title: PMB-2

city: Beijing

address: Dongcheng District, Tiantan Xili

institute: Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences & Peking Union Medical College

data_processing:
In order to guarantee the data quality which was used to analysis, the useful Perl script was used to filter the original data (Raw Data)
Bowtie2 v2.2.3 was used for building the genome index, and Clean Data was then aligned to the reference genome using HISAT2 v2.
The IGV (Integrative Genomics Viewer) was used to view the mapping result by the Heatmap, histogram, scatter plot or other stytle.
Reads Count for each gene in each sample was counted by HTSeq v0.6.0, and FPKM (Fragments Per Kilobase Millon Mapped Reads) was then calculated to estimate the expression level of genes in each sample.
DEGseq was used for differential gene expression analysis
The GO (Gene Ontology, http://geneontology.org/) enrichment of DEGs was implemented by the hypergeometric test, in which p-value is calculated and adjusted as q-value, and data background is genes in the whole genome.
KEGG (Kyoto Encyclopedia of Genes and Genomes, http://www.kegg.jp/) is a database resource containing a collection of manually drawn pathway maps representing our knowledge on the molecular interaction and reaction networks.
Genome_build: Homo_sapiens.GRCh38.89.chr
Supplementary_files_format_and_content: tab-delimited text files include RPKM values for each Sample

extract_protocol:
cells were collected , flash frozen on dry ice, and RNA was harvested using Trizol reagent. A total amount of 2 μg RNA per sample was used as input material for the RNA sample preparations. Sequencing libraries were generated using NEBNext® UltraTM RNA Library Prep Kit for Illumina® (#E7530L, NEB, USA)
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: cell line

value: HepG2 cells

value: hepatocyte

tag: treatment

value: treated withPMB for 24 hours

value: liver


title: 12_AYWB

status: Public on Aug 11 2023

type: SRA

city: Godollo

first: Eva

last: Varallyay

address: 4 Szent-Györgyi Albert Street

department: Institute of Plant Protection

country: Hungary

institute: Hungarian University of Agriculture and Life Sciences

description: small RNA

library_source: transcriptomic

data_processing:
The quality check of the sequenced small RNA libraries was done by FastQC Program and CLC Genomic Workbench 11
Trimming of raw reads was performed using the CLC Genomic Workbench 11
Removing redundant reads using the CLC Genomic Workbench 11
Supplementary files format and content: trimmed redundant list, each read with its number of repeats

molecule: total RNA

organism: Catharanthus roseus

source_name: leaf

extract_protocol:
Phenol chloroform extraction method (White & Kaper, 1989) DOI: 10.1016/0166-0934(89)90122-5
modified Illumina Trueseq small RNA kit protocol: https://doi.org/10.1007/978-1-4939-7683-6_9

tag: tissue

value: leaf

tag: symptoms

value: witches broom

tag: ay phytoplasma presence

value: yes

tag: year of sampling

tag: place of collection

value: glasshouse, with insectproof net


title: PMB-1

first: Yang

last: Gao

country: China

organism: Homo sapiens

source_name: liver

tag: cell type


title: WT-old-LSK-MicroC

city: Houston

first: Jia

last: Li

state: Texas

address: 2121 W HOLCOMBE BLVD

department: CEDP

country: USA

institute: Guangzhou Medical University

library_source: genomic

data_processing:
Micro-C pipeline (https://micro-c.readthedocs.io/en/latest/) was used to analysis the Micro-C data. Specifically, we used bwa (0.17.17) to align raw reads on mm10 genome and the proper mapped read pairs were retained to downstream analysis. For each sample, mapped.pairs file was used as input for juicer_tools_1.22.01 to generate contact maps (hic format file). hicPCA function from HiCExplorer (3.7.2) with H3K4me3 ChIPseq tracks of young (4 month old) mice HSCs as correction for PCA values was applied to determine A/B compartment regions.
Assembly: mm10
Supplementary files format and content: hic format include contact matrix

molecule: genomic DNA

organism: Mus musculus

source_name: Bone marrow

growth_protocol: 6-8 weeks WT mice were used as WT-young; 12-24 months WT mice were used as WT-old;  6-8 weeks Tet2KO mice were used as Tet2KO-young; 12-24 months Tet2KO mice were used as Tet2KO-old;

extract_protocol:
0.5 million LSK (Lin-cKit+Sca1+) cells were harvested from WT-young, WT-old, T2KO-young and T2KO-old mice
micro-C data were generated using Dovetail® Micro-C Kit. In briefly, the cells were crosslinked with FA in the presence of DSG, followed by in situ MNase Nuclease digestion and cell lysis. Then chromatins were captured by chromatin bind beads for future end polishing, bridge ligation, intra-aggregate ligation and reverse crosslinking. The purified DNA were proceeded with library generation.

treatment_protocol: No treatment

value: Bone marrow

tag: genotype

value: WT


title: CON-3

value: untreated


title: biventricular heart, CEK, ZT09, bio rep 2

city: Birmingham

first: Lamario

middle: J

last: Williams

state: AL

address: 1825 University Boulevard, Room 121

department: Medicine

institute: University of Alabama at Birmingham

data_processing:
Basecalls were performed using Illumina's bcl2fastq version 2.18.0.12
Raw sequence reads were aligned with STAR (version 2.6.1d) using parameters --outReadsUnmapped Fastx --outSAMtype BAM SortedByCoordinate --outSAMattributes All
Transcript abundances were calculated from the STAR alignment BAM files using HTSeq-count (version 0.11.1) using parameters -m union -r pos -t exon -i gene_id -a 10 -s no -f bam
Assembly: Gencode GRCm38 p4 Release M11
Supplementary files format and content: tab-delimited text file containing Esnembl ID# and raw counts for each gene in each sample

source_name: biventricular heart

extract_protocol:
RNA was harvested using Rneasy mini plus kit (Qiagen). 1.3 ug of total RNA was used for the construction of sequencing libraries.
RNA libraries for RNA-seq were prepared using SMARTER mRNA-Seq Library Prep Kit following manufacturer's protocols.

tag: strain

tag: Sex

value: male

value: biventricular heart

value: CEK

tag: time


title: WT-Young-LSK-MicroC


title: 241

city: Shanghai

first: Gao

last: Yaqi

address: 145 Middle Shandong Road

institute: Gastroenterology and Hepatology, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China

data_processing:
The raw data was checked by FastQC
Clean reads were aligned to the mouse genome using STAR 2.5
The expression of the transcripts was calculated by FPKM using Cuffnorm
Differentially expression transcripts were determined using DESeq
Assembly: mm10
Supplementary files format and content: FPKM values for each Sample

source_name: colon tissue

extract_protocol:
Total RNA of colon tissues was harvested using Trizol reagent
RNA libraries were prepared for sequencing using standard Illumina protocols

value: colon tissue

value: DSS+ICB+F. prausnitzii gavage


title: biventricular heart, CEK, ZT06, bio rep 4


title: 118_AYP

description: small RNA

value: phyllody


title: 119_AYP

description: small RNA


title: CON-1

value: HepG2 cells


title: 223


title: 214


title: 117_AYP

description: small RNA


title: biventricular heart, CON, ZT06, bio rep 3

value: CON


title: CON-2

value: hepatocyte


title: biventricular heart, CON, ZT03, bio rep 3


title: biventricular heart, CEK, ZT12, bio rep 1


title: TG_ALF_12_125

city: El Cajon

first: Daniel

last: Carlin

state: CA

address: 1250 Mountain View Rd.

institute: UC-San Diego

data_processing:
Sequence alignment of the resulting high-quality reads to the mouse reference genome (build GRCm38) was done using HISAT2 and quantification of gene-level expression was performed using featureCounts. DESEQ2 was used to generate normalized read counts and to determine differentially expressed genes. Genes with an averageexpression of more than 2 counts per minute (CPM) across all samples were considered as expressed.
Assembly: GRCm38
Supplementary files format and content: tab delimited deseq2 normalized counts

source_name: Hippocampus

growth_protocol: APP23 transgenic (TG) mice (B6.Cg-Tg(Thy1-APP)3Somm/J; JAX Stock No: 030504) and non-transgenic (NTG) littermates control mice were housed in light-tight enclosures at the University of California, Los Angeles in the Laboratory of Circadian and Sleep Medicine. APP-SAA KI (B6.Cg-Apptm1.1Dnli/J; JAX Stock No: 034711) were obtained from Jackson Laboratory as heterozygotes and bred to develop heterozygote APP-SAA KI mice (APP-KI) and non-transgenic (NTG) littermates. APP-KI were housed in light-tight enclosures at the University of California, San Diego. The mice were given ad libitum food (Teklad rodent diet 8604; Envigo, Indianapolis, IN) and water access until they were placed in the time-restricted feeding (TRF) or ad libitum feeding (ALF) cohorts. The work presented in this study followed all guidelines and regulations of the UCSD and UCLA Division of Animal Medicine that are consistent with the Animal Welfare Policy Statements and the recommendations of the Panel on Euthanasia of the American Veterinary Medical Association.

extract_protocol:
Mice were euthanized with isoflurane, either in the dark ( ZT12 and ZT18) or in the light (ZT0 and ZT6). Brain hemispheres were collected and placed in either RNAlater Stabilization Solution (Thermo Fisher Scientific, Waltham, MA) or 4% paraformaldehyde. Serum was collected from blood incubated on ice for 30 min, centrifuged at 2000 g for 10 min at 4°C, and then stored until used at -80°C.
Total RNA was isolated from hippocampus tissue dissected from one hemibrain per animal using RNeasy Lipid Tissue Mini kit (Qiagen, Hilden, Germany) as indicated by the manufacturer. Quality of the extracted RNA was assessed using TapeStation (Agilent Technologies, Inc., Santa Clara, CA). All samples showed RIN 8. RNA-Seq library preparation was performed with poly-A enrichment using poly-T oligo-attached magnetic beads employing the PE150 sequencing strategy performed on a NovaSeq 6000 (Illumina, Inc., San Diego, CA) by Novogene Inc. (Sacramento, CA). Raw reads (>20M per sample) were imported to Galaxy platform for analysis 83. Reads were first cleaned by removing adapter sequences, trimming low-quality ends, and filtering reads with low quality using Trimmomatic.

treatment_protocol: Prior to any measurements or beginning any experiments, mice were habituated to a 12:12 light-dark (LD 12:12) cycle and single housing conditions in custom light-tight cabinets. We then measured diurnal rhythms in activity and sleep behavior from the mice. To assess circadian impairments in the mice, some animals were placed in constant darkness (DD), followed by light masking and 6 h phase advance protocols. Feeding manipulations were then performed on mice habituated to LD 12:12. Animals within each sex and genotype were randomly divided into two groups with one group continuing ALF and the other placed on TRF, with food available during 6 h in the middle of the mouse active phase, during zeitgeber time (ZT) 15 to ZT21. By definition, ZT0 is the time when lights go on and ZT12 is the time when the lights go off when the mice are in a 12:12 LD cycle. After habituating to the feeding protocol, metabolites were measured in the mice to ensure the treatment was inducing the expected metabolic effects. The mice remained on TRF or ALF food access until sacrificed for tissue collection. After all tests were completed and the mice had re-entrained to the LD cycle, mice within each genotype and treatment group were randomly assigned for sacrifice at one of four time points: ZT0, ZT6, ZT12, or ZT18. ZT0 and ZT6 tissues were collected in the light; ZT12 and ZT18 were collected in the dark. Brain tissue and blood serum were collected for analysis. This study used a total of 110 mice almost equally distributed across sex/ genotype/treatment throughout the study, and further grouped by ZT collection times. Treated mice started TRF at 6 to 9 months of age and were maintained on TRF until tissue collection, which occurred at 10 to 13 months of age. Experimental mice were housed in standard cages with either normal ad libitum food or with a custom-made programmable food hopper that could temporally control access to food and prevent food consumption during restricted times. Feeders were open for 6 h in the middle of the mouse active phase (ZT15 to ZT21). Food consumption was initially measured twice per week to ensure that all mice acclimated to the new feeding regimen. Mice are coprophagic and were moved to new cages twice per week, as determined empirically, to diminish coprophagic behavior during the fasting period.

