwanak-ro, Gwanak-gu

department: Department of Agricultural Biotechnology and Research Institute of Agriculture and Life Sciences

country: South Korea

institute: Seoul National University

description:
Library name: TD
10x Genomics

data_processing:
The demultiplexing, barcoded processing,gene counting, and aggregation were made using the Cell Ranger (v6.1.1) pipeline
Assembly: GRCh38 human reference genome (Ensemble version 113).
Supplementary files format and content: tab-separated genes,barcodes, and matrix files per sample

source_name: CD4+ T cells

extract_protocol:
Human CD4⁺ T cells were isolated from spleen, and colon tissues of humanized mice. Tissues were dissociated, passed through a 70 μm cell strainer, and red blood cells were lysed using RBC lysis buffer (eBioscience). CD4⁺ T cells were purified by MACS (Miltenyi Biotec) according to the manufacturer’s protocol.
Use LUNA-FL™ Automated Fluorescence Cell Counter (logos biosystems) to consult the 10x Genomics Single Cell Protocols Cell Preparation Guide and the Guidelines for Optimal Sample Preparation flowchart (Documents CG00053 and CG000126 respectively) for more information on preparing cells. Libraries were prepared using the Chromium controller according to the 10x chromium Next GEM Single Cell 3’ v3.1 protocol(CG000315). Briefly, the cell suspensions were diluted in nuclease free  water to achieve a targeted cell count of 15,000. Cell suspension was mixed with master mix and  loaded with Single Cell 3’V3.1 Gel Beads and Partitioning Oil into a chromium Next GEM chip G. RNA transcripts from single cells were uniquely barcoded and reverse transcribed within droplets. cDNA molecules were pooled and the cDNA pool then go through an end repair process, the  addition of a single ‘A’ base, and then ligation of the adapters. The products are then purified andenriched with PCR to create the final cDNA library. The purified libraries were quantified using    qPCR according to the qPCR Quantification Protocol Guide(KAPA) and qualified using the Agilent Technologies 4200 TapeStation . And then the libraries were sequenced using NovaSeq platform (Illumina) according to the read length in the user guide.

