title: sample 2-2-2 BatchA_water extract of Zingiberis Rhizoma Recens (Medium dose)_rep2

value: Zingiberis Rhizoma Recens water extract 100 ug/mL


title: adipocyte, control, rep3

status: Public on Oct 28 2025

type: SRA

city: ANN ARBOR

first: Jorge

middle: L

last: Ruas

state: Michigan

address: 2800 Plymouth RD, B025-2681 NCRC - Ruas Lab NCRC B025

country: USA

institute: University of Michigan

description:
Library name: control_adipocyte_3
K4_MKRN250005866-1A_22LK5CLT4_L8
res_preadipocyte_vs_adipocyte.csv
res.shrink_preadipocyte_vs_adipocyte.csv

library_source: transcriptomic

data_processing:
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to GRCm38 whole genome using STAR 2.7.3a with extra arguments --outFilterMismatchNmax 33 --alignEndsType Local --alignSJoverhangMin 8 --outFilterScoreMinOverLread 0 --outFilterMatchNminOverLread 0 --outFilterMatchNmin 0 --seedSearchStartLmax 12
Reads aligning to gene exons (Mus_musculus.GRCm38.94.gtf) were counted using featureCounts v2.0.3 program from Subread package, summarized at gene level.
Differential gene expression at FDR level 0.05 was obtained using DESeq2 R package
Assembly: GRCm38.p6
Supplementary files format and content: .txt file of raw read counts of all samples
Supplementary files format and content: .txt file containing information about samples
Supplementary files format and content: .csv file of normalized counts of all samples
Supplementary files format and content: .csv file of differential gene expression analysis output from DESeq2 of CGRP-treated adipocytes vs. control, unshrunken
Supplementary files format and content: .csv file of differential gene expression analysis output from DESeq2 of NE-treated adipocytes vs. control, unshrunken
Supplementary files format and content: .csv file of differential gene expression analysis output from DESeq2 of NPY-treated adipocytes vs. control, unshrunken
Supplementary files format and content: .csv file of differential gene expression analysis output from DESeq2 of SP-treated adipocytes vs. control, unshrunken
Supplementary files format and content: .csv file of differential gene expression analysis output from DESeq2 of CGRP-treated preadipocytes vs. control, unshrunken
Supplementary files format and content: .csv file of differential gene expression analysis output from DESeq2 of preadipocytes vs. adipocytes, unshrunken
Supplementary files format and content: .csv file of differential gene expression analysis output from DESeq2 of CGRP-treated adipocytes vs. control, shrunken
Supplementary files format and content: .csv file of differential gene expression analysis output from DESeq2 of NE-treated adipocytes vs. control, shrunken
Supplementary files format and content: .csv file of differential gene expression analysis output from DESeq2 of NPY-treated adipocytes vs. control, shrunken
Supplementary files format and content: .csv file of differential gene expression analysis output from DESeq2 of SP-treated adipocytes vs. control, shrunken
Supplementary files format and content: .csv file of differential gene expression analysis output from DESeq2 of CGRP-treated preadipocytes vs. control, shrunken
Supplementary files format and content: .csv file of differential gene expression analysis output from DESeq2 of preadipocytes vs. adipocytes, shrunken

molecule: polyA RNA

organism: Mus musculus

source_name: Inguinal adipose tissue (iWAT)

growth_protocol: Induction media from day 0 to 4 followed by differentiation media from day 4 to 8 for adipocytes. Preadipocytes maintained in growth media.

extract_protocol:
Total RNA was extracted using TRI reagent (Sigma-Aldrich, T9424) according to the manufacturer's instructions. RNA was further purified, and DNase treated (Zymo, R1013), and integrity confirmed prior to sequencing (Agilent Technologies, 2100 Bioanalyzer), with an RNA integrity number (RIN) greater than 7 as a threshold. Samples were purified by poly-A enrichment before library preparation using the Illumina NovaSeq X Plus Series platform with a paired-end 150 base pair strategy.
Samples were purified by poly-A enrichment before library creation using Illumina TruSeq Stranded mRNA.

treatment_protocol: Vehicle or 100 nM of each treatment.

tag: tissue

value: Inguinal adipose tissue (iWAT)

tag: treatment

value: Control


title: MCF10A cells, sgMETTL3-2, rep2 (ATAC-seq)

city: Boston

first: Xintao

last: Qiu

state: MA

address: 440 brookline ave

institute: Dana Faber Cancer Institute

library_source: genomic

data_processing:
Illumina Casava1.7 software used for basecalling.
Reads were aligned to hg19 using BWA mem.
For peak calling MACS2 was used.
Assembly: hg19
Supplementary files format and content: bigwig

molecule: genomic DNA

organism: Homo sapiens

source_name: breast

growth_protocol: The MCF10A cells were cultured in DMEM/F12 mediaum  supplementing with horse serum,  EGF, hydrocortisone, cholera toxin, insulin  and penicillin-streptomycin

extract_protocol:
1 × 105 control or METTL3 KO MCF10A cells were trypsinised and suspended in 50 ul of ATAC resuspension buffer (10mM Tris-HCl pH 7.4, 10mM NaCl and 3mM MgCl2, 0.1% NP-40, 0.1% Tween-20 and 0.01% digitonin) on ice for 3 mins. After addition of 1 ml ATAC- resuspension buffer containing 0.1% Tween-20, cells were centrifuged at maximum speed for 10 min at 4°C. The supernatant was removed, and nuclei were resuspended in 50 μL transposition mix: 2.5 μL transposase (100 nM final), 16.5 μL PBS, 0.5 μL 1% digitonin, 0.5 μL 10% Tween-20 and 5 μL water. The transposition reactions were carried out at 37 °C for 30 minutes in a thermomixer with shaking at 100 x g followed by sequencing library preparation.
Library concentrations were quantified using the NEBNext Library Quant Kit. 1 µL of the pre-amplified sample is diluted with 99 µL of 1× NEB Dilution Buffer, followed by the creation of qPCR reactions in a 384-well plate. After completing the qPCR, DNA concentration is determined using tape station. The amplified samples are then processed with DNA Binding Buffer, transferred to Zymo-Spin Columns, washed, and finally eluted to obtain the DNA product.

treatment_protocol: Vector expressing CAS9 and sgRNAs targeting either AACS1 or METTL3 was lentivirally transduced into MCF10A cells.

value: breast

tag: cell line

tag: cell type

value: mammary epithelial cells

tag: genotype

value: METTL3 KO


title: adipocyte, control, rep2

description:
Library name: control_adipocyte_2
K3_MKRN250005865-1A_22LMNGLT4_L7
res_preadipocyte_vs_adipocyte.csv
res.shrink_preadipocyte_vs_adipocyte.csv


title: HAP1 MATR3 KO MATR3 PAR-CLIP

city: Bethesda

first: Stefan

last: Muljo

state: MD

address: 10 Center Drive

institute: NIH, NIAID

data_processing:
The PAR-CLIP sequencing data were processed by the PARpipe pipeline (https://ohlerlab.mdc-berlin.de/software/PARpipe_119/).
Adapters and UMI sequences were trimmed using Cutadapt 1.8.3 (parameters: -n 1 -m 16 -u 4) and reads were aligned to the hg38 (GRCh38.p5) genome using Bowtie v1.1.2 allowing for two mismatches and up to 3,600 multimapping alignments tied for the best alignment score (parameters: -v 2 -a -m 3600 –best–strata).
The aligned reads were passed to PARalyzer for clustering and annotation (see Methods section for specific PARalyzer parameters used)
Reads, groups, and clusters were annotated to the features in the Gencode v24 annotation file for reference chromosomes, scaffolds, assembly patches, and alternative loci, with introns given higher precedence in the annotation rank as compared to coding regions.
Assembly: hg38
Supplementary files format and content: The *.clusters.tsv files contain, for each cluster called by PARalyzer, the genomic coordinates, clusterID, annotated feature, annotated gene (if applicable), nucleotide sequence, read count, T2C location/counts and T2C fraction.

molecule: total RNA

growth_protocol: HAP1 cells were cultured in IMDM medium (Quality Biological, cat# 112-035-101) supplemented with 10% (vol/vol) heat-inactivated fetal calf serum (Gibco, cat# 26170-043), 100 U/mL penicillin, 100 µg/mL streptomycin (Gibco, cat# 15140-122), at 37° C in a humidified incubator containing 5% CO2.

extract_protocol:
UV-crosslinked cells were scraped off the dishes, lysed, Rnase T1 treated, and then immunoprecipitated using anti-FLAG magnetic beads. Beads were washed and resuspended in immunoprecipitation wash buffer. RNA residing in the immprecipitate was further trimmed with RNase T1, washed three more times in high-salt lysis buffer, and finally resuspended in one bead volume of dephosphorylation buffer. (see Methods section for exact amounts, concentrations and timings used)
3' ends of RNA fragments were dephosphorylated in calf intestinal phosphatase and RNA was radioactively 5' end-labeled using T4 polynucleotide kinase and P-32-gamma-ATP. Protein-RNA complexes were separated by SDS-PAGE, transferred to nitrocellulose and visualized with autoradiography. Radioactive bands corresponding to the MATR3 rbionucleoprotein complex were recovered, and protein was removed via proteinase K digestion. RNA was then isolated by acidic phenol/chloroform extraction and ethanol precipitation. The recovered RNA was converted into a cDNA library using a standard small RNA library protocol (Hafner et al., Methods 2008) and sequenced on an Illumina NovaSeq 6000 instrument. (see Methods section for exact amounts, concentrations and timings used)

treatment_protocol: Wild Type HAP1 Cells (cat# C631_29281, batch 40661) were purchased from Horizon Discovery and compared with MATR3 KO ((cat# HZGHC007500c001)

value: KBM-7 (CML)-derived near-haploid cell line

value: MATR3 knockout


title: NM145, replicate 1, scRNA-seq, BL

city: dallas

first: Shaheen

last: Khan

state: texas

address: 5323 Harry Hines Blvd

institute: UT Southwestern Medical Center

description: 10X Genomics

data_processing: Data was demultiplexed using Cell ranger by 10X genomics

source_name: PBMCs

extract_protocol:
PBMCs were isolated from blood and frozen in 10% DMSO in liquid Nitrogen tanks. PBMCs were then thawed and RBCs were lysed before preparing the libraries.
Libraries were prepared using 5' prime protocol as per the instructions by 10X Genomics

value: PBMCs

tag: time

value: BL


title: MCF10A cells, sg control, rep2 (ATAC-seq)

value: AAVS1 KO


title: HAP1 WT MATR3 PAR-CLIP rep2

value: Wild Type


title: sample 2-1-4 BatchA_water extract of Zingiberis Rhizoma (High dose)_rep1

city: Daejeon

first: Musun

last: Park

address: 1672 Yuseong-daero, Yuseong-gu

country: South Korea

institute: Korean Institute of Oriental Medicine

data_processing:
Quality of RNA-seq raw data was assessed using FastQC (v0.11.9), and the common regions of MGISEQ adapter sequences were removed using TrimGalore (v0.6.5). The trimmed reads were then aligned to the human reference genome hg38 using the STAR (v2.7.3a) with default settings. Transcript abundance per gene including expected read counts and transcripts per million (TPM) were quantified using RSEM (v1.3.3) with gene annotation GRCh38.84.
Assembly: GRCh38.84
Supplementary files format and content: tab-delimited text files include Expected counts for each Sample

growth_protocol: Human hepatocellular carcinoma cell line, HepG2 (HB-8065) were purchased from the American Type Culture Collection (ATCC, Manassas, VA, USA). HepG2 cells were maintained in DMEM medium supplemented with 10% (v/v) heat-inactivated FBS, 100 IU/mL penicillin, and 100 mg/mL streptomycin at 37 °C, 5% CO2 incubator. The cell was subcultured every 3 or 4 days, depending on the cell density.

extract_protocol:
Total RNA was isolated according to the manufacturer’s protocol. The concentration of the isolated RNA was determined using an Agilent RNA 6000 Nano Kit (Agilent Technologies, Waldbronn, Germany), and RNA quality was evaluated by assessing the RNA integrity number (RIN > 7).
A total RNA (1 microgr) was processed to prepare a mRNA sequencing library using the MGIEasy RNA Directional Library Prep kit (#1000006386, MGI Tech Co., Ltd., China) according to the manufacturer’s instruction. The library was quantified using the QauntiFluor® ssDNA System (E3190, Promega Corporation, WI, USA). The prepared DNA nanoball was sequenced on the MGIseq system (MGI Tech Co., Ltd., China) with 100 bp paired-end reads.

