title: healthy_CD34_4


title: GFPDay3-3

status: Public on Mar 16 2021

type: SRA

city: rochester

first: Yugo

last: Morita

state: New York

address: 601 Elmwood Avenue, Box 665, Center for Musculoskeletal Research

country: USA

institute: University of Rochester Medical Center

library_source: transcriptomic

data_processing:
The libraries were sequenced with the Illumina NovaSeq6000 platform (Illumina).
Quality filtering and adapter removal were performed by fastp version 0.20.0 (78) using the following parameters: “--in1 ../${SAMPLE}_R1.fastq.gz --out1 clt_${SAMPLE}_R1.fastq.gz --length_required 35 --cut_front_window_size 1 --cut_front_mean_quality 13 --cut_front --cut_tail_window_size 1 --cut_tail_mean_quality 13 --cut_tail -w 8 -y -r -j ${SAMPLE}_fastp.json”.
The remaining high quality processed reads were then mapped to the Mus musculus genome reference (GRCm38.p6) with STAR version 2.7.0f using the following parameters: “--twopassMode Basic --runMode alignReads --genomeDir ${GENOME} --readFilesIn ${SAMPLE} --outSAMtype BAM Unsorted --outSAMstrandField intronMotif --outFilterIntronMotifs RemoveNoncanonical”.
The mapped reads were counted within the GRCm38.p6 gene annotations using featureCounts read quantification program in Subread version 1.6.4.
Data normalization were performed using DESeq2 version 1.22.1 within the R version 3.5.1.
Genome_build: GRCm38.p6
Supplementary_files_format_and_content: tab-delimited text files include normarized gene counts for every sample
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

molecule: total RNA

organism: Mus musculus

source_name: Tibia

extract_protocol:
Tibia were pulverized at liquid nitrogen temperature and homogenized using Bullet Blender Gold (Next Advance). Collection of Total RNA from homogenized tibia was performed using TRIzol extraction (ThermoFisherScientific) and RNeasy Mini Kits (Qiagen).
The TruSeq Stranded Total RNA Library Prep Gold (Illumina) was utilized for next generation sequencing library preparation per the manufacture’s instructions.

tag: strain

tag: tissue

value: tibia

tag: genotype

value: expressing GFP

tag: time post-septic surgery

value: day 3


title: CW20R2

city: Irapuato

first: Octavio

last: Martinez de la Vega

state: Guanajuato

address: km 9.6 Libramiento Norte

department: Unidad de Genómica Avanzada (Langebio)

country: Mexico

institute: Cinvestav

description: Whole fruits of accession California Wonder at 20 Days After Anthesis. Biological replicate R2.

data_processing:
Total RNA Sample QC. All samples need to pass through the following four steps before library construction: (1) Nanodrop: tests RNA purity (OD260/OD280). (2) Agarose Gel Electrophoresis: tests RNA degradation and potential contamination. (3) Agilent 2100: checks RNA integrity.
Library Construction. After the QC procedures, mRNA was enriched using oligo(dT) beads. The mRNA was then fragmented randomly in fragmentation buffer, followed by cDNA synthesis using random hexamers and reverse transcriptase. After first-strand synthesis, a custom second-strand synthesis buffer (Illumina) is added with dNTPs, RNase H and Escherichia coli polymerase I to generate the second strand by nick-translation. The final cDNA library is ready after a round of purification, terminal repair, A-tailing, ligation of sequencing adapters, size selection and PCR enrichment.
Library QC and sequencing. Library concentration was first quantified using a Qubit 2.0 fluorometer (Life Technologies), and then diluted to 1 ng/µl before checking insert size on an Agilent 2100 and quantifying to greater accuracy by quantitative PCR (Q-PCR) (library activity >2 nM). Libraries were fed into Illumina NovaSeq 6000 machines for sequencing.
Raw data and quality control. The original raw data from Illumina were transformed to Sequenced Reads by base calling. Raw data were recorded in a FASTQ files. GC content distribution was evaluated. Raw reads were filtered to remove reads containing adapters or reads of low quality.
Mapping to the Capsicum annuum reference genome. For mapping TopHat (v2.0.12) was employed with mismatch parameter set to 2 and other parameters set to defaults. Only filtered reads were used to analyze the mapping status of RNA-seq data to the reference genome. RAW FILES  were created in the binary alignment/map format (".bam").
Genome_build: Capsicum_annuum.Annuum.v.2.0.genome.fa
Supplementary_files_format_and_content: Processed data files are in the cvs format with three columns, 1 - "id" (a unique numeric identifier of each feature), 2 - "Gene.id" (the identifier of the gene or gene feature) and 3 - A column with the same name than the sample which contains the read count for the corresponding feature.

organism: Capsicum annuum

source_name: California Wonder

growth_protocol: Seeds of 10 Capsicum annuum accessions were surface sterilized with a 70% ethanol solution for 10 s before treatment with a 10% hypochlorite solution for 10 s and six rinses with distilled water. Wild accession seeds were similarly treated, after an initial treatment with 50% sulfuric acid solution to break seed dormancy. All accession seeds were germinated in plastic trays containing a mixture of three parts peat moss, one part perlite, one part vermiculite, one part sludge and two parts forest soil in a growth chamber, with 16 h light (photon flux of 70 μmol m-2 s-1) at 28 °C and 66% relative humidity. Three-week-old chili pepper plants were transplanted individually into plastic 5 L pots containing the same soil mixture described above. During transplantation, 15 g of a mycorrhizae fungal and beneficial bacteria mixture were added to optimize root growth and development. Plants were fertilized with Long Ashton solution every two weeks. Flowers and fruits at 0, 10, 20, 30, 40, 50 and 60 DAA were collected and immediately frozen in liquid nitrogen, and stored at -80 °C.

extract_protocol:
Total RNA was extracted from flowers and whole chili pepper fruits at different developmental stages using a NucleoSpin™ RNA Plant kit (MACHEREY-NAGEL) according to the manufacturer’s instructions. RNA was extracted from two biological samples comprising either flowers (time 0 DAA) fruits or seedlings from 2-6 different plants. RNA quality was verified by determining the RNA Integrity Number (RIN) for each sample.
Samples of total RNA were shipped to Novogene (https://en.novogene.com/) for library construction, sequencing and mapping to a reference genome. At Novogene, libraries were prepared and sequenced using the Illumina NovaSeq platform to obtain at least 20 million raw paired-end reads of 150 bp per sample.

treatment_protocol: No treatments were employed. Samples of fruits and seedlings were collected at the times indicated in the "characteristics: collection time" of the "SAMPLES" section.

tag: sample type

value: Domesticated variety

value: Whole fruit

tag: collection time

value: 20 Days After Anthesis (DAA)


title: GFPDay7-2

value: day 7


title: CW20R1

description: Whole fruits of accession California Wonder at 20 Days After Anthesis. Biological replicate R1.


title: OKseq_CCSER1_locus_rep1

city: PARIS

first: Benoit

last: Le Tallec

address: 46 rue d'Ulm

country: France

institute: IBENS

description: Okazaki fragments

library_source: genomic

data_processing:
CASAVA-1.8.2 (demultiplexing)
Fastqc 0.10.1 (read quality)
Cutadapt-2.1 (adaptor trimming)
BWA-0.7.15-r1140 (mapping)
Aligned OK-seq data (BAM files) were imported in R as GenomicAlignments, and reads were converted into GenomicRanges. Replication fork directionality (RFD) was computed using R as the difference between rightward- and leftward-fork coverage normalized by total coverage. Data were binned into nonoverlapping 10-kb windows. RFD profiles were exported into bigwig files using the export function from the rtracklayer package.
Genome_build: galGal4
Supplementary_files_format_and_content: bigWig files with RFD scores

molecule: genomic DNA

organism: Gallus gallus

source_name: Bursa

growth_protocol: DT40 cells were grown in RPMI 1640 medium with 10% fetal bovine serum, 1% chicken serum, 0.05 mM β-mercaptoethanol, 100 U.ml−1 penicillin and 100 μg.ml−1 streptomycin at 37 °C, 20% O2, 5% CO2.

extract_protocol:
Asynchronously growing cells were labelled with 20 μM 5-ethynyl-2'-deoxyuridine (EdU, Jenabioscience) for 2 minutes. After DNA purification and heat-denaturation, Okazaki fragments were size-purified on sucrose gradients, labelled with biotin at EdU sites using click-chemistry, captured on magnetic beads coated with streptavidin and amplified by PCR.
Libraries were prepared using standard Illumina protocols.

tag: cell type

value: Lymphoblast

tag: cell line

tag: treatment

value: Untreated


title: GFPDay3-2


title: CW10R2

description: Whole fruits of accession California Wonder at 10 Days After Anthesis. Biological replicate R2.

value: 10 Days After Anthesis (DAA)


title: OKseq_DMD_locus_rep3

description: Okazaki fragments


title: GFPDay3-1


title: CW30R1

description: Whole fruits of accession California Wonder at 30 Days After Anthesis. Biological replicate R1.

value: 30 Days After Anthesis (DAA)


title: healthy_CD34_20

city: Ulm

first: Susanne

last: Lux

address: Albert-Einstein-Allee 23

country: Germany

institute: University Hospital Ulm

data_processing:
alignment with STAR 2.4.2
normalization with DEseq2
Genome_build: hg19
Supplementary_files_format_and_content: txt with normalized read counts

organism: Homo sapiens

source_name: healthy G-CSF mobilized CD34+ cells

extract_protocol:
QIAGEN AllPrep
TruSeq Stranded Total RNA Ribominus Gold
100 bp paired-end sequencing on HiSeq2000

value: G-CSF mobilized healthy CD34+


title: healthy_CD34_23


title: OKseq_DMD_locus_rep2

description: Okazaki fragments


title: OKseq_CCSER1_locus_rep3

description: Okazaki fragments


title: GFPDay14-2

value: day 14


title: healthy_CD34_21


title: OKseq_DMD_locus_rep1

description: Okazaki fragments


title: healthy_CD34_22


title: CW00R2

description: Whole fruits of accession California Wonder at 0 Days After Anthesis. Biological replicate R2.

value: 0 Days After Anthesis (DAA)


title: GFPDay7-1


title: healthy_CD34_16


title: OKseq_CCSER1_locus_rep2

description: Okazaki fragments


title: GFPDay1-1

value: day 1


title: healthy_CD34_12


title: GFPDay1-2


title: healthy_CD34_13


title: CW00R1

description: Whole fruits of accession California Wonder at 0 Days After Anthesis. Biological replicate R1.


title: CW10R1

description: Whole fruits of accession California Wonder at 10 Days After Anthesis. Biological replicate R1.


title: healthy_CD34_19


title: healthy_CD34_11


title: CQ60R2

description: Whole fruits of accession F1 of cross CM x QU at 60 Days After Anthesis. Biological replicate R2.

source_name: F1 of cross CM x QU

value: F1 cross between Domesticated and Wild

value: 60 Days After Anthesis (DAA)


title: CQ60R1

description: Whole fruits of accession F1 of cross CM x QU at 60 Days After Anthesis. Biological replicate R1.


title: healthy_CD34_10


title: CQ50R1

description: Whole fruits of accession F1 of cross CM x QU at 50 Days After Anthesis. Biological replicate R1.

value: 50 Days After Anthesis (DAA)


title: GFPDay1-3

value: tibia

value: expressing GFP


title: CQ50R2

description: Whole fruits of accession F1 of cross CM x QU at 50 Days After Anthesis. Biological replicate R2.


title: Aged MCAO, biological rep 3

type: RNA

city: Chengdu

first: Yingjia

last: Guo

state: Sichuan

address: 1 Keyuan 4 Road, Gaopeng Avenue

department: West China Hospital

country: China

institute: Sichuan University

description: Gene expression data from MCAO-subjected aged rats

hyb_protocol: Following fragmentation, 10 ug of cRNA were hybridized for 16 hr at 45C on GeneChip Rat Gene 2.0. GeneChips were washed and stained in the Affymetrix Fluidics Station 400.

scan_protocol: GeneChips were scanned using the GeneChip® Scanner 3000 (Affymetrix).

data_processing: The data were analyzed with RMA using Affymetrix default analysis settings and global scaling as normalization method. The trimmed mean target intensity of each array was arbitrarily set to 100.

label: biotin

organism: Rattus norvegicus

source_name: The peri-infarct cortex of  Rattus norvegicus

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 6 ug total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).

growth_protocol: Four animal groups were studied: sham-operated young rats group, sham-operated aged rats group, MCAO-subjected young rats group, and MCAO-subjected aged rats group. Transient middle cerebral artery occlusion (MCAO) for 60 minutes established acute ischemic stroke followed by 3 days reperfusion.

extract_protocol: RNA sample preparations were performed by CapitalBio Corporation (Beijing, China), according to the procedures outlined by Affymetrix.

treatment_protocol: The cortex or peri-infarct cortex was harvested and rinsed with cold PBS containing diethyl pyrocarbonate for eliminating RNase.