value: Hippocampus

value: TG

value: ALF


title: PMB-3

value: treated withPMB for 24 hours

value: liver


title: 245


title: PDX

data_processing:
The Cell Ranger software and Seurat 3.0(R package) were used for basecalling.Alignment, filtering, barcode counting, and UMI counting were performed with cellranger 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.
GO enrichment, KEGG enrichment, Reactome enrichment and Disease enrichment (human only) of cluster markers were performed using KOBAS software with Benjamini-Hochberg multiple testing adjustment, using top 20 markers gene of cluster. The results were visualized using R package.
Protein-protein interaction was obtained from STRING database with combine_score >=400. For each cluster, the interaction of top 20 marker genes was extracted from the database and the results were visualized using Cytoscape software, respectively.
Transcription Factors were predicted within 2000 bp upstream and 500 bp downstream of TSS for top 20 marker genes of each cluster using TFBSTools and JASPAR database. The gene and TF network for each cluster was visualized using Cytoscape software.
GSEA was performed by using GSEA software version 2.2.2.4, which uses predefined gene sets from the Molecular Signatures Database (MSigDB v6.2).All genes detected in all cells of one sample were used. Gene expression data was calculated by mean umi count of gene in one cluster and the rest cluster, respectively. The minimum and maximum criteria for selection of gene sets from the collection were 0 and 500 genes, respectively.
Genome_build: Homo_sapiens.GRCh38.89.chr
Supplementary_files_format_and_content: matrix table with raw gene count for every gene and every cell

molecule: polyA RNA

extract_protocol:
Using single cell 3 'Library and Gel Bead Kit V3 (10x Genomics, 1000075) and Chromium Single Cell B Chip Kit (10x Genomics, 1000074), the cell suspension was loaded onto the Chromium single cell controller (10x Genomics) to generate single-cell gel beads in the emulsion according to the manufacturer’s protocol. In short, single cells were suspended in PBS containing 0.04% BSA. Captured cells were lysed and the released RNA were barcoded through reverse transcription in individual GEMs. Reverse transcription was performed on a S1000TM Touch Thermal Cycler (Bio Rad) at 53°C for 45 min, followed by 85°C for 5 min, and hold at 4°C. The cDNA was generated and then amplified, and quality assessed using an Agilent 4200 (performed by CapitalBio Technology, Beijing).
According to the manufacture’s introduction, Single-cell RNA-seq libraries were constructed using Single Cell 3’ Library andGel Bead Kit V3.

value: sphere-forming culture

value: liver PDX-derived cancer stem cells


title: biventricular heart, CEK, ZT03, bio rep 2


title: Mtb_truB_woCMC

city: Boston

first: Satoshi

last: Kimura

state: Massachusetts

address: 181 Longwood Avenue

institute: Brigham and Women's Hospital

library_source: other

data_processing:
Analysis: 3’ linker sequences and two nucleotides at the 5’ end were trimmed using cutadapt and fastx-trimmer. Bowtie v1.2.2 was used with default settings to map reads to reference Mtb tRNA sequences retrieved from Mycobrowser[40]. Mpileup files were generated using samtools (samtools mpileup -I -A --ff 4 -x -B -q 0 -d 10000000). For analysis of termination frequencies, 5’ end termini of mapped reads were piled up using bedtools genomecov (option, -d -5 -ibam). The number of 5’ termini at each tRNA position was divided by the total number of mapped termini at that position plus all upstream (5’) positions.
Assembly: EC-tRNA.fa, Mtb-tRNA.fa
Supplementary files format and content: Misincporation and Termination frequencies are shown in the csv file format

molecule: other

organism: Mycobacterium tuberculosis

source_name: bacteria

extract_protocol:
Extraction of total RNA: Strains were grown to mid-log phase with the appropriate antibiotics and inducing agents described above. RNA was collected at the same OD600 for each strain (between 0.4-0.6). Cells were left on ice for 20 minutes, then pelleted by centrifuging at 4,000 rpm for 10 minutes at 4 °C. Pellets were resuspended in 0.5-1 mL of TriZol (Life Technologies) and lysed using a BeadBug microtube homogenizer (Millipore Sigma). 200 uL of chloroform was added to each tube, after which samples obtained from Mtb strains were removed from biosafety level 3 precautions. Samples were centrifuged at 15,000 rpm for 15 minutes at 4 °C and the aqueous layer was collected into a fresh tube. To the original tube, 250 uL of sodium acetate buffer (300 mM sodium acetate pH 5.2 and 10 mM EDTA pH 8.0) was added, and samples were vortexed at 4 °C for 5 minutes then centrifuged at 15,000 g for 15 minutes at 4 °C. The aqueous layer was added to the fresh sample-containing tubes. 400 uL chloroform was added, and tubes were briefly vortexed and then centrifuged at 15,000 rpm for 1 minute at 4 °C. The aqueous phase was collected into a fresh tube and RNA recovered by ethanol precipitation. RNA pellets were resuspended in 10 mM sodium acetate pH 5.2 and stored at -80 °C until processed for sequencing. Total RNA samples were alkali-treated prior to tRNA extraction to deacylate all tRNAs (1 hour at 37 °C in 100 mM Tris-HCl pH 9.0).  Isolation of tRNA fraction: 1-2 ug of total RNA was run on a 10% TBE-UREA gel (ThermoFisher Scientific) at 250 V for 1 hour. Gels were stained with SYBR Gold (ThermoFisher Scientific), and tRNA was excised. Excised gels containing tRNA fractions were mashed in RNAse-free tubes, and 300 uL elution buffer (300 mM NaOAc pH 5.5, 1 mM EDTA pH 8.0, 0.10% SDS) was added to each tube. Samples were shaken on a thermoshaker (Eppendorf) for 1-4 hours at 37 °C and supernatant was collected using an Ultrafree filter column (Millipore Sigma). tRNA was recovered by isopropanol precipitation.
tRNA dephosphorylation: tRNA was dephosphorylated using QuickCIP (New England BioLabs) according to manufacturer instructions, and tRNA was collected by phenol-chloroform extraction followed by isopropanol precipitation.   Iodoacetamide (IAA) treatment: IAA treatment was performed as described. Briefly, 500 ng of total RNA is combined with 10 mM of iodoacetamide, 50 mM NaPO4 pH 8.0, and 50% DMSO in a final volume of 50 uL. Reactions were incubated at 50 °C for 15 minutes and quenched with DTT.  CMC treatment: CMC treatment was carried out as described in ref. Briefly, 2.5 ug tRNA fraction in 0.5 ul was mixed with 15 ul CMC-BEU buffer with or without CMC (0.34 M or 0 M CMC, 7 M urea, 4 mM EDTA, and 50 mM bicine pH 7.9) and incubated at 37 °C for 20 min. Adding 100 ul CMC stop solution (0.3 M NaOAc pH 5.2 and 100 mM EDTA) quenched the reaction. RNA was desalted with PD-10 desalting column (Cytiva) and recovered by ethanol precipitation. RNA was dissolved in 40 ul of 50 mM sodium carbonate buffer (pH 10.4) and incubated at 37 °C for 4 hours, followed by ethanol precipitation.  Adapter ligation: 0.5 uL RNase inhibitor was added to 3.5uL dephosphorylated tRNA (200-250 ng tRNA) and samples were boiled at 80 °C for 2 minutes. Boiled tRNA was mixed with 12 uL PEG buffer mix (10 uL 50% PEG8000, 2 uL 10 x buffer B0216S; New England Biolabs). 3 uL of 5’ adenylated linkers (Supplementary Table 4) were added (33 pmol/uL) along with 1 uL T4 RNA ligase 2 truncated (New England BioLabs) and incubated at 25 uC for 2.5 hours. Samples were recovered by isopropanol precipitation and run on a 10% TBE-Urea PAGE gel for 40 minutes at 250 V. Ligated products were recovered by gel excision as described above.  Reverse transcription: Identical quantities of samples with different adapter sequences were pooled for reverse transcription for a total of 200250 ng tRNA. Reverse transcription was performed by combining 2.1 uL dephosphorylated tRNA with 100 mM Tris-HCl pH 7.5, 0.5 mM EDTA, 1.25 uM RT primer, 450 mM NaCl, 5 mM MgCl2, 5 mM DTT, 500 nM TGIRT (InGex), and 15% PEG8000 in a final volume of 9 uL. Samples were incubated at 25 °C for 30 minutes, after which 1 uL 10 mM dNTPs (New England BioLabs) were added and reactions incubated at 60 °C for 1 hour. 1.15 uL NaOH was added, and samples were boiled for 15 minutes and run on a 10% TBE Urea PAGE gel at 250V for 1 hour. Reverse transcription products were excised from gels and cDNA recovered by isopropanol precipitation. Linear single-stranded cDNA was circularized using CircLigase II (Lucigen) in accordance with manufacturer instructions.  PCR of tRNA libraries: PCR reactions were set up using HF Phusion according to the manufacturer’s instructions using a universal reverse primer and a different index primer for each pool of samples. PCR reactions were aliquoted into 4 tubes and collected after 6, 8, 10, and 12 cycles. Samples were run on a Native TBE PAGE gel (ThermoFisher Scientific) at 180 V for 50 minutes, and amplified products were cut from the same cycle for each sequencing run. Samples were recovered by gel excision and isopropanol precipitation.

value: bacteria

value: truB

value: CMC


title: 210


title: biventricular heart, CON, ZT00, bio rep 1

value: male

value: biventricular heart


title: OVCAR88_shEZH2_rep1

first: Jianfeng

last: Chen

state: Shanghai

address: 270 Dong'an Road

institute: Fudan University Shanghai Cancer Center

data_processing:
Raw reads were filtered and cleaned by fastp software.
Aliments were performed followed by STAR instruction
RSEM were used for counting
R were used for visualization
Genome_build: GRCh38
Supplementary_files_format_and_content: TXT file contain counts for each sample

growth_protocol: OVCAR8 cells are cultured in RPMI1640 medium supplemented with 10% fetal bovine serum, 1% penicillin-streptomycin.

extract_protocol:
RNA was harvested using Trizol reagent (Invitrogen).
RNA libraries were prepared using standard Illumina protocols.

treatment_protocol: OVCAR8 cells were treated with DMSO, 5mM DZNep or doxycycline 1 mg/mL for 72h.

tag: tissue source

value: high grade serous ovarian cancer

tag: genotype/treatment

value: doxycycline-induced EZH2 silence


title: 205

value: colon tissue

value: DSS+ICB


title: Mtb_WT_CMC

value: WT


title: OVCAR88_DZNep_rep2

value: DZNep


title: 201


title: Mtb_WT_woCMC


title: OVCAR88_DZNep_rep1


title: Mtb_mnmA_IAA

value: mnmA


title: 274


title: Mtb_mnmA_woIAA

value: IAA


title: 200


title: Gabpb1(hetro_x_hetro)-b3

city: Fukuoka

first: Kei

last: Miyamoto

address: 744 Moto-oka, Nishi-ku

department: Faculty of Agriculture

country: Japan

institute: Kyushu University

description:
One embryo was subjected to cDNA synthesis
KB8

data_processing:
Fastq files from Illumina sequencing were filtered for low quality reads by sliding window trimming (window size 10, <QV20), and low quality bases were trimmed from the ends of the reads (<QV20) using Trimmomatic. Reads less than 20 bases and unpaired reads were removed. Furthermore, adaptor, polyA, polyT and polyG sequences were removed using trim_galore and cutadapt.
The sequencing reads were then mapped to the mouse genome (mm10) using STAR. Reads on annotated genes were counted using featureCounts. FPKM values were calculated from mapped reads by normalizing to total counts and transcript.
Assembly: mm10
Supplementary files format and content: tab-delimited text file includes FPKM values for each sample

source_name: Mouse blastocyst embryo

growth_protocol: After IVF of sperm and oocytes derived from hetero mice (Alyref+/- or Gabpb1+/-), embryos were cultured in KSOM at 37°C under 5% CO2. As a control, IVF was also performed using WT B6D2F1 mice. At 96 hpi, morula and blastocyst embryos were transferred to PBS containing 0.1% BSA for the subsequent extraction step.

extract_protocol:
After 96 hours of culture, embryos were transferred to PBS containing 0.1% BSA for washing. Then, zona pellucida was removed in acid Tyrode and the embryos were washed with PBS containing 0.1% BSA. The washed embryo was transferred to a 0.2 ml tube containing 9.5 μl of reaction buffer to make a final volume of 10.5 μl (one embryo in 10.5 μl reaction buffer). After cell lysis in the buffer, 1 μl of the solution was removed and, instead, 1 μl of the diluted ERCC RNA Spike-In Mix was added (Thermo Fisher Scientific, 4456740). The lysed RNA solution with the spike-in RNA were subjected to reverse transcription.
The produced cDNA was amplified by PCR with 16 cycles. The amplified cDNA was purified using AMPure XP beads (BECKMAN COULTER, A63882). The purified cDNA was measured on the Bioanalyzer (Agilent) using High Sensitivity DNA Kit (5067-4626) and the normalized volume of DNA was subjected to library preparation using Nextera XT DNA Library Preparation Kit (Illumina, FC-131-1024) by following the vendor’s instruction. The obtained library was quality-checked by Bioanalyzer and quantified using Qubit (Thermo Scientific).
Paired end sequencing (50 bp + 25 bp) was carried out using the Illumina NextSeq system.