treatment_protocol: Enginered Evs from WJ-MSCs

value: CD4+ T cells


title: Pool-3-TGACCAAT


title: Dup_73


title: CML shOxr1+saline-3


title: FF2_125


title: FF2_119


title: FF2_116


title: FF2_127


title: FF2_128


title: FF2_120


title: FF2_126


title: FF2_123


title: FF2_105


title: FF2_124


title: FF2_114


title: FF2_117


title: FF2_113


title: FF2_115


title: FF2_118


title: FF2_109


title: FF2_111


title: CML shControl+saline-3


title: FF2_121


title: FF2_112


title: FF2_110


title: FF2_106


title: IBD+EV

description:
Library name: TDE
10x Genomics


title: FF2_107


title: FF2_103


title: FF2_101


title: FF2_102


title: FF2_99


title: FF2_100


title: FF2_98


title: FF2_93


title: FF2_97


title: FF2_96


title: FF2_91


title: FF2_89


title: FF2_95


title: FF2_90


title: FF5_7

value: Control


title: FF2_30


title: FF5_8


title: FF5_1


title: FF2_32


title: FF2_92


title: Pool-6-CGATGTAT


title: FF5_5


title: Pool-2-GATCAGAT


title: FF2_29


title: FF2_37


title: FF2_33


title: FF2_31


title: FF5_6


title: FF5_2


title: FF2_39


title: FF2_104


title: Pool-2-ATTCCTTT


title: FF2_94


title: FF5_4


title: FF2_40


title: FF2_34


title: FF2_35


title: FF2_36


title: FF5_3


title: Pool-4-ATCACGAT


title: FF2_24


title: Pool-3-CGATGTAT


title: FF2_27


title: FF2_28


title: FF2_26


title: FF2_21


title: Dup_17


title: FF2_15


title: FF2_16


title: FF2_17


title: Pool-1-AGTCAACA


title: FF2_20


title: FF2_23


title: FF2_13


title: FF2_18


title: Pool-6-ATCACGAT


title: Dup_19


title: Dup_18


title: FF2_38


title: Dup_24


title: Dup_22


title: FF2_14


title: Pool-4-GAGTGGAT


title: Pool-2-TGACCAAT


title: Pool-4-GTGAAACG


title: FF2_25


title: FF2_22


title: Pool-2-TTAGGCAT


title: Pool-4-TTAGGCAT


title: Pool-4-ATTCCTTT


title: Pool-4-ACTTGAAT


title: Pool-4-GTCCGCAC


title: Pool-2-CCGTCCCG


title: FF2_136


title: FF2_135


title: FF2_134


title: FF5_23


title: FF5_28


title: FF5_25


title: FF5_21


title: FF2_19


title: FF4_84


title: FF5_24


title: FF4_18


title: FF4_81


title: FF5_22


title: Dup_71


title: FF4_17


title: FF4_78


title: FF4_12


title: Dup_70


title: FF4_46


title: Pool-6-GATCAGAT


title: FF5_74


title: Dup_67


title: FF4_61


title: FF5_76


title: FF5_81


title: Pool-6-TGACCAAT


title: Pool-1-TTAGGCAT


title: FF5_27


title: FF5_26


title: FF5_104


title: FF5_105


title: FF5_80


title: FF5_77


title: FF5_107


title: FF5_101


title: FF5_100


title: FF5_106


title: FF5_103


title: FF5_82


title: FF5_83


title: FF3_101

value: PBMC


title: FF3_99


title: FF5_75


title: FF3_97


title: FF3_95


title: FF3_100


title: FF3_102


title: FF3_96


title: FF3_91


title: FF3_92


title: FF3_88


title: FF3_87


title: FF5_78


title: FF3_89


title: FF3_93


title: FF3_94


title: FF5_113


title: FF5_112


title: FF5_108


title: FF5_114


title: FF5_109


title: FF5_102


title: FF5_111


title: FF5_115


title: FF3_131


title: FF3_136


title: FF3_98


title: FF3_137


title: FF3_135


title: FF3_138


title: FF3_133


title: FF3_130


title: FF3_126


title: FF3_134


title: FF3_124


title: FF3_127


title: FF3_123


title: FF3_128


title: FF3_113


title: FF3_125


title: FF5_110


title: FF3_112


title: FF3_129


title: FF3_114


title: FF3_109


title: FF3_90


title: FF3_110


title: FF5_99


title: FF3_108


title: FF5_97


title: FF3_132


title: FF5_93


title: FF5_92


title: FF5_95


title: FF5_96


title: FF5_94


title: FF5_98


title: FF3_85


title: FF3_86


title: FF3_84


title: FF3_83


title: FF5_85


title: FF5_84


title: FF5_87


title: FF3_106


title: FF3_105


title: Pool-1-GTGAAACG


title: FF3_107


title: FF3_104


title: FF5_91


title: Pool-5-CTTGTAAT


title: Dup_57


title: FF5_89


title: Pool-3-GTGGCCTT


title: Pool-1-TGACCAAT


title: Pool-2-ATGTCAGA


title: Pool-2-AGTTCCGT


title: Pool-5-AGTCAACA


title: FF5_90


title: Pool-3-CGTACGTA


title: Pool-5-GATCAGAT


title: Pool-2-AGTCAACA


title: Pool-4-CGATGTAT


title: Dup_46


title: Pool-2-ACTGATAT


title: Pool-5-ACAGTGAT


title: Pool-2-ATCACGAT

value: TS


title: Pool-1-TAGCTTAT


title: FF5_86


title: FF3_7


title: FF3_8


title: FF2_9


title: FF2_10


title: Pool-5-TAGCTTAT


title: Pool-5-AGTTCCGT


title: FF3_5


title: FF2_12


title: FF3_6


title: Pool-3-TTAGGCAT


title: Dup_7


title: Pool-1-CAGATCAT


title: Pool-1-ACAGTGAT


title: Pool-2-CAGATCAT


title: FF2_11


title: Pool-4-CAGATCAT


title: Pool-4-GCCAATAT


title: FF2_141


title: Pool-2-GTTTCGGA


title: Pool-6-ACAGTGAT


title: FF2_139


title: Pool-5-ACTTGAAT


title: FF2_142


title: FF2_140