treatment_protocol: Dulbecco's Modified Eagle Medium (DMEM), phosphate-buffered saline (PBS), TrypLE Express, penicillin-streptomycin, and fetal bovine serum (FBS) were purchased from Gibco (Grand Island, NY, USA). Cell culture flasks and multiwell culture plates were purchased from Thermo Fisher Scientific (Waltham, MA, USA). Dimethyl sulfoxide (DMSO) and QIAzol lysis reagents were purchased from Sigma-Aldrich (St. Louis, MO, USA) and Qiagen (Germantown, MD, USA), respectively. The dried medicinal plants, prepared in compliance with the Korean Pharmacopoeia, were supplied from Kwangmyung-dang Medicinal Herbs Co. (Ulsan, Republic of Korea). The samples used in the experiment underwent organoleptic examination by Dr. Choi Goya, an herbal medicine organoleptic examination expert appointed by the Korea Food and Drug Administration, and were additionally identified at the species level by analyzing the DNA barcode region sequences. Voucher specimen were deposited in the Korean Herbarium of Standard Herbal Resources, Herbal Medicine Resources Research Center, Korea Institute of Oriental Medicine, Naju, Republic of Korea . All herbs and extracts can be obtained from the Oriental Medicine Resources Research Center (KIOM, https://oasis.kiom.re.kr/herblib). A hot water and 70% ethanol extract of each plant was prepared and supplied by KOC Biotech Co. (Daejeon, Republic of Korea). Briefly, dried plants (1,000 g) were pulverized and extracted in hot distilled water (10 L) for 3 h under a reflux extraction system (MS-DM609; MTOPS, Seoul, Republic of Korea) or in 70% ethanol (15L) for 1 h under ultrasonication system (VCP-20, Lab companion, Dajeon, Republic of Korea) twice. The extract solution was filtered through a 5 µm cartridge filter, concentrated in a rotary evaporator (Ev-1020, SciLab, Seoul, Republic of Korea), and then lyophilized in a freeze dryer (LP-20, Ilshin-Bio-Base, Dongduchen, Republic of Korea) to obtain the final extract. The extract was finely homogenized and packed in a glass bottle containing a desiccant silica gel. The decoction was prepared by mixing each extract, considering the composition ratio and extract yield of a single medicinal herb, and homogenized, according to the Korean Pharmacopoeia. For the in vitro treatment, extracts (100 mg) were vigorously vortexed for 30 min in 10 mL phosphate-buffered saline (PBS; Thermo Fisher Scientific, Rockford, IL, USA) containing 2% DMSO and then sterilized by filtering through a 0.22 µm membrane to obtain a stock solution (10 mg/mL); the stock solution was aliquoted in small volumes and stored at -80°C until use.

value: Zingiberis Rhizoma water extract 500 ug/mL


title: adipocyte, control, rep1

description:
Library name: control_adipocyte_1
K1_MKRN250005864-1A_22LMNGLT4_L7
res_preadipocyte_vs_adipocyte.csv
res.shrink_preadipocyte_vs_adipocyte.csv


title: MCF10A cells, sgMETTL3-4, rep1 (ATAC-seq)


title: sample 2-1-3 BatchA_water extract of Zingiberis Rhizoma Recens (Low dose)_rep1

value: Zingiberis Rhizoma Recens water extract 20 ug/mL


title: HAP1 WT MATR3 PAR-CLIP rep1


title: sample 2-1-2 BatchA_water extract of Zingiberis Rhizoma Recens (Medium dose)_rep1


title: MCF10A cells, sgMETTL3-2, rep1 (ATAC-seq)


title: adipocyte_day_4, CGRP_day2-4, rep2

description: Library name: cgrp_d4_2

data_processing:
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to GRCm38 whole genome using STAR 2.7.3a with extra arguments --outFilterMismatchNmax 33 --alignEndsType Local --alignSJoverhangMin 8 --outFilterScoreMinOverLread 0 --outFilterMatchNminOverLread 0 --outFilterMatchNmin 0 --seedSearchStartLmax 12
Reads aligning to gene exons (Mus_musculus.GRCm38.94.gtf) were counted using featureCounts v2.0.3 program from Subread package, summarized at gene level.
Differential gene expression at FDR level 0.05 was obtained using DESeq2 R package, genes with total raw counts less then 10 across all samples were excluded from analysis
Assembly: GRCm38.p6
Supplementary files format and content: .txt file of raw read counts of all samples
Supplementary files format and content: .txt file containing information about samples
Supplementary files format and content: .csv file of normalized counts of all samples
Supplementary files format and content: .csv file of differential gene expression analysis output from DESeq2 of day 4 CGRP treated versus control
Supplementary files format and content: .csv file of differential gene expression analysis output from DESeq2 of HFD samples

growth_protocol: Induction media from day 0 to 4 followed by differentiation media from day 4 to 8

extract_protocol:
Total RNA was extracted using TRI reagent (Sigma-Aldrich, T9424) according to the manufacturer's instructions. RNA was further purified, and DNase treated (Zymo, R1013), and integrity confirmed prior to sequencing (Agilent Technologies, 2100 Bioanalyzer), with an RNA integrity number (RIN) greater than 7 as a threshold. Samples were purified by poly-A enrichment before library preparation using the Illumina NovaSeq X Plus Series platform with a paired-end 150 base pair strategy.
Samples were purified by poly-A enrichment before library creation using Illumina TruSeq Stranded mRNA

treatment_protocol: Differentiation media or 100 nM CGRP

value: CGRP


title: skin tissue-SCC3

city: Shanghai

first: Tao

last: Xu

state: Shanghai

address: Fengyang road

department: dermatology

country: China

institute: Changzheng hospital

description: single-cell RNA

data_processing:
Raw data were processed using the Cellranger (10X Genomics version 2.1.0).
Resulting FastQ files were aligned to the human reference genome using Spliced Transcripts Alignment to a Reference (STAR).
Gene-Barcode matrices were generated using (UMIs) counter and filtering non-cell associated barcodes by default parameters.
A normalized gene-barcode matrix was generated for further analysis, which contained the barcoded cells and gene expression counts.
Assembly: hg19
Supplementary files format and content: barcodes.tsv (cell barcode files), genes.tsv (gene files), matrix.mtx (gene-barcode counts).

source_name: skin tissue

extract_protocol:
All the samples were incubated with dispase for 2 h at 37 °C. The epidermis was peeled from the dermis and minced with scissors to pieces < 1 mm3, incubation with 5mL of 0.25% trypsin-EDTA for 30 minutes. The dissociated cell suspension was washed with 10% PBS, filtered through a 70 mm filter, centrifuged at 1500 rpm for 5 min. Dermis were incubated with 5mL of 0.35% collagenase for 30 minutes. All steps other than incubation were performed on ice.
Cells were loaded according to standard protocol of the Chromium single cell 3' kit and the v2 chemistry from 10X Genomics.
The library was pooled and sequenced on the Illumina HiSeq 6000 platform to get ~150-200 million reads.

value: cutaneous squamous cell carcinoma

value: yes

tag: histopathology

value: cutaneous squamous cell carcinoma (moderately differentiated)


title: adipocyte_day_4, CGRP_day2-4, rep3

description: Library name: cgrp_d4_3

value: Inguinal adipose tissue (iWAT)


title: skin tissue-SCC2

description: single-cell RNA

value: no

value: cutaneous squamous cell carcinoma (well differentiated)


title: MCF10A cells, sg control, rep1 (ATAC-seq)

value: breast

value: mammary epithelial cells


title: Piwi-KD_RNA_rep2

city: Tokyo

first: Kensaku

last: Murano

address: 35 Shinanomachi, Shinjuku

department: Department of Molecular Biology

country: Japan

institute: Keio University School of Medicine

description: Library name: RNA-seq_Sample 12

data_processing:
After adaptor sequence filtering by TrimGalore 0.6.7, RNA-seq reads were mapped to Drosophila melanogaster reference genome (dm6) using STAR 2.7.10a with following options, --outSAMmode NoQS; --alignEndsType Local; --outReadsUnmapped Fastx; --outMultimapperOrder Random; --outFilterMultimapNmax 1000; --winAnchorMultimapNmax 2000; --outFilterMismatchNmax 0; --seedSearchStartLmax 30; --alignSoftClipAtReferenceEnds No; -outFilterType BySJout; --alignSJoverhangMin 15; --alignSJDBoverhangMin 1. Adaptor trimmed ChIP-seq reads were mapped to Drosophila melanogaster reference genome (dm6) and Luc reporter plasmid (pSUb-14Box-Luc) using Bowtie2 2.2.5 with --very-sensitive option.
RNA-seq reads mapped on TE loci were quantified using featureCounts v2.0.1 and multi-mapped reads were counted fractionally for TEs. TE annotation data (RepeatMasker) was obtained from University of California, Santa Cruz (UCSC) Genome Browser.
For visualization of ChIP-seq reads on reference genome, alignment track were generated using bamCoverage program of deepTools 3.5.1 with --normalizeUsing RPKM; -bs 1 parameters.
Assembly: dm6
Supplementary files format and content: Read count data (.txt) from RNA-seq analysis. Read coverage data (.bw) from ChIP-seq analysis.

organism: Drosophila melanogaster

source_name: Ovary

growth_protocol: Cells were grown at 26 degrees in OSC medium (M3 medium supplemented with fly extract)

extract_protocol:
For RNA-seq, total RNAs were isolated by RNeasy kit (Qiagen). Poly(A)+ RNAs were obtained using oligo-dT beads. For ChIP experiments targeting RNA polymerase II, OSC were fixed in 0.3% formaldehyde (methanol-free, Thermo Fisher) for 10 min at room temperature and quenched with a final concentration of 125 mM glycine for 2 min. After washing twice with PBS (containing 125 mM glycine), nuclei were isolated using 200 µl Low salt lysis buffer (10 mM HEPES-KOH pH 7.4, 10 mM NaCl, 1.5 mM MgCl2, 1% Nonidet P-40) supplemented with cOmplete (Protease inhibitor cocktail, Roche) and PhosSTOP (Phosphatase inhibitor cocktail, Roche). The isolated nuclei were resuspended in 200 µl Sonication buffer (50 mM Tris-HCl pH 7.9, 150 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate) supplemented with cOmplete and PhosSTOP, and sonicated using a probe-sonicator (BRANSON, SFX150) at 30% amplitude, with 2 sec pulses and 3 sec intervals, for a total of 2 min, in an ice-cold water bath. After centrifugation, the supernatant was passed through a 0.22-μm syringe filter (Membrane Solutions) and used for immunoprecipitation.To elucidate Pol II occupancy on the reporter gene, we utilized an anti-Pol II ser5-P antibody (Cosmobio, CMA603).The chromatin lysate was incubated with 1 µg of anti-Pol II ser5-P antibody at 4 °C for 3 h with rotation. The antibody was captured using Dynabeads Protein G.The captured chromatin was extracted in 100 µl of Elution buffer (50 mM Tris-HCl pH 7.9, 10 mM EDTA, and 1% SDS) at 65 °C for 10 min. The supernatant was supplemented with 4 µl of 5 M NaCl, and cross-link were reversed at 65 °C overnight in a Thermomixer at 1400 rpm (Eppendorf). DNA was purified using a spin column and eluted in Milli-Q water.
For RNA-seq, Poly(A)+ RNA was isolated from 2 µg total RNA using Dynabeads Oligo (dT)25 (Thermo Fisher), followed by heat-fragmentation at 94ºC for 6 min. cDNA was synthesized from the fragmented RNA by random hexamers and SuperScript III (Thermo Fisher), and then purified by AMPure XP (Beckman). Second strand DNA was synthesized by DNA polymerase I (NEB) in the presence of 1 mM dUTP at 16ºC for 2.5 h, and then purified by AMPure XP. Library preparation was carried out using the NEBNext Ultra II DNA Library Prep kit for illumina (NEB), according to the manufacturer’s instructions. For ChIP-seq, ChIPed DNAs were used for library preparation by NEBNext Ultra II DNA Library Prep Kit for Illumina (NEB) following the manufacture’s protocol.

value: Ovary

value: OSC

value: Ovarian somatic cell

value: Piwi-KD


title: skin tissue-SCC1

description: single-cell RNA

value: carcinoma in situ


title: sample 2-1-6 BatchA_water extract of Zingiberis Rhizoma (Low dose)_rep1

value: Zingiberis Rhizoma water extract 20 ug/mL


title: adipocyte_day_4, CGRP_day2-4, rep1

description: Library name: cgrp_d4_1


title: sample 2-1-5 BatchA_water extract of Zingiberis Rhizoma (Medium dose)_rep1

value: Zingiberis Rhizoma water extract 100 ug/mL


title: adipocyte_day_6, control, rep3

description: Library name: control_d6_3

value: Control


title: sample 2-1-1 BatchA_water extract of Zingiberis Rhizoma Recens (High dose)_rep1

value: Zingiberis Rhizoma Recens water extract 500 ug/mL


title: sample 1-3-4 BatchA_Wortmannin_rep3

value: wortmannin 10 uM


title: skin tissue-AK8

description: single-cell RNA

value: actinic keratosis

value: AK I


title: adipocyte_day_6, control, rep1

description: Library name: control_d6_1

value: Inguinal adipose tissue (iWAT)