value: cortex

value: Sprague-Dawley

tag: Sex

value: male

tag: age

value: 18-20 months, body weight 585-685 g

value: MCAO


title: U2AF1_4

source_name: AML leukemic blasts

tag: mutation

value: diagnosis


title: CO-CM_BMDM_Rep2

city: Aurora

first: Sushil

last: Kumar

institute: University of Colorado Anschutz medical campus

data_processing:
QC of sequenced read were checked by FastQC
Sequenced reads were trimmed for adaptor sequence by using AfterQC,  then mapped to mouse GRCm38v91. The RNA-Seq FASTQ reads were aligned using the following STAR parameters: --genomeDir INDEX; --sjdbGTFfile GTF; --runThread 16 ; --outSAMstrandField intronMotif ; --outFilterIntronMotifs RemoveNoncanonical ; --readFilesIn fastq1 fastq2 ; --readFilesCommand zcat ; -- outSAMtype BAM Unsorted ;--outReadsUnmapped Fastx ;--outSAMmode Full.
Applied featureCounts v1.6.2 (parameters used: -T 16; -t exon; -g gene_id; -a GTF; bam) to assign reads to genes using mouse transcript annotations as a guide.
EdgeR (version 3.14) in R (version 3.4.3) was used for data normalization and differential expression analysis of RNA-Seq expression profiles with biological replication.
Genome_build: GRCm38v91
Supplementary_files_format_and_content: Gene expression levels were normylized by RPKM.

source_name: bone marrow derived macrophages

growth_protocol: Mouse bone marrow-derived macrophages (BMDMs) were prepared by incubating extracted bone marrow from male wild-type C57B6 mice in “Macrophage Growth Medium”, DMEM media containing 5mM glutamine, 25mM HEPES, 1mM pyruvate supplemented with 10% FBS (Endotoxin levels below 0.3 IU) and 50ng/ml CSF1, for 5-10 days. For CM treatment experiments, 0.5 million BMDMs were plated per well of a 24well dish.  BMDMs were treated with fibroblast CM for 16 hours for RNA isolation

extract_protocol:
Total RNA was purified using NucleoSpin RNA Isolation kit (Machery-Nagel, www.mn-net.com) according to the manufacturer’s instructions. RNA quantity and quality were evaluated using NanoDrop and Bioanalyzer.
RNA-Seq library preparation and sequencing were conducted using a NuGen universal plus mRNA-Seq kit. Sequencing was performed on Illumina NovaSEQ6000 system, using the paired-read 2x150 cycle option. RNA-Seq reads were generated using Illumina NovaSEQ6000 analysis pipeline. The read quality of all the samples were checked using FastQC v0.11.5. Adapter trimming, quality control, and base correction were performed by AfterQC (Chen et al., 2017). The sequencing reads in the FASTQ files of all the samples were aligned to the Mus musculus reference genome (Mus_musculus.GRCm38v91) with Spliced Transcripts Alignment to a Reference (STAR) version 2.6.0c. We next applied featureCounts v1.6.2 to assign reads to genes using mouse transcript annotations as a guide. EdgeR in R was used for data normalization and differential expression analysis of RNA-Seq expression profiles with biological replication. The Benjamini-Hochberg procedure was used to control the false discovery rate (FDR), and a cut-off criterion of FDR ≤ 0.05 was applied to identify differentially expressed genes. Individual gene expression was calculated as ‘reads per kilobase per million mapped reads’ (RPKM).

treatment_protocol: BMDMs were treated with either CO-CM or PH-CM for 24hrs or left untreated

value: bone marrow derived macrophages

value: treated with CO-CM


title: PH-CM_BMDM_Rep1

value: treated with PH-CM


title: PH-CM_BMDM_Rep2


title: CO-CM_BMDM_Rep3


title: U2AF1_6


title: Aged MCAO, biological rep 1


title: U2AF1_5


title: CO-CM_BMDM_Rep1


title: U2AF1_3


title: CQ40R2

description: Whole fruits of accession F1 of cross CM x QU at 40 Days After Anthesis. Biological replicate R2.

value: Whole fruit

value: 40 Days After Anthesis (DAA)


title: CQ40R1

description: Whole fruits of accession F1 of cross CM x QU at 40 Days After Anthesis. Biological replicate R1.


title: U2AF1_2


title: UNX_BMDM_Rep3

value: untreated


title: U2AF1_1


title: SRSF2_SF3B1


title: Aged MCAO, biological rep 2


title: Young MCAO, biological rep 2

description: Gene expression data from MCAO-subjected young rats

value: 3-4 months, body weight 340-400 g


title: Aged sham, biological rep 3

description: Gene expression data from sham-operated aged rats

source_name: The cortex of  Rattus norvegicus

value: sham


title: Young MCAO, biological rep 3


title: CQ30R2

description: Whole fruits of accession F1 of cross CM x QU at 30 Days After Anthesis. Biological replicate R2.

value: F1 cross between Domesticated and Wild

value: 30 Days After Anthesis (DAA)


title: CQ20R2

description: Whole fruits of accession F1 of cross CM x QU at 20 Days After Anthesis. Biological replicate R2.

value: 20 Days After Anthesis (DAA)


title: PH-CM_BMDM_Rep3


title: Aged sham, biological rep 2

value: cortex


title: Young MCAO, biological rep 1

value: Sprague-Dawley


title: hnCD16/CD38KO/IL-15RF iNK cells

city: Minneapolis

first: Frank

middle: Martin

last: Cichocki

state: Minnesota

address: 420 Delaware Street SE

department: Department of Medicine

institute: University of Minnesota

data_processing:
cellranger-4.0.0
Genome_build: GRCh38
Supplementary_files_format_and_content: Each matrix is stored in the Market Exchange Format (MEX) for sparse matrices. It also contains gzipped TSV files with feature and barcode sequences corresponding to row and column indices respectively.

source_name: Natural killer cells

extract_protocol:
Cells were prepared per standard 10x Genomics protocol using Chromium Single Cell 3' Reagent Kits (v3 Chemistry)
Libraries were prepared per standard 10x Genomics protocol.

value: iPSC-derived NK cells

tag: transduction


title: J_S10

city: Memphis

first: Beisi

last: Xu

state: Tennessee

address: 262 Danny Thomas Pl

department: Center for Applied Bioinformatics

institute: St Jude Children's Research Hosipital

data_processing:
Library strategy: TCR-seq
VDJ gene assignments were calculated and filtered with MIGEC software. VDJtools v1.2.1 software was applied for filtering non-functional TCR and decontaminate samples
Immunarh package (http://doi.org/10.5281/zenodo.3367200) was used to explore the T cell repertoires
Genome_build: mm10
Supplementary_files_format_and_content: Processed data contains CDR3 nucleotide sequence, CDR3 amino acid sequence, V gene usage, D gene usage and J gene usage. And different TCR counts and frequency.

source_name: Lymphocytes

growth_protocol: Regulatory T cells isolated ex vivo were directly used for RNA extraction and TCR sequencing.

extract_protocol:
Total RNA was extracted by TRIzol according to the manufacturer’s instructions. cDNA was synthesized by SMARTScribe Reverse Transcriptase with primers targeting mouse TCRa or TCRb chains and template switch adapter
TCRa and TCRb chains were further amplified by a two-step protocol. Sequencing adaptors were then added by Kapa Hyper Prep Kit (Kapa Biosystems). Samples were pooled at the same molar ratios and sequenced by Nova Seq with at least 20x coverage relative to the numbers of Treg cell used.

treatment_protocol: No treatment.

value: Regulatory T cells isolated from the spleen

value: WT


title: CQ30R1

description: Whole fruits of accession F1 of cross CM x QU at 30 Days After Anthesis. Biological replicate R1.


title: UNX_BMDM_Rep2


title: SRSF2_5

value: diagnosis


title: UNX_BMDM_Rep1

value: bone marrow derived macrophages


title: CQ10R2

description: Whole fruits of accession F1 of cross CM x QU at 10 Days After Anthesis. Biological replicate R2.

value: 10 Days After Anthesis (DAA)


title: CQ20R1

description: Whole fruits of accession F1 of cross CM x QU at 20 Days After Anthesis. Biological replicate R1.


title: J_S6

value: KO


title: Non-transduced iNK cells

value: non-transduced


title: Aged sham, biological rep 1

value: male

value: 18-20 months, body weight 585-685 g


title: SF3B1_4


title: SRSF2_4


title: Young sham, biological rep 3

description: Gene expression data from sham-operated young rats

value: 3-4 months, body weight 340-400 g


title: Young sham, biological rep 2

value: sham


title: CQ10R1

description: Whole fruits of accession F1 of cross CM x QU at 10 Days After Anthesis. Biological replicate R1.

value: F1 cross between Domesticated and Wild

value: Whole fruit


title: SRSF2_3


title: Young sham, biological rep 1


title: hnCD16/CD38KO iNK cells


title: J_S8


title: CQ00R2

description: Whole fruits of accession F1 of cross CM x QU at 0 Days After Anthesis. Biological replicate R2.

value: 0 Days After Anthesis (DAA)


title: SF3B1_3


title: J_S4


title: SRSF2_1


title: J_S9


title: CO50R2

description: Whole fruits of accession Piquin Coahuila at 50 Days After Anthesis. Biological replicate R2.

source_name: Piquin Coahuila

value: Wild accession (C. a. var. glabriusculum)

value: 50 Days After Anthesis (DAA)


title: SRSF2_2


title: J_S3


title: tbc-11(ok2567)rep1

city: Riverside

first: Weifeng

last: Gu

state: California

address: 900 University Ave

department: Molecular, Cell and Systems Biology

institute: University of California at Riverside

description: Small RNA analysis

data_processing:
The samples are debarcoded using the 8 nt barcodes
The 3' AGATCGGAA and beyond are removed and only reads of size >16 are kept
Small RNAs are mapped to genome and cDNA using Bowtie 0.12.7 and at most 2 mismatches are considered
Genome_build: C. elegans WS215
Supplementary_files_format_and_content: sequence and read number for the analysis

organism: Caenorhabditis elegans

source_name: Cloning tbc-11(ok2567) small RNA

growth_protocol: Worms were grown on NGM plates with OP50 at 20 °C.

extract_protocol:
RNAs were extracted with TRIzol
RNAs were linked with Truseq LT/V1/V2 linkers using the standard 5' and 3' ligation methods. Amplicons with ~20 nt inserts were selected for sequencing. The samples were multiplexed with 8 nt indexes in the 3' linker.
5' RNA ligation and 3' RNA ligation

treatment_protocol: For cloning 22G-RNA, the sample is treated with 0.5 μM PIR-1 at 20 °C for 2 hours in the ligation condition

tag: developmental stage

value: young adult worms

tag: genotype/variation


title: SF3B1_2


title: CO60R2

description: Whole fruits of accession Piquin Coahuila at 60 Days After Anthesis. Biological replicate R2.

value: 60 Days After Anthesis (DAA)


title: J_S5

value: Regulatory T cells isolated from the spleen


title: J_S2


title: tbc-11(ok2567)rep3

description: Small RNA analysis


title: tbc-11(ok2567)rep2

description: Small RNA analysis


title: CQ00R1

description: Whole fruits of accession F1 of cross CM x QU at 0 Days After Anthesis. Biological replicate R1.


title: J_S7

value: WT


title: tbc-11(qbc24)rep1

description: Small RNA analysis

source_name: Cloning tbc-11(qbc24) small RNA


title: SF3B1_1

value: diagnosis


title: CO50R1

description: Whole fruits of accession Piquin Coahuila at 50 Days After Anthesis. Biological replicate R1.


title: tbc-11(qbc24)rep2

description: Small RNA analysis


title: CO30R2

description: Whole fruits of accession Piquin Coahuila at 30 Days After Anthesis. Biological replicate R2.

value: 30 Days After Anthesis (DAA)


title: CO40R2

description: Whole fruits of accession Piquin Coahuila at 40 Days After Anthesis. Biological replicate R2.

value: 40 Days After Anthesis (DAA)


title: CO60R1

description: Whole fruits of accession Piquin Coahuila at 60 Days After Anthesis. Biological replicate R1.


title: NPM1_8


title: EKLF/GFP- 2

city: New York

first: Kaustav

last: Mukherjee

state: NY

address: 1 Gustave Levy Place

department: Cell, Developmental and Regenerative Biology

institute: Icahn School of Medicine at Mount Sinai

description: polyA mRNA

data_processing:
Basecalling and demultiplexing: standard Illumina software to generate fastq files
Alignments (Samples 1-6): STAR --runThreadN 16 --outFilterMultimapNmax 20 --alignSJoverhangMin 8 --alignSJDBoverhangMin 1 --outFilterMismatchNmax 999 --alignIntronMin 20 --alignIntronMax 500000 --alignMatesGapMax 500000 -- --outSAMstrandField intronMotif --outFilterIntronMotifs RemoveNoncanonical --outSAMattributes All --outSAMtype BAM SortedByCoordinate
Counts (Samples 1-6): htseq-count -f bam -r pos -s yes
Mapping (Samples 7-12): Salmon --libType SR --threads 8 --fldMean 250 --fldSD 50 –validateMappings
Count normalization and DEGs: DESeq2
Genome_build: STAR: mm10. Salmon: Ensembl Grcm38.v79
Mapping (Sample 13): Alevin –chromiumV3 -p 12 –dumpMtx
Cell Filtering, Clustering, Differential expression: Seurat R package
Genome_build: Gencode GRCm38.vM24
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample ...
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample
Supplementary_files_format_and_content: Text file with barcodes and genes for Sample 14

molecule: polyA RNA

source_name: embryonic fetal liver

extract_protocol:
Trizol
At Embryonic Day E13.5 fetal livers were dissected out of mouse embryos and homogenized by gentle pipetting in 1X PBS + 2% FBS. Cells were strained through a 70μm nylon cell strainer (Thermo Fisher) into 1X PBS + 2% FBS. After staining for appropriate antibodies, cells were sorted by FACS and collected in Trizol solution immediately for RNA-Seq, or purified by EZ-Sep (Cell Signaling technologies #17656) and processed for single cell sequencing using the Chromium V3 single cell kit (10X Genomics). RNA was extracted using ethanol precipitation, except for samples 7, 8, and 9 where we used the Agilent Absolutely RNA Nanoprep kit (#400753)
Libraries were constructed using Illumina Truseq stranded RNA kit
Single-end second strand RNA-Seq
Paired-end single cell sequencing

value: wild type (pEKLF-GFP)


title: J_S1

value: KO


title: NPM1_9


title: CO20R2

description: Whole fruits of accession Piquin Coahuila at 20 Days After Anthesis. Biological replicate R2.