title: Pool-3-GTCCGCAC


title: Pool-1-ACTGATAT


title: Pool-4-GTTTCGGA


title: Dup_86


title: FF2_137


title: Pool-4-CCGTCCCG


title: FF2_138


title: Pool-1-GTTTCGGA


title: FF4_22


title: IgG_SV_HFD_1

city: Kuopio

first: Juho

middle: Mikael

last: Mononen

address: Yliopistonranta 1E

department: Faculty of Health Sciences

country: Finland

institute: University of Eastern Finland

data_processing:
Reads were then trimmed with Trimmomatic (v0.36) using long TruSeq3-SE.v2 adapters and as essential settings: ILLUMINACLIP:0:30:10, LEADING:3, TRAILING:3, MINLEN:35.
Alignment was done using to strain-specific genomes, generated with g2gtools (v0.2.9) and MGPv5 SNPs and InDels, using STAR (v2.6.1) with ‘--alignIntronMax 1 --alignEndsType "EndToEnd"’.
Reads were filtered for true pairs and MAPQ>20 with Samtools (v1.9) and duplicates were removed using Picard (v2.8.13) MarkDuplicates.
Peaks were called with MACS2 (v2.2.7.1) using ‘-g mm -B --call-summits --keep-dup auto -q 0.05’.
Assembly: mm10
Supplementary files format and content: Sample-wise peaks: H3K27ac summits were widened to peaks with Bedtools (v2.27.1) using ‘slop -b 75’. Peaks were shifted to mm10 coordinates using g2gtools. ENCODE blacklisted regions were removed from the regions.
Supplementary files format and content: BigWigs: BAM files shifted to mm10 coordinates were downsampled after and merged by group using Samtools. BigWigs were generated using Deeptools (v3.4.2) bamCoverage with '--normalizeUsing CPM --outFileFormat bigwig --binSize 10'.

source_name: eWAT

extract_protocol:
Frozen tissue was Dounce homogenized and fixed with 20 strokes using the loose pestle A followed by 20 strokes with the tight pestle B in Cell Lysis Buffer (5 mM PIPES pH 8, 85 mM KCl, 1% Igepal, 1× protease inhibitor) containing 0.5% formaldehyde. The mixture was rotated for 8 minutes at room temperature. The cross-linking reaction was quenched by adding 150 mM glycine for 5 minutes at room temperature. Samples were washed twice with ice-cold Cell Lysis Buffer supplemented with 1× protease inhibitors. The pellet was suspended in 300 µl of SDS Lysis Buffer (50 mM Tris-Cl, 0.5% SDS, 10 mM EDTA, 1× protease inhibitor) and incubated on ice for 1 hour. Chromatin was sheared using a Bioruptor Plus sonicator (Diagenode) for 35 cycles at a high setting (30s ON, 30s OFF per cycle), allowing the sonicator to cool down for 5 minutes after every 10 cycles. Triton-X-100 was added to a final concentration of 1% along with 1× protease inhibitors. Samples were centrifuged at 13,000 rpm for 20 minutes, and the supernatant was collected for immunoprecipitation.
ChIP and library preparation were performed using the recently described ChIPmentation protocol (Gustafsson C et al. BMC Genomics. 2019;20(1):59) with minor modifications. For ChIP, 2 µg of rabbit polyclonal anti-CTCF antibody, 5µl of rabbit monoclonal anti-TCF4/TCF7L2 antibody or 2 µg of Anti-Histone H3 (acetyl K27) antibody was added to 50 µl Protein G-coupled Dynabeads (Thermo Fisher Scientific) in 1 × PBS with 0.5% bovine serum albumin (BSA) and rotated at 40 rpm for 4 h at 4°C. Dynabeads were washed three times with PBS with 0.5% BSA and then mixed with 190 µl chromatin samples in 1.5 ml tubes. The samples were rotated at 40 rpm overnight at 4°C. Chromatin was washed with 150 µl of low-salt buffer (50 mM Tris/HCl, 150 mM NaCl, 0.1% SDS, 0.1% sodium deoxycholate, 1% Triton X-100, and 1 mM EDTA), high-salt buffer (50 mM Tris/HCl, 500 mM NaCl, 0.1% SDS, 0.1% sodium deoxycholate, 1% Triton X-100, and 1 mM EDTA) and LiCl buffer (10 mM Tris/HCl, 250 mM LiCl, 0.5% IGEPAL CA-630, 0.5% sodium deoxycholate, and 1 mM EDTA), followed by two washes with TE buffer (10 mM Tris/HCl and 1 mM EDTA) and two washes with ice-cold Tris/HCl pH 8. Chromatin was resuspended in 30 µl of 2 × TD buffer and 1 µl of transposase (Nextera, Illumina) for tagmentation. Samples were incubated at 37°C for 10 min followed by two washes with low-salt buffer. Bead bound tagmented chromatin was diluted in 23 µl of water and 25 µl of 2 × Ultra II Q5 Master Mix (New England BioLabs, M0544S) and 1 µl of both amplification primers (Buenrostro et al, 2013; 0.125 µM final concentration) were added. The libraries were incubated as follows: 72 °C 5 min (adapter extension); 95 °C 5 min (reverse cross-linking); followed by 11 cycles of 98 °C 10s, 63 °C 30s and 72 °C 3 min. After PCR amplification, double-sided purification was performed using SPRIselect beads (Beckman Coulter). To prepare input controls, 2 μl of 50 mM MgCl2 was added to 10 μl sonicated lysate to neutralize the EDTA in the SDS lysis buffer. Tagmentation and amplification was done as described above.