tag: strains

tag: developmental stage

value: Blastocyst

value: WT or hetero


title: Mtb_WT_IAA


title: WT-b1

description:
One embryo was subjected to cDNA synthesis
WB1


title: WT-b2

description:
One embryo was subjected to cDNA synthesis
WB2


title: 258


title: WT-b3

description:
One embryo was subjected to cDNA synthesis
WB3


title: Mtb_WT_woIAA


title: Mtb_truB_CMC


title: OVCAR88_shEZH2_rep2


title: Gabpb1(hetro_x_hetro)-b2

description:
One embryo was subjected to cDNA synthesis
KB7


title: p13plus8_Mel1_ESC

city: Perth

first: Ryan

last: Lister

state: WA

address: 35 Stirling Highway

country: Australia

institute: The University of Western Australia

description: Human embryonic stem cell line

data_processing:
Sequencing adapters were trimmed with BBduk with the options mink = 3, qtrim = r, trimq = 10 minlength = 20 before alignment to hg19 with Bowtie and BSseeker2 with the option -n 1. PCR duplicates were removed using Sambamba and DNA methylation levels at base resolution calculated using CGmap tools.
Genome_build: hg19
Supplementary_files_format_and_content: CGmap file generated with CGmap tools. File has 8 columns: [1]chromosome [2] nucleotide on Watson (+) strand [3] position [4] context (CG/CHG/CHH) [5] dinucleotide-context (CA/CC/CG/CT) [6] methylation-level = #_of_C / (#_of_C + #_of_T) [7] #_of_C (methylated C, the count of reads showing C here) [8] = #_of_C + #_of_T (all Cytosines, the count of reads showing C or T here).

source_name: Human embryonic stem cells

growth_protocol: To generate human primed and naïve iPSCs used for this study, somatic cell reprogramming with the Cytotune 2.0 kit, experiments were performed according to the manufacturer’s instructions (Invitrogen). Briefly, human dermal fibroblast (adult) were seeded at ~5x104 cells per well of a 6-well plate in fibroblast medium containing DMEM (Gibco), 10% FBS (Hyclone), 1% nonessential amino acids (Gibco), 1mM GlutaMAX (Gibco), Pen-strep (Gibco), 0.1mM 2-mercaptoethanol (Gibco) and 1mM sodium pyruvate (Gibco). After 2 days, cells were transduced with Sendai viruses in fibroblast medium with multiplicity of infections (MOIs) as follows, KOS MOI=5, c-MYC MOI=15, KLF4 MOI=6. After 24 hours, the medium was replaced with fresh fibroblast medium and media were changed every other day thereafter. On day 7, cells were trypsinized and seeded onto a layer of iMEF feeders at ~0.5x10^5~1x10^5 per flask in fibroblast medium. The next day, the cells were switched to primed or naive media. 21-24 days after transduction, cells were passaged and expanded. Reprogramming intermediates were collected at the indicated time and isolated by FACS.

extract_protocol:
Genomic DNA was isolated with the Qiagen Blood and Tissue Kit according to manufacturer's instructions. 0.5% (w/w) of unmethylated lambda phage DNA (Promega) was added to the sample genomic DNA for the purpose of an unmethylated control to measure the bisulfite non-conversion frequency in each sample.
Genomic DNA was fragmented with a Covaris S2 sonicator to a mean length of 200 bp, then end-repaired, A-tailed, ligated to methylated Illumina TruSeq adapters, and subjected to PCR amplification with KAPA HiFi Uracil+ DNA polymerase (KAPA Biosystems). Single-end sequencing was performed on a HiSeq1500, NextSeq500 or NovaSeq.

treatment_protocol: On day 8, the cells were switched to primed, naive, naive-to-primed (NtP) or transient-naive-treatment (TNT) reprogramming protocols. For primed: Cells were transient primed essential8 (E8) culture media after the initial 7 days culturing in fibroblast media. For Naive: Naive hiPSCs colonies were established through extended naive culturing. For TNT: cells were transient naive culture for 5 days after the initial 7 days culturing in fibroblast media, followed by culturing in primed E8 media. For NtP: Naive hiPSCs colonies were established through extended naive culturing and then transitioned the cells to a primed pluripotent state in E8 media to give rise to naive-to-primed hiPSCs.

value: hESC

tag: background

tag: pluripotent state

value: Primed


title: biventricular heart, CEK, ZT00, bio rep 1

value: CEK


title: GP5d, H3K36me3,  replicate 2

city: Helsinki

first: Konsta

last: Karttunen

address: Haartmaninkatu 8

country: Finland

institute: University of Helsinki

data_processing:
FastQC v.0.11.9 was used for quality control and determination of read lengths of raw data.
Raw reads were mapped with bowtie2 v.2.4.1 (bowtie2 --very-sensitive) to the reference genome.
Picard v.2.23.4 was used to remove duplicates (MarkDuplicates -REMOVE_DUPLICATES false -ASSUME_SORT_ORDER coordinate).
Samtools v.1.7 was used to filter reads with MAPQ smaller than 20 and remove marked duplicates (samtools view -F 1024 -b -q 20).
Peaks were called with MACS2 v.2.2.7.1 (macs2 callpeak -f BAMPE -g hs --keep-dup all).
Peaks overlapping with ENCODE blacklisted regions were removed from the narrowPeak and summit files with bedtools v.2.29.2 (bedtools subtract -A).
A RPKM normalized bigwig file was created with deepTools v.3.5.0 (bamCoverage --binSize 50 --normalizeUsing RPKM --effectiveGenomeSize 2913022398).
Assembly: hg38
Supplementary files format and content: bigWig, broadPeak/narrowPeak (except for the IgG samples)

growth_protocol: GP5d cells were grown in DMEM supplemented with 10% FBS, 2 mM L-glutamine and 1% penicillin-streptocmycin. Cells were checked for mycoplasma contamination and were passaged at 80% confluency.

extract_protocol:
GP5d cells were formaldehyde cross-linked and sonicated chromatin was immunoprecipitated by using Dynal-bead coupled antibodies. Immunoprecipitated DNA was purified and used to prepare ChIP-seq library for Illumina sequencing.
Immunoprecipitated DNA was end-repaired, dA tailed and adapter ligated. Adaptor ligated DNA was PCR enriched as described earlier (Sahu et al., Nat. Gen. 2022).

tag: cell line

value: Colon adenocarcinoma

tag: chip antibody


title: p32_Mel1_iPSC_TNT

description: TNT-hiPSC reprogrammed from secondary fibroblasts

source_name: Human pluripotent stem cells

tag: treatment


title: Gabpb1(hetro_x_hetro)-b1

description:
One embryo was subjected to cDNA synthesis
KB6


title: p33_Mel1_iPSC_primed

description: Primed-hiPSC reprogrammed from secondary fibroblasts


title: GP5d, TFAP2A, replicate 2


title: 202

tag: tissue

value: DSS


title: 100


title: OVCAR88_Ctrl_rep2

value: control


title: H9_ESC_E8

data_processing:
Sequencing adapters were trimmed with BBduk with the options mink = 3, qtrim = r, trimq = 10 minlength = 20 before alignment to hg19 with Bowtie and BSseeker2 with the option -n 1. PCR duplicates were removed using Sambamba and DNA methylation levels at base resolution calculated using CGmap tools. The non–conversion rate was calculated using the DNA methylation levels for the spiked–in lambda phage genome. When DNA methylation levels were calculated for regions such as promoters, enhancers, DMRs or ICRs, the methylation level was calculated as a coverage–weighted average by summing the number of methylated C calls (mCG) and dividing that by the total number of reads with either a C or T call (CG). This is represented in the figures and text as mCG/CG. To calculate methylation in CH contexts (where H = A, T or C), the level of methylation was calculated as above (mCH/CH) with the non–conversion rate subtracted from this value. When CH methylation was calculated for individual contexts, for example CA methylation, the non–conversion rate for that context was subtracted from the calculated methylation levels. For CA methylation browser tracks, mCA/CA was calculated for 5kb sliding windows (1kb slide), with the CA methylation non–conversion rate for that library subtracted from each window. To calculate per-read methylation, reads classified as methylated had methylation calls at every CG position in the read; unmethylated reads had zero methylation calls at CG positions; partially methylated reads had at least one CG methylation call  and one non-methylated CG call.
Genome_build: hg19
Supplementary_files_format_and_content: CGmap file generated with CGmap tools. File has 8 columns: [1]chromosome [2] nucleotide on Watson (+) strand [3] position [4] context (CG/CHG/CHH) [5] dinucleotide-context (CA/CC/CG/CT) [6] methylation-level = #_of_C / (#_of_C + #_of_T) [7] #_of_C (methylated C, the count of reads showing C here) [8] = #_of_C + #_of_T (all Cytosines, the count of reads showing C or T here).

growth_protocol: To generate human primed and naïve iPSCs used for this study, somatic cell reprogramming with the Cytotune 2.0 kit, experiments were performed according to the manufacturer’s instructions (Invitrogen). Briefly, human dermal fibroblast (adult) were seeded at ~5x104 cells per well of a 6-well plate in fibroblast medium containing DMEM (Gibco), 10% FBS (Hyclone), 1% nonessential amino acids (Gibco), 1mM GlutaMAX (Gibco), Pen-strep (Gibco), 0.1mM 2-mercaptoethanol (Gibco) and 1mM sodium pyruvate (Gibco) . After 2 days, cells were transduced with Sendai viruses in fibroblast medium with multiplicity of infections (MOIs) as follows, KOS MOI=5, c-MYC MOI=15, KLF4 MOI=6. After 24 hours, the medium was replaced with fresh fibroblast medium and media were changed every other day thereafter. On day 7, cells were trypsinized and seeded onto a layer of iMEF feeders at ~0.5x105~1x105 per flask in fibroblast medium. The next day, the cells were switched to primed or naive media. 21-24 days after transduction, cells were passaged and expanded. Reprogramming intermediates were collected at the indicated time and isolated by FACS.


title: OVCAR88_Ctrl_rep1


title: 14397X18 THP-1 KO 3

city: Salt Lake City

last: O'Connell

state: UT

address: Emma Eccles Jones, Medical Research Building, 15 North Medical Drive East

department: Pathology, Microbiology & Immunology

institute: University of Utah

data_processing:
The human GRCh38 genome and gene annotation files were downloaded from Ensembl release 102 and a reference database was created using STAR version 2.7.6a
Optical duplicates were removed from the paired end FASTQ files using clumpify v38.34 and reads were trimmed of adapters using cutadapt 1.16
The trimmed reads were aligned to the reference database using STAR in two pass mode to output a BAM file sorted by coordinates
Mapped reads were assigned to annotated genes using featureCounts version 1.6.3
Differentially expressed genes were identified using a 5% false discovery rate with DESeq2 version 1.30.1
Assembly: GRCh38
Supplementary files format and content: featureCount output files combined into a single tab-delimited raw count matrix

source_name: AML cell line

extract_protocol:
mRNA was isolated by following manufacturer protocols using the Qiagen miRNeasy Mini kit (Cat.: 217084). The cell extracts were treated withQiagen RNase free DNase by following manufacturer protocol (Cat.: 79254).
Illumina Stranded Total RNA Library Prep w Ribo-Zero Plus

value: AML cell line

value: KO


title: 14397X17 THP-1 KO 2


title: 213


title: 14397X16 THP-1 KO 1


title: 208


title: 32F_iPSC_E8media


title: ATAC-seq_iPSC_TNT_p43

description:
TNT-hiPSC reprogrammed from secondary fibroblasts
RL2457

data_processing:
Sequencing adapters were trimmed with BBduk with the options mink = 3, ktrim = r, before alignment to hg19 with Bowtie2 with the option -X 2000. Reads were filtered for proper pairs, and PCR duplicates and mitochondrial reads removed using samtools. Bigwig browser tracks were normalised for library size using the counts per million method at single base resolution.
Genome_build: hg19
Supplementary_files_format_and_content: Bigwig formatted files of aligned ATAC-seq read counts. Data are normalised using the counts per million (CPM) method. Files were generated using the DeepTools bamCoverage function.

extract_protocol:
Approximately one million freshly harvested cells were pelleted and washed in PBS, then resuspended in 1 ml of RSB buffer (10 mM Tris-HCl, 10 mM NaCl, 3 mM MgCl2, 0.1% NP-40, 0.1% Tween-20, 0.01% Digitonin). After 10 min incubation on ice, samples were spun at 500 g for 5 min and resuspended in 500 µl RSB without NP40 or digitonin, then strained through a 30 µm filter and pelleted again. Resulting nuclei were counted using trypan blue and 50,000 nuclei were resuspended in 25 µl of 2x TD buffer (20 mM Tris-HCl, 10 mM MgCl2, 20% dimethyl formamide)
Tagmentation mix was completed by adding 100 U of loaded Tn5, 16.5 µl PBS, 0.5 µl of 1% digitonin and 0.5 µl of Tween-20 to a final volume of 50 µl, followed by incubation for 30 minutes at 37°C with 1,000 RPM mixing on a thermo block. After tagmentation, samples were cleaned up using the Qiagen MinElute PCR purification kit. Eluat was amplified using NEBNext 2x MasterMix and Nextera-based adapters as primers. After 10 PCR cycles, a double-sided bead purification was performed using 0.5x and 1.8x Ampure XP beads. Libraries were quantified by qPCR on a CFX96/C1000 cycler (Biorad) and sequenced on a Novaseq 6000 (Illumina) in 2x 61 bp paired-end format.

value: TNT


title: Gabpb1(hetro_x_hetro)-m2

description:
One embryo was subjected to cDNA synthesis
KM10

source_name: Mouse morula embryo

value: Morula


title: TBX5 OE rep2

first: William

middle: T.