value: 20 Days After Anthesis (DAA)


title: NPM1_7


title: NPM1_8_rl

value: relapse


title: CO40R1

description: Whole fruits of accession Piquin Coahuila at 40 Days After Anthesis. Biological replicate R1.


title: NPM1_6_rl


title: N2rep3

description: Small RNA analysis

source_name: Cloning N2 small RNA


title: tbc-11(qbc24)rep3

description: Small RNA analysis


title: KB_DC_3

city: Urbana

last: Zhao

state: IL

address: 1201 W. Gregory Dr.

department: Crop Sciences

institute: University of Illinois

data_processing:
Fastq files were generated and demultiplexed with with the bcl2fastq v1.8.4 Conversion Software (Illumina)
Sequenced reads were trimmed for adaptor sequence, then mapped to PstDC3000 whole genome using bowtie2 v2.3.2
The number of reads mapped to CDS of PstDC3000 were counted using samtools and bedtools
Normalization and statistical analysis were carried out using the R package DESeq2 as described in Love et al. Genome biology, 2014.
Genome_build: bowtieNC_013961
Supplementary_files_format_and_content: tab-delimited text files include read counts for each sample

organism: Pseudomonas syringae

source_name: bacterial cells

growth_protocol: Overnight cultures of wild-type and mutant strains grown in KB were harvested and part of the cells were washed twice with hrp-inducing minimal media.

extract_protocol:
Bacterial cells were harvested, and RNA was isolated using Qiagen RNeasy bacteria Mini kit
The stranded RNAseq libraries were prepared with Illumina's 'TruSeq Stranded RNA Sample Prep kit' (illumina)

treatment_protocol: Bacterial cells were adjusted to 2 x 10^8 cfu/ml in 5 mL of hrp-inducing minimal media or KB medium; and incubated for 6 h at 18 °C or in KB at 28 °C, respectively

value: wild-type

tag: media

value: KB medium


title: EKLF/GFP- 3

description: polyA mRNA


title: N2rep2

description: Small RNA analysis


title: CO30R1

description: Whole fruits of accession Piquin Coahuila at 30 Days After Anthesis. Biological replicate R1.

value: Whole fruit


title: EKLF/GFP+ 1

description: polyA mRNA

extract_protocol:
Agilent Absolutely RNA Nanoprep kit (#400753)
At Embryonic Day E13.5 fetal livers were dissected out of mouse embryos and homogenized by gentle pipetting in 1X PBS + 2% FBS. Cells were strained through a 70μm nylon cell strainer (Thermo Fisher) into 1X PBS + 2% FBS. After staining for appropriate antibodies, cells were sorted by FACS and collected in Trizol solution immediately for RNA-Seq, or purified by EZ-Sep (Cell Signaling technologies #17656) and processed for single cell sequencing using the Chromium V3 single cell kit (10X Genomics). RNA was extracted using ethanol precipitation, except for samples 7, 8, and 9 where we used the Agilent Absolutely RNA Nanoprep kit (#400753)
Libraries were constructed using Illumina Truseq stranded RNA kit
Single-end second strand RNA-Seq
Paired-end single cell sequencing


title: NPM1_7_rl


title: N2rep1

description: Small RNA analysis


title: CO00R2

description: Whole fruits of accession Piquin Coahuila at 0 Days After Anthesis. Biological replicate R2.

value: 0 Days After Anthesis (DAA)


title: EKLF/GFP- 1

description: polyA mRNA


title: CO20R1

description: Whole fruits of accession Piquin Coahuila at 20 Days After Anthesis. Biological replicate R1.

value: Wild accession (C. a. var. glabriusculum)


title: KB_A3_3

value: rsmA3 mutant


title: EKLF/GFP+ 3

description: polyA mRNA


title: WT F4/80+

description: total mRNA

extract_protocol:
Chromium single cell 3’ reagent kit V3 (10X Genomics)
At Embryonic Day E13.5 fetal livers were dissected out of mouse embryos and homogenized by gentle pipetting in 1X PBS + 2% FBS. Cells were strained through a 70μm nylon cell strainer (Thermo Fisher) into 1X PBS + 2% FBS. After staining for appropriate antibodies, cells were sorted by FACS and collected in Trizol solution immediately for RNA-Seq, or purified by EZ-Sep (Cell Signaling technologies #17656) and processed for single cell sequencing using the Chromium V3 single cell kit (10X Genomics). RNA was extracted using ethanol precipitation, except for samples 7, 8, and 9 where we used the Agilent Absolutely RNA Nanoprep kit (#400753)
Libraries were constructed using Illumina Truseq stranded RNA kit
Single-end second strand RNA-Seq
Paired-end single cell sequencing

value: wild type


title: CO00R1

description: Whole fruits of accession Piquin Coahuila at 0 Days After Anthesis. Biological replicate R1.


title: KB_DC_2


title: CO10R2

description: Whole fruits of accession Piquin Coahuila at 10 Days After Anthesis. Biological replicate R2.

value: 10 Days After Anthesis (DAA)


title: EKLF/GFP+ 2

description: polyA mRNA


title: NPM1_6


title: KB_DC_1


title: CO10R1

description: Whole fruits of accession Piquin Coahuila at 10 Days After Anthesis. Biological replicate R1.


title: NPM1_5


title: KB_A23_2

value: rsmA2/rsmA3 double mutant


title: KB_A23_1


title: NPM1_20_rl


title: NPM1_4


title: HM_DC_3

value: HMM medium


title: keratinocytes monolayer 2 - KC34wod8

city: Vienna

first: Julia

last: Lachner

address: Waehringer Guertel 18-20

country: Austria

institute: Medical University of Vienna

data_processing:
fastq generation, bcl2fastq v2.19.1.403
alignment and extraction of counts/gene, STAR aligner v2.6.1a_08-27, command: STAR --runThreadN 16 --genomeDir ggal5/RSEM_index --readFilesCommand zcat --readFilesIn $FASTQ1 $FASTQ2 --outSAMattributes NH HI NM MD AS nM jM jI XS --outSAMtype BAM SortedByCoordinate --outSAMattrRGline ID:"$sample" SM:"$sample" CN:MUW_CF PG:STAR –quantMode TranscriptomeSAM GeneCounts --twopassMode Basic
count normalization and differential gene expression analysis, DESeq2 v1.22.2
Genome_build: ggal5, ensembl 91 annotations plus custom annotations described in this paper
Supplementary_files_format_and_content: plain text, raw counts per gene, one file per sample

source_name: confluent keratinocytes 2

growth_protocol: Primary chicken fibroblasts and keratinocytes were grown in monolayer culture or in 3-dimensional skin equivalents

extract_protocol:
RNA was isolated with Trizol reagent
Sequencing libraries were be prepared using the NEBNext Poly(A) mRNA Magnetic Isolation Module and the NEBNext Ultra™ II Directional RNA Library Prep Kit for Illumina according to the manufacturer's protocols (New England Biolabs). Libraries were QC-checked on a Bioanalyzer 2100 (Agilent) using a High Sensitivity  DNA Kit for correct insert size and quantified using Qubit dsDNA HS Assay (Invitrogen). Pooled libraries were  sequenced on a NextSeq500 instrument (Illumina) in 2x75bp paired-end sequencing mode.

value: primary keratinocytes

tag: diff. stage

value: confluent


title: skin equivalent 3 - SE56wo

source_name: skin equivalent 3

value: 3D differentiation in skin equivalent


title: skin equivalent 2 - SE34wo

source_name: skin equivalent 2


title: NPM1_3_rl


title: KB_A3_2

tag: strain


title: keratinocytes subconfluent - KC20prol

source_name: subconfluent keratinocytes

value: subconfluent


title: KB_A3_1


title: skin equivalent 1 - SE20d7

source_name: skin equivalent 1


title: HM_DC_2


title: CM60R2

description: Whole fruits of accession Criollo de Morelos (CM334) at 60 Days After Anthesis. Biological replicate R2.

source_name: Criollo de Morelos (CM334)

value: Domesticated variety

tag: tissue

value: 60 Days After Anthesis (DAA)


title: NPM1_3

source_name: AML leukemic blasts

value: diagnosis


title: NPM1_1


title: KB_A23_3

tag: genotype


title: NPM1_2


title: CM60R1

description: Whole fruits of accession Criollo de Morelos (CM334) at 60 Days After Anthesis. Biological replicate R1.


title: CM30R2

description: Whole fruits of accession Criollo de Morelos (CM334) at 30 Days After Anthesis. Biological replicate R2.

value: 30 Days After Anthesis (DAA)


title: keratinocytes monolayer 3 - KC56wod8

source_name: confluent keratinocytes 3


title: CM50R2

description: Whole fruits of accession Criollo de Morelos (CM334) at 50 Days After Anthesis. Biological replicate R2.

value: 50 Days After Anthesis (DAA)


title: CM30R1

description: Whole fruits of accession Criollo de Morelos (CM334) at 30 Days After Anthesis. Biological replicate R1.

value: Whole fruit


title: CM50R1

description: Whole fruits of accession Criollo de Morelos (CM334) at 50 Days After Anthesis. Biological replicate R1.


title: keratinocytes monolayer 1 - KC20d8

source_name: confluent keratinocytes 1


title: NPM1_20


title: NPM1_17


title: NPM1_16


title: BW25113 slyA Mutant_ECMS2_5

city: dover

first: Derrick

last: Scott

state: Delaware

address: 1200 n dupont hwy

institute: Delaware State University

data_processing:
FastQC used to examine quality of raw reads
Trimmomatic was used to trim the Illumina specific adaptors and primers
STAR Aligner was used to index and align the reads to the BW25113 reference genome
HTSeq was used to count the numbers of times that the gene reads aligned to the reference
DESeq2 was used to compare the difference in the number of times the genes showed up in each sample
Genome_build: Genbank Reference sequence CP009273.1 was used in the indexing and alignment step
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample
processed data files contains data for 121 genes and contains the following:
__alignment_not_unique  1808510

organism: Escherichia coli

source_name: E coli BW25113 slyA Mutant

growth_protocol: Each strain of E coli BW25113 and  E coli BW25113 slyA Mutant were grown to log phase (16hrs) overnight in Nutrient broth at 37 degree celcius.

extract_protocol:
Total RNA was extracted using the RNeasy minikit (Qiagen), and residual DNA was digested using the RNase-free DNase kit (Qiagen).
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: no treatment

value: slyA Mutant

value: overnight log phase growth


title: BW25113 slyA Mutant_ECMS2_7


title: CM10R2

description: Whole fruits of accession Criollo de Morelos (CM334) at 10 Days After Anthesis. Biological replicate R2.


title: NPM1_1_rl

value: relapse


title: BW25113 slyA Mutant_ECMS2_3


title: CM10R1

description: Whole fruits of accession Criollo de Morelos (CM334) at 10 Days After Anthesis. Biological replicate R1.


title: CM40R1

description: Whole fruits of accession Criollo de Morelos (CM334) at 40 Days After Anthesis. Biological replicate R1.

value: 40 Days After Anthesis (DAA)


title: HM_DC_1

value: wild-type


title: NPM1_18


title: fibroblasts - FB

source_name: fibroblasts

value: primary fibroblasts


title: HM_A3_2

value: rsmA3 mutant

value: HMM medium


title: CM00R2

description: Whole fruits of accession Criollo de Morelos (CM334) at 0 Days After Anthesis. Biological replicate R2.

value: 0 Days After Anthesis (DAA)


title: BW25113 slyA Mutant_ECMS2_8


title: NPM1_18_rl


title: BW25113 slyA Mutant_ECMS2_2


title: NPM1_13_rl


title: CM40R2

description: Whole fruits of accession Criollo de Morelos (CM334) at 40 Days After Anthesis. Biological replicate R2.


title: HM_A3_1


title: HM_A3_3


title: CM20R2

description: Whole fruits of accession Criollo de Morelos (CM334) at 20 Days After Anthesis. Biological replicate R2.

value: 20 Days After Anthesis (DAA)


title: HM_A23_3

value: rsmA2/rsmA3 double mutant


title: NPM1_13


title: CM20R1

description: Whole fruits of accession Criollo de Morelos (CM334) at 20 Days After Anthesis. Biological replicate R1.