value: eWAT

value: IgG (Diagenode, #C15410206)

value: HFD


title: Pool-2-GTCCGCAC


title: IgG_B6_HFD_1


title: IgG_B6_Chow_1

value: Chow


title: IgG_SV_Chow_1


title: FF4_89


title: H3K27ac_SV_HFD_3


title: FF4_93


title: H3K27ac_SV_Chow_3


title: H3K27ac_B6_HFD_2


title: H3K27ac_B6_Chow_3


title: FF4_29


title: Wnt3a_knockout_3

city: Frederick

first: Terry

middle: P

last: Yamaguchi

state: Maryland (MD)

address: Building 560,  1050 Boyles St

department: Cancer and Developmental Biology

institute: NIH/NCI

description:
Library name: Sample 6
Libraries were generated using Illumina TruSeq (FC-122-1001) kit and sequenced on an Illumina HiSeq2000 sequencer using PhiX as sequencing control.

data_processing:
Base calling was performed with the RTA 1.9.35.0 software before raw count matrices were generated from raw fastq files using the RENEE (CCBR.github.io/RENEE/) v2.5 pipeline.
Briefly, following adaptor removal and read quality trimming with Cutadapt v1.18, reads were aligned to GRCm38.p6 with STAR v2.7.6a in two pass mode.
FastQC was used to examine read quality before and after trimming. Picard v2.17.11 and RSeQC v2.6.4 were similarly used to generate quality metrics post-alignment. Kraken v2.0.8-beta and Krona v2.7 were used to determine levels of contamination.
Expected and isoform counts, according to the GENCODE vM25 primary assembly annotation reference, were collected using the RSEM v1.3.0 package.
DESeq2 v1.42.0 was used to conduct differential expression analysis. The clusterProfiler R/Bioconductor package v4.10.0 was used to conduct Gene Ontology term (Biological Processes) over-representation analysis.
Assembly: GRCm38.p6 M25 (mm10): The genome reference files used throughout the analysis were built using the renee build executable from the RENEE pipeline.
Supplementary files format and content: RSEM.genes.results were obtained from the DEG_ALL output folder following analysis using RENEE. These raw counts can be loaded into the DESeq2 pipeline using tximport for downstream analysis.