title: Panx-KD_RNA_rep2

description: Library name: RNA-seq_Sample 10

value: Panx-KD


title: Piwi-KD_RNA_rep1

description: Library name: RNA-seq_Sample 11


title: skin tissue-AK6

description: single-cell RNA

value: no


title: adipocyte_day_4, control, rep3

description: Library name: control_d4_3


title: Panx-KD_RNA_rep1

description: Library name: RNA-seq_Sample 9


title: sample A1-3-1 BatchA_Vehicle(DMSO)_rep3

value: 0.1% DMSO


title: adipocyte_day_6, control, rep2

description: Library name: control_d6_2


title: skin tissue-AK7

description: single-cell RNA


title: sample 1-1-4 BatchA_Wortmannin_rep1


title: Suv210-KD_RNA_rep2

description: Library name: RNA-seq_Sample 8


title: skin tissue-AK5

description: single-cell RNA


title: sample A1-2-1 BatchA_Vehicle(DMSO)_rep2


title: adipocyte_day_4, control, rep2

description: Library name: control_d4_2


title: Suv210-KD_RNA_rep1

description: Library name: RNA-seq_Sample 7


title: skin tissue-AK4

description: single-cell RNA

value: AK II


title: sample A1-1-1 BatchA_Vehicle(DMSO)_rep1


title: adipocyte_day_4, control, rep1

description: Library name: control_d4_1


title: sample 1-2-4 BatchA_Wortmannin_rep2


title: Smt3_2-KD_RNA_rep2

description: Library name: RNA-seq_Sample 6

value: Ovary


title: adipocyte_day_2, control, rep3

description: Library name: control_d2_3


title: Smt3_2-KD_RNA_rep1

description: Library name: RNA-seq_Sample 5

value: OSC


title: skin tissue-AK3

description: single-cell RNA


title: Smt3_1-KD_RNA_rep2

description: Library name: RNA-seq_Sample 4

value: Ovarian somatic cell


title: adipocyte_day_2, control, rep2

description: Library name: control_d2_2


title: Orthotopic implanted tumor, WT mouse,  rep1

city: Kyoto-City

first: Kosuke

last: Iwane

address: 54 Shogoin-Kawahara-Cho, Sakyo-Ku

department: Department of Gastroenterology and Hepatology

institute: Kyoto University Graduated School of Medicine

description:
Library name: WT_1
10xGenomics

library_source: transcriptomic single cell

data_processing:
The Cell Ranger software suite (version 7.1.0; 10X Genomics) was used to process, align, and summarize unique molecular identifier (UMI) counts.
Assembly: mm10
Supplementary files format and content: H5 archives: Thbs1KO_1_filtered_feature_bc_matrix.h5, Thbs1KO_2_filtered_feature_bc_matrix.h5, Thbs2KO_1_filtered_feature_bc_matrix.h5, Thbs2KO_2_filtered_feature_bc_matrix.h5, WT_1_filtered_feature_bc_matrix.h5, WT_2_filtered_feature_bc_matrix.h5

source_name: orthotopic implanted tumor in the rectum of the mice

extract_protocol:
Tumors generated by orthotopic implantation of MTO in the rectum of WT, Thbs1-/- and Thbs2-/- mice (n = 2 per group) were processed into single-cell suspensions using the Tumor Dissociation Kit (Miltenyi Biotec). Red blood cells were removed from the dissociated tumors using the Red Blood Cell Lysis Solution (Miltenyi Biotec), and dead cells were eliminated with the EasySep Dead Cell Removal (Annexin V) Kit (STEMCELL Technologies).
scRNA-seq libraries were prepared using the Chromium Next GEM Single Cell 3’ v3.1Kit (10X Genomics). Cells were loaded onto the 10X Chromium Controller (10X Genomics) with the Chromium Next GEM Chip G Single Cell kit (10X Genomics) at a concentration of 1,000 cells per µL, as described in the manufacturer’s protocol (10X Chromium Single Cell 3' Reagent Kits User Guide (v3.1 Chemistry Dual Index)). A total of 16,500 cells from each mouse were loaded. Generation of gel beads in emulsion (GEMs), barcoding, GEM-RT clean-up, complementary DNA amplification and library construction were performed according to the manufacturer’s protocol. Individual sample quality was checked using a TapeStation (Agilent Technology). The final library pool was sequenced using the DNBSEQ-G400RS (MGI).

value: WT


title: Orthotopic implanted tumor, WT mouse,  rep2

description:
Library name: WT_2
10xGenomics

value: orthotopic implanted tumor in the rectum of the mice


title: Orthotopic implanted tumor, Thbs2KO mouse,  rep2

description:
Library name: Thbs2KO_2
10xGenomics


title: skin tissue-AK1

description: single-cell RNA


title: adipocyte_day_2, control, rep1

description: Library name: control_d2_1


title: Smt3_1-KD_RNA_rep1

description: Library name: RNA-seq_Sample 3


title: adipocyte_day_0, control, rep3

description: Library name: control_d0_3


title: Orthotopic implanted tumor, Thbs2KO mouse,  rep1

description:
Library name: Thbs2KO_1
10xGenomics


title: Orthotopic implanted tumor, Thbs1KO mouse,  rep2

description:
Library name: Thbs1KO_2
10xGenomics


title: skin tissue-NS2

description: single-cell RNA

value: normal sun-exposed skin

value: normal skin


title: LacZ-KD_RNA_rep2

description: Library name: RNA-seq_Sample 2


title: adipocyte_day_0, control, rep2

description: Library name: control_d0_2

value: Control


title: skin tissue-NS3

description: single-cell RNA


title: LacZ-KD_RNA_rep1

description: Library name: RNA-seq_Sample 1


title: Orthotopic implanted tumor, Thbs1KO mouse,  rep1

description:
Library name: Thbs1KO_1
10xGenomics


title: adipocyte_day_0, control, rep1

description: Library name: control_d0_1


title: Mouse Thymus (APC), Perinatal Day 28, Wild-type, Sample 1

city: Nashville

first: Bryan

middle: Robert

last: Helm

state: Tennessee

address: 2525 West End Avenue, Suite 1100

department: Center for Quantitative Science

institute: Vanderbilt University Medical Center

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were performed with Cell Ranger software v6.0.1 (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger)
Assembly: mm10
Supplementary files format and content: Tab-separated value files and matrix files with gene names, cell barcodes, and raw expression counts

source_name: Thymus

extract_protocol:
To sort hematopoietic APCs for scRNA-sequencing, thymi were harvested and enzymatically digested with an enzymatic solution comprising  LiberaseTM (Roche) and DNAse I in PBs at 37 degrees celsius. Isolated cells were spun down and stained with the corresponding panel of antibodies. The sort samples were resuspended in FWB+ EDTA buffer plus propidium iodide (Enzo). Sorted hematopoietic APCsells were then spun down in FWB without EDTA, counted and resuspended at a concentration of 700-1200 cells/µL prior to submission to the sequencing core.
Library prep was performed per the manufacturer's protocol for scRNA-seq samples (Chromium Next GEM Single Cell 3' v3.1).

value: Thymus

value: Antigen Presenting Cells

value: Wild Type


title: skin tissue-NS1

description: single-cell RNA


title: skin tissue-AK2

description: single-cell RNA

value: actinic keratosis

value: AK I


title: 95D cells, Control，rep1

status: Public on Oct 27 2025

city: Guangzhou

first: tianyu

last: tao

state: Guangdong

address: No.74, Zhongshan Er Road

department: Zhongshan School of Medicine

institute: Sun Yat-sen University

description:
Library name: Sample1
Column name in raw_All Samples.txt: 95D_Control_rep1

data_processing:
Raw data (raw reads) of fastq format were firstly processed through in-house perl scripts. In this step, clean data (clean reads) were obtained by trimming reads containing adapter, ploy-N or with low quality from raw data. At the same time, Q20, Q30 and GC content of the clean data were calculated. All the downstream analyses were performed based on the clean data with high quality. Reference genome and gene model annotation files were downloaded from genome website directly. Index of the reference genome was built using Hisat2 v2.0.5 and paired-end clean reads were aligned to the reference genome using Hisat2 v2.0.5. We selected Hisat2 as the mapping tool for that Hisat2 can generate a database of splice junctions based on the gene model annotation file and thus a better mapping result than other non-splice mapping tools. (Stringtie- 1.3.3b) was used to count the read numbers mapped to each gene. And then FPKM was calculated, which expected number of Fragments Per Kilobase of transcript sequence per Millions base pairs sequenced. FPKM is a simple and mostly common used expression level normalization method. It normalizes for both sequencing depth and genome size.
Assembly: hg38
Supplementary files format and content: tab-delimited text file includes raw counts for each Sample
Supplementary files format and content: txt files include FPKM values for each Sample

growth_protocol: The human lung cancer cell line 95D was cultured in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 10% (v/v) fetal bovine serum (FBS) and 1% (v/v) penicillin/streptomycin.

extract_protocol:
Total RNA was isolated from the cultured cells using TRIzol reagent (Invitrogen) following the manufacturer’s instructions. 1-2 µg of total RNA was reverse transcribed using the HiScript III RT SuperMix with gDNA wiper (Vazyme).
Libraries were constructed from the extracted total RNA using standard protocols, including rRNA depletion, fragmentation, cDNA synthesis, and adapter ligation. The Qualified libraries were pooled and sequenced on Illumina platforms with PE150 strategy in Novogene Bioinformatics Technology Co., Ltd (Beijing, China), according to effective library concentration and data amount required.

treatment_protocol: For nutrient deprivation experiments, cells were seeded for 24 hours in complete medium. Subsequently, the medium was replaced with pre-warmed glucose-free DMEM or glutamine-free DMEM, each supplemented with 10% dialyzed FBS, and cells were cultured for 48 hours at 37°C with 5% CO2.

value: human lung cancer cell line

value: cultured in control DMEM conditions for 48 hours


title: 95D cells, Control，rep2

description:
Library name: Sample2
Column name in raw_All Samples.txt: 95D_Control_rep2


title: 95D cells, glucose deficiency，rep1

description:
Library name: Sample3
Column name in raw_All Samples.txt: 95D_Glc_deficiency_rep1

value: cultured in glucose-deficient conditions for 48 hours


title: 95D cells, glutamine deficiency，rep1

description:
Library name: Sample5
Column name in raw_All Samples.txt: 95D_Gln_deficiency_rep1

value: cultured in glutamine-deficient conditions for 48 hours


title: 95D cells, glutamine deficiency，rep2

description:
Library name: Sample6
Column name in raw_All Samples.txt: 95D_Gln_deficiency_rep2


title: ATF4, out DAMER, 1M7

description:
Library name: Sample27
SHAPE-MAP
ATF4_with_DAMER_1M7

library_source: other

data_processing:
Process the raw sequencing reads using ShapeMapper2 (v2.3) software. Use a read count cutoff of 1000 to calculate the SHAPE reactivity profile for each nucleotide. Use the calculated SHAPE reactivity data as constraints for secondary structure modeling with the Superfold (v1.0) software. Visualize the resulting secondary structure models using ViennaRNA Web Services (http://rna.tbi.univie.ac.at/forna/) to reproduce the structural diagrams.
Assembly: hg38
Supplementary files format and content: tab-delimited text file includes raw counts for each Sample
Supplementary files format and content: txt files include reactivitysignal for each Sample

molecule: other

source_name: In vitro transcribed RNA

extract_protocol:
500 ng of in vitro transcribed DAMER and/or 500 ng of ATF4 mRNA were used.RNA was denatured at 95°C for 2 minutes and snap-cooled on ice.RNA was then refolded by incubating at 37°C for 20 minutes in 3.3× folding buffer (333 mM HEPES pH 8.0, 333 mM KCl, 33.3 mM MgCl2).Folded RNAs were treated with 100 nM of the SHAPE reagent 1-methyl-7-nitroisatoicanhydride (1M7) (as a negative control) for 5 minutes at 37°C. The modified RNA was purified using RNeasy Mini columns (Qiagen).Reverse transcription was performed using SuperScript II Reverse Transcriptase with specific primers in MaP buffer.
Sequencing libraries were generated from the resulting cDNA using the Nextera XT DNA Library Preparation Kit (Illumina). Sequencing was performed on a NovaSeq 6000 system with PE250 reads.

value: In vitro transcribed RNA

value: co-incubated with DAMER, and treated with 1M7


title: ATF4, out DAMER, DMSO

description:
Library name: Sample28
SHAPE-MAP
ATF4_with_DAMER_DMSO

value: co-incubated with DAMER, and treated with DMSO


title: ATF4, without DAMER, 1M7

description:
Library name: Sample25
SHAPE-MAP
ATF4_without_DAMER_1M7

value: treated with 1M7


title: DAMER_M1/2, with ATF4, DMSO

description:
Library name: Sample24
SHAPE-MAP
DAMER_M12_with_ATF4_DMSO

value: co-incubated with ATF4 mRNA, and treated with DMSO


title: ATF4, without DAMER, DMSO

description:
Library name: Sample26
SHAPE-MAP
ATF4_without_DAMER_DMSO

value: treated with DMSO


title: DAMER_M1/2, without ATF4, DMSO

description:
Library name: Sample22
SHAPE-MAP
DAMER_M12_without_ATF4_DMSO


title: DAMER_M1/2, with ATF4, 1M7

description:
Library name: Sample23
SHAPE-MAP
DAMER_M12_with_ATF4_1M7

value: co-incubated with ATF4 mRNA, and treated with 1M7


title: DAMER_M1/2, without ATF4, 1M7

description:
Library name: Sample21
SHAPE-MAP
DAMER_M12_without_ATF4_1M7


title: DAMER_M2, with ATF4, 1M7

description:
Library name: Sample19
SHAPE-MAP
DAMER_M2_with_ATF4_1M7


title: DAMER_M2, without ATF4, 1M7

description:
Library name: Sample17
SHAPE-MAP
DAMER_M2_without_ATF4_1M7


title: DAMER_M2, with ATF4, DMSO

description:
Library name: Sample20
SHAPE-MAP
DAMER_M2_with_ATF4_DMSO


title: DAMER_M1, with ATF4, DMSO

description:
Library name: Sample16
SHAPE-MAP
DAMER_M1_with_ATF4_DMSO


title: DAMER_M1, without ATF4, DMSO

description:
Library name: Sample14
SHAPE-MAP
DAMER_M1_without_ATF4_DMSO


title: DAMER_M2, without ATF4, DMSO

description:
Library name: Sample18
SHAPE-MAP
DAMER_M2_without_ATF4_DMSO


title: DAMER_M1, with ATF4, 1M7

description:
Library name: Sample15
SHAPE-MAP
DAMER_M1_with_ATF4_1M7


title: 95D cells, glucose deficiency，rep2

description:
Library name: Sample4
Column name in raw_All Samples.txt: 95D_Glc_deficiency_rep2