last: Pu

state: Ma

address: 300 Longwood Ave

department: Cardiology

institute: Boston Children's Hospital

data_processing:
Reads were aligned to mouse genome version mm10 with STAR using parameter --quantMode GeneCounts to retrieve read counts.
Deseq2 with default settings was used to assess differential expression between conditions.
Assembly: MM10
Supplementary files format and content: DEseq2 and counts output files

source_name: Left Ventricle

extract_protocol:
RNA was extracted using Qiagen Rneasy Plus Micro Kit, cat. No. 74034
Libraries were generated using the Lexogen Corral total RNA-seq library Prep kit (cat. 095)

value: Left Ventricle

value: Myh6-Cre Tbx5OE


title: TBX5 OE rep3


title: ATAC-seq_iPSC_primed_p44

description:
Primed-hiPSC reprogrammed from secondary fibroblasts
RL2458


title: 14397X15 THP-1 EV 3

value: EV


title: TBX5 OE rep1


title: 14397X14 THP-1 EV 2


title: Gabpb1(hetro_x_hetro)KO-m3

description:
One embryo was subjected to cDNA synthesis
KM8

value: Gabpb1 KO


title: ATAC-seq_iPSC_TNT_p44

description:
TNT-hiPSC reprogrammed from secondary fibroblasts
RL2449


title: Control rep5

value: WT


title: 14397X12 Molm-14 KO 3


title: ATAC-seq_iPSC_primed_p42

description:
Primed-hiPSC reprogrammed from secondary fibroblasts
RL2454


title: Alyref(hetro_x_hetro)-b3

description:
One embryo was subjected to cDNA synthesis
KB3

value: Blastocyst


title: ATAC-seq_iPSC_N2P_p9_22

description:
NTP-hiPSC reprogrammed from secondary fibroblasts
RL2450

value: NtP


title: 14397X13 THP-1 EV 1


title: Gabpb1(hetro_x_hetro)KO-m2

description:
One embryo was subjected to cDNA synthesis
KM7


title: Control rep4


title: Alyref(hetro_x_hetro)-b2

description:
One embryo was subjected to cDNA synthesis
KB2

value: WT or hetero


title: Gabpb1(hetro_x_hetro)KO-m1

description:
One embryo was subjected to cDNA synthesis
KM6


title: Alyref(hetro_x_hetro)-b1

description:
One embryo was subjected to cDNA synthesis
KB1


title: 248

value: colon tissue


title: ATAC-seq_ESC_p13_35

description:
Human embryonic stem cell line
RL2448

value: ESC

value: n/a (Human embryonic stem cell line)


title: 14397X11 Molm-14 KO 2


title: 14397X10 Molm-14 KO 1


title: ATAC-seq_iPSC_N2P_p9_21

description:
NTP-hiPSC reprogrammed from secondary fibroblasts
RL2443

value: Primed


title: Gabpb1(hetro_x_hetro)-m1

description:
One embryo was subjected to cDNA synthesis
KM9


title: Control rep2


title: Alyref(hetro_x_hetro)-m2

description:
One embryo was subjected to cDNA synthesis
KM4


title: ATAC-seq_ESC_p13_34

description:
Human embryonic stem cell line
RL2442


title: 14397X9 Molm EV 3


title: Control rep1


title: Alyref(hetro_x_hetro)-m1

description:
One embryo was subjected to cDNA synthesis
KM3


title: 14397X8 Molm EV 2


title: ATAC-seq_iPSC_N2P_p13_73

description:
NTP-hiPSC reprogrammed from secondary fibroblasts
RL2439


title: ATAC Control left atria Rep. 2

description:
snATAC library
single nucleus ATAC library

data_processing:
Raw fastq files were taken as input into cellranger ARC to generate files for later analysis
Files from CellRanger ARC were read into R and analyzed using the Seurat/Signac pipeline
Assembly: mm10
Supplementary files format and content: 10x Genomics output files: barcodes.tsv.gz, features.tsv.gz, matrix.mtx.gz

source_name: Left Atrium

extract_protocol: snRNA libraries and snATAC libraries were prepared according to the 10x multiome protocol

value: Left Atrium

value: complex tissue

value: Control


title: Alyref(hetro_x_hetro)KO-m2

description:
One embryo was subjected to cDNA synthesis
KM2

value: Alyref KO


title: 14397X7 Molm EV 1

value: AML cell line


title: ATAC TBX5 KO left atria rep. 2

value: TBX5 KO


title: Input for LSK cells

data_processing:
Chromium 10x v2 was processed using Cell Ranger (v6.0.0) with default parameters to generated single cell RNA sequencing data. Single cell RNAseq was sequenced on Novaseq S4 platform. Raw fastq files were demultiplexed using cellranger mkfastq from 10x Genomics. Then, the demultiplexed fastq files were aligned on mouse genome (refdata-gex-mm10-2020-A) provided by 10x Genomics. Scanpy (https://scanpy.readthedocs.io/en/stable/) was used to filter out cells with low UMI (<1,000) and cells with mitochondria reads percentage > 5%. Cells with less than 200 genes detected and genes expressed in less than 10 cells were also filtered out. Normalization and clustering analysis were using scanpy packages with default settings.
scATACseq data was prepared using Chromium 10x v2 kits and was demultiplexed using cellranger-atac mkfastq (cellranger-atac-1.2.0). The fastq files were aligned to mm10 (refdata-cellranger-atac-mm10-1.2.0) using Cellranger-atac count. ArchR (https://www.archrproject.com) was used to do quality control and data analysis. Specifically, the size of fragments and TSS enrichment were estimated, and doublet was removed. Clustering and gene scores calculation were performed based on the tutorial (https://www.archrproject.com/bookdown/index.html).
Bulk RNAseq data was mapped to mm10 using STAR (version=2.7.7a). Uniquely mapped reads was used to calculate raw counts for each gene and sample were inputted to DESeq2 R package to identify significantly differential expressed genes among groups.
For the WGBS analysis, raw FASTQ files were mapped to the NCBI Human Reference Genome Build GRCh37 (hg19) using BSMAP. Fastp was used to trim adaptor sequences from fastq data and remove 15 bp from the two ends of the reads based on Swift protocol. MCALL module in MOABS software was used to calculate DNA methylation ratio and coverage for each CpG site. Differentially methylated CpGs and regions were identified using the MCOMP module considering variance among samples (--withVariacne 1) in the MOABS software. Bisulfite conversion efficiencies were estimated using spike-in unmethylated lambda phage DNA. The output bedGraph files from MCALL include single base resolution DNA methylation ratios, which were transformed to a bigwig file format. The bigwig files were uploaded to the UCSC genome browser for visualization.
For CMS-IP-seq analysis, raw FASTQ files were mapped to mm10 using BSMAP . Only uniquely mapped non duplicated reads were kept as input for MACS2  to call 5hmC enriched regions (peaks). Differential hydroxymethylation regions (DHMRs) were identified using DESeq2 with default parameters. Bam files were transformed to bigwig format, which can be used to visualize 5hmC signals in the UCSC genome browser.. H3K9me3 and H3K9me2 ChIPseq data was aligned on mm10 using bowtie2. The bam files were inputted to EDD (https://github.com/CollasLab/edd) to identify ChIPseq enriched regions.
Genome_build: mm10
Supplementary_files_format_and_content: bigwig; csv; txt

source_name: LSK cells from bone marrow

extract_protocol:
RNA and DNA were purified using Qiagen AllPrep DNA/RNA micro kit.
1. Single-cell ATAC-seq libraries were generated by Chromium Single Cell ATAC Reagent Kits (10× Genomics); 2. Low amount RNA seq libraries were generated using Takara SMART-Seq Single Cell Kit; 3. CMSIP: 5hmC-containing genomic DNA were immunoprecipitated by CMS antibody after being bisulfite treated.  Immunoprecipitated DNA and input were proceed for Library generation by using zymo pico methyl kit. 4. CHIP seq: H3K9me2/3 binded chromatins were immunoprecipitated by H3K9me3 (ab8898) and H3K9me2 (ab1220). immunoprecipitated DNA and input were proceeded for library generation by using Takara ThruPLEX DNA library kit. 5. WGBS: Bisulfite conversions were performed by using Zymo lighting bisulfite kit and libraries were generated by using Swift Methyl kit for BS converted DNA.

treatment_protocol: mxCre_Dnmt3af/f mice were injected with 300 ul pIpC every other day for total 5 times

value: LSK

value: Dnmt3a KO

tag: age

value: not provided


title: 14397X6 MV4-11 KO 3

value: KO


title: ATAC-seq_iPSC_TNT_p42

description:
TNT-hiPSC reprogrammed from secondary fibroblasts
RL2437

value: TNT


title: Alyref(hetro_x_hetro)KO-m1

description:
One embryo was subjected to cDNA synthesis
KM1

value: Morula


title: ATAC-seq_iPSC_primed_p40

description:
Primed-hiPSC reprogrammed from secondary fibroblasts
RL2438


title: 14397X5 MV4-11 KO 2


title: Control rep3

value: Left Ventricle

value: WT


title: ATAC TBX5 KO left atria rep. 1


title: ATAC-seq_ESC_p13_33

description:
Human embryonic stem cell line
RL2436

value: ESC


title: H3K9me2 for LSK cells from Dnmt3a KO1 young mouse

value: young


title: 14397X4 MV4-11 KO 1


title: Alyref(hetro_x_hetro)-m3

description:
One embryo was subjected to cDNA synthesis
KM5

value: WT or hetero


title: RNA Control left atria Rep. 2

description:
3' mRNA library: read1 file contains cell barcode and UMI; read2 file contains transcript
single nuclues polyA RNA

value: Left Atrium


title: 14397X3 MV4-11 EV 3

value: EV


title: H3K9me2 for LSK cells from Dnmt3a KO2 young mouse


title: P4_plus_10_TNT_MEL1_Input

description: RL2009_2020_02_06_P4_plus_10_TNT_MEL1_H3K9me3_ChIP_S5_merged_lanes_dedup_FE.bigwig

data_processing:
Adapters were trimmed using fastp with default parameters, and mapped to hg19 using bowtie2 with the option -X 2000. H3K9me3 fold enrichment was calculated for each ChIP and associated input library using the MACS2 bdgcmp function with the option -FE.
Genome_build: hg19

source_name: Human pluripotent stem cell

extract_protocol:
Cells were crosslinked for 10 minutes in 1% formaldehyde and quenched in 125 mM glycine. Prior to ChIP, antibodies were bound to beads by mixing 3 µg H3K9me3 antibody (Abcam, ab8898) with 50 µl washed Dynabead M-280 Sheep Anti-Rabbit IgG  (Thermofisher) in 500 µl RIPA-150 buffer (50 mM Tris-HCl pH 8.0, 0.15 M NaCl, 1 mM EDTA , 0.1% SDS, 1% Triton X-100 and 0.1% sodium deoxycholate) and incubated at 40C for 6 hours on a rotator . Crosslinked cells were lysed on ice for 10 min in 15 ml ChIP lysis buffer (50 mM HEPES pH 7.9, 140 mM NaCl, 1 mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100) supplemented with 1x EDTA-free Protease Inhibitor Cocktail (Roche). Lysed cells were centrifuged at 3,200xg for 5 min, supernatant removed and followed by two washes with 10ml ChIP wash buffer (10 mM Tris-Cl pH 8.0, 200 mM NaCl and 1 mM EDTA pH 8.0). Lysed cells were resuspended in 130 µl nuclei lysis buffer (50mM  Tris-HCl pH 8.0, 10mM EDTA and 1% SDS) supplemented with 1x EDTA-free Protease Inhibitor Cocktail (Roche), transferred to Covaris tubes (microTUBE AFA Fiber 6x16mm) and sheared with the Covaris (S220) for 5 min (5% duty cycle, 200 cycles per burst and 140 watts peak output at 40C). Sheared chromatin was transferred to 1.5 ml eppendorf tubes, centrifuged at 10,000xg for 10 min. The supernatant was transferred to 2 ml low-bind tubes containing 1.2 ml ChIP dilution Buffer (50 mM Tris-HCl pH 8.0, 0.167 M NaCl, 1.1% Triton X-100 and 0.11% sodium deoxycholate) and 0.65 ml RIPA-150 buffer, and incubated with the previously prepared H3K9me3 antibody bound Dynabeads at 40C overnight on a rotator. Chromatin bound beads were subsequently washed one time with 1 ml RIPA-150 buffer, two times with 1 ml RIPA-500 buffer (50 mM Tris-HCl pH 8.0, 0.5 M NaCl, 1 mM EDTA , 0.1% SDS, 1% Triton X-100 and 0.1% sodium deoxycholate), two times with 1ml RIPA-LiCl buffer (50 mM Tris-HCl pH 8.0, 1 mM EDTA , 1% NP-40, 0.7% sodium deoxycholate and 0.5 M LiCl2) and two times with TE buffer (10 mM Tris-HCl, pH 8.0, 0.1 mM EDTA). After wash steps, DNA was eluted, crosslinks were reversed, and immunoprecipitated DNA was purified by Agencourt AMPure XP beads (Beckman Coulter, Cat no: A63880).
Libraries were prepared from ChIP eluate containing 10 ng DNA using the SMARTer ThruPLEX DNA-Seq Kit (Takara, R400675) with SMARTer DNA unique dual index (Takara, R400665). After limited PCR amplification, libraries were purified using Agencourt AMPure XP beads (Beckman Coulter, Cat no: A63880), and eluted in a final volume of 20 ul. Libraries were sequenced on the Novaseq 6000.