source_name: Posterior region of E8.25 mouse embryos+2 somites

extract_protocol: Libraries were constructed according to Illumina's Truseq RNA library protocol. About 200-500ng of total RNA was used for mRNA capture using oligo dT coated magnetic beads. After mRNA fragmentation, cDNA synthesis was performed using random primers. This is followed by standard Illumina library preparation using cDNA with end repair, adapter ligation and PCR amplification before sequencing.


title: FF4_96


title: FF4_11


title: FF4_50


title: C57BL6_control_2

description:
Library name: Sample 2
Libraries were generated using Illumina TruSeq (FC-122-1001) kit and sequenced on an Illumina HiSeq2000 sequencer using PhiX as sequencing control.

value: Posterior region of E8.25 mouse embryos+2 somites


title: Wnt3a_knockout_2

description:
Library name: Sample 5
Libraries were generated using Illumina TruSeq (FC-122-1001) kit and sequenced on an Illumina HiSeq2000 sequencer using PhiX as sequencing control.


title: FF4_62


title: C57BL6_control_3

description:
Library name: Sample 3
Libraries were generated using Illumina TruSeq (FC-122-1001) kit and sequenced on an Illumina HiSeq2000 sequencer using PhiX as sequencing control.


title: FF4_71


title: C57BL6_control_1

description:
Library name: Sample 1
Libraries were generated using Illumina TruSeq (FC-122-1001) kit and sequenced on an Illumina HiSeq2000 sequencer using PhiX as sequencing control.


title: FF4_88


title: Sp5Sp8_knockout_3

description: Libraries were generated using Illumina TruSeq (FC-122-1001) kit and sequenced on an Illumina HiSeq2000 sequencer using PhiX as sequencing control.

data_processing:
Base calling was performed with the RTA 1.9.35.0 software before raw count matrices were generated from raw fastq files using the RENEE (CCBR.github.io/RENEE/) v2.5 pipeline.
Briefly, following adaptor removal and read quality trimming with Cutadapt v1.18, reads were aligned to GRCm38.p6 with STAR v2.7.6a in two pass mode.
FastQC was used to examine read quality before and after trimming. Picard v2.17.11 and  RSeQC v2.6.4 were similarly used to generate quality metrics post-alignment.  Kraken v2.0.8-beta and Krona v2.7 were used to determine levels of contamination.
Expected and isoform counts, according to the GENCODE vM25 primary assembly annotation reference, were collected using the RSEM v1.3.0 package.
DESeq2 v1.42.0 was used to conduct differential expression analysis. The clusterProfiler R/Bioconductor package v4.10.0 was used to conduct Gene Ontology term (Biological Processes) over-representation analysis.
Assembly: GRCm38.p6 M25 (mm10): The genome reference files used throughout the analysis were built using the renee build executable from the RENEE pipeline.
Supplementary files format and content: Sp58_KO_main_count_matrix.csv contains the count data loaded in using the tximport function and generated using the write.csv function in R.

extract_protocol: Libraries were constructed according to Illumina's Truseq RNA library protocol. About 200-500ng of total RNA was used for mRNA capture using oligo dT coated magnetic beads.  After mRNA fragmentation, cDNA synthesis was performed using random primers.  This is followed by standard Illumina library preparation using cDNA with end repair, adapter ligation and PCR amplification before sequencing.