value: Domesticated variety


title: BW25113 Wild Type_ECWT1_7

source_name: E coli BW25113 Wild Type

value: Wild Type


title: NPM1_12_rl


title: NPM1_12

value: diagnosis


title: BW25113 slyA Mutant_ECMS2_4


title: NPM1_15_rl


title: NPM1_15


title: HM_A23_2


title: BW25113 Wild Type_ECWT1_5


title: NPM1_11


title: BW25113 Wild Type_ECWT1_3


title: BW25113 Wild Type_ECWT1_2


title: NPM1_10


title: BW25113 slyA Mutant_ECMS2_6


title: HM_A23_1


title: SZC RNA Seq  Replicate1

city: Boston

first: Ramesh

last: Shivdasani

state: MA

address: 450 Brookline Ave. Dana 720D

department: Medical Oncology

institute: Dana Farber Cancer Institute

description: SZC RNA Seq  Replicate1

data_processing:
ATAC-seq:bowtie2 version 2.3.4.3. For 75bp paired-end data, RNA-seq: STAR aligner version 2.5.2b
ATAC-seq peak calling: MACS 1.4
RNA-Seq - Read counts were normalized using Trimmed Means of M-values (TMM) in edgeR package. Differntial expression was determined in edgeR using Exact Test (q<0.05).
Genome_build: UMD 3.1
Supplementary_files_format_and_content: ATAC-Seq: bigwig file for visualizing aligned sequence tags
Supplementary_files_format_and_content: RNA-Seq: Table with raw read counts for all cell types

organism: Bos taurus

source_name: bovine superficial zone articular chondrocytes (SZC)

extract_protocol:
Newly isolated bovine superficial zone chondrocytes and deep zone chondrocytes were cultured (in DMEM/F12 plus 10% FBS) for three days prior to performing RNA-Seq analysis. RNA from superficial zone chondrocytes and deep zone chondrocytes were purified using Trizol reagent (Life Technologies).
Total RNA was extracted with TRIzol reagent (Life Technologies) followed by DNase treatment with the TURBO DNA-free™ Kit (Thermo Fisher Scientific, Cat#: AM1907)
SMART-Seq v4 Ultra Low Input RNA Kit (Clontech) was used according to the manufacturer's instructions and 75 bp single-end reads were sequenced on an Illumina NextSeq 500 instrument.

value: Superficial zone articular chondrocytes from knee joints from 1-2 week old bovine calves


title: CM00R1

description: Whole fruits of accession Criollo de Morelos (CM334) at 0 Days After Anthesis. Biological replicate R1.


title: mock_gamma_ChIPSeq_Uninduced

status: Public on Mar 15 2021

city: Pavia

first: Sofia

last: Francia

address: Via Abbiategrasso, 207

country: Italy

institute: Istituto di Genetica Molecolare - Consiglio Nazionale delle Ricerche (IGM-CNR)

data_processing:
quality assesment: FastQC (http://www.bioinformatics.babraham.ac.uk/ projects/fastqc/).
aligment: The samples passing the literature quality standards were aligned on the human genome (GRCh37/hg19) using BWA (Li and Durbin, 2009) using default parameters. In order to mantain the collinearity between the read signal and the protein occupancy on the genome, multiple-matching reads were eliminated.
Peak calling: Peak calling was performed with MACS 1.4.2 (Zhang et al., 2008). The output was intersected with the AsiSI sites database using BEDtools (Quinlan and Hall, 2010). Quantitative analysis of induced (+4OHT) and uninduced (-4OHT) dataset was carried out via PscanChIP (Zambelli et al., 2013) using default parameters, for producing a final list of AsiSI sites ranked for the imbalance of γH2AX signal via χ2 test.
Data visualization: Data visualization was obtained with ngs.plot package (Shen et al., 2014).
BigWig files were created from bam files using bamCoverage from Deeptools suite (Ramírez et al., 2016)
Genome_build: GRCh37/hg19
Supplementary_files_format_and_content: Coverage track (bigWig format) of aligned reads.

source_name: DiVA (AsiSI-ER-U2OS) cells

growth_protocol: DIvA (AsiSI-ER-U20S) cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM, Gibco) without phenol red, supplemented with glutamine, pyruvate, HEPES and 10% fetal bovine serum (FBS, Euroclone). Cells were grown at 37°C under a humidified atmosphere with 5% CO2 and selected on a semi regular basis with puromycin at a final concentration of 1 μg/ml.

extract_protocol:
Lysates were clarified from sonicated nuclei and protein-DNA complexes were isolated with specific antibody against gH2AX (Abcam 2893) or DROSHA (Cell Signaling 3364).
Libraries were prepared using standard Illumina TruSeq for HiSeq 2000 reagents

treatment_protocol: For AsiSI-dependent DSB induction, DiVA cells were treated with 300 nM 4-Hydroxytamoxifen (4OHT, Sigma; H7904) for 4h.

value: DiVA (AsiSI-ER-U2OS) cells

value: U2OS-derived cells

value: Uninduced (-4OHT)

tag: chip antibody

value: No antibody


title: mock_gamma_ChIPSeq_Induced

value: Induced (+4OHT)


title: Bone marrow_ALL_rep2

city: Nanjing

first: Ting

middle: Yu

last: Zhu

address: No. 72 Guangzhou Road

institute: Children's Hospital of Nanjing Medical University

description: Gene expression  of bone marrow from ALL cases

hyb_protocol: DNA in hybridization solution was denatured at 95℃ for 3 min prior to loading onto a microarray. Arrays were hybridized was preformed in a Agilent Hybridization Oven overnight at a rotation speed of 20 rpm at 42℃ and washed with two consecutive solutions (0.2% SDS, 2× SSC at 42℃ for 5 min, and 0.2× SSC for 5 min at room temperature).

scan_protocol: Scanned on an Agilent G2565CA scanner.

data_processing: Feature Extraction v10.7 (Agilent Technologies, CA) software was used to extract all features of the data obtained from the scanned images and genespring software was used to analyze the raw data, which are normalized by percentile normalization.

source_name: Bone marrow, ALL, rep2

label_protocol: cDNA labeled with a fluorescent dye (Cy3-dCTP) was produced by Eberwine’s linear RNA amplification method and subsequent enzymatic reaction. This procedure has been previously described, and the procedure has been improved by using CapitalBio cRNA Amplification and Labeling Kit (CapitalBio) for producing higher yields of labeled cDNA.

extract_protocol: RNA was prepared using the TRIzol reagent following the manufacturer's recommendations.  RNA was quantified using a NanoDrop-1000 spectrophotometer.

treatment_protocol: Bone marrow RNA was extracted from samples of ALL children and healthy controls.

tag: diagnosis

value: Acute lymphoblastic leukemia (ALL)

value: Bone marrow tissue

tag: gender

value: male


title: BW25113 slyA Mutant_ECMS2_1

value: slyA Mutant

value: overnight log phase growth


title: Bone marrow_non-ALL_rep3

description: Gene expression  of bone marrow from healthy control

source_name: Bone marrow, non-ALL, rep3

value: healthy

value: female


title: Input_gamma_ChIPSeq_Uninduced

tag: cell type


title: DZC ATAC Seq  Replicate1

description: DZC ATAC Seq  Replicate1

source_name: bovine deep zone articular chondrocytes (DZC)

extract_protocol:
Newly isolated bovine superficial zone articular chondrocytes and deep zone articular chondrocytes were cultured (in DMEM/F12 plus 10% FBS) for three days prior to performing ATAC-Seq analysis. ATAC-Seq (Buenrostro et al., 2013; Buenrostro et al., 2015) was performed on replicate samples of 8,000 to 35, 000 superficial zone chondrocytes or deep zone chondrocytes.
Nuclear pellets were isolated and treated with Nextera Tn5 Transposase.Transposed DNA was immediately isolated using a Qiagen MinElute PCR Purification Kit, and then PCR amplified in a 50 μl reaction using a common forward primer and different reverse primers with unique barcodes for each sample as per (Buenrostro et al., 2015).
ThruPLEX DNA-seq Kit (Rubicon Genomics) was used to prepare libraries, and 75 bp single-end reads were sequenced on an Illumina NextSeq 500 instrument.

value: Deep zone articular chondrocytes from knee joints from 1-2 week old bovine calves


title: BW25113 Wild Type_ECWT1_8


title: SZC ATAC Seq  Replicate2

description: SZC ATAC Seq  Replicate2


title: Input_gamma_ChIPSeq_Induced

tag: cell line


title: gamma_ChIPSeq_Induced

tag: treatment

value: gH2AX antibody (Abcam 2893, Rabbit polyclonal)


title: NPM1_14


title: BW25113 Wild Type_ECWT1_6

value: Wild Type


title: mock_DROSHA_ChIPSeq_Uninduced


title: SZC ATAC Seq  Replicate1

description: SZC ATAC Seq  Replicate1


title: DZC ATAC Seq  Replicate2

description: DZC ATAC Seq  Replicate2


title: NPM1_11_rl

value: relapse


title: DZC RNA Seq  Replicate2

description: DZC RNA Seq  Replicate2


title: DROSHA_ChIPSeq_Induced

value: DROSHA antibody (Cell Signaling 3364, Rabbit monclonal)


title: DROSHA_ChIPSeq_Uninduced

value: DiVA (AsiSI-ER-U2OS) cells


title: Sample_75 [Cat_HPC-005]

city: Seoul

first: Jin-Wu

last: Nam

address: Seongdong-Gu Hangdang-dong

department: Department of Life Science

country: South Korea

institute: Hanyang University

data_processing:
Illumina Casava1.7 software was used for base-calling
Low quality reads were detected using  FastQC and trimmed using Sickle
Filtered reads were aligned to hg19 using STAR aligner
Gene expression levels were quantified using featureCounts
Genome_build: hg19
Supplementary_files_format_and_content: read counts; FPKM

source_name: Gastric cancer, liver

extract_protocol:
Liver tissues were removed, flash frozen on dry ice, and RNA was harvested using Trizol reagent. Illumina TruSeq RNA Sample Prep Kit (Cat#FC-122-1001) was used with 1 ug of total RNA for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: patient id

tag: patient age

tag: patient gender

value: M

tag: hepatectomy

value: Gastric cancer, liver

tag: tumor stage

value: combined HCC and intrahepatic cholangiocarcinoma (ICC)

tag: paired nontumor


title: Sample_76 [Cat_HPC-006]


title: Bone marrow_ALL_rep3

source_name: Bone marrow, ALL, rep3

tag: age


title: BW25113 Wild Type_ECWT1_4


title: Sample_72 [Cat_HPC-002]


title: Sample_70 [Cat_TG3-024]


title: Sample_61 [Cat_TG3-012]


title: Sample_67 [Cat_TG3-021]

value: F


title: DZC RNA Seq  Replicate1

description: DZC RNA Seq  Replicate1


title: Sample_73 [Cat_HPC-003]


title: NPM1_10_rl

extract_protocol:
QIAGEN AllPrep
TruSeq Stranded Total RNA Ribominus Gold
100 bp paired-end sequencing on HiSeq2000

tag: mutation


title: Bone marrow_ALL_rep1

source_name: Bone marrow, ALL, rep1

value: Acute lymphoblastic leukemia (ALL)


title: Sample_51 [Cat_TG3-001]


title: SZC RNA Seq  Replicate2

description: SZC RNA Seq  Replicate2

value: Superficial zone articular chondrocytes from knee joints from 1-2 week old bovine calves


title: BW25113 Wild Type_ECWT1_1

tag: strain


title: Sample_64 [Cat_TG3-016]

tag: tissue


title: Sample_56 [Cat_TG3-006]


title: Input_DROSHA_ChIPSeq_Uninduced

value: U2OS-derived cells

value: Uninduced (-4OHT)

value: No antibody


title: Sample_58 [Cat_TG3-008]


title: Sample_54 [Cat_TG3-004]


title: Sample_74 [Cat_HPC-004]


title: Bone marrow_non-ALL_rep2

source_name: Bone marrow, non-ALL, rep2

value: healthy

value: Bone marrow tissue

value: male


title: Bone marrow_non-ALL_rep1

source_name: Bone marrow, non-ALL, rep1


title: gamma_ChIPSeq_Uninduced

value: gH2AX antibody (Abcam 2893, Rabbit polyclonal)


title: Sample_71 [Cat_HPC-001]


title: Sample_47 [Cat_TG2-035]


title: Sample_53 [Cat_TG3-003]


title: mock_DROSHA_ChIPSeq_Induced

value: Induced (+4OHT)


title: Sample_69 [Cat_TG3-023]


title: Sample_60 [Cat_TG3-010]


title: Sample_55 [Cat_TG3-005]


title: Sample_52 [Cat_TG3-002]


title: Input_DROSHA_ChIPSeq_Induced


title: Sample_43 [Cat_TG2-030]


title: Sample_50 [Cat_TG2-038]


title: Sample_41 [Cat_TG2-028]


title: Sample_40 [Cat_TG2-026]


title: Sample_88 [Cat_HPC-003r]


title: Sample_66 [Cat_TG3-020]


title: Sample_39 [Cat_TG2-025]


title: Sample_68 [Cat_TG3-022]


title: Sample_35 [Cat_TG2-014]


title: Sample_65 [Cat_TG3-019]


title: Sample_32 [Cat_TG2-010]


title: Sample_29 [Cat_TG2-005]


title: Sample_63 [Cat_TG3-015]


title: Sample_28 [Cat_TG2-004]


title: Sample_59 [Cat_TG3-009]


title: Sample_57 [Cat_TG3-007]


title: Sample_27 [Cat_TG2-002]


title: Sample_49 [Cat_TG2-037]


title: Sample_48 [Cat_TG2-036]


title: Sample_62 [Cat_TG3-013]


title: Sample_23 [Cat_TG1-021]


title: Sample_25 [Cat_TG1-025]


title: Sample_87 [Cat_TG2-035r]


title: Sample_84 [Cat_TG1-017]


title: Sample_20 [Cat_TG1-009]


title: Sample_42 [Cat_TG2-029]


title: Sample_44 [Cat_TG2-032]


title: Sample_46 [Cat_TG2-034]


title: Sample_45 [Cat_TG2-033]


title: Sample_37 [Cat_TG2-022]


title: Sample_38 [Cat_TG2-023]