value: NtP

tag: antibody

value: Input


title: ATAC Control left atria Rep. 1

tag: cell type

value: complex tissue

value: Control


title: P17_MEL1_D13_TNT_Input

description: RL2008_2020_02_06_P17_MEL1_D13_TNT_H3K9me3_ChIP_S4_merged_lanes_dedup_FE.bigwig


title: 14397X2 MV4-11 EV 2

value: AML cell line


title: H3K9me2 for LSK cells from Tet2 KO1 old mouse

value: LSK

tag: genotype

value: Tet2 KO

value: old


title: H3K9me2 for LSK cells from Tet2 KO2 old mouse


title: RNA TBX5 KO left atria rep. 2

value: TBX5 KO


title: 14397X1 MV4-11 EV 1


title: RNA TBX5 KO left atria rep. 1


title: P18_MEL1_HDF_to_E8_Input

description: RL2007_2020_02_06_P18_MEL1_HDF_to_E8_H3K9me3_ChIP_S3_merged_lanes_dedup_FE.bigwig

value: Primed


title: GP5d, rIgG

value: Colon adenocarcinoma

value: Santa Cruz Biotechnology, sc-2027


title: H3K27ac TBX5AKO atria 2

description: HiChIP

data_processing:
Intial processing of FASTQ files was performed with HiCPro.
The outputs of HiCPro were then used by Hichipper to call loops, using the mm10 genome.
Assembly: mm10
Supplementary files format and content: bigWig, Mango (significant interactions of each sample)
Library strategy: HiChIP-seq

source_name: Atria- complex tissue

extract_protocol:
Nuclei were extracted and fixed for 15 mins. In 1% FA.
Libraries were prepped using the Arima HiChIP kit with the H3K27ac antibody.

treatment_protocol: Nppa-cre or Nppa-EGFP was injected at P2 into neonatal mice. Atria were collected at P20 and flash-frozen in liquid nitrogen until the experiment.

value: Atria- complex tissue

value: H3K27ac (Active motif, 39133)


title: GP5d, mIgG

value: Santa Cruz Biotechnology, sc-2025


title: P33_32F_Naive_SR_clone1_Input

description: RL2011_2020_02_06_P33_32F_Naive_SR_clone1_H3K9me3_ChIP_S7_merged_lanes_dedup_FE.bigwig

value: Naive


title: H3K9me2 for LSK cells from Tet2 KO1 young mouse

value: young


title: H3K9me2 for LSK cells from Tet2 KO2 young mouse


title: H3K9me2 for LSK cells from WT1 old mouse

value: WT


title: P17_MEL1_HDF_to_SR_Input

value: Input


title: P13_plus_20_MEL1_to_E8_Input

description: RL2006_2020_02_06_P13_plus_20_MEL1_to_E8_H3K9me3_ChIP_S2_merged_lanes_dedup_FE.bigwig

value: ESC


title: GP5d, TFAP2A, rep 1


title: H3K9me2 for LSK cells from WT2 old mouse


title: P33_32F_Naive_SR_clone1_H3K9me3_ChIP


title: H3K27ac TBX5AKO atria 1

description: HiChIP


title: P9_plus_6_TNT_MEL1_H3K9me3_ChIP

description: RL2010_2020_02_06_P9_plus_6_TNT_MEL1_H3K9me3_ChIP_S6_merged_lanes_dedup_FE.bigwig

value: NtP


title: RNA Control left atria Rep. 1

value: Left Atrium

value: complex tissue

value: Control


title: H3K27ac control atria 2

description: HiChIP


title: H3K9me2 for LSK cells from WT1 young mouse

value: LSK


title: P4_plus_10_TNT_MEL1_H3K9me3_ChIP


title: GP5d, H3K36me3, rep 1


title: H3K9me2 for LSK cells from WT2 young mouse


title: Spatial transcriptomics, 12 month DES uterus

city: RTP

first: Wendy

last: Jefferson

state: NC

address: 111 Alexander Drive

institute: NIH

data_processing:
base calling by bcl2fastq v2.20.0.422
scRNA processed by Cell Ranger v 3.0.1
Visium processed by Space Ranger v 1.2.2
Assembly: mm10
Supplementary files format and content: Raw count matrix provided by 10X pipeline.
Supplementary files format and content: CSV of relevant metadata for all cells in count matrix.

source_name: Uterus

extract_protocol:
Extraction was performed using Chromium Single Cell 3’ Reagent Kit v3 (10X Genomics) or Visium Spatial Gene Expression Slide kit (10X Genomics)
Library prep was performed using Chromium Single Cell 3’ Reagent Kit v3 (10X Genomics) or Visium Spatial Gene Expression Slide kit (10X Genomics)
Library strategy was performed using Chromium Single Cell 3’ Reagent Kit v3 (10X Genomics) or Visium Spatial Gene Expression Slide kit (10X Genomics)

value: Uterus

tag: strain

value: Neonatal DES (1mg/kg/day)


title: P17_MEL1_D13_TNT_H3K9me3_ChIP

value: TNT


title: 14398X12 Molm14.KO3

data_processing:
The human GRCh38 genome was downloaded from Ensembl release 87 and a reference database was created using STAR version 2.5.2.
Reads were aligned to the reference database using custom options for small RNAs to output a BAM file sorted by coordinates.
Mapped reads were assigned to mirBase annotations using featureCounts version 1.5.1.
Assembly: GRCh38
Supplementary files format and content: featureCount output files combined into a single tab-delimited raw count matrix.

extract_protocol:
mRNA was isolated by following manufacturer protocols using the Qiagen miRNeasy Mini kit (Cat.: 217084). The cell extracts were treated with Qiagen RNase free DNase by following manufacturer protocol (Cat.: 79254).
NEBNext Multiplex Small RNA Library Prep Set for Illumina.

value: AML cell line

value: KO


title: P18_MEL1_HDF_to_E8_H3K9me3_ChIP

value: Primed


title: H3K9me3 for LSK cells from Dnmt3a KO1 young mouse

value: Dnmt3a KO


title: Spatial transcriptomics, 12 month control uterus

value: Control (corn oil)


title: 14398X11 Molm14.KO2


title: 14398X10 Molm14.KO1


title: scRNA-seq, 12 month DES uterus

description:
DES
CONDES.rawcounts.csv.gz
CONDES.metadata.csv.gz


title: H3K9me3 for LSK cells from Tet2 KO1 old mouse

value: Tet2 KO

value: old


title: H3K9me3 for LSK cells from Dnmt3a KO2 young mouse

value: young


title: scRNA-seq, 12 month control uterus

description: CON


title: P13_plus_20_MEL1_to_E8_H3K9me3_ChIP

value: ESC


title: 14398X9 Molm14.EV3

value: EV


title: H3K27ac control atria 1

description: HiChIP

value: Atria- complex tissue

value: H3K27ac (Active motif, 39133)


title: P17_MEL1_HDF_to_SR_H3K9me3_ChIP

description: RL2005_2020_02_06_P17_MEL1_HDF_to_SR_H3K9me3_ChIP_S1_merged_lanes_dedup_FE.bigwig

value: Naive


title: P24_32F_primed_in_E8_Input

value: Input


title: H3K9me3 for LSK cells from Tet2 KO1 young mouse

value: LSK


title: GP5d, H3K4me3

value: Colon adenocarcinoma


title: D13_plus_10_32F_N2P_Input

value: TNT


title: H3K9me3 for LSK cells from Tet2 KO2 old mouse


title: 14398X8 Molm14.EV2

value: AML cell line


title: CreAlbScarb2F/FMYC-3

city: Bei Jing

address: Tiantan Xili 2

department: Institute of Medicinal Biotechnology

institute: Chinese Academy of Medical Sciences & Peking Union Medical College

data_processing:
In order to guarantee the data quality which was used to analysis, the useful Perl script was used to filter the original data (Raw Data).
Bowtie2 v2.2.3 was used for building the genome index, and Clean Data was then aligned to the reference genome using HISAT2 v2.
The IGV (Integrative Genomics Viewer) was used to view the mapping result by the Heatmap, histogram, scatter plot or other style.
Reads Count for each gene in each sample was counted by HTSeq v0.6.0, and FPKM (Fragments Per Kilobase Million Mapped Reads) was then calculated to estimate the expression level of genes in each sample.
DEGseq was used for differential gene expression analysis between two samples with non biological replicates.
The GO (Gene Ontology, http://geneontology.org/) enrichment of DEGs was implemented by the hypergeometric test, in which p-value is calculated and adjusted as q-value, and data background is genes in the whole genome.
KEGG (Kyoto Encyclopedia of Genes and Genomes, http://www.kegg.jp/) is a database resource containing a collection of manually drawn pathway maps representing our knowledge on the molecular interaction and reaction networks.
Assembly: Homo_sapiens.GRCh38.89.chr
Supplementary files format and content: Tab-delimited text files include FPKM values for each Sample.

source_name: liver

extract_protocol:
Livers were removed, flash frozen on dry ice, and RNA was harvested using Trizol reagent. A total amount of 2 μg RNA per sample was used as input material for the RNA sample preparations.
Sequencing libraries were generated using NEBNext® UltraTM RNA Library Prep Kit for Illumina® (#E7530L, NEB, USA).

value: liver

value: post natal day 60


title: 14398X7 Molm14.EV1


title: CreAlbScarb2F/FMYC-2


title: CreAlbScarb2F/FMYC-1


title: H3K9me3 for LSK cells from Tet2 KO2 young mouse


title: D13_plus_7_38F_N2P_H3K9me3_ChIP


title: 14398X6 MV4-11.KO3

value: KO


title: CreAlbMYC-3

value: CreAlbMYC


title: H3K9me3 for LSK cells from WT1 old mouse

value: WT


title: D13_plus_7_32F_N2P_H3K9me3_ChIP


title: 14398X5 MV4-11.KO2


title: CreAlbMYC-2


title: P11_plus_11_38F_N2P_H3K9me3_ChIP

value: NtP


title: H3K9me3 for LSK cells from WT2 old mouse

value: old


title: 14398X4 MV4-11.KO1


title: H3K9me3 for LSK cells from WT1 young mouse

value: young


title: CreAlbMYC-1


title: P16_38F_primed_in_E8_H3K9me3_ChIP

value: Primed


title: 14398X3 MV4-11.EV3

value: EV


title: CreAlbScarb2F/FMYC

data_processing:
In order to guarantee the data quality which was used to analysis, the useful Perl script was used to filter the original data (Raw Data)
Bowtie2 v2.2.3 was used for building the genome index, and Clean Data was then aligned to the reference genome using HISAT2 v2.
The IGV (Integrative Genomics Viewer) was used to view the mapping result by the Heatmap, histogram, scatter plot or other stytle.
Reads Count for each gene in each sample was counted by HTSeq v0.6.0, and FPKM (Fragments Per Kilobase Millon Mapped Reads) was then calculated to estimate the expression level of genes in each sample.
DEGseq was used for differential gene expression analysis between two samples with non biological replicates.
The GO (Gene Ontology, http://geneontology.org/) enrichment of DEGs was implemented by the hypergeometric test, in which p-value is calculated and adjusted as q-value, and data background is genes in the whole genome.
KEGG (Kyoto Encyclopedia of Genes and Genomes, http://www.kegg.jp/) is a database resource containing a collection of manually drawn pathway maps representing our knowledge on the molecular interaction and reaction networks.
Genome_build: mm8
Supplementary_files_format_and_content: tab-delimited text files include RPKM values for each Sample ...