value: WT


title: DAMER_M1, without ATF4, 1M7

description:
Library name: Sample13
SHAPE-MAP
DAMER_M1_without_ATF4_1M7


title: DAMER_ΔA5B, with ATF4, DMSO

description:
Library name: Sample12
SHAPE-MAP
DAMER_dA5B_with_ATF4_DMSO


title: DAMER_ΔA5B, with ATF4, 1M7

description:
Library name: Sample11
SHAPE-MAP
DAMER_dA5B_with_ATF4_1M7


title: DAMER_ΔA5B, without ATF4, 1M7

description:
Library name: Sample9
SHAPE-MAP
DAMER_dA5B_without_ATF4_1M7


title: DAMER_ΔA5B, without ATF4, DMSO

description:
Library name: Sample10
SHAPE-MAP
DAMER_dA5B_without_ATF4_DMSO


title: DAMER_ΔA4B, with ATF4, DMSO

description:
Library name: Sample8
SHAPE-MAP
DAMER_dA4B_with_ATF4_DMSO


title: DAMER_ΔA4B, with ATF4, 1M7

description:
Library name: Sample7
SHAPE-MAP
DAMER_dA4B_with_ATF4_1M7


title: DAMER_ΔA4B, without ATF4, 1M7

description:
Library name: Sample5
SHAPE-MAP
DAMER_dA4B_without_ATF4_1M7


title: DAMER_ΔA4B, without ATF4, DMSO

description:
Library name: Sample6
SHAPE-MAP
DAMER_dA4B_without_ATF4_DMSO


title: DAMER_WT, with ATF4, DMSO

description:
Library name: Sample4
SHAPE-MAP
DAMER_WT_with_ATF4_DMSO


title: DAMER_WT, with ATF4, 1M7

description:
Library name: Sample3
SHAPE-MAP
DAMER_WT_with_ATF4_1M7


title: DAMER_WT, without ATF4, 1M7

description:
Library name: Sample1
SHAPE-MAP
DAMER_WT_without_ATF4_1M7


title: DAMER_WT, without ATF4, DMSO

description:
Library name: Sample2
SHAPE-MAP
DAMER_WT_without_ATF4_DMSO


title: RNA-Seq, 95D cells, glutamine deficiency

description:
Library name: Sample3
Column name in raw_All Samples.txt: 95D_glucose deficiency

data_processing:
Raw data (raw reads) of fastq format were firstly processed through in-house perl scripts. In this step, clean data (clean reads) were obtained by trimming reads containing adapter, ploy-N or with low quality from raw data. At the same time, Q20, Q30 and GC content of the clean data were calculated. All the downstream analyses were performed based on the clean data with high quality. Reference genome and gene model annotation files were downloaded from genome website directly. Index of the reference genome was built using Hisat2 v2.0.5 and paired-end clean reads were aligned to the reference genome using Hisat2 v2.0.5. We selected Hisat2 as the mapping tool for that Hisat2 can generate a database of splice junctions based on the gene model annotation file and thus a better mapping result than other non-splice mapping tools.
Assembly: hg38
Supplementary files format and content: tab-delimited text file includes raw counts for each Sample
Supplementary files format and content: txt files include FPKM values for each Sample

extract_protocol:
The full-length DAMER lncRNA sequence was transcribed in vitro using the T7 High Yield RNA Synthesis Kit. The transcribed RNA was 3'-end biotinylated using the Pierce RNA 3' End Biotinylation Kit to create the bait probe. Cell lysates were prepared from 95D cells using Pierce IP Lysis Buffer. 50 pmol of biotinylated RNA probe was added to streptavidin magnetic beads and incubated at room temperature for 30 minutes. The probe-bound beads were then incubated with 100 µl of a Master Mix containing the cell lysate for 1 hour at 4°C. RNA bound to the DAMER probe was eluted and purified using TRIzol reagent.
Libraries were constructed from the extracted total RNA using standard protocols, including rRNA depletion, fragmentation, cDNA synthesis, and adapter ligation. The Qualified libraries were pooled and sequenced on Illumina platforms with PE150 strategy in Novogene Bioinformatics Technology Co., Ltd (Beijing, China), according to effective library concentration and data amount required.


title: HC151T

type: RNA

city: Kyoto

first: Makoto

middle: Mark

last: Taketo

state: Kyoto

address: Yoshida-Honmachi, Sakyo

department: Kyoto University Hospital

institute: Kyoto University

description: Gene expression in the serum-containing medium

hyb_protocol: Each aliquot (600 ng) of labeled cRNA was fragmented by adding 5 μl 10x blocking agent and 1 μl of 25x fragmentation buffer, and then heated at 60 °C for 30 min. Finally, 25 μl of 2x GE hybridization buffer was added to dilute the labeled cRNA. Forty microliter of hybridization solution was dispensed into the gasket slide and loaded on the Agilent SurePrint G3 Human GE 8x 60K, V3 Microarrays (Agilent). The slides were incubated for 17 h at 65 °C in an Agilent hybridization oven, and then washed at room temperature by using the Agilent One-Color Microarray-Based Gene Expression Analysis protocol (cited above).

scan_protocol: The hybridized array was immediately scanned with an Agilent Microarray Scanner D (Agilent Technologies, Inc.)

data_processing: Raw data were extracted using the Agilent Feature Extraction Software (v11.0.1.1). The raw data for the same gene was then summarized automatically using the Agilent feature extraction protocol to generate a raw text file, providing expression data for each gene probed on the array.

source_name: CRC-SC

label_protocol: One-hundred ng of total RNA from each sample was linearly amplified and labeled with Cy3-dCTP. The labeled cRNAs were purified by RNAeasy Mini Kit (Qiagen). The concentration and specific activity of the labeled cRNA preparations (pmol Cy3/μg cRNA) were determined by NanoDrop ND-1000 (NanoDrop, Wilmington, DE, USA).

growth_protocol: The method has been described previously (Miyoshi H, Oncotarget 2018;9(31):21950-64.). Briefly, two pieces of the tumor (~1 cm3 each) and one piece of normal mucosa (~1 cm2) were isolated from the mound of a colorectal cancer lesion and the surrounding normal colonic tissue, respectively. They were transferred to the research laboratory in an ice-cold washing medium [DMEM/ F12 with HEPES and L-glutamine (Nacalai Tesque, Kyoto, Japan), 100 units/ml penicillin (Nacalai), 0.1 mg/ml streptomycin (Nacalai), and 10% calf serum (Sigma)]. Tissues were processed within 24 h after surgery. Pieces of the colorectal cancer specimens and fragments of normal colonic mucosa were minced separately using scissors in 60-mm Petri dishes. Each sample was digested in 2 ml of collagenase solution [the washing medium supplemented with 0.2% collagenase type I (Thermo Fisher Scientific, Waltham, MA) and 50 μg/ml gentamicin (Thermo Fisher)] at 37 °C for 40–60 min, and cells were dissociated by pipetting. Colorectal cancer and normal colonic epithelial cells were then filtered once through 100 μm-diameter cell strainers (Corning Inc., Corning, NY), and collected by centrifugation. Isolated cells were resuspended in Matrigel (Corning), and placed in the center of each well of a 12-well cell culture plate (30 μl per well; TPP, Trasadingen, Switzerland). After gellation of Matrigel at 37 °C, epithelial cells were cultured at 37 °C in 5% CO2 in Advanced DMEM/F-12 (Thermo Fisher) supplemented with penicillin (100 units/ml, Nacalai), streptomycin (0.1 mg/ml, Nacalai), L-glutamine (2 mM), Y27632 (10 μM) and SB431542 (1 μM). NCE-SC cells were cultured in the eL-WRN medium as described previously (Miyoshi H, Oncotarget 2018;9(31):21950-64.).

extract_protocol: NCE-SC or CRC-SC were cultured as descried previously (Miyoshi H, Oncotarget 2018;9(31):21950-64.). After aspiration of the medium, lysis buffer (TaKaRa Bio, Kusatsu, Japan) was directly added to each well of the spheroid culture. Total RNA of NCE-SC and CRC-SC spheroid lines was prepared using NucleoSpin RNA II kit (TaKaRa Bio, Kusatsu, Japan) according to the manufacturer’s protocol. RNA was quantified using a NanoDrop-1000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA). Microarray analysis was performed by Macrogen Inc. (Seoul, Republic of Korea).

treatment_protocol: Epithelial cells were cultured at 37 °C in 5% CO2 in Advanced DMEM/F-12 (Thermo Fisher) supplemented with penicillin (100 units/ml, Nacalai), streptomycin (0.1 mg/ml, Nacalai), L-glutamine (2 mM), Y27632 (10 μM), SB431542 (1 μM). NCE-SC cells were cultured in the eL-WRN medium as described previously (Miyoshi H, Oncotarget 2018;9(31):21950-64.).

tag: pathology

value: Cancer


title: HC149T


title: RNA-Seq, 95D cells, glucose deficiency

description:
Library name: Sample2
Column name in raw_All Samples.txt: 95D_glucose deficiency


title: A subline for data normalization


title: RNA-Seq, 95D cells, control

description:
Library name: Sample1
Column name in raw_All Samples.txt: 95D_Control

value: cultured in control conditions for 48 hours


title: 95D cells, glutamine deficiency


title: 95D cells, glucose deficiency

description:
Library name: Sample1
Column name in raw_All Samples.txt: 95D_glucose deficiency


title: HC146T


title: HC122T


title: HC101T


title: HC62T


title: HC129T


title: Synchronous S2, TRESLIN-MTBP over-expression, LIG1/LIG3-mAID, 5-Ph-IAA

city: Mishima

first: Masato

last: Kanemaki

state: Shizuoka

address: Yata 1111

department: Department of Chromosome Science

institute: National Institute of Genetics

description: Library name: Sync LD-OK-seq TRESLIN-MTBP over-ex S2

data_processing:
Raw sequencing reads (FASTQ) were initially quality-checked using FastQC (v0.11.9). Data was processed using Trim Galore (v0.6.6) to remove Illumina sequencing adapters (AGATCGGAAGAGC) with the '--illumina' parameter, and 10 bp were trimmed from the 5' end of both read pairs using the parameters '--clip_R1 10' and '--clip_R2 10'. Cleaned reads were aligned to the GRCh38 (NCBI) reference genome using Bowtie2 (v2.2.5) with parameters: --end-to-end --very-sensitive --no-mixed --no-discordant -I 0 -X 1000. The resulting alignments were converted to BAM format and filtered using SAMtools (v1.12) 'view' with three criteria: mapping quality score MAPQ ≥ 20 (-q 20), proper pair mapping (-f 0x2), and removal of unmapped reads (-F 0x4). PCR duplicates were marked and removed using SAMtools 'sort', 'fixmate', and 'markdup' functions. Filtered reads were separated by strand orientation using SAMtools 'view': Crick strand reads were captured using flags '-f 128 -F 16' and '-f 64 -F 32', while Watson strand reads were captured using flags '-f 144' and '-f 96'. Read coverage was calculated in 1 kb genomic bins using BEDtools (v2.30.0) 'intersect'. Raw bin counts were normalised by sequencing depth using a custom R function (norm.by.depth, available on GitHub) and converted to bigWig format using bedGraphToBigWig (v2.8).
Assembly: GRCh38
Supplementary files format and content: bigwig (Okazaki reads in 1 kb bins), bed (initiation zones)

extract_protocol:
Approximately 7 million cells were collected for each sample and processed to purify genomic DNA using Blood & Cell Culture DNA Midi Kit (QIAGEN, #13343) with 100/G Genomic-tips. To examine single-stranded Okazaki fragments, 30 µg of genomic DNA was mixed with 2×RNA loading dye (95% formamide, 0.02% SDS, 0.04% Orange G, 1 mM EDTA) and denatured at 70°C for 10 min followed by 10 min incubation on ice. The DNA samples were loaded onto a 1% alkaline agarose gel and electrophoresed for 45 min at 70 V at 4°C. Gels were neutralised in 1×TAE buffer for 30 min and then stained with SYBR Gold (Invitrogen, #S11494) for 1 h at room temperature. Single-stranded DNA ranging from 100 to 650 bp was excised from the gel, which was subsequently processed for ssDNA purification using NucleoSpin Gel and PCR Clean-up Kit (Takara, #740609) with NTC buffer (Takara, #U0654A). Quantification of ssDNA was performed using Quant-iT OliGreen ssDNA Assay Kit (Invitrogen, #O7582). To convert purified ssDNA to dsDNA, 50-100 ng of ssDNA in 30 µl was mixed with 5 µl of 10×NEBuffer 2 (NEB, #B7002S) and 5 µl of 3 mg/ml 8N random primers (Eurofins). The mixture was incubated at 95°C for 5 min followed by cooling for 5 min on ice, and then supplemented with 5 µl of 2 mM dNTP mix (without dTTP, Roche, #11969064001) plus dUTP (Roche, #11934554001), 2.5 µl of 5,000 U/ml Klenow DNA polymerase (NEB, #M0210S), and nuclease-free H2O to a final volume of 50 µl. The supplemented mixture was incubated at 37°C for 20 min. Reactions were stopped by adding 5 µl of 0.5 M EDTA. The dsDNA was purified using AMPure XP beads (Beckman Coulter, #A63881) and quantified with Quant-iT PicoGreen dsDNA Assay Kit (Invitrogen, #P7589) before proceeding to library preparation.
Sequencing libraries were prepared using NEBNext Ultra II DNA Library Prep Kit for Illumina (NEB, #E7645S) according to the manufacturer's instructions. Sequencing was performed using the Illumina NovaSeq 6000 platform with 100 bp paired-end reads for approximately 100 million read pairs per sample.