title: Sample_16 [Cat_TG1-003]


title: Sample_36 [Cat_TG2-019]


title: Sample_80 [Cat_DH009]

source_name: Dysplastic nodule-high tissue, liver

value: Dysplastic nodule-high tissue, liver

value: Dysplastic nodule high grade


title: Sample_79 [Cat_DH008]


title: Sample_34 [Cat_TG2-013]


title: Sample_33 [Cat_TG2-012]


title: Sample_TN1507L0593 [Cat_DL026]

source_name: Dysplastic nodule-low tissue, liver

value: Dysplastic nodule-low tissue, liver

value: Dysplastic nodule low grade


title: Sample_31 [Cat_TG2-008]

value: M

value: Gastric cancer, liver


title: Sample_30 [Cat_TG2-006]


title: Sample_26 [Cat_TG2-001]


title: Sample_TN1507L0592 [Cat_DL025]


title: Sample_TN1507L0590 [Cat_DL023]


title: Sample_18 [Cat_TG1-005]


title: Sample_22 [Cat_TG1-013]

value: F


title: Sample_86 [Cat_TG1-017r]


title: Sample_21 [Cat_TG1-012]


title: Sample_19 [Cat_TG1-007]


title: Sample_83 [Cat_DH013]


title: Sample_82 [Cat_DH011]


title: Sample_TN1507L0587 [Cat_DL019]


title: Sample_TN1507L0585 [Cat_DL011]


title: Sample_TN1507L0584 [Cat_DL002]


title: Sample_TN1507L0579 [Cat_CS022]

source_name: Cirrhosis tissue, liver

value: Cirrhosis tissue, liver

value: Cirrhosis


title: Sample_TN1507L0586 [Cat_DL016]


title: Sample_77 [Cat_DH003]


title: Sample_TN1507L0582 [Cat_CS038]


title: Sample_17 [Cat_TG1-004]


title: Sample_TN1507L0575 [Cat_CS010]


title: Sample_TN1507L0574 [Cat_CS002]


title: Sample_24 [Cat_TG1-023]


title: Sample_TN1507L0589 [Cat_DL021]


title: Sample_TN1507L0583 [Cat_CS039]


title: Sample_TN1507L0591 [Cat_DL024]


title: Sample_78 [Cat_DH004]


title: Sample_TN1507L0580 [Cat_CS028]


title: Sample_TN1507L0581 [Cat_CS029]


title: Sample_81 [Cat_DH010]


title: Sample_TN1507L0588 [Cat_DL020]


title: Sample_TN1507L0578 [Cat_CS019]


title: Sample_TN1507L0577 [Cat_CS017]


title: Sample_TN1507L0569 [Cat_FH010]

source_name: Fibrosis-high tissue, liver

value: Fibrosis-high tissue, liver

value: Fibrosis high grade


title: Sample_TN1507L0576 [Cat_CS016]


title: Sample_TN1507L0565 [Cat_FH005]


title: Sample_TN1507L0562 [Cat_FL013]

source_name: Fibrosis-low tissue, liver

value: Fibrosis-low tissue, liver

value: Fibrosis low grade


title: Sample_TN1507L0572 [Cat_FH017]


title: Sample_TN1507L0570 [Cat_FH014]


title: Sample_TN1507L0568 [Cat_FH009]


title: Sample_TN1507L0571 [Cat_FH016]


title: Sample_TN1507L0563 [Cat_FL014]


title: Sample_TN1507L0558 [Cat_FL007]


title: Sample_TN1507L0567 [Cat_FH008]


title: Sample_TN1507L0561 [Cat_FL012]


title: Sample_TN1507L0564 [Cat_FH002]


title: Sample_TN1507L0559 [Cat_FL008]


title: Sample_TN1507L0557 [Cat_FL004]


title: Sample_TN1507L0573 [Cat_FH018]


title: Sample_TN1507L0556 [Cat_FL003]


title: Sample_TN1507L0554 [Cat_FL001]


title: Sample_15 [Cat_N018]

source_name: Normal tissue, liver

value: Normal tissue, liver


title: Sample_12 [Cat_N015]


title: Sample_13 [Cat_N016]


title: Sample_11 [Cat_N014]


title: Sample_14 [Cat_N017]


title: Sample_85 [Cat_N013r]


title: Sample_TN1507L0555 [Cat_FL002]


title: Sample_TN1507L0566 [Cat_FH007]


title: Sample_10 [Cat_N013]


title: Sample_TN1507L0560 [Cat_FL010]


title: Sample_9 [Cat_N012]


title: Sample_6 [Cat_N006]


title: Sample_7 [Cat_N007]


title: Sample_5 [Cat_N005]


title: Sample_4 [Cat_N004]


title: Sample_3 [Cat_N003]


title: Sample_8 [Cat_N011]


title: Sample_1 [Cat_N001]


title: GABAergic CB1-KO - open field - rep 5 [2015_01_48_dlx_ko_OF_659]

city: Mainz

first: Anna

last: Wierczeiko

address: Hanns-Dieter-Hüsch-Weg 15

department: Institute for Human Genetics

institute: University Medical Center, Johannes Gutenberg University Mainz

data_processing:
Quality check of the raw sequencing data was done via FastQC version (v0.11.8).
Bbduk.sh from the BBMap suite of tools (version 38.06) was employed to perform adapter trimming and quality filtering of the raw sequence reads (q30 cutoff).
The trimmed and cleaned sequences were mapped against the mouse reference genome mm10 from UCSC (downloaded via Illumina iGenome: http://support.illumina.com/sequencing/sequencing_software/igenome.html, download date 06/06/18), using the STAR aligner (version 2.6.0a) with default options. Summarized quality results were created by MultiQC (version 1.8.dev0).
Counting of reads per gene was done via featureCounts (version 1.6.2), with the -s 2 option, using the annotation file for the mm10 mouse genome from iGenomes.
Genome_build: mm10
Supplementary_files_format_and_content: raw gene counts for every gene and every sample

source_name: GABAergic CB1-KO – open field

growth_protocol: Conditional CB1-KO mice were generated via crossing CB1f/f mice with animals expressing Cre recombinase under the control of the regulatory elements of the genes dlx5/6 (for deletion in GABAergic terminals; MGI Cat# 3758334, RRID:MGI:) and nex1 (for deletion in glutamatergic terminals; 3758334MGI Cat# 3758333, RRID:MGI:3758333). This use of the Cre/LoxP system allowed for high specificity in the genetic deletion as shown previously (Monory et al., 2006). Genotyping was performed as previously described (Massa et al., 2010). Male mice had food and water ad libitum during their 12-hour light-dark cycle (07:00-19:00) and were 8-10 weeks old when they were sacrificed.

extract_protocol:
In order to microdissect only the Cornus Ammonis from the hippocampus, we first removed the dentate gyrus (DG) from the rest of the hippocampus according to the protocol of Hideo Hagihara (Hagihara et al., 2009). The remaining hippocampal tissue was then dissected out in one-piece and snap-frozen on dry ice. In order to sustain the amount and quality of the hippocampal RNA for sequencing, the extraction and purification of RNA was performed as previously described (Lerner et al., 2018). To summarize, we followed the instructions from the RNeasy Mini kit (Qiagen) with a slight modification during the homogenization step. Frozen tissue was homogenized in 600μLRLT buffer with 1% β-mercaptoethanol (according to the manufacturer ́s protocol), though with the addition of 200 μL chloroform. Apart from this step, therewere no other changes made. Obtained RNA samples were treated with DNase I to degrade any possible contamination by genomic DNA. Samples were then eluted in 30 μL RNase-free water. The working bench and the tools used were cleaned before and during the extraction with RNase away plus (MßP, San 128Diego, CA, USA) to avoid any RNA degradation.
The hippocampal transcriptome sequencing occurred at the Core Facility Genomics (Institute of Molecular Biology gGmbH, IMB; Mainz). Next generation sequencing (NGS) library preparation was performed in two steps. First, cDNA was generated using NuGEN ́s Ovation RNA-seq system v2, from an input amount of 10 ng of total RNA, following the kit ́s instructions from the year 2012 (NuGEN, The Netherlands). Samples were amplified using the single primer isothermal amplification (SPIA) method from NuGEN. The resulting purified cDNA was quantified using the Qubit dsDNA HS assay kit in a Qubit 2.0 Fluorometer (Life 141Technologies, Germany). Afterward, the cDNA was profiled on a high sensitivity DNA chip using a 2100 Bioanalyzer (Agilent Technologies, Germany) as quality control. From the total cDNA, 1.5 μg were fragmented using a Covaris S2 focused-ultrasonicator (Covaris, UK), with the following parameters: 1) Duty cycle = 10%; 2) Intensity = 5; 3) Cycles/Burst = 200; 4) Time 145= 160 seconds; and 5) Water level = 15. After the fragmentation, the resulting material was once again quantified and profiled using a Qubit 2.0 and a 2100 Bioanalyzer, respectively, as described above. This process was repeated as an extra quality control point to ensure an optimal fragmentation. Secondly, NGS libraries were generated from 100 ng of fragmented cDNA usingNuGEN ́s Ovation ultralow system v2, following the kit ́s manual from the year 2014 (NuGEN, The 151Netherlands). Libraries were amplified in 7 PCR cycles and purified using beads. These purified libraries were quantified as described above, and profiled on a DNA 1000 Chip using a 2100 Bioanalyzer as the last quality control. Libraries representing all the experimental groups were pooled into individual pools containing 12 libraries, all of them in equimolar ratio. Each pool was loaded into four lanes of an Illumina's HiSeq flowcell and ran on a HiSeq 2500 in High-output mode (Illumina, USA), generating single-reads 50 base pairs long with an average yield of 52 million reads per library.

treatment_protocol: In order to study how the hippocampus of conditional CB1-KO mice react upon stimulation, we induced neuronal activity by exposing the animals to an open field arena (40x40x30 cm white box), a new environment that the mouse could explore undisturbed for 5 minutes (light intensity: y: 20-30 lux). This sufficient yet straightforward stimulus-induced neuronal activity throughout the hippocampus during arena exploration induced gene expression changes to be investigated.

value: dlx5/6-cre x CB1f/f

tag: genotype

value: 8 weeks

tag: Sex

value: male

value: open field exposure

value: Brain - hippocampal Cornus Ammonis

tag: treatment synonym

value: dlx-KO-OF


title: GABAergic CB1-KO - open field - rep 1 [2015_01_43_dlx_ko_OF_491]


title: GABAergic CB1-KO - open field - rep 3 [2015_01_45_dlx_ko_OF_533]


title: GABAergic CB1-KO - open field - rep 2 [2015_01_44_dlx_ko_OF_507]


title: GABAergic CB1-WT - open field - rep 7 [2015_01_46_dlx_wt_OF_525]

source_name: GABAergic CB1-WT – open field

value: dlx-WT-OF


title: Sample_2 [Cat_N002]

value: M


title: GABAergic CB1-KO - open field - rep 4 [2015_01_47_dlx_ko_OF_613]


title: GABAergic CB1-WT - open field - rep 6 [2015_01_42_dlx_wt_OF_663]


title: GABAergic CB1-WT - open field - rep 5 [2015_01_41_dlx_wt_OF_489]


title: GABAergic CB1-WT - open field - rep 3 [2015_01_39_dlx_wt_OF_603]


title: GABAergic CB1-KO - basal condition - rep 5 [2015_01_35_dlx_ko_ctr_605]

source_name: GABAergic CB1-KO – basal condition

value: basal condition (control)

value: dlx-KO-ctr


title: GABAergic CB1-WT - open field - rep 4 [2015_01_40_dlx_wt_OF_655]


title: CARS2 siRNA 2

city: Ottawa

first: Anh-Thu

last: Dang

state: ON

address: 40 Ruskin St

department: Biochemistry

country: Canada

institute: University of Ottawa Heart Institute

hyb_protocol: Samples were hybridized with HuGene-2_0-st arrays (Affymetrix) and scanned at the Centre for Applied Genomics (TCAG), Toronto.

scan_protocol: Array scanning was performed according to the manufacturer's instruction (Affymetrix).

data_processing: Relative expression changes assessed by Expression Console.

label: Biotin

source_name: THP-1 monocytic leukaemia cell line

label_protocol: Standard Affymetrix Protocol.

growth_protocol: 5.5 mM glucose RPMI supplemented with 10% serum.

extract_protocol: Total RNA was isolated using the High Pure RNA Tissue Kit (Sigma) following manufacturer's instructions.

treatment_protocol: THP-1 macrophages were transfected for 72 hours with CARS2 siRNA or NT2 control siRNA at a final concentration of 16.6 nM using Lipofectamine® RNAiMAX Reagent.