extract_protocol:
Livers were removed , flash frozen on dry ice, and RNA was harvested using Trizol reagent. A total amount of 2 μg RNA per sample was used as input material for the RNA sample preparations. Sequencing libraries were generated using NEBNext® UltraTM RNA Library Prep Kit for Illumina® (#E7530L, NEB, USA)
RNA libraries were prepared for sequencing using standard Illumina protocols


title: P24_32F_primed_in_E8_H3K9me3_ChIP


title: CreAlbMYC


title: 10x scATACseq for Lin- cells from Tet2 KO old mouse

source_name: Lin- cells from bone marrow

value: Tet2 KO


title: H3K9me3 for LSK cells from WT2 young mouse


title: 14398X1 MV4-11.EV1


title: 14398X2 MV4-11.EV2


title: P12_plus_13_38F_N2P_H3K9me3_ChIP


title: 10x scATACseq for Lin- cells from Tet2 KO young mouse


title: CMSIP for LSK cells from Tet2 KO2 old mouse

data_processing:
Library strategy: 5hmC mapping
Chromium 10x v2 was processed using Cell Ranger (v6.0.0) with default parameters to generated single cell RNA sequencing data. Single cell RNAseq was sequenced on Novaseq S4 platform. Raw fastq files were demultiplexed using cellranger mkfastq from 10x Genomics. Then, the demultiplexed fastq files were aligned on mouse genome (refdata-gex-mm10-2020-A) provided by 10x Genomics. Scanpy (https://scanpy.readthedocs.io/en/stable/) was used to filter out cells with low UMI (<1,000) and cells with mitochondria reads percentage > 5%. Cells with less than 200 genes detected and genes expressed in less than 10 cells were also filtered out. Normalization and clustering analysis were using scanpy packages with default settings.
scATACseq data was prepared using Chromium 10x v2 kits and was demultiplexed using cellranger-atac mkfastq (cellranger-atac-1.2.0). The fastq files were aligned to mm10 (refdata-cellranger-atac-mm10-1.2.0) using Cellranger-atac count. ArchR (https://www.archrproject.com) was used to do quality control and data analysis. Specifically, the size of fragments and TSS enrichment were estimated, and doublet was removed. Clustering and gene scores calculation were performed based on the tutorial (https://www.archrproject.com/bookdown/index.html).
Bulk RNAseq data was mapped to mm10 using STAR (version=2.7.7a). Uniquely mapped reads was used to calculate raw counts for each gene and sample were inputted to DESeq2 R package to identify significantly differential expressed genes among groups.
For the WGBS analysis, raw FASTQ files were mapped to the NCBI Human Reference Genome Build GRCh37 (hg19) using BSMAP. Fastp was used to trim adaptor sequences from fastq data and remove 15 bp from the two ends of the reads based on Swift protocol. MCALL module in MOABS software was used to calculate DNA methylation ratio and coverage for each CpG site. Differentially methylated CpGs and regions were identified using the MCOMP module considering variance among samples (--withVariacne 1) in the MOABS software. Bisulfite conversion efficiencies were estimated using spike-in unmethylated lambda phage DNA. The output bedGraph files from MCALL include single base resolution DNA methylation ratios, which were transformed to a bigwig file format. The bigwig files were uploaded to the UCSC genome browser for visualization.
For CMS-IP-seq analysis, raw FASTQ files were mapped to mm10 using BSMAP . Only uniquely mapped non duplicated reads were kept as input for MACS2  to call 5hmC enriched regions (peaks). Differential hydroxymethylation regions (DHMRs) were identified using DESeq2 with default parameters. Bam files were transformed to bigwig format, which can be used to visualize 5hmC signals in the UCSC genome browser.. H3K9me3 and H3K9me2 ChIPseq data was aligned on mm10 using bowtie2. The bam files were inputted to EDD (https://github.com/CollasLab/edd) to identify ChIPseq enriched regions.
Genome_build: mm10
Supplementary_files_format_and_content: bigwig; csv; txt


title: CMSIP for LSK cells from Tet2 KO1 old mouse


title: CMSIP for LSK cells from Tet2 KO1 young mouse


title: CMSIP for LSK cells from WT1 old mouse


title: CMSIP for LSK cells from Tet2 KO2 young mouse


title: 10x scATACseq for Lin- cells from WT old mouse


title: NtP_hiPSC_P4_MEL1

description: NTP-hiPSC reprogrammed from secondary fibroblasts


title: Fibroblast_MEL1

description: Secondary fibroblasts differentiated from human embryonic stem cells

source_name: Human secondary fibroblasts

value: Fibroblast

value: Differentiated


title: NtP_hiPSC_P9_MEL1


title: D13_plus_10_32F_N2P_H3K9me3_ChIP


title: 10x scRNAseq for Lin- cells from WT 30 month old mouse

value: LSK


title: TNT_hiPSC_MEL1

description: TNT-hiPSC reprogrammed from secondary fibroblasts


title: 10x scRNAseq for Lin- cells from WT 18 month old mouse


title: CMSIP for LSK cells from WT1 young mouse


title: zebrafish, AR42_TFA_4, 3dpf

city: Chongqing

first: Sijia

last: Fan

address: Chongqing Medical University, Medical College Road, Yuzhong District

institute: Chongqing medical university

data_processing:
Sequence reads were trimmed for adaptor sequence/low-quality sequence using Trimmomatic (version 0.36)
Clean Reads were further treated with UMI_soft (in-house) to eliminate duplication bias introduced in library preparation and sequencing on using UMI. （Default Parameter developed by Wuhan Seqhealth Co., Ltd.）
The de-duplicated consensus sequences were mapped to GRCh38 using STAR software (version 2.5.3a) (parameters- --outSAMtype BAM SortedByCoordinate)
Read count extraction and normalization were performed using featureCounts（Version 1.5.1）（parameter -T 10 -d 30 -D 1000 -C -s 1 -t {exon} -g {geneid} --primary -O -a {gff_annotation_file}）
Assembly: GRCz10
Supplementary files format and content: tab-delimited text files include raw counts for each Sample
Supplementary files format and content: tab-delimited text files include RPKM values for each Sample

organism: Danio rerio

source_name: tail

extract_protocol:
RNA was harvested using TRIzol Reagent (Invitrogen, cat. NO 15596026）. 2 ug of total RNA was used for the construction of sequencing libraries.
total RNAs were used for stranded RNA sequencing library preparation using KCTM Stranded mRNA Library Prep Kit (Catalog NO. DR08402, Wuhan Seqhealth Co., Ltd. China) following the manufacturer’s instruction.

value: tail

value: AR-42 treated larvae with TFA modeling


title: Primed_hiPSC_P17_MEL1

description: Primed-hiPSC reprogrammed from secondary fibroblasts


title: zebrafish, AR42_TFA_3, 3dpf


title: WGBS for LSK cells from MxCre Ctrl young mouse

value: MxCre Ctrl


title: zebrafish, AR42_TFA_2, 3dpf


title: hESC_MEL1

description: Human embryonic stem cell line

value: hESC


title: CMSIP for LSK cells from WT2 young mouse


title: WGBS for LSK cells from Dnmt3a KO young mouse

value: Dnmt3a KO


title: zebrafish, AR42_TFA_1, 3dpf


title: INPUT-CTRLcas9_ChIPSeq

data_processing:
Illumina bcl2fq software was used for basecalling.
Low-quality reads (Q≤19 bases over 50%), Ns reads (N%>5%) and adapter polluted reads (adapter bases>5 bp) are filtered out.
ChIP-seq reads were aligned to the human genome assembly using Bowtie2.
Peaks were called using MACS2.
Genome_build: Homo_sapiens.GRCh38.89.chr

extract_protocol:
Lysates were clarified from sonicated nuclei and histone-DNA complexes were isolated with antibody.
Libraries were prepared according to Illumina's instructions.Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 15 cycles and library fragments of ~150 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Illumina HiSeq X following the manufacturer's protocols.

value: HepG2 cells

value: hepatocyte

value: liver


title: INPUT-SCARB2cas9_ChIPSeq


title: WGBS for LSK cells from Tet2 KO old mouse


title: N2P_MEL1_P15

description:
RL1979
MEL1_gene_and_TE_counts.txt.gz

data_processing:
Adapters were trimmed using fastp with default parameters, and mapped to hg19 using HISAT2 with the options --no-mixed --dta --rna-strandness RF -k 2. Alignments were then filtered to keep only unique mapping read pairs using Samtools view -F "[NH]==1". Gene and transposable element read counts were calculated using TEtranscripts and the TElocal script and the curated transposable element GTF files for hg19 that accompany this software. Elements with zero counts across all samples were removed from count table to reduce size.
Genome_build: hg19
Supplementary_files_format_and_content: Tab-seperated count table with samples as columns and features (genes and transposable elements) as rows. First column is feature identifier (ID).

source_name: NtP induced pluripotent stem cell

extract_protocol:
RNA was extracted using the Agencourt RNAdvance Cell v2 (Beckman Coulter) system following the manufacturer's instruction with one additional DNAse (NEB) treatment step. RNA amounts and RINe scores were assessed on a TapeStation using RNA Screen Tape (Agilent), and 500 ng of total RNA were used per sample to generate RNAseq libraries. ERCC ExFold RNA Spike-In mixes (Thermo Scientific) were added as internal control.
Libraries were prepared using the TruSeq Stranded mRNA library prep kit (Illumina), using TruSeq RNA unique dual index adapters (Illumina). Libraries were quantified by qPCR on a CFX96/C1000 cycler (Biorad) and sequenced on a Novaseq 6000 (Illumina) for 2x 53 bp as paired-end.

value: NtP


title: WGBS for LSK cells from Tet2 KO young mouse


title: zebrafish, DMSO_TFA_3, 3dpf

value: DMSO treated larvae with TFA modeling


title: WGBS for LSK cells from WT old mouse

value: WT

value: old


title: Naive_hiPSC_P17_MEL1

description: Naïve-hiPSC reprogrammed from secondary fibroblasts

data_processing:
Sequencing adapters were trimmed with BBduk with the options mink = 3, qtrim = r, trimq = 10 minlength = 20 before alignment to hg19 with Bowtie and BSseeker2 with the option -n 1. PCR duplicates were removed using Sambamba and DNA methylation levels at base resolution calculated using CGmap tools.
Genome_build: hg19
Supplementary_files_format_and_content: CGmap file generated with CGmap tools. File has 8 columns: [1]chromosome [2] nucleotide on Watson (+) strand [3] position [4] context (CG/CHG/CHH) [5] dinucleotide-context (CA/CC/CG/CT) [6] methylation-level = #_of_C / (#_of_C + #_of_T) [7] #_of_C (methylated C, the count of reads showing C here) [8] = #_of_C + #_of_T (all Cytosines, the count of reads showing C or T here).

value: Naïve


title: zebrafish, DMSO_TFA_4, 3dpf


title: IP-SCARB2cas9_ChIPSeq

value: c-Myc antibody (CST, #9402)


title: Fibroblast_MEL1_P4plus19

description:
RL1977
MEL1_gene_and_TE_counts.txt.gz

source_name: In vitro derived fibroblast

value: Fibroblast


title: IP-CTRLcas9_ChIPSeq


title: 9120G, liver, treatment

city: Auburn University

first: Xu

last: Wang

address: 273 CASIC Building, Auburn Research Park, 559 Devall Dr

department: Pathobiology, College of Veterinary Medicine

institute: Auburn University

data_processing:
FastQC (version 11.5) software was used for quality check.
Sequencing adapter sequences and low-quality bases were trimmed using Trimmomatic v0.36
On average, 87% of reads were mapped to the cattle reference genome (GenBank: GCA_002263795.2) by Tophat-2.1.1
Gene reads counts were performed in three software packages, Cufflinks-2.2.1, Bedtools -2.30.0, and HT-seq
DESeq2 packages in R were used to perform normalization and detect significant differentially expressed genes (DEGs) [23; 24], at an FDR (False Discovery Rate) cutoff of 0.05. The log2 fold change (LogFC) was determined for each gene.
Assembly: Cattle reference genome (GenBank: GCA_002263795.2)
Supplementary files format and content: Gzipped, tab-delimited text file including raw gene counts for every gene in all samples

organism: Bos taurus

source_name: Bos taurus liver biopsy

extract_protocol:
The total RNA of liver samples was extracted using the ZYMO Quick DNA/RNA Miniprep Plus Kit (Zymo Research, CA). For homogenization, tissue samples with DNA/RNA Shield were mechanically homogenized by Qiagen TissueRuptor II (Qiagen, MD). RNA concentrations were measured by Qubit fluorometer 3.0 (Thermo Fisher Scientific, MA) with Qubit RNA BR Assay Kit.
RNA sequencing libraries were constructed using the NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (New England Biolabs, MA) and NEBNext Poly(A) mRNA Magnetic Isolation Module (New England Biolabs, MA), with a 1500 ng total RNA input. The concentrations and the size distribution of the libraries were checked by the LabChip GX Touch HT machine using the HT DNA NGS 3K Assay (Perkin Elmer, MA). The fragment size of final libraries ranged from 343 to 409 bp. The libraries were individually barcoded and pooled. The libraries were sequenced on an Illumina NovaSeq 6000 sequencing machine to generate 150-bp paired-end reads

treatment_protocol: The diet offered was ad libitum Bermudagrass hay combined with a nutritional supplement composed of 1.61 kg of endophyte-free tall fescue seeds, and 1.61 kg of pellets composed of 46.5% ground corn, 46.5% soybean meal, 5% wheat middlings, and 2% soybean oil; and 0.1 kg of molasses per animal per day. The control group animals received endophyte free fescue seeds supplements, and the treatment group was fed endophyte-infected tall fescue seeds for 30 days.