value: Colonic epithelial cells

value: CMV-OsTIR1 (F74G) LIG1-mAC LIG3-mAC Tet-TRESLIN Tet-MTBP

value: Doxycycline for 48 hours, nocodazole for 15 hours and release for 8 hours, 5-Ph-IAA for 8 hours


title: HC120T


title: HC42T


title: HC55T


title: HC72T


title: Synchronous S1, TRESLIN-MTBP over-expression, LIG1/LIG3-mAID, 5-Ph-IAA

description: Library name: Sync LD-OK-seq TRESLIN-MTBP over-ex S1

value: Doxycycline for 48 hours, nocodazole for 15 hours and release for 6 hours, 5-Ph-IAA for 6 hours


title: HC90T


title: HC89T


title: Asynchronous, RIF1-KO + rescue with RIF1-ppC, LIG1/LIG3-mAID, 5-Ph-IAA, DMSO

description: Library name: Async LD-OK-seq RIF1-KO rescue RIF1-ppC DMSO

value: CMV-OsTIR1 (F74G) LIG1-mAC LIG3-mAC RIF1-KO Tet-RIF1-ppC

value: DMSO for 48 hours, 5-Ph-IAA for 4 hours


title: HC102T


title: HC94T


title: HC14T


title: HC93T


title: HC127T


title: Asynchronous, RIF1-KO + rescue with RIF1-WT, LIG1/LIG3-mAID, 5-Ph-IAA, Dox

description: Library name: Async LD-OK-seq RIF1-KO rescue RIF1-WT Doxycycline

value: CMV-OsTIR1 (F74G) LIG1-mAC LIG3-mAC RIF1-KO Tet-RIF1-WT

value: Doxycycline for 48 hours, 5-Ph-IAA for 4 hours


title: HC97T


title: HC83T


title: HC40T


title: Synchronous S4, RIF1-KO, LIG1/LIG3-mAID, 5-Ph-IAA

description: Library name: Sync LD-OK-seq RIF1-KO S4

value: CMV-OsTIR1 (F74G) LIG1-mAC LIG3-mAC RIF1-KO

value: Nocodazole for 15 hours and release for 12 hours, 5-Ph-IAA for 4 hours


title: HC80T


title: Asynchronous, RIF1-KO + rescue with RIF1-ppC, LIG1/LIG3-mAID, 5-Ph-IAA, Dox

description: Library name: Async LD-OK-seq RIF1-KO rescue RIF1-ppC Doxycycline


title: Asynchronous, RIF1-KO + rescue with RIF1-WT, LIG1/LIG3-mAID, 5-Ph-IAA, DMSO

description: Library name: Async LD-OK-seq RIF1-KO rescue RIF1-WT DMSO


title: Synchronous S2, RIF1-KO, LIG1/LIG3-mAID, 5-Ph-IAA

description: Library name: Sync LD-OK-seq RIF1-KO S2

value: Nocodazole for 15 hours and release for 8 hours, 5-Ph-IAA for 8 hours


title: Synchronous S3, RIF1-KO, LIG1/LIG3-mAID, 5-Ph-IAA

description: Library name: Sync LD-OK-seq RIF1-KO S3

value: Nocodazole for 15 hours and release for 10 hours, 5-Ph-IAA for 4 hours


title: HC91T


title: Synchronous S6, RIF1-KO, LIG1/LIG3-mAID, 5-Ph-IAA

description: Library name: Sync LD-OK-seq RIF1-KO S6

value: Nocodazole for 15 hours and release for 16 hours, 5-Ph-IAA for 4 hours


title: Synchronous S1, RIF1-KO, LIG1/LIG3-mAID, 5-Ph-IAA

description: Library name: Sync LD-OK-seq RIF1-KO S1

value: Nocodazole for 15 hours and release for 6 hours, 5-Ph-IAA for 6 hours


title: Synchronous S5, RIF1-KO, LIG1/LIG3-mAID, 5-Ph-IAA

description: Library name: Sync LD-OK-seq RIF1-KO S5

value: Nocodazole for 15 hours and release for 14 hours, 5-Ph-IAA for 4 hours


title: Asynchronous, RIF1-KO, LIG1/LIG3-mAID, 5-Ph-IAA, Rep2

description: Library name: Async LD-OK-seq RIF1-KO Rep2

value: 5-Ph-IAA for 4 hours


title: HCX1T

value: Cancer


title: HC36T


title: HC77T


title: Synchronous S5, LIG1/LIG3-mAID, 5-Ph-IAA

description: Library name: Sync LD-OK-seq S5

value: Colonic epithelial cells

value: CMV-OsTIR1 (F74G) LIG1-mAC LIG3-mAC


title: Synchronous S6, LIG1/LIG3-mAID, 5-Ph-IAA

description: Library name: Sync LD-OK-seq S6


title: Synchronous S3, LIG1/LIG3-mAID, 5-Ph-IAA

description: Library name: Sync LD-OK-seq S3


title: Synchronous S2, LIG1/LIG3-mAID, 5-Ph-IAA

description: Library name: Sync LD-OK-seq S2


title: Synchronous S1, LIG1/LIG3-mAID, 5-Ph-IAA

description:
Library name: Sync LD-OK-seq S1
IZs were selected from the six time-course samples


title: Asynchronous, LIG1/LIG3-mAID, ATRi, 5-Ph-IAA, Rep2

description: Library name: Async LD-OK-seq ATRi 5-Ph-IAA Rep2

value: 5-Ph-IAA for 4 hours, ATRi for 5 hours


title: Asynchronous, RIF1-KO, LIG1/LIG3-mAID, 5-Ph-IAA, Rep1

description: Library name: Async LD-OK-seq RIF1-KO Rep1


title: HC168T


title: HC11T


title: Synchronous S4, LIG1/LIG3-mAID, 5-Ph-IAA

description: Library name: Sync LD-OK-seq S4


title: HC144T


title: HC54T


title: Asynchronous, LIG1/LIG3-mAID, ATRi, DMSO

description: Library name: Async LD-OK-seq ATRi DMSO

value: DMSO for 4 hours, ATRi for 5 hours


title: HC20T


title: Asynchronous, LIG1/LIG3-mAID, 5-Ph-IAA, Rep1

description:
Library name: Async LD-OK-seq 5-Ph-IAA Rep1 (LIG1-mAID LIG3-mAID)
IZs were selected from the two replicates


title: HC15T


title: HC19T


title: HC3T


title: HC108T


title: HC117T


title: HC70T


title: HC58T


title: HC74T


title: Asynchronous, LIG1/LIG3-mAID, ATRi, 5-Ph-IAA, Rep1

description:
Library name: Async LD-OK-seq ATRi 5-Ph-IAA Rep1
IZs were selected from the two replicates


title: HC73T


title: Asynchronous, LIG3-mAID, DMSO

description: Library name: Async LD-OK-seq DMSO (LIG3-mAID)

value: CMV-OsTIR1 (F74G) LIG3-mAC

value: DMSO for 4 hours


title: Asynchronous, LIG1/LIG3-mAID, 5-Ph-IAA, Rep2

description: Library name: Async LD-OK-seq 5-Ph-IAA Rep2 (LIG1-mAID LIG3-mAID)


title: Asynchronous, LIG1-mAID, 5-Ph-IAA

description: Library name: Async LD-OK-seq 5-Ph-IAA (LIG1-mAID)

value: CMV-OsTIR1 (F74G) LIG1-mAC


title: HC50T

description: Gene expression in the serum-containing medium


title: HC59T


title: HC70N

source_name: NCE-SC

value: Normal


title: HC73N


title: HC59N


title: HC17T


title: TRESLIN FLAG-IP with GINS4-mAID, Early-S, DDKi (Input)

description: Library name: TRESLIN FLAG-IP Early-S GINS4-mAID DDKi Input

data_processing:
Raw FASTQ reads were processed using Trim Galore with default parameters to remove adapter sequences. The trimmed reads were then aligned to the GRCh38 reference genome using Bowtie2, applying the same parameters used for LD-OK-seq data processing. After alignment, reads were filtered, and PCR duplicates were removed using SAMtools. Signal track generation was performed using MACS2 (v2.2.7.1) to produce treatment (IP DNA) and control (Input) bedGraph files, which were subsequently converted to bigWig format using bedGraphToBigWig. Finally, IP signals were normalised against input using deepTools (v3.5.5) ‘bigwigCompare’ with the following parameters: --operation ratio --pseudocount 0 -bs 100. This normalization process divided the genome into 100 bp bins and calculated the IP/Input ratio within each bin. The input-normalized signal was used to generate all plots in this study.
Assembly: GRCh38
Supplementary files format and content: bigwig (input normalized IP ration)

extract_protocol:
Approximately 2×107 cells were harvested and lysed in 1 ml of IP lysis buffer (20 mM HEPES pH 7.5, 150 mM NaCl, 1 mM EDTA, 0.5% NP-40, 5% glycerol, 3 mM MgCl2, 100 U benzonase (Millipore, #70746), complete protease inhibitor cocktail (Roche, #11873580001), and phosphatase inhibitor cocktail (Roche, #490684500)) at 10°C for 30 min. The lysate was centrifuged twice at 14,000 rpm for 15 min at 4°C to remove insoluble debris, followed by a pre-clearing step with 0.75 mg of Protein G Dynabeads (Invitrogen, #10004D) at 4°C for 1 h. Pre-cleared lysate was incubated with 1.5 µg of Protein G Dynabeads pre-bound with anti-FLAG M2 antibody (Sigma, #F1804) for 2 h at 4°C. Beads were washed twice with 1ml of IP wash buffer (20 mM HEPES pH 7.5, 150 mM NaCl, 1 mM EDTA, 0.5% NP-40, 5% glycerol, complete protease inhibitor cocktail, and phosphatase inhibitor cocktail). Bound DNA was eluated twice by adding 90 µl of elution buffer (TE + 1% SDS) and subsequently by incubating at 45°C for 10 min. The combined eluate was treated with 20 µg RNase A for 1 h at 37°C, followed by treatment with 40 µg Proteinase K for 2 h at 50°C. DNA was purified using NucleoSpin Gel and PCR Clean-up Kit and quantified.
NGS libraries were prepared from 2 ng of DNA using NEBNext Ultra II DNA Library Prep Kit for Illumina, and 50 bp paired-end sequencing was performed on the Illumina NovaSeq 6000 platform.

value: HCT116 CMV-OsTIR1 (F74G) GINS4-mAID Stag-3xFLAG-TRESLIN

value: Nocodazole for 15 hours and release for 6 hours, 5-Ph-IAA for 4 hours, DDKi for 4 hours

tag: chip antibody


title: TRESLIN FLAG-IP with GINS4-mAID, Early-S, DDKi (IP)

description: Library name: TRESLIN FLAG-IP Early-S GINS4-mAID DDKi IP

value: FLAG M2 (Sigma, F1804)


title: Asynchronous, LIG1/LIG3-mAID, DMSO

description: Library name: Async LD-OK-seq DMSO (LIG1-mAID LIG3-mAID)


title: Asynchronous, LIG3-mAID, 5-Ph-IAA

description: Library name: Async LD-OK-seq 5-Ph-IAA (LIG3-mAID)


title: HC50N


title: HC3N


title: TRESLIN FLAG-IP with GINS4-mAID, Early-S, 5phIAA (IP)

description: Library name: TRESLIN FLAG-IP Early-S GINS4-mAID 5phIAA IP

value: Nocodazole for 15 hours and release for 6 hours, 5-Ph-IAA for 4 hours


title: TRESLIN FLAG-IP with GINS4-mAID, Early-S, DMSO (Input)

description: Library name: TRESLIN FLAG-IP Early-S GINS4-mAID DMSO Input

value: Nocodazole for 15 hours and release for 6 hours, DMSO for 4 hours


title: Asynchronous, LIG1-mAID, DMSO

description: Library name: Async LD-OK-seq DMSO (LIG1-mAID)


title: HC3Nd

source_name: Differentiated 3N


title: TRESLIN FLAG-IP with GINS4-mAID, Early-S, 5phIAA (Input)

description: Library name: TRESLIN FLAG-IP Early-S GINS4-mAID 5phIAA Input


title: pre-leukemic HSPCs, multiplexed scRNA-Seq, sublibrary 8

city: Stanford

first: Thomas

last: Koehnke

state: CA

address: 265 Campus Drive

institute: Stanford University

description: Library name: sublibrary 8

data_processing:
Read trimming, quality control, alignment and count matrix generation was performed using the Parse Pipeline v.1.1.2. Data was processed using Seurat v5.0.3, pre-processing and normalization was performed using the SCTransformv2, putative doublets were removed using DoubletFinder v2.0.4 and samples were integrated using HarmonyIntegration. Cell types were assigned using Cell Marker Accordion and GSEA was performed using fgsea. For genotyping of the DNMT3A 882 codon from single-cell RNA-Seq data, all reads containing any 20-mers motifs found in the final DNMT3A exon, both for the endogenous allele as well as for our codon modified HDR alleles were extracted from raw FASTQ files using fxtract (https://github.com/ctSkennerton/fxtract). These reads were then re-aligned to the DNMT3A locus and genotypes determined at codon 882. Cell barcodes for each read were extracted and genotyping information was added to the Seurat object as metadata.
Assembly: human GRCh38 reference genome (release 104)
Supplementary files format and content: Seurat object containing expression values and metadata

source_name: Blood

growth_protocol: residual HSPCs were purified from patient samples and cells were cultured at 37°C, 5% CO2 and 5% O2 in in growth media comprised of StemSpan SFEM II (STEMCELL Technologies) base media supplemented with 20 ng/mL of TPO, SCF, FLT3L, and IL-6 (Peprotech) and 35nM of UM-171 (Selleckchem) for 48h.  Synthetic, chemically modified sgRNAs targeting DNMT3A:R882H (5’-CGTCTCCAACATGAGCCACT-3') and Cas9 protein (Alt-R HiFi CRISPR-Cas9) were precomplexed at a molar ratio of 1:2.5 at 25°C for 10 min immediately prior to electroporation. Cells were electroporated using the Lonza Nucleofector 4D (program DZ-100) using 150 µg/mL Cas9 protein, with the addition of 20 pmol siTP53 in P3 Primary Cell Nucleofector Solution (Lonza). Immediately after electroporation, cells were plated with rAAV6 particles at a multiplicity of infection of 5,000 in growth media as above, supplemented with 0.5 µM AZD7648. rAAV6 contain two homology arms of 400 bp flanking the remaining, codon-optimized corrected (R882R) or re-mutated (R882H) coding sequence of the final DNMT3A exon, as well as a cassette containing the UBC promoter and either turboGFP (for DNMT3A correction) or mTagBFP2 (for DNMT3A re-mutation). Cells were washed after 8h and cultured in growth media for a total of 3 days after electroporation before isolation of fluorescent protein positive populations by FACS. 200-500 edited residual HSCs were added to 4 mL of MethoCult H4435 (Stemcell Technologies) and differentiated for 14 days. Cells were washed and live cells were sorted by FACS.