value: PMA differentiated THP-1 macrophages

tag: sirna

value: CARS2 siRNA


title: GABAergic CB1-WT - basal condition - rep 4 [2015_01_28_dlx_wt_ctr_657]

source_name: GABAergic CB1-WT – basal condition

value: dlx-WT-ctr


title: GABAergic CB1-WT - basal condition - rep 5 [2015_01_29_dlx_wt_ctr_503]


title: GABAergic CB1-KO - basal condition - rep 4 [2015_01_34_dlx_ko_ctr_615]


title: GABAergic CB1-WT - basal condition - rep 6 [2015_01_30_dlx_wt_ctr_671]


title: GABAergic CB1-KO - basal condition - rep 6 [2015_01_36_dlx_ko_ctr_665]


title: GABAergic CB1-WT - open field - rep 1 [2015_01_37_dlx_wt_OF_531]


title: GABAergic CB1-WT - open field - rep 2 [2015_01_38_dlx_wt_OF_515]


title: NT2 Control siRNA 1

tag: cell type

value: NT2 Control siRNA


title: CARS2 siRNA 1

tag: cell line


title: NT2 Control siRNA 3


title: NT2 Control siRNA 2


title: CARS2 siRNA 3


title: GABAergic CB1-KO - basal condition - rep 3 [2015_01_33_dlx_ko_ctr_611]

tag: treatment


title: GABAergic CB1-KO - basal condition - rep 2 [2015_01_32_dlx_ko_ctr_529]


title: GABAergic CB1-WT - basal condition - rep 3 [2015_01_27_dlx_wt_ctr_511]


title: GABAergic CB1-WT - basal condition - rep 1 [2015_01_25_dlx_wt_ctr_513]


title: glutamatergic CB1-KO - open field - rep 6 [2015_01_23_nex_ko_OF_893]

source_name: glutamatergic CB1-KO – open field

value: nex-cre x CB1f/f

value: nex-KO-OF


title: GABAergic CB1-KO - basal condition - rep 1 [2015_01_31_dlx_ko_ctr_509]


title: glutamatergic CB1-KO - open field - rep 7 [2015_01_24_nex_ko_OF_1167]


title: GABAergic CB1-WT - basal condition - rep 2 [2015_01_26_dlx_wt_ctr_535]


title: glutamatergic CB1-KO - open field - rep 4 [2015_01_21_nex_ko_OF_801]


title: glutamatergic CB1-KO - open field - rep 2 [2015_01_19_nex_ko_OF_757]


title: glutamatergic CB1-KO - open field - rep 3 [2015_01_20_nex_ko_OF_763]


title: glutamatergic CB1-WT - open field - rep 5 [2015_01_18_nex_wt_OF_747]

source_name: glutamatergic CB1-WT – open field

value: nex-WT-OF


title: glutamatergic CB1-WT - open field - rep 4 [2015_01_17_nex_wt_OF_1191]


title: glutamatergic CB1-WT - open field - rep 1 [2015_01_14_nex_wt_OF_751]


title: glutamatergic CB1-WT - open field - rep 3 [2015_01_16_nex_wt_OF_1147]


title: glutamatergic CB1-WT - open field - rep 2 [2015_01_15_nex_wt_OF_895]


title: glutamatergic CB1-KO - basal condition - rep 7 [2015_01_12_nex_ko_ctr_1181]

source_name: glutamatergic CB1-KO – basal condition

value: nex-KO-ctr


title: glutamatergic CB1-KO - open field - rep 1 [2015_01_13_nex_ko_OF_795]

value: 8 weeks


title: glutamatergic CB1-KO - basal condition - rep 5 [2015_01_10_nex_ko_ctr_891]

value: male


title: glutamatergic CB1-KO - basal condition - rep 4 [2015_01_09_nex_ko_ctr_887]


title: glutamatergic CB1-KO - basal condition - rep 3 [2015_01_08_nex_ko_ctr_793]


title: glutamatergic CB1-KO - basal condition - rep 6 [2015_01_11_nex_ko_ctr_1171]


title: glutamatergic CB1-KO - basal condition - rep 2 [2015_01_07_nex_ko_ctr_789]


title: glutamatergic CB1-WT - basal condition - rep 5 [2015_01_06_nex_wt_ctr_1177]

source_name: glutamatergic CB1-WT – basal condition

value: nex-WT-ctr


title: glutamatergic CB1-WT - basal condition - rep 4 [2015_01_05_nex_wt_ctr_1169]

value: Brain - hippocampal Cornus Ammonis


title: glutamatergic CB1-WT - basal condition - rep 3 [2015_01_04_nex_wt_ctr_767]


title: glutamatergic CB1-KO - open field - rep 5 [2015_01_22_nex_ko_OF_885]

value: open field exposure


title: H2Aub Morula Eed MatKO Rep1

city: Doha

first: Mohamed Nadhir

last: Djekidel

state: Doha

address: Gharafa

department: Translational Medicine Department

country: Qatar

institute: Sidra Medicine

description:
H2Aub_Morula_EedKO_Mat.bw
H2Aub_Morula_EedKO_Pat.bw

data_processing:
Low quality reads and adaptor sequences of the CUT&RUN data were trimmed using Trimgalore (v0.4.5) with the parameter “--paired”. The trimmed reads were then aligned to mm10 genome using Bowtie2 (v2.3.4.3)  with parameters “‘--no-unal --no-mixed --no-discordant -I 10 -X 700”. To minimize mapping bias toward reference genome (B6) for the F1 hybrid data, SNPs between PWK and B6/129 or B6/DBA were masked as “N”, depending the maternal genetic background. Unmapped and low quality reads were removed using sambamba (v0.5.9) with parameters “-f bam -F ‘mapping_quality >= 20 and not (unmapped or mate_is_unmapped)’”. Only uniquely mapped reads were kept for downstream analyses. PCR duplicates were removed using the “MarkDuplicates” command from Picard tools (v2.8.0) (https://broadinstitute.github.io/picard/). SNPsplit (v0.3.2) was used to determine the parental origin of the CUT&RUN read pairs. The bigwig files were generated using the “bamCoverage” command from deepTools (v3.0.2) with parameters “--binSize 25 --extendReads 150 --normalizeUsing RPKM --outFileFormat bigwig --scaleFactor 1”. Reproducibility were assessed by calculating Pearson correlation coefficient of RPKM in 5-kb bins between replicates. The biological replicates were pooled using the ‘merge’ command from sambamba (v0.5.9). CUT&RUN reads within the mm10 black list regions (https://github.com/Boyle-Lab/Blacklist/tree/master/lists) were not used for subsequent analyses.
Genome_build: mm10 with SNPs masked as "N"
Supplementary_files_format_and_content: Bigwig files that can be uploaded on a genome browser to view H3K27me3 and H2Aub at different genomic locations.

source_name: Morula embryo

extract_protocol:
As all libraries generated in this study involved low-input techniques,  samples collected were directly put into lysis buffer for further library construction.
H3K27me3 and H2Aub CUT&RUN libraries were constructed as previously described (Skene and Henikoff, 2017 eLife. 2017; 6: e21856, PMID: 28079019).
CUT&RUN

tag: background

value: B6/129 mixed x PWK

tag: manipulation

value: Gdf9-Eed (fl/fl) x WT

tag: embryos/oocytes

value: 22 Morula


title: H3K27me3 GV WT Rep1

source_name: germinal vesicle oocyte

value: WT

value: 100 GV oocytes


title: H2Aub Morula Eed MatKO Rep2

value: 20 Morula


title: glutamatergic CB1-WT - basal condition - rep 1 [2015_01_02_nex_wt_ctr_755]

value: basal condition (control)


title: H2Aub Morula WT Rep1

description:
H2Aub_Morula_WT.bw
H2Aub_Morula_Mat.merged.bw
H2Aub_Morula_Pat.merged.bw

value: Eed (fl/fl) x WT

value: 27 Morula


title: H2Aub Morula WT Rep2

value: 26 Morula


title: glutamatergic CB1-KO - basal condition - rep 1 [2015_01_01_nex_ko_ctr_799]

value: nex-cre x CB1f/f


title: H2Aub 2cell WT Rep2

description:
H2Aub_2C_WT.bw
H2Aub_2Cell_Mat.merged.bw
H2Aub_2Cell_Pat.merged.bw

source_name: 2-cell embryo

value: B6/DBA x PWK

value: WT x WT

value: 90  2-cell


title: H2Aub 2cell WT Rep1


title: H2Aub GV Eed MatKO Rep2

value: B6/129 mixed

value: Gdf9-Eed (fl/fl)

value: 80 GV oocytes


title: H2Aub GV WT

value: Eed (fl/fl)

value: 70 GV oocytes


title: H2Aub GV Eed MatKO Rep1

value: 50 GV oocytes


title: glutamatergic CB1-WT - basal condition - rep 2 [2015_01_03_nex_wt_ctr_1153]

value: 8 weeks

value: male

value: Brain - hippocampal Cornus Ammonis

value: nex-WT-ctr


title: H2Aub 4cell WT Rep1

description:
H2Aub_4C_WT.bw
H2Aub_4Cell_Mat.merged.bw
H2Aub_4Cell_Pat.merged.bw

source_name: 4-cell embryo

value: B6/129 mixed x PWK

value: Eed (fl/fl) x WT

value: 80 4-cell


title: HiC_WH7_d7-10_dTAG13

first: Andrea

middle: June

last: Kriz

address: 185 Cambridge St

department: Molecular Biology

institute: Massachusetts General Hospital, Harvard Medical School

data_processing:
ChIP-seq and Calibrated ChIP-seq: For uncalibrated ChIP-seq, input and ChIP-seq reads were aligned to the 129S1/SvJm (mus) and CAST/Eih (cas) genome as previously described (Kung et al., 2015; Minajigi et al., 2015). PCR duplicates were removed and uniquely mapped reads used for further analysis. Fpm-normalized bigwig tracks were generated for all uniquely mapped (comp), mus-specific (mus), and cas-specific (cas) reads. Heatmaps and metaplots were generated using deepTools functions computeMatrix and plotHeatmap with the reference point as the summit of the called ChIP peaks.Calibrated ChIP-seq analysis was performed as previously described (Hu et al., 2015). Briefly, paired-end reads were aligned to a concatenated mouse and human genome (mm9 + hg19) using Bowtie2 with ?--no-mixed? and ?--no-discordant? options. PCR duplicates were filtered out with SAMBAMBA (Tarasov et al., 2015). Uniquely aligned mouse reads were normalized to human spike-in by subsampling mm9 reads so that total number of hg19 reads was matched between samples and bigWig tracks generated from normalized samples. For calibration of RAD21 cChIP between RAD21-FKBPdegron clones, only reads within hg19 peaks (hg19 tracks combined) were counted for normalization purposes. Subsampling factors were further adjusted based on ratio of hg19 to mm9 reads in matched input samples. After normalization factor was determined, paired end reads were aligned to mus and cas genomes as previously described, except using Bowtie2 with the same parameters as above (with ?--no-mixed? and ?--no-discordant?) instead of Novoalign. PCR duplicates were filtered out with SAMBAMBA and allelic reads normalized using the same normalization factor as above.Input subtracted H3K27me3 calibrated ChIP coverage profiles were generated with SPP (Kharchenko et al., 2008) with smoothing using 1kb windows recorded every 500 bp, as previously described (Simon et al., 2013). For input subtraction, inputs from DMSO and dTAG-13 treated samples were combined to ensure equal profiles were subtracted from each calibrated IP.Heatmaps and metaplots were generated using deepTools functions computeMatrix and plotHeatmap with the reference point as the summit of the called ChIP peaks. Plots of calibrated H3K27me3 signal over upregulated or downregulated genes were calculated using ?scale-regions? with the entire gene scaled to 3kb. To compare the allelic densities of H3K27me3 over different classes of X-linked genes, we computed average densities over gene bodies using multiBigwigSummary from deepTools with default parameters. Non-expressed genes were defined as X-linked genes have 0 RPKM in at least one RNA-seq replicate of DMSO control. Genes subject to XCI were identified as the set of genes have allelic information (see RNA-seq analysis above), with escapees, Xist and Tsix excluded. Escapees were as defined previously (Wang et al., 2018). To compare allelic densities of H3K27me3 over S1 and S2 compartments, we defined S1 compartments as regions having positive value in the first Principal Component calculated in the WAPL-FKBPdegron DMSO control days 4-7 sample and S2 compartments as regions have negative value. S1 and S2 compartments were then binned into 100kb windows using bedtools makewindows, excluding windows that overlapped with unmappable regions as defined previously (Wang et al., 2018). We then we computed average densities over these 100kb bins using multiBigwigSummary from deepTools with default parameters.
RNA-seq: RNA-seq reads were aligned allele-specifically to the 129S1/SvJm (mus) and CAST/Eih (cas) genomes using TopHat2 (Kim et al., 2013) as previously described (Kung et al., 2015; Minajigi et al., 2015). PCR duplicates were removed and unique reads mapping to exons of each gene quantified by Homer (Heinz et al., 2010). DESeq 2 was used to call differentially expressed genes with a cut off of adjusted p-value < 0.1. An expression threshold of FKPM > 0.5 was also applied. Expressed genes with a minimum of 13 allelic-specific reads in at least one sample were counted as genes with allelic information (Colognori et al., 2019). For a given gene, percent mus was calculate as the percentage of allelic reads that were mus-specific (mus / (mus+cas) reads).
in situ Hi-C: Reads were trimmed using cutadapt with the options --adapter=GATCGATC (MboI ligation junction) and --minimum-length=20. Reads of each pair were individually mapped to the mus (Xi) and cas (Xa) reference genomes using novoalign and merged into Hi-C summary files and filtered using HOMER as previously described (Minajigi et al., 2015). Hi-C contact maps were generated from reads in the filtered HOMER mus and cas tag directories using the ?pre? command of Juicer tools with the genomeID mm9 (Durand et al., 2016b). The resulting Hi-C contact maps in .hic format were visualized and normalized with the ?Coverage Square Root? option in Juicebox version 1.8.8 (Durand et al., 2016a). Filtered reads from separate RAD21-FKBPdegron clones were combined before generating Hi-C contact maps.
CHART-seq: Input and ChIP-seq reads were aligned to the 129S1/SvJm (mus) and CAST/Eih (cas) genome as previously described (Kung et al., 2015). PCR duplicates were removed and uniquely mapped reads used for further analysis. Fpm-normalized bigwig tracks were generated for all uniquely mapped (comp), mus-specific (mus), and cas-specific (cas) reads. CHART libraries were sampled to equal read depth and input subtracted CHART coverage profiles were generated with SPP (Kharchenko et al., 2008) with smoothing using 1kb windows recorded every 500 bp, as previously described (Simon et al., 2013). After scaling, we computed base 2 logarithm of the ratio of the coverage of DMSO control divided by that of dTAG-13 using bigWigCompare from deepTools with --binSize 1000 and default parameters otherwise.
PRO-seq: Paired-end reads were aligned to a concatenated mouse and fly genome (mm9 + dm6) using Bowtie2 with ?--no-mixed? and ?--no-discordant? options. DMSO and dTAG-13 samples for each clone were normalized to drosophila spike-in by subsampling mm9 reads so that total number of dm6 reads was matched between samples. For RAD21-FKBPdegron clone G9, downsampling factors corresponded to library size (i.e., were equivalent to downsampling based on total number of reads). For RAD21-FKBPdegron clone C4, downsampling factors were 1 for DMSO and 0.73 for dTAG-13 although library sizes were similar, indicating that there were overall lower levels of nascent transcription in the dTAG-13 sample. Duplicates were not removed to preserve reads corresponding to TSS and promoter proximal PolII positions. Reads mapping to genes (full gene bodies) were quantified by Homer (Heinz et al., 2010), removing miRNA, snoRNA and scaRNAs. DESeq 2 was used to call differentially expressed genes with a cut off of adjusted p-value < 0.1. For analyses on all genes, no FPKM threshold was applied, but genes were required to have at least 1 count in one sample. An expression threshold of FKPM > 0.5 was applied for analyses on expressed genes. Log2FoldChange and log2baseMean from DESeq2 were plotted for indicated classes of genes. H3K27me3 genes were defined as genes overlapping H3K27me3 cChIP-seq peaks in their respective samples (DMSO and dTAG-13 peaks combined). H3K27me3 loop genes were defined as H3K27me3 genes overlapping H3K27me3-bound loop anchors called in RAD21-FKBPdegron dTAG-13 Hi-C maps (i.e., cohesin-independent H3K27me3 anchored loops). For overlap with RAD21 loop anchors, loops were called in respective DMSO Hi-C maps.For allelic analysis, PRO-seq reads not mapping to the fly genome (dm6) were aligned to mus and cas genomes using Bowtie2 with the same parameters as above (?--no-mixed? and ?--no-discordant?) and downsampled using the same factors as calculated above. Expressed genes with a minimum of 13 allelic-specific reads in at least one sample were counted as genes with allelic information (Colognori et al., 2019). For CDPs, normalized mus and cas counts divided by gene length were plotted.
Genome_build: mm9
Supplementary_files_format_and_content: ChIP-seq: bigWig (bw) files for all reads (.bw), regardless of whether they are allele-specific or not, cas-specific (.cas.bw) and mus-specific (.mus.bw). All bigWig files are fpm normalized and H3K27me3 bigWig files are additionally input normalized.
Supplementary_files_format_and_content: Calibrated ChIP-seq: bigWig (bw) files for all reads (.bw), regardless of whether they are allele-specific or not, cas-specific (.cas.bw) and mus-specific (.mus.bw). All bigWig files are normalized to spike-in HEK293 control and H3K27me3 bigWig files are additionally input normalized.
Supplementary_files_format_and_content: RNA-seq: Strand specific (cwig, Crick strand, wwig, Watson strand) bigWig (bw) files for all reads (.bw), regardless of whether they are allele-specific or not, cas-specific (.cas.bw) and mus-specific (.mus.bw). All bw files are fpm normalized. Raw exonic reads in each RNA-seq library were quantified and summarized in .txt files.
Supplementary_files_format_and_content: PRO-seq: Strand specific (cwig, Crick strand, wwig, Watson strand) bigWig (bw) files for all reads (.bw), regardless of whether they are allele-specific or not, cas-specific (.cas.bw) and mus-specific (.mus.bw). All bw files are downsampled according to Drosophila spike-in. Gene body counts in each PRO-seq library were quantified and summarized in .txt files.
Supplementary_files_format_and_content: In situ Hi-C: Compressed Hi-C matrix (.hic) files for all contacts genome-wide (.hic), regardless of whether they are allele-specific or not, cas-specific chrX (chrX.cas.his) contacts and mus-specific chrX (chrX.mus.hic) contacts. File can be loaded onto Juicebox to view either raw or normalized Hi-C interaction matrices at 5kb to 2.5Mb resolutions.