value: Sp5L/L; Sp8d/d


title: FF4_94


title: Sp5Sp8_knockout_2


title: FF4_36


title: FF4_51


title: FF4_33


title: Sp5Sp8_knockout_1


title: Long TFR-patient 2 TKI-D

description: Library name: Sample10

data_processing:
The quality of reads, in FASTQ format, was evaluated using FastQC (Andrews S, 2010) version 0.11.7.
Reads were trimmed to remove Illumina adapters from the 3’ ends using cutadapt version 1.14 (Martin, 2011) with a minimum Q score cutoff of 25.
Trimmed reads were aligned to the amplicon sequence using the Burrows Wheeler Aligner (BWA version 0.7.12) (Li H et al., 2009).
Resulting alignment files in BAM format were cleaned, sorted, and marked for duplicates using the Picard Tools version 1.85 CleanSam, SortSam, and Mark Duplicates, respectively.
Variants were detected from the processed BAM files using BCFtools (Li H et al., 2011) mpileup with minimum base quality and maximum depth parameters set to 20 and 8000, respectively.
Assembly: Transcript: ENST00000318560.6 ABL1-201 (cgagttggttcatcatcatt caacggtggc cgacgggctc atcaccacgc tccattatcc agccccaaagcgcaacaagc ccactgtcta tggtgtgtcc cccaactacg acaagtggga gatggaacgcacggacatca ccatgaagca caagctgggc gggggccagt acggggaggt gtacgagggcgtgtggaaga aatacagcct gacggtggcc gtgaagacct tgaaggagga caccatggaggtggaagagt tcttgaaaga agctgcagtc atgaaagaga tcaaacaccc taacctggtgcagctccttg gggtctgcac ccgggagccc ccgttctata tcatcactga gttcatgacctacgggaacc tcctggacta cctgagggag tgcaaccggc aggaggtgaa cgccgtggtgctgctgtaca tggccactca gatctcgtca gccatggagt acctggagaa gaaaaacttcatccacagag atcttgctgc ccgaaactgc ctggtagggg agaaccactt ggtgaaggtagctgattttg gcctgagcag gttgatgaca ggggacacct acacagccca tgctggagccaagttcccca tcaaatggac tgcacccgag agcctggcct acaacaagtt ctccatcaagtccgacgtct gggcatttgg agtattgctt tgggaaattg ctacctatgg catgtccccttacccgggaa ttgacctgtc ccaggtgtat gagctgctag agaaggacta ccgcatggagcgcccagaag gctgcccaga gaaggtctat gaactcatgc gagcatgttg gcagtggaatccctctgacc ggccctcctt tgctgaaatc caccaagcct ttgaaacaat gttccaggaatccagtatct cagacgaagt gga)
Supplementary files format and content: VCF Files; variants in ENST00000318560.6 ABL1-201 transcript in each sample

source_name: Peripheral blood

growth_protocol: Peripheral blood collected from CML patients at or post-TKI discontinuation; mononuclear cells isolated byFicoll-Paque PLUS (GE Healthcare) density gradient centrifugation.

extract_protocol:
RNA was isolated from the patients' PBMCs using the RNeasy plus micro kit (Qiagen 74034). Nested PCR was performed to enrich the bcr-abl kinase domain (KD) with bcr-abl specific primer set 1 (bcr-F: ACAGCATTCCGC TGACCATCAATAAG; abl-R1: ATGGTCCAGAGGATCGCTCTCT) and abl kinase domain primer set 2 (abl-F:  TGGTTCATCATCATTCAACGGTGG; abl-R2: TCCACTTCGTCTGAGATACTGGATT).
The library was constructed with Nextera XT DNA Library Prep kit( Illumina, FC-131-1024  ) and purified with SPRIselect beads (Beckman Coulter, A66514). The multiplexed libraries were loaded for next-generation sequencing (NGS) on Illumina HiSeq 4000 using single-ended 50bp mode.

treatment_protocol: TKI discontinuation per ELN 2020

value: Peripheral blood

tag: subject group/id

value: Long TFR-patient 2

value: PBMCs

value: TKI discontinuation

value: TKI-D


title: Long TFR-patient 3 TKI-D

description: Library name: Sample11

value: Long TFR-patient 3


title: FF4_55

value: cortical organoid

value: fibroblast


title: Long TFR-patient 1 TKI-D

description: Library name: Sample9

value: Long TFR-patient 1


title: FF4_10


title: FF4_42


title: FF4_83


title: FF2_78


title: Short TFR-patient 4 TKI-D

description: Library name: Sample7

value: Short TFR-patient 4


title: Short TFR-patient 3 TKI-D

description: Library name: Sample5

value: Short TFR-patient 3


title: Wnt3a_knockout_1

description:
Library name: Sample 4
Libraries were generated using Illumina TruSeq (FC-122-1001) kit and sequenced on an Illumina HiSeq2000 sequencer using PhiX as sequencing control.