value: Angus x Simmental

value: endophyte-infected tall fescue seeds


title: CMSIP for LSK cells from WT2 old mouse

value: LSK


title: Naive_MEL1_P20plus3

description:
RL1976
MEL1_gene_and_TE_counts.txt.gz

source_name: Naïve induced pluripotent stem cell


title: zebrafish, DMSO_TFA_1, 3dpf

value: tail


title: RNAseq for LSK cells from Tet2 KO1 old mouse

value: Tet2 KO


title: 9056G, liver, treatment


title: zebrafish, AR42_4, 3dpf

value: AR-42 treated larvae without TFA modeling


title: 9036G, liver, treatment


title: zebrafish, AR42_3, 3dpf


title: 9093G, liver, tolerant, control

tag: phenotype

value: tolerant

value: control


title: N2P_MEL1_P9plus13

description:
RL1978
MEL1_gene_and_TE_counts.txt.gz


title: RNAseq for LSK cells from Tet2 KO1 young mouse

value: young


title: zebrafish, AR42_2, 3dpf


title: TNT_MEL1_P17

description:
RL1973
MEL1_gene_and_TE_counts.txt.gz

source_name: TNT induced pluripotent stem cell

value: TNT


title: RNAseq for LSK cells from Tet2 KO2 young mouse


title: RNAseq for LSK cells from WT1 old mouse


title: zebrafish, AR42_1, 3dpf


title: WGBS for LSK cells from WT young mouse


title: zebrafish, DMSO_TFA_2, 3dpf


title: Primed_MEL1_P18

description:
RL1972
MEL1_gene_and_TE_counts.txt.gz

source_name: Primed indiced pluripotent stem cell

value: Primed


title: 9001G, liver, tolerant, control

value: liver


title: zebrafish, DMSO_4, 3dpf

value: DMSO treated larvae without TFA modeling


title: 9085G, liver, susceptible, control

value: susceptible


title: 9008G, liver, tolerant, control


title: Primed_MEL1_P17

description:
RL1971
MEL1_gene_and_TE_counts.txt.gz


title: Naive_MEL1_P15

description:
RL1970
MEL1_gene_and_TE_counts.txt.gz

value: Naïve


title: RNAseq for LSK cells from WT1 young mouse

value: WT


title: 9050G, liver, susceptible, control


title: Fibroblast_MEL1_P8

description:
RL1975
MEL1_gene_and_TE_counts.txt.gz

value: Fibroblast


title: RNAseq for LSK cells from Tet2 KO2 old mouse

value: old


title: RNAseq for LSK cells from WT2 old mouse


title: zebrafish, DMSO_3, 3dpf


title: zebrafish, DMSO_2, 3dpf


title: TNT_MEL1_P16

description:
RL1974
MEL1_gene_and_TE_counts.txt.gz


title: 9009G, liver, susceptible, control

value: Angus x Simmental


title: ESC_MEL1_P13plus14

description:
RL1967
MEL1_gene_and_TE_counts.txt.gz

source_name: Human embryonic stem cell

value: Control


title: ESC_MEL1_P13plus20

description:
RL1966
MEL1_gene_and_TE_counts.txt.gz


title: zebrafish, DMSO_1, 3dpf

value: tail


title: WT_EToH_set_4_S17

status: Public on Aug 10 2023

city: San Francisco

first: Kristen

last: Seim

address: 201 Haskins Way

institute: GraphiteBio

data_processing:
Fastq files from the different runs were merged using “cat”. The merged paired FASTQ files for each sample were run through FastQC (version 0.11.09) to obtain general QC metrics. Adapters were trimmed by Trim Galore (version 0.6.6). Salmon3 (version 1.4.0) was used for our alignment-free pipeline. Adapter-trimmed reads were used as input.
We quantified gene expression using raw counts and kept the genes that showed at least 5 reads in 80% of the samples.
We performed differential expression gene testing with DESeq2 (v.1.24.0 R package) using default settings. Replicate was used as covariates within the DESeq2 model. Statistical significance was set a 5% FDR (Benjamini-Hochberg).
The Bioconductor package fgsea (v 1.10.1 R package), was used for gene set enrichment analysis (GSEA).
Assembly: Danio Rerio reference genome (GRCz11) with associated transcript annotations (Gencode version GRCz11.103) was used.
Supplementary files format and content: Salmon quant files for liver samples from WT and abcb11b mutants
Supplementary files format and content: Raw counts for the 40 livers per replicate

extract_protocol:
Total mRNAs were extracted from pools of dissected livers from 6 dpf abcb11b mutants and WT siblings, 20 livers per replicate and 3 replicates per genotype, by using Arcturus PicoPure RNA isolation kit (Applied Biosystems, KIT0204). For the 40 livers per replicate (4-5 replicates per group) were dissected under a brightfield microscope at 7dpf and then pooled prior to RNA extraction using the Qiagen RNAeasy Micro kit. Animals in the overfed group were provided with six times the normal amount of food at each feeding from day 5 to 7 while EtOH treated animals were exposed to 2% ethanol for 32 hours prior to sample collection. The presence of hepatic steatosis was confirmed in both groups.
WT and abcb11b mutants samples were submitted to Novogene Co for library preparation and RNA-sequencing using the vendor’s standard protocol.  The 40 samples were  submitted to the Centre for Applied Genomics (TCAG, Toronto, Canada). SMART-Seq v4 Ultra Low Input RNA kit was used for the cDNA conversion. Library preparation followed the protocol of Nextera XT library prep kit. Libraries were quality controlled (QC)ed (run on the Bioanalyzer DNA High Sensitivity chip to check for size and quantified by qPCR using Kapa Library Quantification Illumina/ABI Prism Kit protocol). All libraries were pooled and sequenced on 2 lanes on the NovaSeq SP flowcell paired-end 2x100bp.

tag: time

value: EToH treated animal


title: WT_EToH_set_2_S15


title: WT_EToH_set_3_S16


title: WT_EF_set_3_S7

value: overfed animal


title: PerturbSci-Kinetics pooled screen

city: New York

first: Zihan

last: Xu

state: NY

address: 1230 York Ave

institute: The Rockefeller University

description: PerturbSci-Kinetics library

data_processing:
For PerturbSci-Kinetics transcriptome reads processing and whole-transcriptome/nascent transcriptome gene counting, the pipeline was developed based on EasySci10 and Sci-fate14 with minor modifications. After demultiplexing on index 7, Read1 were matched against a constant sequence on the sgRNA capture primer to remove unspecific priming, and cell barcodes and UMI sequences sequenced in Read1 were added to the headers of the fastq files of Read2, which were retained for further processing. After potential polyA sequences and low-quality bases were trimmed from Read2 by Trim Galore55, reads were aligned to a customized reference genome consisting of a complete hg38 reference genome and the dCas9-KRAB-MeCP2 sequence from Lenti-idCas9-KRAB-MECP2-T2A-mCherry-Neo using STAR. Unmapped reads and reads with mapping score < 30 were filtered by samtools57. Then deduplication at the single-cell level was performed based on the UMI sequences and the alignment location, and retained reads were split into SAM files per cell. These single-cell sam files were converted into alignment tsv files using the sam2tsv function in jvarkit58. Only reads with FLAG values of 0 or 16 and high-quality mismatches with QUAL scores > 45 and CIGAR of M in them were maintained. All mutations were transformed onto the plus strand and were further filtered against background SNPs called by VarScan using our in-house EasySci data on HEK293 cells. Reads in which at least 30% of mutations were T to C mismatches were identified as nascent reads, and the list of reads were extracted from single-cell whole transcriptome sam files by Picard59. Finally single-cell whole transcriptome gene x cell count matrix and nascent transcriptome gene x cell count matrix were retrieved by assigning reads to genes if the aligned coordinates overlapped with the gene locations on the genome. At the same time, single cell exonic/intronic read numbers were also counted by checking whether reads were mapped to the exonic or the intronic regions of genes. To quantify dCas9-KRAB-MECP2 expression, a customized gtf file consisting of the complete hg38 genomic annotations and additional annotations for dCas9 was used in this step.
Read1 and read2 of PerturbSci-Kinetics sgRNA libraries were matched against constant sequences respectively with a maximum of 1 mismatch allowed. For each filtered read pair, cell barcode, sgRNA sequence, and UMI were extracted from designed positions. Extracted sgRNA sequences with a maximum of 1 mismatch from the sgRNA library were accepted and corrected, and the corresponding UMI was used for deduplication. Duplicates were removed by collapsing identical UMI sequences of each individual corrected sgRNA under a unique cell barcode. Cells with overall sgRNA UMI counts higher than 10 were maintained and the sgRNA x cell count matrix was constructed.
For compatibility with the GEO paired-end sequencing database, Index5 barcodes of each read pair originally in separate fastq files have been attached to the beginning of headers.
Assembly: human reference genome (hg38) with two additional customized lentiviral sequences
Supplementary files format and content: Processed data files includes 3 mtx files consisting of the whole transcriptome gene count sparse matrix of on-target cells, the nascent transcriptome gene count sparse matrix of on-target cells, and the sgRNA count sparse matrix of on-target cells. The meta-data of these on-target cells are stored in a csv file.
Supplementary files format and content: on_target_whole_tx_count_matrix.mtx: the whole transcriptome gene count sparse matrix of on-target cells
Supplementary files format and content: on_target_nascent_tx_count_matrix.mtx: the nascent transcriptome gene count sparse matrix of on-target cells
Supplementary files format and content: on_target_sgRNA_count_matrix.mtx: the sgRNA count sparse matrix of on-target cells
Supplementary files format and content: on_target_cell_metadata.csv: the meta-data of on-target cells
Supplementary files format and content: whole_txome_sgRNA_sample_name_barcode_table.csv: the cross-reference table of whole transcriptome fastq file names and sgRNA readout fastq file names
Supplementary files format and content: on_target_whole_tx.Genes.tsv: the tsv file including the rownames (gene symbols) annotation for on_target_whole_tx_count_matrix.mtx
Supplementary files format and content: on_target_nascent_tx.Genes.tsv: the tsv file including the rownames (gene symbols) annotation for on_target_nascent_tx_count_matrix.mtx
Supplementary files format and content: on_target_sgRNA.Genes.tsv: the tsv file including the rownames (gene symbols) annotation for on_target_sgRNA_count_matrix.mtx
Supplementary files format and content: on_target_whole_tx.Barcodes.tsv: the tsv file including the colnames (cell barcodes) annotation for on_target_whole_tx_count_matrix.mtx
Supplementary files format and content: on_target_nascent_tx.Barcodes.tsv: the tsv file including the colnames (cell barcodes) annotation for on_target_nascent_tx_count_matrix.mtx
Supplementary files format and content: on_target_sgRNA.Barcodes.tsv: the tsv file including the colnames (cell barcodes) annotation for on_target_sgRNA_count_matrix.mtx

source_name: cell line

growth_protocol: HEK293-idCas9 cells were transduced with a sgRNA library at a low MOI and were selected by Puromycin. Then the dCas9-KRAB-MeCP2 expression was induced by adding Dox to the culture medium. Cells were cultured for additional 7 days and were passed every other day with 4000x coverage/sgRNA.

extract_protocol:
After trypsinization, cells in each 10cm dish were collected into a 15ml falcon tube and kept on ice. Cells were spun down at 300xg for 5 minutes (4 °C) and washed once in 3ml ice-cold PBS. Cells were fixed with 5ml ice-cold 4% PFA in PBS for 15 minutes on ice. PFA was then quenched by adding 250ul 2.5M Glycine, and cells were pelleted at 500xg for 5 minutes (4 °C). Fixed cells were washed once with 1ml PBSR (PBS, 0.1% SUPERase In, and 10mM dithiothreitol (DTT)), and were then resuspended, permeabilized, and further fixed in 1ml PBSR-triton-BS3 (PBS, 0.1% SUPERase In, 0.2% Triton-X100 (Sigma X100-500ML), 2mM bis(sulfosuccinimidyl)suberate (BS3), 10mM DTT) for 5 minutes. Additional 4ml of PBS-BS3 (PBS, 2mM BS3, 10mM DTT) was then added, and cells were incubated on ice for 15 minutes. Cells were pelleted at 500xg, 4 °C for 5 minutes and resuspended in 500ul nuclease-free water supplemented with 0.1% SUPERase In and 10mM DTT. 3ml of 0.05N HCl was added for further permeabilization. After 3 minutes of incubation on ice, 3.5ml Tris-HCl, pH 8.0, and 35ul of 10% Triton X-100 were added to each tube to neutralize the HCl. After spinning down at 4 °C, 500xg for 5 minutes, cells were finally resuspended in 400ul PSB-DTT at the concentration of ~2e6 cells/100ul (PBS, 1% SUPERase In, 1% Bovine Serum Albumin (BSA), 1mM DTT), mixed with 10% DMSO, and were slow-frozen and stored in -80 °C.
Fixed cells were thawed, then mRNA and sgRNA within each individual cell were indexed by indexing reverse transcription, indexing ligation, and finally indexing PCR. SgRNA libraries were further separated and enriched by another round of PCR. A step-by-step experimental protocol of PerturbSci-Kinetics library preparation is included in the supplementary material of the publication.

treatment_protocol: On day7, before the end of the screen, 6ml of the original media from each plate was mixed with 6uL of 200mM 4sU dissolved in DMSO and was put back for nascent RNA metabolic labeling. After 2 hours of treatment, cells were harvested for single-cell PerturbSci-Kinetics profiling.