extract_protocol:
Samples were fixed using the Parse Biosciences Cell Fixation kit (Parse Biosciences).
Barcoding and library preparation was performed using the the Evercode Whole Transcriptome (v2) kit (Parse Biosciences). Sample loading (multiplexing) is detailed in supplementary file sample_load_table.txt. 8 sublibraries were prepared and sequenced.

value: Blood

value: pre-leukemic HSPCs

value: DNMT3A Correction (R882R) or ReMutation (R882H)


title: TRESLIN FLAG-IP with GINS4-mAID, Early-S, DMSO (IP)

description: Library name: TRESLIN FLAG-IP Early-S GINS4-mAID DMSO IP


title: pre-leukemic HSPCs, multiplexed scRNA-Seq, sublibrary 7

description: Library name: sublibrary 7


title: IP control with untagged cells, Early-S (IP)

description: Library name: FLAG-IP untagged control IP

value: HCT116 CMV-OsTIR1 (F74G) LIG1-mAC LIG3-mAC

value: Nocodazole for 15 hours and release for 6 hours


title: IP control with untagged cells, Early-S (Input)

description: Library name: FLAG-IP untagged control Input

value: Colonic epithelial cells


title: pre-leukemic HSPCs, multiplexed scRNA-Seq, sublibrary 6

description: Library name: sublibrary 6


title: 3xFLAG-TRESLIN GINS4-mAID cells, Early-S, 5phIAA, DDKi (ChIP)

description: Library name: TRESLIN Early-S GINS4-mAID 5phIAA DDKi ChIP

data_processing:
Raw FASTQ data quality was assessed using FastQC, followed by Illumina adapter removal using Trim Galore with default settings. Reads were aligned to the GRCh38 reference genome using Bowtie2. Unmapped reads and reads with mapping quality lower than 20 were filtered out. Duplicated reads were removed using SAMtools. Signal tracks were generated using MACS2 for both Input and ChIP samples. ChIP signals were normalized against Input signals using deepTools across the genome in 100 bp bins.
Assembly: GRCh38
Supplementary files format and content: bigwig (input normalized signal)

growth_protocol: HCT116 cells were maintained in McCoy's 5A medium supplemented with 10% fetal bovine serum (FBS), 2 mM L-glutamine, 100 U/ml penicillin, and 100 µg/ml streptomycin at 37°C in an incubator with 5% CO₂.

extract_protocol:
20 million cells were collected by trypsinization and resuspended in 1 ml PBS. Cells were crosslinked at room temperature for 10 min by adding an equal volume of 2% PFA. Crosslinked cells were washed with PBS and 0.5% NP-40 in PBS and then lysed in 1 ml ChIP lysis buffer. Chromatin was fragmented using Covaris S2. Lysate was centrifuged at top speed and then pre-cleared with empty protein G dynabeads for at least 1 hour. Cleared lysate was then incubated with FLAG antibody conjugated protein G dynabeads overnight. Beads were washed with a series of washing buffers and then proteins were eluted using 1% SDS in TE buffer. Eluates were reverse crosslinked with proteinase K at 65 ºC for at least 4 hours. DNA was purified using the NucleoSpin Gel and PCR Clean-up Kit.
NGS libraries were prepared from 5-10 ng DNA using the NEBNext Ultra II DNA Library Prep Kit for Illumina following the manufacturer's instructions.

value: Nocodazole for 15 hours and release for 6 hours, 5-Ph-IAA for 6 hours, DDKi for 5 hours


title: 3xFLAG-TRESLIN GINS4-mAID cells, Early-S, 5phIAA, DDKi (Input)

description: Library name: TRESLIN Early-S GINS4-mAID 5phIAA DDKi Input


title: HC23T

value: Cancer


title: HC18T


title: HC10T


title: 3xFLAG-TRESLIN RIF1-KO GINS4-mAID cells, Early-S, 5phIAA (Input)

description: Library name: TRESLIN Early-S RIF1-KO GINS4-mAID 5phIAA Input

value: HCT116 CMV-OsTIR1 (F74G) GINS4-mAID Stag-3xFLAG-TRESLIN RIF1-KO

value: Nocodazole for 15 hours and release for 6 hours, 5-Ph-IAA for 6 hours


title: 3xFLAG-TRESLIN RIF1-KO GINS4-mAID cells, Early-S, 5phIAA (ChIP)

description: Library name: TRESLIN Early-S RIF1-KO GINS4-mAID 5phIAA ChIP


title: 3xFLAG-TRESLIN RIF1-KO GINS4-mAID cells, Early-S, DMSO (Input)

description: Library name: TRESLIN Early-S RIF1-KO GINS4-mAID DMSO Input

value: Nocodazole for 15 hours and release for 6 hours, DMSO for 6 hours


title: HC22T

description: Gene expression in the serum-containing medium


title: 3xFLAG-TRESLIN RIF1-KO GINS4-mAID cells, Early-S, DMSO (ChIP)

description: Library name: TRESLIN Early-S RIF1-KO GINS4-mAID DMSO ChIP


title: HC16T


title: HC13T


title: HC35T


title: pre-leukemic HSPCs, multiplexed scRNA-Seq, sublibrary 4

description: Library name: sublibrary 4


title: pre-leukemic HSPCs, multiplexed scRNA-Seq, sublibrary 3

description: Library name: sublibrary 3


title: pre-leukemic HSPCs, multiplexed scRNA-Seq, sublibrary 2

description: Library name: sublibrary 2


title: HC8T


title: MTBP-3xFLAG CDC45-mAID cells, Early-S, 5phIAA (ChIP)

description: Library name: MTBP Early-S CDC45-mAID 5phIAA ChIP

value: HCT116 CMV-OsTIR1 (F74G) CDC45-mAID MTBP-Stag-3xFLAG


title: 3xFLAG-TRESLIN RIF1-KO cells, Early-G1 (Input)

description: Library name: TRESLIN Early-G1 RIF1-KO Input

value: Nocodazole for 15 hours and release for 2 hours


title: MTBP-3xFLAG CDC45-mAID cells, Early-S, DMSO (ChIP)

description: Library name: MTBP Early-S CDC45-mAID DMSO ChIP


title: MTBP-3xFLAG CDC45-mAID cells, Early-S, 5phIAA (Input)

description: Library name: MTBP Early-S CDC45-mAID 5phIAA Input


title: MTBP-3xFLAG CDC45-mAID cells, Early-S, DMSO (Input)

description: Library name: MTBP Early-S CDC45-mAID DMSO Input


title: 3xFLAG-TRESLIN RIF1-KO cells, Early-G1 (ChIP)

description: Library name: TRESLIN Early-G1 RIF1-KO ChIP


title: pre-leukemic HSPCs, multiplexed scRNA-Seq, sublibrary 5

description: Library name: sublibrary 5


title: pre-leukemic HSPCs, multiplexed scRNA-Seq, sublibrary 1

description: Library name: sublibrary 1


title: MTBP-3xFLAG cells, Early-G1 (ChIP)

description: Library name: MTBP Early-G1 ChIP


title: HC6T


title: HC9T


title: MTBP-3xFLAG cells, Early-G1 (Input)

description: Library name: MTBP Early-G1 Input


title: RNA-seq with RIF1-KO cells (HCT116 LIG1/3-mAID2 RIF1-KO), Rep3

description: Library name: RNA-seq RIF1-KO cells Rep3

data_processing:
Raw FASTQ data quality was assessed using FastQC, followed by Illumina adapter removal using Trim Galore with default settings. Clean reads were aligned to the GRCh38 reference genome using HISAT2 (v2.2.1) with parameters '--no-mixed --no-discordant --rna-strandness RF'. The aligned reads were converted to BAM format and filtered using the same criteria as in the LD-OK-seq data processing. Read counts per transcript were quantified from the filtered BAM files using featureCounts (v2.0.6) with options '-s 2 -F GTF -g gene_id -t exon -B -C -p -countReadPairs', which enables strand-specific counting while treating each read pair as a single fragment. The annotation GTF file was obtained from the NCBI MANE (Matched Annotation from NCBI and EMBL-EBI) project14 and filtered to include only protein-coding genes. Raw read counts were normalized to transcripts per million (TPM) using R.
Assembly: GRCh38
Supplementary files format and content: csv files (including raw read counts, TPM and log2TPM for protein-coding genes)

extract_protocol:
Total RNA was extracted from cells using illustra RNAspin columns (Cytiva, #25050071). Ribosomal RNA was removed using NEBNext rRNA Depletion Kit v2 (NEB, #E7405L).
RNA libraries were prepared with NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (NEB, #E7760S), following the manufacturer's instructions. Sequencing was performed on the HiSeq 2500 platform with paired-end 50 bp reads to obtain at least 40 million read pairs per sample.

value: HCT116 CMV-OsTIR1 (F74G) LIG1-mAC LIG3-mAC RIF1-KO


title: HC7T


title: 3xFLAG-TRESLIN GINS4-mAID cells, Early-S, 5phIAA (ChIP)

description: Library name: TRESLIN Early-S GINS4-mAID 5phIAA ChIP

value: HCT116 CMV-OsTIR1 (F74G) GINS4-mAID Stag-3xFLAG-TRESLIN

value: FLAG M2 (Sigma, F1804)


title: HC2T


title: 3xFLAG-TRESLIN GINS4-mAID cells, Early-S, 5phIAA (Input)

description: Library name: TRESLIN Early-S GINS4-mAID 5phIAA Input


title: RNA-seq with RIF1-KO cells (HCT116 LIG1/3-mAID2 RIF1-KO), Rep2

description: Library name: RNA-seq RIF1-KO cells Rep2


title: SU575, DNMT3A ReMutation, Xenograft 156_5

description: Library name: SU575_156_5_ReMut

data_processing:
Sequencing reads were trimmed and quality control was performed using TrimGalore v.0.6.6 (https://doi.org/10.5281/zenodo.5127899). The cDNA index was built using Kallisto Bustools v.0.40.0 using gene annotations downloaded from Ensembl (release 110) and the GRCh37 reference genome. Pseudo-alignment was performed using Kallisto v0.44.0. Data was imported into R using tximport v.1.30.0 using gene mode. Differential testing was performed using DESeq2 v1.42.1.
Assembly: human GRCh37 reference genome (release 110)
Supplementary files format and content: txt file includes estimated counts per sample

growth_protocol: Purified AML blasts were cultured at 37°C, 5% CO2 and 21% O2 in StemSpan SFEM II (STEMCELL Technologies) base media supplemented with 20 ng/mL of TPO, SCF, FLT3L, GM-CSF and IL-6 (Peprotech) for 48h.  Synthetic, chemically modified sgRNAs targeting DNMT3A:R882H (5’-CGTCTCCAACATGAGCCACT-3') and Cas9 protein (Alt-R HiFi CRISPR-Cas9) were precomplexed at a molar ratio of 1:2.5 at 25°C for 10 min immediately prior to electroporation. Cells were electroporated using the Lonza Nucleofector 4D (program DZ-100) using 150 µg/mL Cas9 protein, with the addition of 20 pmol siTP53 in P3 Primary Cell Nucleofector Solution (Lonza). Immediately after electroporation, cells were plated with rAAV6 particles at a multiplicity of infection of 5,000 in growth media as above, supplemented with 0.5 µM AZD7648. rAAV6 contain two homology arms of 400 bp flanking the remaining, codon-optimized corrected (R882R) or re-mutated (R882H) coding sequence of the final DNMT3A exon, as well as a cassette containing the UBC promoter and either turboGFP (for DNMT3A correction) or mTagBFP2 (for DNMT3A re-mutation). Cells were washed after 8h and cultured in growth media for a total of 3 days after electroporation before isolation of fluorescent protein positive populations by FACS. Isolated cells were engrafted into 6 to 8 week old NOD.Cg-Prkdcscid Il2rgtm1WjlTg(CMV-IL3,CSF2,KITLG)1Eav/MloySzJ (NSGS) mice 2 to 24h after sublethal irradiation (200 rad) by intrafemoral (primary AML studies) injection. 4 months later, mice were humanely euthanized, and femurs, tibias, hip bones, sternum, and spine were harvested and crushed. Mononuclear cells were isolated by Ficoll-Paque PLUS (GE Healthcare) density gradient centrifugation, red blood cells were removed with ACK lysis (ACK Lysing Buffer, ThermoFisher Scientific), and stained for flow cytometry. Human, fluorescent protein positive populations were isolated by FACS and subjected to DNA and RNA isolation.