title: FF2_77


title: Short TFR-patient 3 Relapse post-DC

description: Library name: Sample6

value: Relapse post-DC


title: Short TFR-patient 4 Relapse post-DC

description: Library name: Sample8


title: Short TFR-patient 2 Relapse post-DC

description: Library name: Sample4

value: Short TFR-patient 2


title: Short TFR-patient 2 TKI-D

description: Library name: Sample3


title: FF2_80


title: FF2_73


title: FF2_74


title: Short TFR-patient 1 TKI-D

description: Library name: Sample1

value: Short TFR-patient 1


title: Short TFR-patient 1 Relapse post-DC

description: Library name: Sample2


title: FF2_79


title: Hypoxia_BMI1_3

description: Library name: Hyp_BMI_3

data_processing:
cutadapt
HISAT2
featureCounts
Assembly: GRCh38
Supplementary files format and content: tab-delimited text file with raw count from CD4 cells
Supplementary files format and content: tab-delimited text file with raw count from MSCs

source_name: WJ-MSCs

extract_protocol:
Trizol
Libraries were generated using the Illumina TruSeq strand mRNA sample preparation kit (Illumina, San Diego, CA, USA) and sequenced on a NovaSeq 6000 (2 x 150 paired end sequencing, Illumina) according to the manufacturer's protocol.

value: WJ-MSCs


title: Hypoxia_BMI1_2

description: Library name: Hyp_BMI_2


title: FF2_54


title: FF4_60


title: Normoxia_GFP_3

description: Library name: Nor_GFP_3


title: Hypoxia_BMI1_1

description: Library name: Hyp_BMI_1


title: FF2_56


title: FF2_53


title: Normoxia_GFP_2

description: Library name: Nor_GFP_2


title: Normoxia_GFP_1

description: Library name: Nor_GFP_1


title: FF4_67


title: FF4_85


title: TDE_1

description: Library name: TDE_1

source_name: CD4 cells

value: CD4 cells


title: TDE_2

description: Library name: TDE_2


title: FF2_55


title: FF4_79


title: FF2_75


title: Penicillium oxalicum, ∆PoxKu70, Glucose, 24 h, rep3

city: Nanning

first: Di

last: Tian

address: DaXue east road

institute: College of Life Science and Technology,GuangXi university

data_processing:
Data filtering: The raw data was filtered by software SOAPnuke v1.5.2 to generate clean data,clean reads were obtained and stored in FASTQ format
Reference genome mapping: The clean data were mapped to the reference genome by HISAT (v2.1.0)
Reference gene mapping: The clean data were mapped to the assembled unique gene by Bowtie2 (v2.2.5) .The expression level of genes was calculated by RSEM (v1.2.8)
Differential Gene Expression: The differentially expressed genes (DEGs) were calculated by DESeq2 tool with |Log2 fold change| ≥ 1 and q-value ≤ 0.05 as thresholds
Assembly: P. oxalicum HP7-1 genome (accession number JRVD00000000)
Supplementary files format and content: tab-delimited text files include FPKM values for each sample

organism: Penicillium oxalicum

source_name: Mycelium

extract_protocol:
RNA was extracted with Trizol,  the purity of RNA (OD260/280) was detected by Nanodrop2000. The integrity of RNA were accurately detected by Agilent 2100,the detection indicators include: RIN value, 28S/18S, whether the map baseline is raised, 5S peak, and electrophoresis to detect whether the RNA sample is diffuse or has a genome DNA contamination.
After the sample is qualified, the library is constructed. The main process is as follows: 1) mRNA enrichment for total RNA by mRNA enrichment method: Enrich mRNA with polyA tail using magnetic beads with Oligo dT. 2) Fragment the RNA obtained by interrupting the buffer, perform reverse transcription with random N6 primers, and synthesize the double-stranded cDNA to form double-stranded DNA. 3) Fill in the end of the synthesized double-stranded DNA and phosphorylate the 5' end, form a sticky end with a protruding "A" at the 3' end, and then connect a bubbling linker with a protruding "T" at the 3' end. 4) The ligation product is amplified by PCR with specific primers. 5) The PCR product is thermally denatured into single-stranded DNA, and then a single-stranded circular DNA library is obtained by circularizing the single-stranded DNA with a bridge primer.