source_name: Hi-C, clone H7

growth_protocol: Undifferentiated mESCs were grown on γ-irradiated MEF feeders in medium containing DMEM, high glucose, GlutaMAX supplement, pyruvate, 15% Hyclone FBS (Sigma), 25 mM HEPES pH 7.2-7.5, 1x MEM non-essential amino acids, 1x Pen/Strep, 0.1 mM βME, and 500 U/mL ESGRO recombinant mouse Leukemia Inhibitory Factor (LIF) protein (Sigma, ESG1107) at 37°C with 5% CO2. To induce differentiation in EBs, mESCs were trypsinized and resuspended in the same media without LIF. For the first 3 days of differentiation, EBs were grown in suspension. On day 4 and subsequent days, EBs were grown on gelatinized plates until harvested at day 7 or day 10.

extract_protocol:
ChIP-seq: ChIP-seq was performed as described previously with minor modifications (Del Rosario et al., 2019). Briefly, cells were trypsinized, resuspended in 1 million cells/mL of media and crosslinked with 1% formaldehyde for 10 min at room temperature with rotation. Cells were then quenched with 0.2 M glycine for 5 min. Crosslinked cells were washed once with cold PBS, pelleted, and snap frozen in an EtOH/dry ice bath. Cell pellets were stored at -80C until further processing. 5-10 million cross-linked cells per IP were thawed on ice and resuspended in 300ul Lysis Buffer 1 (25mM HEPES pH 7.2-7.5, 25mM EDTA, 0.5% SDS). Nuclei were sonicated (Qsonica Q800 Sonicator) in polystyrene tubes at 40% power setting, 30s ON/30s OFF for a total sonication (ON) time of 6 min. After sonication, 1.2 mL of Lysis Buffer 2 (25mM HEPES pH 7.2-7.5, 187.5mM NaCl, 1.25% Triton X-100, 0.625% Na-deoxycholate, 6.25% glycerol, 1x cOmplete EDTA-free protease inhibitor cocktail) was added. 30ul of Dynabeads Protein G (Thermo Fisher Scientific) per IP were washed 3 times with ChIP Wash Buffer 1 (25mM HEPES pH 7.2-7.5, 150mM NaCl, 5mM EDTA, 1% Triton X-100, 0.5% Na-deoxycholate, 5% glycerol, 1x cOmplete EDTA-free protease inhibitor cocktail) and resuspended in 60ul ChIP Wash Buffer 1. Each IP was blocked with 20ul washed beads for 2 hours at 4C with rotation. The other 40ul of washed beads were incubated with antibody (5ul of RAD21 antibody or 2ul of H3K27me3 antibody/IP) for 2 hours at 4C. 5% input was taken from blocked lysates. Beads conjugated to antibody were added to blocked lysates and IP allowed to proceed overnight at 4C with rotation. IPs were then washed 3 times with cold ChIP Wash Buffer 1, 3 times with cold ChIP Wash Buffer 2 (25mM HEPES pH 7.2-7.5, 300mM NaCl, 5mM EDTA, 1% Triton X-100, 0.5% Na-deoxycholate, 5% glycerol, 1x cOmplete EDTA-free protease inhibitor cocktail). And 1 time with cold TENG (50mM Tris-HCl pH 8, 1mM EDTA, 50mM NaCl, 5% glycerol, 1x cOmplete EDTA-free protease inhibitor cocktail). All washes were performed by inverting tubes and setting aside for ~2 minutes per wash. IPs were then resuspended in 500ul TENG with 100ug RNAse A (Thermo Fisher Scientific) and incubated for 30 min at 37C. IPs were then washed 2 times with cold ChIP Wash Buffer 2 and 2 times with cold TENG without protease inhibitors. IPs were resuspended in 200ul elution buffer (50mM Tris-HCl pH 7.5, 10mM EDTA, 1% SDS) and incubated 15 min at 65C. Eluate was then transferred to a new tube and 198ul TE buffer (Ambion) and 40ug Proteinase K (Sigma) added. Inputs were thawed, TE buffer added to equalize volume to IPs and 200ul elution buffer and 40ug Proteinase K added. Inputs and IPs were then incubated for 2h at 45C and crosslinks reversed overnight at 65C. DNA was purified with phenol-chloroform extraction and resuspended in 50ul TE. Efficiency of ChIP was assayed via qPCR, with samples displaying over 10-fold enrichment of positive regions over negative control prepared for sequencing. Libraries were prepared from ~500ng of input or IP DNA using the NEBNext® Ultra? II DNA Library Prep Kit for Illumina® (NEB) according to kit instructions. Experiments were performed in two biological replicates. Input and ChIP-seq libraries were sequenced on Illumina HiSeq, generating 20-50 million paired end 50 or 150 nucleotide reads per sample.
Calibrated ChIP-seq: Calibrated ChIP-seq was performed using the same protocol as ChIP-seq except that 50,000 HEK293 cells per 5 million mouse cells were mixed in at the initial lysis step. Cells were counted and equal numbers of cells used per IP.
in situ Hi-C: As described in Rao et al. (2014), 3-5 million cells were crosslinked with 1% formaldehyde for 10min. Crosslinked cell pellets were lysed with 300uL ice-cold HiC lysis buffer (10mM Tris HCl pH 8, 10mM NaCl, 0.2% Igepal CA630) with protease inhibitors (Roche). Cell suspension was incubated for 30min at 4C with rotation, then pelleted at 2500xg for 5min. Pelleted nuclei were then washed with 500ul ice-cold HiC lysis buffer and resuspended in 50uL 0.5% SDS solution and incubated at 62C for 10min. 145ul H2O and 25ul 10% Triton X-100 were added to quench SDS and nuclei incubated at 37C for 15min. 26.1ul of 10X NEB Buffer 2 and 100U of MboI restriction enzyme were added and nuclei digested overnight at 37C with rotation. MboI was then heat activated at 62C for 20min, then nuclei cooled to room temperature. To fill in MboI fragment overhangs and mark DNA ends with biotin, 50ul of fill-in master mix was added (37.5ul 0.4mM biotin-14-dATP, 1.5ul 10mM dCTP, 1.5ul 10mM dGTP, 1.5ul 10mM dTTP, 8ul 5U/ul DNA Polymerase I, Large (Klenow) Fragment) and mixed by pipetting. Nuclei were then incubated at 37C for 1h with rotation. To ligate biotinylated DNA ends together, 900ul ligation master mix was added (649ul H2O, 120ul 10X NEB T4 ligase buffer, 100ul 10% Triton X-100, 6ul of 20mg/ml BSA, 25ul of 400U/ul T4 DNA ligase). Reaction was mixed by inverting and incubated at room temperature for 4h with rotation. Reaction was then centrifuged at 1700xg for 5min at room temperature. Pelleted nuclei were resuspended in 750ul TES buffer (1mM EDTA, 10mM Tris-HCl pH 8, 0.5% SDS), 50ul Proteinase K (20mg/ml) added and incubated at 55C for 1h with rotation. To reverse crosslinks, 88.9ul 3M NaCl was added and crosslinks reversed by incubated overnight at 65C with shaking (Eppendorf thermomixer, 950rpm). Samples were cooled to room temperature and DNA extracted with phase-lock gel tubes (5Prime). RNA was digested from purifying DNA by adding 2ul RNAse A (20mg/ml) and incubating at 37C for 1h. DNA was then extracted with phase-lock gel tubes (5Prime) and resuspended in 130ul 1X Tris buffer (10mM Tris-HCl, pH 8). To ensure complete resuspension, DNA was then incubated at 37C for 15min. DNA was then sonicated to a size of 300-500bp using a Covaris with the following parameters: 140W, 10% duty cycle, 200 bursts/cycle, 100s. Samples were then size selected to 300-500bp using AMPure XP beads and eluted in 300ul 1X Tris buffer. To pulldown biotinylated ligation products, 30ul (per sample) of MyOne Streptavidin C1 beads were washed with 400ul of 1X Tween washing buffer (5mM Tris-HCl pH 7.5, 0.5mM EDTA, 1M NaCl, 0.05% Tween 20) and resuspended in 300ul 2X binding buffer (10mM Tris-HCl pH 7.5, 1mM EDTA, 2M NaCl). Beads were then added to purified DNA and sample incubated at room temperature for 15min with rotation. Beads were washed twice with 600ul 1X Tween Washing buffer. Subsequent library preparation steps were then done directly on beads.
CHART-seq: Xist CHART-seq was performed as previously described (Wang et al., 2018). Briefly, cells were trypsinized, resuspended in 1 million cells/mL of media and crosslinked with 1% formaldehyde for 10 min at room temperature with rotation. Cells were then quenched with 0.2 M glycine for 5 min. Crosslinked cells were washed once with cold PBS, pelleted, and snap frozen in an EtOH/dry ice bath. Cell pellets were stored at -80C until further processing. 25 million cross-linked cells per CHART were thawed on ice and resuspended in 1 mL cold sucrose buffer. (10 mM HEPES pH 7.5, 0.3 M sucrose, 1% Triton X-100, 100 mM potassium acetate, 0.1 mM EGTA) supplemented with 0.5 mM spermidine, 0.15 mM spermine, 1 mM DTT, 1x protease inhibitor cocktail (Sigma, P8340), 10 U/mL RNase inhibitor (Roche). The cell suspension was rotated for 10 min at 4°C, after which it was diluted with additional 2 mL cold sucrose buffer. This was added to a pre-chilled 15-mL glass Wheaton dounce tissue grinder (Thermo Fisher Scientific) that had been pre-rinsed once with RNaseZAP RNase decontamination solution (Thermo Fisher Scientific), three times with DEPC-treated water, and once with 2 mL sucrose buffer. Release of nuclei was accomplished by douncing cells 20 times with a tight pestle, and verified by staining with Trypan Blue Solution (Thermo Fisher Scientific). The nuclear suspension was then layered atop a cushion of 7.5 mL cold glycerol buffer (10 mM HEPES pH 7.5, 25% glycerol, 1 mM EDTA, 0.1 mM EGTA, 100 mM potassium acetate) supplemented with 0.5 mM spermidine, 0.15 mM spermine, 1 mM DTT, 1x cOmplete EDTA-free protease inhibitor cocktail (Roche), and 5 U/mL RNase inhibitor, and centrifuged at 1500 g for 10 min at 4°C. The purified nuclear pellet was resuspended in 6 mL PBS and further cross-linked with 3% formaldehyde for 30 min at room temp with rotation. Afterward, nuclei were pelleted by centrifugation at 1,000 g for 5 min at 4°C and washed three times with cold PBS. Nuclei were resuspended in in 1 mL cold nuclear extraction buffer (50 mM HEPES pH 7.5, 250 mM NaCl, 0.1 mM EGTA, 0.5% sarkosyl, 0.1% sodium deoxycholate, 5 mM DTT, 10 U/mL RNase inhibitor) and incubated for 10 min at 4°C with rotation. Nuclei were then centrifuged at 400 g for 5 min at 4°C and resuspended in 230 uL cold sonication buffer (50 mM HEPES pH 7.5, 75 mM NaCl, 0.1 mM EGTA, 0.5% sarkosyl, 0.1% sodium deoxycholate, 0.1% SDS, 5 mM DTT, 10 U/mL RNase inhibitor) to a final volume of ?270 uL. Nuclei were sonicated for 5 min at 4°C using a Covaris E220e (140W peak incident power, 10% duty factor, 200 cycles/burst). Sonicated chromatin was centrifuged at 16,000 g for 20 min at 4°C to remove debris. The supernatant (?220 uL) was mixed with 110 uL sonication buffer to a final volume of ?330 uL, which was split