title: WT_EF_set_4_S8


title: WT_EToH_set_1_S14


title: WT_EF_set_2_S6


title: MTM1_set_2_S12

value: mtm mutant


title: WT_EF_set_1_S5


title: MTM1_set_3_S13


title: MTM1_set_1_S11


title: MTM1_3R_resub_Jan18_21_S22


title: MTM1_R1_resub_Jan11_21_S20


title: RNAseq for LSK cells from WT2 young mouse

address: 2121 W HOLCOMBE BLVD

institute: Guangzhou Medical University

value: LSK


title: AB_set_4_S4


title: MTM1_2R_resub_Jan18_21_S21


title: 121_AYWB

first: Eva

last: Varallyay

address: 4 Szent-Györgyi Albert Street

institute: Hungarian University of Agriculture and Life Sciences

description: small RNA

data_processing:
The quality check of the sequenced small RNA libraries was done by FastQC Program and CLC Genomic Workbench 11
Trimming of raw reads was performed using the CLC Genomic Workbench 11
Removing redundant reads using the CLC Genomic Workbench 11
Supplementary files format and content: trimmed redundant list, each read with its number of repeats

source_name: leaf

extract_protocol:
Phenol chloroform extraction method (White & Kaper, 1989) DOI: 10.1016/0166-0934(89)90122-5
modified Illumina Trueseq small RNA kit protocol: https://doi.org/10.1007/978-1-4939-7683-6_9

value: leaf

tag: symptoms

value: witches broom

tag: ay phytoplasma presence

value: yes

tag: year of sampling

tag: place of collection

value: glasshouse, with insectproof net


title: AB_set_1_S1

value: liver


title: WT rep3

value: 6 dpf


title: AB_set_2_S2


title: WT rep1


title: WT rep2


title: abcb11b mutants  rep1

value: abcb11b mutant


title: abcb11b mutants  rep2


title: Del14-4N24 DnybW PkasOP::nybF::tkt, 70 h , B

city: Bielefeld

first: Christian

last: Ruckert-Reed

address: Sequenz 1

department: Medical School OWL

country: Germany

institute: Bielefeld University

data_processing:
Basecalls were performed using RTA v 1.18.66.3
Sequenced reads were mapped to the C113229.1 whole genome using bowtie2 v2.3.2 with parameters -X 600
Raw read counts per gene were calculated using featureCounts v2.0.0 with parameters -O -M -t gene -g locus_tag -s 1
Read cound normalization and data analysis were performed using DESeq2
Assembly: CP113229.1
Supplementary files format and content: tab-delimited matrix table with raw read counts for every gene and every sample
Supplementary files format and content: tab-delimited matrix table with normalized read counts derived via DESeq2 for each gene

organism: Streptomyces albidoflavus

source_name: liquid culture

growth_protocol: Main cultures to overproduce nybomycin were grown in chemically defined minimal medium containing per liter: 10 g of D-mannitol, 15 g of (NH4)2SO4, 20.9 g of MOPS, 0.5 g of K2HPO4, 1 g of NaCl, 0.55 g of  CaCl2, 0.2 g of MgSO4·7H2O, 0.02 g of FeSO4·7H2O, 2 mg of FeCl3·6H2O, 2 mg of MnSO4·H2O, 0.5 mg of ZnSO4·H2O, 0.5 mg of CuCl2·2H2O, 0.2 mg of Na2B4O7·10H2O, 0.1 mg of (NH4)6Mo7O24·4H2O, 1 mg of nicotinamide, 1 mg of riboflavin, 0.5 mg of thiamine hydrochloride, 0.5 mg of pyridoxine hydrochloride, 0.2 mg of biotin and 0.1 mg of p-aminobenzoic acid. Liquid cultures were incubated in 500 mL baffled shake flasks (10% filing volume) on an orbital shaker at 230 rpm, 75% relative humidity, and 28 °C (5 cm shaking diameter). The medium was supplemented with 30 g 5mm soda-lime glass beads. Samples were taken after 12, 46, and 70 hours of cultivation.

extract_protocol:
Total RNA was isolated using the Quick-RNA Miniprep Plus kit (Zymo Research, Freiburg, Germany). After DNase treatment, the obtained RNA was purified (RNA Clean & Concentrator-5 kit, Zymo Research) and quantified (DropSense 16, Trinean NV, Gent, Belgium).
To construct whole transcriptome cDNA libraries, 2.5 μg of total RNA each was depleted of rRNA (Ribo-Zero rRNA Removal Kit (Bacteria), Illumina, San Diego, CA, USA). Successful rRNA removal was validated (Agilent RNA Pico 6000 kit, Agilent 2100 Bioanalyzer, Agilent Technologies). The obtained mRNA was converted into a cDNA library according to the TruSeq Stranded mRNA Sample Preparation guide (Illumina). Appropriate cDNA quality and quantity was validated (Agilent High Sensitivity DNA kit, Agilent 2100 Bioanalyzer, Agilent Technologies). Sequencing was performed on an Illumina NextSeq 500 instrument using 75 bases read length (Illumina).

value: liquid culture

value: Del14-4N24 DnybW PkasOP::nybF::tkt


title: Del14-4N24 DnybW PkasOP::nybF::tkt, 70 h , C


title: Del14-4N24 DnybW PkasOP::nybF::tkt, 70 h , A


title: Del14-4N24 DnybW PkasOP::nybF::tkt, 46 h, B


title: Del14-4N24 DnybW PkasOP::nybF::tkt, 12 h, B


title: Del14-4N24 DnybW PkasOP::nybF::tkt, 12 h, C


title: Del14-4N24 DnybW PkasOP::nybF::tkt, 46 h, A


title: Del14-4N24 DnybW PkasOP::nybF::tkt, 12 h, A


title: Del14-4N24 DnybW PkasOP::nybF::zwf2, 70 h , C

value: Del14-4N24 DnybW PkasOP::nybF::zwf2


title: Del14-4N24 DnybW PkasOP::nybF::zwf2, 70 h , B


title: AB_set_3_S3

value: WT


title: Del14-4N24 DnybW PkasOP::nybF::zwf2, 70 h , A


title: abcb11b mutants  rep3


title: Del14-4N24 DnybW PkasOP::nybF::zwf2, 46 h, C


title: Del14-4N24 DnybW PkasOP::nybF::zwf2, 46 h, B


title: Del14-4N24 DnybW PkasOP::nybF::zwf2, 46 h, A


title: Del14-4N24 DnybW PkasOP::nybF::zwf2, 12 h, C


title: Del14-4N24 DnybW PkasOP::nybF::zwf2, 12 h, B


title: Del14-4N24 PkasOP::nybF::zwf2, 70 h , A

value: Del14-4N24 PkasOP::nybF::zwf2


title: Del14-4N24 DnybW PkasOP::nybF::zwf2, 12 h, A


title: Del14-4N24 DnybW PkasOP::nybF::tkt, 46 h, C


title: Del14-4N24 PkasOP::nybF::zwf2, 70 h , B


title: Del14-4N24 PkasOP::nybF::zwf2, 46 h, B


title: Del14-4N24 PkasOP::nybF::zwf2, 12 h, C


title: Del14-4N24 PkasOP::nybF::zwf2, 70 h , C


title: Del14-4N24 PkasOP::nybF::zwf2, 46 h, C


title: Del14-4N24 PkasOP::nybF::zwf2, 12 h, A


title: Del14-4N24 PkasOP::zwf2, 70 h , C

value: Del14-4N24 PkasOP::zwf2


title: Del14-4N24 PkasOP::nybF::zwf2, 46 h, A


title: Del14-4N24 PkasOP::nybF::zwf2, 12 h, B


title: Del14-4N24 PkasOP::zwf2, 12 h, C


title: Del14-4N24 PkasOP::zwf2, 70 h , A


title: Del14-4N24 PkasOP::zwf2, 12 h, B


title: Del14-4N24 PkasOP::zwf2, 70 h , B


title: Del14-4N24, 70 h , B


title: Del14-4N24, 70 h , A


title: Del14-4N24, 12 h, C


title: Del14-4N24, 12 h, B


title: Del14-4N24, 12 h, A


title: Del14-4N24 PkasOP::zwf2, 12 h, A


title: Del14-4N24, 70 h , C


title: cDNA_BR2_EEP_Ap1m1_Run2

city: Amsterdam

first: Bas

last: van Steensel

address: Plesmanlaan 121

country: Netherlands

institute: The Netherlands Cancer Institute (NKI)

data_processing:
Basecalling, filtering and demultiplexing were performed using standard software of the Illumina MiSeq, NextSeq or NovaSeq platforms
For cDNA and pDNA samples, the barcode sequence was extracted from the read (using a custom python scripts) and counted.
For iPCR samples the insert sequence was aligned to a custom genome (using bowtie2) containing only the target sequences extracted from the mm10 genome, and the barcode was extracted from the ead using a custom python script.
Barcodes were clustered using Starcode (version 1.1) {Zorita, 2015, 25638815} to remove errors from barcode sequencing.
barcodes  matched with multiple element combinations were removed from the data
Per  combination an activity value was calculated as the ratio of the mean readcount in the RNA samples and the mean readcount in the pDNA library samples across all barcodes assigned to the singlet or combination.
Baselines were calculated across control-control combinations and then used to calculate boost indices as the log2 ratio between the activity of a combination and the baseline
Assembly: mm10
Supplementary files format and content: tsv format, each row contains a unique EEP combination, with fragment identifiers, calculated activities, baselines and Boost indices

growth_protocol: E14TG2a mouse embryonic stem cells (mESC) (ATCC CRL-1821) were cultured in 2i+LI. 2i+LIF was made according to the 4DN nucleome protocol for culturing mESCs (https://data.4dnucleome.org/protocols/cb03c0c6-4ba6-4bbe-9210-c430ee4fdb2c/).

extract_protocol:
For iPCR samples, each library was digested overnight with I-CeuI (#R0699S, NEB), barcode-insert fragments were then circularized, remaining linear fragments digested, and barcode-insert fragments were linearized again with I-SceI (#R0694S; NEB). Barcode-insert fragments were then amplified by PCR with Illumina adapters and sequenced on an Illumina NextSeq 550 platform using 150 bp paired-end sequencing. For cDNA samples, cells were collected 24 hours after transfection in 5 ml of TRIsure. From each sample, the aqueous phase containing the total RNA of the TRIsure solution was extracted and purified on an RNA extraction column (#K0732, Thermo Scientific). Total RNA was digested with 10 units of DNase I (#04716728001; Roche) for 30 minutes in 10-8 10 µl reactions containing 5 µg of RNA. DNase I was inactivated by addition of 1 µl of 25 mM EDTA and incubation at 70°C for 10 min. cDNA was produced in 20 µl reactions (1 per 10 µDNAse reaction) using Maxima Reverse transcriptase (#EP0743; ThermoFisher Scientific) and a gene specific (targeting the GFP reporter) primer (JvA304, see Supplementary table 3 for sequence). dNTPs and the primer were mixed with the DNAse digested RNA and incubated at 65°C for 5 minutes. The RT buffer, enzyme and RNAse inhibitor were added and the reaction was incubated at 50 °C for 1 hour. cDNA was then amplified by two nested PCRs to make it strand specific. The first PCR reaction uses (index variants of 285JvA containing the S2, index and p7 adaptor)  and 305JvA (targeting the adapter introduced by 304JvA). Each 20 µl RT reaction was amplified in a 100-µl PCR reaction with MyTaq Red mix (#BIO-25043; Bioline). The second PCR reaction was performed using 10 µl of the product of the previous reaction in a 100 µl reaction using the same index variant primer and index variants of 437JvA (containing the S1, index and p5 adaptor) (see Supplementary table 3 for primer sequences). The first PCR was run for 10 cycles and the second one for 8 cycles using the recommended Mytaq Red mix conditions ( for both PCRs: 1 min 96 °C, then each cycle 15 s 96 °C, 15 s 60 °C, 15 s 72 °C). PCR products were purified with magnetic beads and mixed for sequencing in an Illumina NovaSeq 6000 platform using 100bp single end reads. For pDNA samples, 1 µg of each plasmid library was digested with I-SceI in order to linearise the plasmid. Barcodes were amplified by PCR for 9 cycles from 50 ng of material. Primers and reaction conditions were the same as in the amplification of cDNA. PCR products were purified using magnetic beads and mixed for sequencing.
PCR

treatment_protocol: Each EEP library was transfected separatedly. Cells were transiently transfected using Amaxa nucleofector II, program A-30, and Mouse Embryonic Stem Cell NucleofectorTM Kit (#VPH-1001, Lonza). Three independent biological replicates were done for each library mix. For each biological replicate 20 million cells were transfected (5 million cells with 4 µg plasmid per cuvette).

value: mESCs


title: cDNA_BR3_EEP_Tbx3_Run2


title: cDNA_BR3_EEP_Ap1m1_Run2


title: pDNA_EEP_Nanog_Run2


title: cDNA_BR3_EEP_Lefty1_Run2


title: pDNA_EEP_Ap1m1_Run2


title: cDNA_BR2_EEP_Nanog_Run2


title: cDNA_BR3_EEP_Nanog_Run2


title: pDNA_EEP_Tbx3_Run2


title: cDNA_BR1_EEP_Lefty1_Run2


title: cDNA_BR2_EEP_Tbx3_Run2


title: cDNA_BR1_EEP_Tbx3_Run2


title: cDNA_BR1_EEP_Nanog_Run2


title: pDNA_EEP_Lefty1_Run2


title: cDNA_BR3_EEP_Fgf5_Run2