value: Mycelium

value: Wild type

value: Cultivated in medium containing glucose


title: FF4_2


title: FF4_43


title: FF4_59


title: TD_3

description: Library name: TD_3


title: TD_2

description: Library name: TD_2


title: FF4_95


title: FF4_21


title: Penicillium oxalicum, ∆PoxKu70, Glucose, 24 h, rep2


title: TD_1

description: Library name: TD_1


title: Penicillium oxalicum, ∆PoxKu70, Glucose, 24 h, rep1


title: FF2_76


title: Pool-1-ACTTGAAT

value: Control


title: FF4_38


title: AsPC-1 cells, Palbociclib+MRTX1133, 24 h, rep3

city: Göttingen

first: Maj-Britt

last: Paulsohn

address: Justus-von-Liebig-Weg 11

department: Department of Molecular Oncology

institute: University Medical Center Göttingen

description:
Library name: Palbociclib+MRTX1133_3
PM.3

data_processing:
Galaxy environment provided by the GWDG (Gesellschaft für wissenschafltiche Datenverarbeitung mbH Göttingen, https://galaxy.gwdg.de/)
Sequences were trimmed for 11 nucleotides (Trim, version 0.0.2).
Alignment was performed on GRCh38 using HISAT2 (version 2.0.5.2).
Read counts were acquired via featureCounts (version 1.6.3+galaxy2).
DeSeq2 (version 2.11.40.6+galaxy1) was used to perform differential gene expression analysis and to obtain normalized counts.
Assembly: CRCh38
Supplementary files format and content: tab-delimited file contains normalized count data

growth_protocol: AsPC-1 cells were maintained in RPMI 1640 supplemented with 10% FCS, 50 U/mL Penicillin, 50 μg/mL Streptomycin and 2 mM Glutamin. AsPC-1 RRID: CVCL_0152

extract_protocol:
Total RNA was extracted from cells using TRIzol®. 200 µL Chloroform was added per 1000 µL TRIzol. For phase separation, samples were centrifuged, and the aqueous phase was used to precipitate the RNA with 800 µL isopropanol. Two washing steps with 75 % ethanol were performed before the RNA was resuspended in nuclease-free water. 400 ng RNA was used for sequencing.
mRNA library preparation (poly A enrichment) and reverse transcription into cDNA followed by QC at Novogene

treatment_protocol: AsPC-1 cells were seeded. Next day, treatment was applied as follows: DMSO, Palbociclib (5 µM), MRTX1133 (0.5 µM), or the combination (Palbociclib + MRTX1133). After 24 h of treatment RNA was harvested.

value: pancreatic cancer cells

value: Palbociclib (5 µM) + MRTX1133 (0.5µM)

tag: time


title: FF2_6


title: Penicillium oxalicum, ∆PoxKu70,  Avicel, 24 h, rep3

value: Cultivated in medium containing Avicel


title: FF2_4

value: cortical organoid


title: TDE_3

description: Library name: TDE_3


title: AsPC-1 cells, Palbociclib+MRTX1133, 24 h, rep2

description:
Library name: Palbociclib+MRTX1133_2
PM.2


title: Penicillium oxalicum, ∆PoxKu70,  Avicel, 24 h, rep2


title: FF4_87

value: fibroblast


title: AsPC-1 cells, Palbociclib+MRTX1133, 24 h, rep1

description:
Library name: Palbociclib+MRTX1133_1
PM.1


title: Dup_4


title: FF4_20


title: FF2_3


title: AsPC-1 cells, MRTX1133, 24 h, rep3

description:
Library name: MRTX1133_3
MRTX.3


title: FF2_2


title: Penicillium oxalicum, ∆PoxCxrE, Glucose, 24 h, rep3

value: PoxCxrE knockout in Wild type


title: AsPC-1 cells, MRTX1133 24 h, rep2

description:
Library name: MRTX1133_2
MRTX.2


title: FF3_1


title: FF2_8


title: AsPC-1 cells, MRTX1133, 24 h, rep1

description:
Library name: MRTX1133_1
MRTX.1


title: FF2_5


title: FF4_13


title: FF3_2