extract_protocol:
Total RNA was extracted from >100,000 cells using DNA/RNA microprep plus kit (Zymo) following manufacturer’s protocol.
Illumina TruSeq stranded mRNA library preparation was performed according to the manufacturer's protocol

value: AML blasts

value: DNMT3A ReMutation (R882H)


title: 3xFLAG-TRESLIN GINS4-mAID cells, Early-S, DMSO (Input)

description: Library name: TRESLIN Early-S GINS4-mAID DMSO Input


title: SU575, DNMT3A ReMutation, Xenograft 156_4

description: Library name: SU575_156_4_ReMut

value: Blood


title: RNA-seq with RIF1-KO cells (HCT116 LIG1/3-mAID2 RIF1-KO), Rep1

description: Library name: RNA-seq RIF1-KO cells Rep1


title: 3xFLAG-TRESLIN GINS4-mAID cells, Early-S, DMSO (ChIP)

description: Library name: TRESLIN Early-S GINS4-mAID DMSO ChIP


title: 3xFLAG-TRESLIN cells, Early-G1 (ChIP)

description: Library name: TRESLIN Early-G1 ChIP


title: SU575, DNMT3A ReMutation, Xenograft 157_1

description: Library name: SU575_157_1_ReMut


title: MCM4-3xFLAG RIF1-KO cells, Late-G1 (ChIP)

description: Library name: MCM4 G1 RIF1-KO ChIP

value: HCT116 CMV-OsTIR1 (F74G) LIG1-mAC LIG3-mAC MCM4-Stag-3xFLAG RIF1-KO

value: Nocodazole for 15 hours and release for 4 hours


title: 3xFLAG-TRESLIN cells, Early-G1 (Input)

description: Library name: TRESLIN Early-G1 Input


title: SU575, DNMT3A Correction, Xenograft 156_2

description: Library name: SU575_156_2_Corr

value: DNMT3A Correction (R882R)


title: SU575, DNMT3A Correction, Xenograft 156_3

description: Library name: SU575_156_3_Corr


title: MCM4-3xFLAG RIF1-KO cells, Late-G1 (Input)

description: Library name: MCM4 G1 RIF1-KO Input

value: Colonic epithelial cells


title: MCM2-3xFLAG RIF1-KO cells, Late-G1 (ChIP)

description: Library name: MCM2 G1 RIF1-KO ChIP

value: HCT116 CMV-OsTIR1 (F74G) LIG1-mAC LIG3-mAC MCM2-Stag-3xFLAG RIF1-KO


title: HC5T

value: Cancer


title: RNA-seq with control cells (HCT116 LIG1/3-mAID2), Rep3

description: Library name: RNA-seq Control cells Rep3

value: HCT116 CMV-OsTIR1 (F74G) LIG1-mAC LIG3-mAC


title: SU575, DNMT3A Correction, Xenograft 156_1

description: Library name: SU575_156_1_Corr


title: HC1T


title: MCM2-3xFLAG RIF1-KO cells, Late-G1 (Input)

description: Library name: MCM2 G1 RIF1-KO Input


title: 3xFLAG-ORC1 RIF1-KO cells, Late-G1 (ChIP)

description: Library name: ORC1 G1 RIF1-KO ChIP

value: HCT116 CMV-OsTIR1 (F74G) Stag-3xFLAG-ORC1 RIF1-KO


title: RNA-seq with control cells (HCT116 LIG1/3-mAID2), Rep1

description: Library name: RNA-seq Control cells Rep1


title: SU540, DNMT3A ReMutation, Xenograft 155_4

description: Library name: SU540_155_4_ReMut


title: 3xFLAG-ORC1 RIF1-KO cells, Late-G1 (Input)

description: Library name: ORC1 G1 RIF1-KO Input


title: SU540, DNMT3A ReMutation, Xenograft 155_3

description: Library name: SU540_155_3_ReMut


title: nucleus pulposus,treat

city: Beijing

first: Lixiang

last: Ding

state: None Selected

address: No. 10, Tieyi Road, Yangfangdian, Haidian District, Beijing

institute: Beijing Shijitan Hospital, Capital Medical University

description: Library name: Sample1

data_processing:
Raw reads were demultiplexed and mapped to the reference genome by CelScene® pipeline using default parameters. Alignment, filtering, barcode counting, and UMI counting were performed with CelScene® count module to generate feature-barcode matrix and determine clusters.Dimensionality reduction was performed using PCA and the first ten principle compoments were used to generate clusters by K-means algorithm and graph-based algorithm, respectively.
Assembly: mm10-2020-A
Supplementary files format and content: Tab-separated values fles and matrix files

source_name: nucleus pulposus

extract_protocol:
The nucleus pulposus tissue was cut into pieces and washed with rpmi-1640 (Gibco C11875500137) before being dissociated with a vascular tissue dissociation kit (Bestopcell ， BA3310). The whole tissue dissociation process was performed according to the protocol provided with the kit. At the end of digestion, the cell suspension was passed through a 70μm nylon cell strainer (ThermoFisher Scientifific, 22-363-548) to remove clumps and debris,centrifuge the cells at 300 g for 5 min at 4 ℃ , discard the supernatant and the cells were resuspended in PBS (Bioss，C7033) containing 5% FBS (Solarbio，s9020). Then cells were counted at count star. We observed that cell contained red blood cells in the count star instrument, next, we used red blood cell lysis buffer to remove red blood cell. Following a 15 min incubation at 4℃,samples were centrifuged at 450 g for 5 min at 4℃, discard the supernatant and wash twice times use PBS containing 5% FBS, Cells were then re-suspended in PBS containing 5% FBS. Next, cells were counted at count star and adjust the cell suspension concentration (600-1200 living cells permicroliter).
Single-cell capturing and downstream library constructions were performed using CelCode® Single Cell 3' transcriptome Kit v1.0, according to the manufacturer's protocol. Briefly, a total 70μl of Master Mix and Cell Suspension, 50μl of barcoded gel beads and 45μl partitioning oil were loaded to CelCode® Driver to generate single-cell gel beadin-emulsion (GEM). Immediately following GEM generation, the Gel Bead is dissolved, primers are released. The polyadenylated mRNA were reverse-transcribed into cDNA. Full-length cDNA along with cell barcode identifiers were PCR-amplified and sequencing libraries were prepared and normalized. The libraries were finally sequenced using a MGI DNBSEQ-T7 sequencer with a sequencing depth of at least 50,000 reads  per cell with pair-end 150 bp (PE150) reading strategy performed by Bestopcell, Beijing.

value: nucleus pulposus

tag: age

value: 5-6 weeks old

value: semaglutide


title: MCM4-3xFLAG cells, Late-G1 (ChIP)

description: Library name: MCM4 G1 ChIP

value: HCT116 CMV-OsTIR1 (F74G) MCM4-Stag-3xFLAG


title: SU540, DNMT3A Correction, Xenograft 155_2

description: Library name: SU540_155_2_Corr


title: nucleus pulposus,control

description: Library name: Sample2

value: no


title: RNA-seq with control cells (HCT116 LIG1/3-mAID2), Rep2

description: Library name: RNA-seq Control cells Rep2


title: Male KO CV, leptin, bio rep 4

city: Saint Louis

first: Erica

last: Scheller

state: MO

address: 660 S Euclid Ave

institute: Washington University

description:
CV_691
Read_Counts.txt
TPM_values.txt

data_processing:
The DNBSEQ platform was used to perform paired-end 100 bp sequence reads, generating an average of 4.84 to 6.87Gb per sample.
Sequencing data was filtered using SOAPnuke, a filtering algorithm that removes reads containing the adaptor, reads with N content >5%, and reads with base quality score <15 as the proportion of total bases in the reads that are greater than 20% as low-quality reads.
After filtering, HISAT and Bowtie2 were used to align clean reads to the Mus musculus reference genome version GCF_000001635.26_GRCm38.p6 with an average mapping ratio of 98% and 81-86%, respectively --> read counts.
Read counts were normalized per sample to generate TPM values
Assembly: GRCm39
Supplementary files format and content: tab-delimited text file includes raw counts for each Sample
Supplementary files format and content: tab-delimited text file includes TPM values for each Sample

source_name: Caudal vertebrae

growth_protocol: All mice were housed in a specific pathogen-free facility at a controlled temperature of 22–23 °C on a 12-hour light/dark cycle.

extract_protocol:
After endpoint dissection, tissues were homogenized and preserved in Trizol at -80 °C before RNA extraction. To purify RNA, samples were processed with PureLink™ RNA Mini Kit (Invitrogen, 12183025) according to the manufacturer’s instructions, and the RNA concentration and quality in the samples were checked with a spectrophotometer (Thermo Scientific, NanoDrop 2000).
After being enriched by oligo dT and fragmented, a cDNA library was generated by reverse-transcribing mRNA using random N6-primed RT.

treatment_protocol: Mice were treated with either ICV PBS or 100 ng/hr ICV leptin for 9 days. A group of mice were also subcutaneously supplemented with insulin for 9 days.

value: Caudal vertebrae

value: ICV leptin


title: MCM4-3xFLAG Bromo-RNAPol2 cells, Late-G1, DMSO (Input)

description: Library name: MCM4 G1 Bromo-RNAPol2 DMSO Input

value: HCT116 CMV-OsTIR1 (F74G) MCM4-Stag-3xFLAG BC-POLR2A

value: Nocodazole for 15 hours and release for 4 hours, DMSO for 4 hours


title: MCM4-3xFLAG Bromo-RNAPol2 cells, Late-G1, AGB1 (ChIP)

description: Library name: MCM4 G1 Bromo-RNAPol2 AGB1 ChIP

value: Nocodazole for 15 hours and release for 4 hours, 5-Ph-IAA for 4 hours

value: FLAG M2 (Sigma, F1804)


title: MCM4-3xFLAG cells, Late-G1 (Input)

description: Library name: MCM4 G1 Input

value: Nocodazole for 15 hours and release for 4 hours


title: 3xFLAG-ORC2 cells, Late-G1 (Input)

description: Library name: ORC2 G1 Input

value: Colonic epithelial cells

value: HCT116 CMV-OsTIR1 (F74G) Stag-3xFLAG-ORC2


title: 3xFLAG-ORC1 cells, Late-G1 (ChIP)

description: Library name: ORC1 G1 ChIP

value: HCT116 CMV-OsTIR1 (F74G) Stag-3xFLAG-ORC1


title: SU372C, DNMT3A ReMutation, Xenograft 153_5

description: Library name: SU372.C_153_5_ReMut

value: Blood

value: AML blasts

value: DNMT3A ReMutation (R882H)


title: Male WT CV, leptin, bio rep 8

description:
CV_686
Read_Counts.txt
TPM_values.txt


title: 3xFLAG-ORC1 cells, Late-G1 (Input)

description: Library name: ORC1 G1 Input


title: SU372C, DNMT3A ReMutation, Xenograft 153_4

description: Library name: SU372.C_153_4_ReMut


title: Male KO CV, leptin, bio rep 3

description:
CV_690
Read_Counts.txt
TPM_values.txt


title: SU540, DNMT3A Correction, Xenograft 155_1

description: Library name: SU540_155_1_Corr

value: DNMT3A Correction (R882R)


title: 3xFLAG-ORC2 cells, Late-G1 (ChIP)

description: Library name: ORC2 G1 ChIP


title: Male WT CV, leptin, bio rep 7

description:
CV_685
Read_Counts.txt
TPM_values.txt


title: MCM4-3xFLAG Bromo-RNAPol2 cells, Late-G1, DMSO (ChIP)

description: Library name: MCM4 G1 Bromo-RNAPol2 DMSO ChIP


title: MCM4-3xFLAG Bromo-RNAPol2 cells, Late-G1, AGB1 (Input)

description: Library name: MCM4 G1 Bromo-RNAPol2 AGB1 Input


title: MCM2-3xFLAG cells, Late-G1, pre-extracted (ChIP)

description: Library name: MCM2 G1 pre-extracted ChIP

value: HCT116 CMV-OsTIR1 (F74G) LIG1-mAC LIG3-mAC MCM2-Stag-3xFLAG

value: Nocodazole for 15 hours and release for 4 hours, pre-extracted before ChIP


title: MCM2-3xFLAG cells, Late-G1, pre-extracted (Input)

description: Library name: MCM2 G1 pre-extracted Input


title: Male WT CV, leptin, bio rep 6

description:
CV_684
Read_Counts.txt
TPM_values.txt


title: MCM2-3xFLAG cells, Late-G1, Rep2 (Input)

description: Library name: MCM2 G1 Rep2 Input


title: SU372C, DNMT3A Correction, Xenograft 153_2

description: Library name: SU372.C_153_2_Corr


title: MCM2-3xFLAG cells, Late-G1, Rep2 (ChIP)