into two CHART reactions (Xist capture using antisense probes and control capture using sense probes). For two CHART reactions, 600 uL MyOne Streptavidin C1 beads (Thermo Fisher Scientific) were used, which were pre-washed with 1 mL DEPC-treated water, blocked with 600 uL blocking buffer (33% sonication buffer, 67% 2x hybridization buffer, 500 ng/uL yeast RNA [Thermo Fisher Scientific], 1% BSA) at room temp for 1 hr, washed with 1 mL DEPC-treated water again, and resuspended in 610 uL 1x hybridization buffer (33% sonication buffer, 67% 2x hybridization buffer [see below for composition]). For each CHART, 160 uL chromatin was mixed with 320 iL 2x hybridization buffer (50 mM Tris pH 7.0, 750 mM NaCl, 1% SDS, 1 mM EDTA, 15% formamide, 1 mM DTT, 1 mM PMSF, 1x cOmplete EDTA-free protease inhibitor cocktail, 100 U/mL RNase inhibitor) and pre-cleared with 60 uL pre-blocked MyOne Streptavidin C1 beads at room temp for 1 hr with rotation. Pre-cleared chromatin was separated from beads, with 1% saved as ?input? sample. The remaining pre-cleared chromatin (?500 uL) was mixed with 36 pmol of either antisense (Xist-targeting) or sense (control) biotinylated capture probes. We used a pool of 9 capture probes, which were selected from the 11 probes described previously (Simon et al., 2013), excluding probes X.3 and X.8. Hybridization was carried out at 37°C for 4 hr followed by incubation with 240 uL blocked MyOne Streptavidin C1 beads at 37°C for 1 hr. The beads were washed once with 1x hybridization buffer (33% sonication buffer, 67% 2x hybridization buffer) at 37°C for 10 min, five times with wash buffer (10 mM HEPES pH 7.5, 150 mM NaCl, 2% SDS, 2 mM EDTA, 2 mM EGTA, 1 mM DTT) at 37°C for 5 min, and twice with elution buffer (10 mM HEPES pH 7.5, 150 mM NaCl, 0.5% NP-40, 3 mM MgCl2, 10 mM DTT) at 37°C for 5 min. 10% of the final wash was saved as an ?RNA capture? sample to estimate the efficiency of Xist RNA capture afterward by qRT-PCR. CHART-enriched DNA was eluted twice in 200 uL elution buffer supplemented with 5 U/uL RNase H (New England BioLabs) at 37°C for 30 min. An ?input? sample and CHART DNA were treated with 0.5 mg/mL RNase A (Thermo Fisher Scientific) at 37°C for 1 hr and then with 1% SDS, 10 mM EDTA, and 0.5 mg/mL proteinase K (Sigma) at 55°C for 1 hr. Reversal of crosslinks was performed by addition of 150 mM NaCl (final 300 mM) and incubation at 65°C overnight. DNA was purified by phenol-chloroform extraction and further sheared to < 500-bp fragments using Covaris E220e (140W peak incident power, 10% duty factor, 200 cycles/burst). Sonicated DNA was purified using 1.8x Agencourt AMPure XP beads (Beckman Coulter). Libraries were sequenced on Illumina, generating ?50 million 150-nt paired-end reads per sample
RNA-seq: Total RNA was extracted using TRIzol Reagent (Thermo Fisher Scientific) and mRNA isolated using the NEBNext Poly(A) Magnetic Isolation Module (NEB) according to the kit instructions. Libraries were sequenced on the Illumina HiSeq2500 (rapid run), generated 30-50 million 50 nt paired end reads per sample.
PRO-seq: PRO-Seq library construction was performed by the Nascent Transcriptomics Core at Harvard Medical School, Boston, MA. Aliquots of frozen (-80°C) permeabilized cells were thawed on ice and pipetted gently to fully resuspend. An aliquot was removed and diluted 1:10 in 1xPBS for cell counting using an automatic counter (Biorad TC-20) with counts spot checked by manual recount using a hemocytometer as needed. An aliquot of each dilution was also stained with Trypan Blue and counting repeated to determine permeabilization efficiency. For each sample, 1 million permeabilized cells were used for nuclear run-on. For normalization, 50,000 permeabilized Drosophila S2 cells were added to each sample of 1 million cells. Nuclear run on assays and library preparation were then performed essentially as described [Elrod, N.D., et al. (2019) Molecular Cell 76: 738-752.e7] with modifications noted below. The nuclear run-on buffer was prepared as 4X stock (20mM Tris (pH 8), 20mM MgCl2, 2mM DTT, 600mM KCl, 40uM/ea biotin-11-NTPs (Perkin Elmer), 20U SuperaseIN (Thermo)). The 4X stock was mixed 1:1 with 2% sarkosyl (Sigma) to yield 2X complete run-on mix. The 3' adapter (RNA oligo: 5'P-GAUCGUCGGACUGUAGAACUCUGAAC-3'InvdT) was pre-adenylated prior to use (5' DNA adenylation kit, NEB, according to the manufacturer's instructions). Adenylated oligo was purified by ethanol precipitation, resuspended in water and quality was modification confirmed by electrophoresis on a 15% TBE-Urea gel (Novex) next to appropriate controls. The adapter concentration was adjusted to 10uM and 1uL was used for each ligation which used T4 RNA ligase 2, truncated KQ (NEB) with 15% PEG-8000 in the reaction mix. Reactions were incubated at 16°C overnight. Each ligation reaction was then diluted with 180uL of betaine binding buffer (1.42g of betaine brought to 10mL with binding buffer and sterile filtered) containing 1uL of 100uM blocking oligo (TCCGACGATCCCACGTTCCCGTGG/3InvdT/). The blocking oligo is complementary to and overhangs the 3' end of the 3' adapter and reduces 5'/3' adapter dimer formation during the 5' adapter ligation step. The blocking oligo has modifications at its 3? end, to prevent extension by reverse transcriptase. Blocking oligo was also included in final wash solution prior to the 5' adapter ligation and in the ligation reaction itself, both at 3-fold molar excess over the initial amount of 3' adapter (i.e. 1uL of 30uM per reaction). The 5' adapter ligation was performed as in Elrod except PEG-8000 was increased to 15%. Libraries were prepared using the NEBNext® Ultra? II DNA Library Prep Kit for Illumina® (NEB) according to kit instructions. PRO-seq: After reverse transcription, cDNA was immediately amplified for 5-cycles (preCR step). A PCR cocktail consisting of NEBNext Ultra II Q5 master mix (NEB) and Illumina TruSeq PCR primers (RP-1, common; and RPI-X, indexing) was added to each cDNA and amplification was done according to manufacturer's suggested 2-step cycling conditions for NGS applications. To determine optimal library amplification, the preCRs were thawed on ice and an aliquot serially diluted. Each dilution was mixed with a PCR cocktail (Q5 DNA polymerase with optional high GC enhancer) containing 10uM/ea of Illumina universal P5 and P7 primers and amplified 15-cycles. The reactions were electrophoresed on a 2.2% agarose gel (SeaKem) cast with SYBR gold (Thermo) and analyzed to determine the optimal number of cycles for final amplification. preCRs were transferred from ice to a pre-heated thermal cycler block and amplified for the appropriate number of additional cycles to reach the total determined in the test amplification.

treatment_protocol: Degradation of RAD21 or WAPL in FKBPdegron lines during differentiation was induced by adding a 1:20,000 dilution of 10mM dTAG-13 in DMSO to cell culture media (500nM final concentration) for the indicated time frames. The small molecule dTAG-13 was synthesized as previously described (Erb et al., 2017; Nabet et al., 2018). For DMSO vehicle controls, the same volume of DMSO alone was added to cell culture media.

value: CAST/Ei x 129/Sv/Jae

tag: genotype/variation

value: WAPL-FKBP(degron)

value: ES cell (Tsix-stop)

value: 3 days dTAG-13 (500nM)

tag: differentiation day

value: day 10


title: HiC_WH7_d7-10_DMSO

value: 3 days DMSO


title: HiC_RG9_d108h_dTAG13

description:
processed data file: RAD21_13.all.hic
processed data file: RAD21_13.chrX.cas.hic
processed data file: RAD21_13.chrX.mus.hic

source_name: Hi-C, clone G9


title: HiC_RG9_d108h_DMSO

description:
processed data file: RAD21_D.all.hic
processed data file: RAD21_D.chrX.cas.hic
processed data file: RAD21_D.chrX.mus.hic

value: 8h DMSO


title: HiC_RC4_d108h_dTAG13

source_name: Hi-C, clone C4


title: ChIP_TST_d8_H3K27me3

source_name: H3K27me3 ChIP, parental

value: Wild-type

value: day 8

tag: chip antibody

value: H3K27me3 (GeneTex, GTX60892)


title: ChIP_TST_d8_H2AK119Ub

source_name: H2AK119Ub ChIP, parental

value: H2AK119ub (Cell Signaling, CST8240)


title: ChIP_input_TST

source_name: ChIP input, parental


title: HiC_RC4_d108h_DMSO


title: RNA_WH7_d7-10_DMSO.rep1

description: processed data file: RNA_WH7_d7-10_rep1.rep2.counts.txt

source_name: RNAseq, clone H7, rep1


title: RNA_WH7_d7-10_dTAG13.rep2

source_name: RNAseq, clone H7, rep2


title: RNA_WH7_d7-10_DMSO.rep2


title: ChIP_input_RC4_d102h

source_name: ChIP input, clone C4

value: 2h DMSO


title: Liver lysate 4

city: Quebec

first: Luca

last: Pellegrini

address: 1050 Rue de la Médecine

department: Department of Molecular Biology

institute: Université Laval

data_processing:
Trimming: fastp v0.12.4 with --length_required 30, -qualified_quality_phred 20, --correction, --cut_by_quality3, --cut_window_size 10 and --cut_mean_quality 30
Pseudoalignment: kallisto quant v0.44.0 with default parameters values.
Differential expression: DESeq2 v1.20.0 with R v3.5.0 and Bioc v3.7.
Genome_build: Ensembl release 94
Supplementary_files_format_and_content: Excel files containing raw counts, tpm counts and supplementary table.

source_name: Lysate

extract_protocol:
1 g of liver was homogenized with a modified teflon pestle. The liver lysate was obtained by removing from this homogenate all the tissue debris and cell nuclei through 3 rounds of low speed centrifugation. WAM fractions were prepared from this lysate usi
NA was isolated from mouse liver homogenates using the Dynabeads mRNA Direct Micropurification Kit (Thermo Fisher Scientific); of this, 250 ng were then used to prepare sequencing libraries using the NEBNext Ultra II directional RNA library prep kit for I

value: Liver

tag: fraction

value: liver postnuclear lysate

tag: age

value: 9-12 weeks old

value: male

tag: feeding state

value: 3 hours postprandial, after overnight fasting


title: WAM fraction 4

source_name: WrappER-associated mitochondria (WAM) fraction

tag: strain

tag: tissue

value: wrappER-associated mitochondria (WAM) fraction