description: Library name: MCM2 G1 Rep2 ChIP


title: Male WT CV, PBS, bio rep 5

description:
CV_683
Read_Counts.txt
TPM_values.txt

value: ICV PBS


title: MCM2-3xFLAG cells, Late-G1, Rep1 (ChIP)

description: Library name: MCM2 G1 Rep1 ChIP


title: MCM2-3xFLAG cells, Late-G1, Rep1 (Input)

description: Library name: MCM2 G1 Rep1 Input


title: Mouse Nodose 96

city: New York

first: Robert

last: Lovelett

state: New York

address: 430 E 29th Street, 10th Floor

institute: Kallyope

description:
Library name: K_10X_mNOD096
10X Genomics

data_processing:
Cell Ranger 8.0.0 mkfastq, count, and multi utilties were used to demultiplex and quantify the reads using pipelines implemented in nf-core demultiplex v1.5.3 and nf-core/scrnaseq v3.0.0.
Assembly: Custom version of mm10 where 3' region was extended up to 5Kbp when it did not collide with another known gene on the same strand
Supplementary files format and content: h5ad files with gene expression count matrices, 10X h5 matrices with gene expression and antibody counts for the antibody tagged libraries (K_10X_mGUT035 and K_10X_mGUT036)

source_name: Nodose ganglion

extract_protocol:
EEC protocol: Samples included Neurod1-Cre and Neurog3-Cre mice that were crossed with Ai9 reporters, Chga-GFP, Tph1-CFP, Ffar3-mRFP, and wild type mice. Intestines (stomach, duodenum, jejunum, ileum, and colon) were digested to single cell suspensions following the protocol described in Roberts et al (Diabetes, 2019). They were then FACSed sorted to the cells that emit the fluorophore or wild type mice were sorted with antibodiesto Epcam+/Cldn- cells. For two libraries (K_10X_mGUT035 and K_10X_mGUT036), cell hashing was employed to multiplex multiple samples following the protocol at https://www.protocols.io/view/cite-seq-5qpvo4rxg4o1/v1  Nodose protocol: Wild type mice were used. Nodose ganglia were dissected and as much nerve as possible we trimmed off. 10-11 ganglia were pooled and dissociated with 500 ul of dissociation solution, (5 mg/ml Collegenase II,1 mg/ml Dispace II, in Hank's buffered salt solution) and stored on ice. After dissections were complete, ganglia in dissociation solution were placed in a 37 C water bath for 1 hour with gentle mixing and pipetting every 10 minutes. Triteration was performed with a 1 ml pipette. Undissociated tissue was visualized by microscope to check for the presence of neurons. Dissociation continued until all neurons were visibly dissociated. Then, 500 ul of nodose medium (10% heat inactivated fetal bovine serum in DMEM) was added to the dissociated tissue, mixed 5 times with a 1 ml pippette and spun at 420 RCF for 5 minutes at 4 C. Supernatatants were discarded and pellets were resuspended in nodose medium. Resuspended cells were layered on top of 1ml of BSA gradient in DPBS on ice. Tubes with BSA and cell suspections were spun at 200 RCF for 5 minutes at 4 C. Supernatants were discarded; pellets were resuspended in 500 ul DPBS with 0.4% BSA and filtered through a 70 um cell strainer. Cells were counted and diluted following the 10X Genomics protocol
Single cell suspensions were loaded on to a 10x Chromium Controller as quickly as possible. Approximately 20,000 cells were loaded per reaction. LIbraries were generated with V2 or V3 chemistry following 10x Genomics protocols.

value: Nodose ganglion

tag: library type

value: GEX


title: ChIP control with untagged cells, Late-G1, Rep2 (Input)

description: Library name: ChIP untagged control Rep2 Input

value: HCT116 CMV-OsTIR1 (F74G) LIG1-mAC LIG3-mAC


title: ChIP control with untagged cells, Late-G1, Rep2 (ChIP)

description: Library name: ChIP untagged control Rep2 ChIP


title: Mouse Nodose 95

description:
Library name: K_10X_mNOD095
10X Genomics


title: Male KO CV, leptin, bio rep 2

description:
CV_5488
Read_Counts.txt
TPM_values.txt

value: Caudal vertebrae

value: ICV leptin


title: SU372C, DNMT3A Correction, Xenograft 153_3

description: Library name: SU372.C_153_3_Corr


title: HUVECs, Ad-SRSF1, 2 (IP)

first: Wenting

last: Zhu

address: No.95 Yong'an Road, Xicheng District

institute: Beijing Friendship Hospital, Capital Medical University

description:
Library name: Sample4, IP
library strategy: eCLIP-seq
SRSF1_eCLIP.reproducible_enriched_windows.tsv

data_processing:
Raw Reads were first clustered according to the UMI sequences, in which reads with the same UMI sequence were grouped into the same cluster.Reads in the same cluster were compared to each other by pairwise alignment, and then reads with sequence identity over 95% were extracted to a new sub-cluster. After all sub-clusters were generated, multiple sequence alignment was performed to get one consensus sequence for each sub-clusters. After these steps, the errors and biases introduced by PCR amplification and sequencing were eliminated. De-duplicated consensus sequences mapped to rRNAs were removed with SortMeRNA (version 4.3.3).
Raw sequencing data quality was checked with FastQC (version 0.11.9). Reads was then filtered by fastp (version 0.22.0), of which low-quality reads were discarded and the reads contaminated with adaptor sequences were trimmed.
Filtered reads were aligned to GRCh38_release110 reference genome using STAR (v2.7.4a) with the following configurations --alignEndsType EndToEnd --genomeLoad NoSharedMemory --outBAMcompression 10 --winAnchorMultimapNmax 100 --outFilterMultimapNmax 100 --outFilterMultimapScoreRange 1 --outSAMmultNmax 1 --outMultimapperOrder Random --outFilterScoreMin 10 --outFilterType BySJout --limitOutSJcollapsed 5000000 --outReadsUnmapped None --outSAMattributes All --outSAMmode Full --outSAMtype BAM SortedByCoordinate --outSAMunmapped Within --readFilesCommand zcat --outStd Log --runMode alignReads.
BAM files were filtered to retain uniquely mapped reads (NH:i:1) with mapping quality ≥10 using samtools. Command:  samtools view -h "$bam_file" |  awk '($0 ~ /^@/ || ($5 >= 10 && $0 ~ /NH:i:1/))' | samtools view -Sb -o "$filtered_bam". Coverage at 5' positions was calculated using bedtools genomecov with scaling factors, excluding non-canonical chromosomes. Command:  factor=$(samtools idxstats "$filtered_bam" | cut -f 3 | paste -sd+ | bc | xargs -I {{}} echo 'scale=6; 10^6 / {{}}' | bc);  bedtools genomecov -scale $factor -5 -bg -ibam "$filtered_bam" | sort -k1,1 -k2,2n | grep -v -E "EBV|KI|GL" | sed 's/^/chr/g' > "${SCALED_DIR}/${base_name}_unique_scaled.bg"
Differential peaks between CLIP and INPUT samples were identified using Skipper with default parameters.
Assembly: GRCh38
Supplementary files format and content: Tab-delimited text files include differential enriched peaks for each Sample
Supplementary files format and content: Bigwig files for each Sample

source_name: Human Umbilical Vein Endothelial Cells

growth_protocol: Human umbilical vein endothelial cells (HUVECs) were cultured using endothelial cell medium containing Endothelial Cell Growth Supplement, 5% fetal bovine serum and 1% penicillin/streptomycin.Cells were cultured at 37℃ at 5% CO2.

extract_protocol:
The samples were irradiated twice with 254 nm UV-C light on ice at 400mJ, and lysed on ice for 15 min with lysis buffer (20 mM Tris, pH 8.0, 1 mM EDTA, 400 mM NaCl, 0.1% SDS, 0.5% Triton X-100 (Merck, cat. no. X100), 1×Protease Inhibitor (Thermo Fisher, cat. no. 87785), 200 U/mL RNasin Plus (Promega, cat. no. N2611) and 20 U/mL DNase I (NEB, cat. no. M0303S)). The lysate was further treated with 40 U RNase I (Thermo Fisher, cat. no. EN0601) and rotary incubated at 37°C for 5 min. The reaction was quenched by adding RNasin® Plus. The mixture was centrifuged at 4°C for 15 minutes at 15,000 g, and 2% of the supernatant was was taken for Input sample. The remain of supernatant was incubated with Invitrogen 32-4500 antibody for immunoprecipitation, rotating slowly at 4°C for 4 h. 10 μL protein A/G magnetic beads (Thermo Fisher, cat. no. 88802) was added and incubated with immune complexes for 2 h followed by successive washes. The 3' phosphate groups on the RNA were removed using FastAP enzyme (Thermo Fisher, cat. no. EF0654) to facilitate subsequent ligation steps. A 3' App-DNA linker is ligated to the RNA molecules using T4 RNA ligase (NEB, cat. no. M0204). The RNA-protein complexes were then separated by Nupage 4-12% Bis-Tris protein gels (Thermo Fisher, cat. no. WG1401BX10) electrophoresis and transferred to a nitrocellulose (NC) membrane. The NC membrane was cut to extract the RNA bound to the protein of interest. The protein-bound RNA was extracted from the NC membrane by incubation with proteinase K (NEB, cat. no. P8107) at 37°C for 20 min and subsequent TRIzol (Thermo Fisher, cat. no. 15596018CN) treatment.
The purified RNA was subsequently concentrated, desalted, reverse transcribed. The resulted cDNA was recovered, ligated with a 3' UMI (Unique Molecular Identity) adapter, and amplified by PCR to generate sequencing libraries, which was sequenced on a Novaseq X Plus sequencer (Illumina) using PE150 mode.

value: Human Umbilical Vein Endothelial Cells


title: Male KO CV, PBS, bio rep 4

description:
CV_5486
Read_Counts.txt
TPM_values.txt


title: Mouse Gut EEC 37

description:
Library name: K_10X_mGUT037
10X Genomics

source_name: Small Intestine - EEC enriched

value: Small Intestine - EEC enriched

tag: marker sort


title: Mouse Nodose 4

description:
Library name: K_10X_mNOD004
10X Genomics


title: Mouse Nodose 5

description:
Library name: K_10X_mNOD005
10X Genomics


title: Male KO CV, PBS, bio rep 5

description:
CV_5487
Read_Counts.txt
TPM_values.txt


title: ChIP control with untagged cells, Late-G1, Rep1 (Input)

description: Library name: ChIP untagged control Rep1 Input

value: Colonic epithelial cells

value: Nocodazole for 15 hours and release for 4 hours


title: HUVECs, Ad-GFP, 1 (IP)

description:
Library name: Sample1, IP
library strategy: eCLIP-seq
GFP_eCLIP.reproducible_enriched_windows.tsv


title: ChIP control with untagged cells, Late-G1, Rep1 (ChIP)

description: Library name: ChIP untagged control Rep1 ChIP

value: FLAG M2 (Sigma, F1804)


title: Male KO CV, PBS, bio rep 3

description:
CV_5485
Read_Counts.txt
TPM_values.txt


title: Male WT CV, leptin + insulin, bio rep 6

description:
CV_454
Read_Counts.txt
TPM_values.txt

value: ICV leptin + SubQ insulin


title: Male WT CV, leptin + insulin, bio rep 5

description:
CV_453
Read_Counts.txt
TPM_values.txt


title: HUVECs, Ad-SRSF1, 1 (Input)

description:
Library name: Sample3, Input
library strategy: eCLIP-seq
SRSF1_eCLIP.reproducible_enriched_windows.tsv


title: HUVECs, Ad-GFP, 1 (Input)

description:
Library name: Sample1, Input
library strategy: eCLIP-seq
GFP_eCLIP.reproducible_enriched_windows.tsv


title: HUVECs, Ad-GFP, 2 (Input)

description:
Library name: Sample2, Input
library strategy: eCLIP-seq
GFP_eCLIP.reproducible_enriched_windows.tsv


title: HUVECs, Ad-GFP, 2 (IP)

description:
Library name: Sample2, IP
library strategy: eCLIP-seq
GFP_eCLIP.reproducible_enriched_windows.tsv


title: HUVECs, Ad-SRSF1, 2 (Input)

description:
Library name: Sample4, Input
library strategy: eCLIP-seq
SRSF1_eCLIP.reproducible_enriched_windows.tsv


title: Mouse Gut EEC 43

description:
Library name: K_10X_mGUT043
10X Genomics

source_name: Stomach - EEC enriched

value: Stomach - EEC enriched


title: Mouse Gut EEC 36 ADT

description:
Library name: K_10X_mGUT036_ADT
10X Genomics

data_processing:
Cell Ranger 8.0.0 mkfastq, count, and multi utilties were used to demultiplex and quantify the reads using pipelines implemented in nf-core demultiplex v1.5.3 and nf-core/scrnaseq v3.0.0.
Assembly: Custom version of mm10 where 3' region was extended up to 5Kbp when it did not collide with another known gene on the same strand
Supplementary files format and content: h5ad files with gene expression count matrices, 10X h5 matrices with gene expression and antibody counts for the antibody tagged libraries (K_10X_mGUT035 and K_10X_mGUT036)
Library strategy: CITE-seq

molecule: protein

source_name: Gut - EEC enriched

value: Gut - EEC enriched

value: ADT

tag: adt

value: EpCAM antibody (GeneTex GTX80323)